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Journal Cover Journal of Cellular Physiology
  [SJR: 1.842]   [H-I: 139]   [6 followers]  Follow
   Hybrid Journal Hybrid journal (It can contain Open Access articles)
   ISSN (Print) 0021-9541 - ISSN (Online) 1097-4652
   Published by John Wiley and Sons Homepage  [1589 journals]
  • Generation of Insulin-Producing Cells From Human-Induced Pluripotent Stem
           Cells Using a Stepwise Differentiation Protocol Optimized With
           Platelet-Rich Plasma
    • Authors: Seyed Ehsan Enderami; Yousef Mortazavi, Masoud Soleimani, Samad Nadri, Alireza Biglari, Reyhaneh Nassiri Mansour
      Abstract: Studies on patient-specific human-induced pluripotent stem cells (hiPSCs) as well as a series of autologous growth factors presumably will reveal their many benefits for cell base replacement therapy in type 1 diabetes mellitus (TIDM) patients in the future. For this purpose, we established a multistep protocol by adding platelet-rich plasma (PRP) that induce the hiPSCs into insulin-producing cells (IPCs). We present here a differentiation protocol consisting of five stages in two groups including protocol with PRP and without PRP. Charac­teristics of derived IPCs in both groups were evaluated at the mRNA and protein levels, cell cycle and viability in the end stage of cell differentiation. The in vitro studies indicated the treatment of hiPSCs in the protocol with PRP resulting in differentiated cells with strong characteristics of IPCs including islet-like cells, the expression of mature and functional pancreatic beta cell specific marker genes. In addition to these pancreatic specific markers were detected by immunochemistry and Western blot. Our differentiated cells in two groups secreted insulin and C-peptide in a glucose stimulation test by ELISA showing in vitro functional. The results of the cell cycle assay confirmed that differentiation has been done. We reported for the first time that PRP might be ideal additive in the culture medium to induce pancreatic differentiation in the hiPSCs. This study provides a new approach to investigate the role of PRP in pancreatic differentiation protocols and enhance the feasibility of using patient-specific iPSCs and autologous PRP for future beta cells replacement therapies for T1DM. J. Cell. Physiol. 9999: 1–9, 2016. © 2016 Wiley Periodicals, Inc.We established a multistep protocol by adding platelet-rich plasma (PRP) that induce the hiPSCs into insulin-producing cells (IPCs). We reported for the first time that PRP might be ideal additive in the culture medium to induce pancreatic differentiation in the hiPSCs.
      PubDate: 2017-04-21T10:24:12.254003-05:
      DOI: 10.1002/jcp.25721
  • CRISPR/Cas9: an RNA-guided highly precise synthetic tool for plant genome
    • Authors: Yeliz Demirci; Baohong Zhang, Turgay Unver
      Abstract: CRISPR/Cas 9 is a newly developed and naturally occurred genome editing tool, which is originally used by bacteria for immune defence. In the past years, it has been quickly employed and modified to precisely edit genome sequences in both plants and animals. Compared with the well-developed zinc finger nucleases (ZFNs) and transcription activator-like effector nucleases (TALENs), CRISPR/Cas 9 has lots of advantages, including easier to design and implement, higher targeting efficiency, and less expensive. Thus, it is becoming one of the most powerful tools for knockout of an individual gene as well as insertion of one gene and/or control of gene transcription. Studies have shown that CRISPR/Cas 9 is a great tool to edit many genes in a variety of plant species, including the model plant species as well as agriculturally important crops, such as cotton, maize and rice. CRISPR/Cas 9-based genome editing can be used for plant functional studies and plant improvement to yield, quality and tolerance to environmental stress. This article is protected by copyright. All rights reserved
      PubDate: 2017-04-21T08:55:39.923648-05:
      DOI: 10.1002/jcp.25970
  • Osteoblast role in osteoarthritis pathogenesis
    • Authors: Nicola Maruotti; Addolorata Corrado, Francesco Paolo Cantatore
      Abstract: Even if osteoarthritis pathogenesis is still poorly understood, numerous evidences suggest that osteoblasts dysregulation plays a key role in osteoarthritis pathogenesis. An abnormal expression of OPG and RANKL has been described in osteoarthritis osteoblasts, which is responsible for abnormal bone remodeling and decreased mineralization. Alterations in genes expression are involved in dysregulation of osteoblast function, bone remodeling and mineralization, leading to osteoarthritis development. Moreover, osteoblasts produce numerous transcription factors, growth factors and other proteic molecules which are involved in osteoarthritis pathogenesis. This article is protected by copyright. All rights reserved
      PubDate: 2017-04-20T06:21:37.936124-05:
      DOI: 10.1002/jcp.25969
  • Genetic variants as potential predictive biomarkers in advanced colorectal
           cancer patients treated with oxaliplatin-based chemotherapy
    • Authors: Afsane Bahrami; Forouzan Amerizadeh, Seyed Mahdi Hassanian, Soodabeh ShahidSales, Majid Khazaei, Mina Maftouh, Majid Ghayour-Mobarhan, Gordon A. Ferns, Amir Avan
      Abstract: Chemotherapy regimen containing oxaliplatin is often the first-line treatment for patient with advanced colorectal cancer. Oxaliplatin binds to DNA, leading to the formation of crosslinks and bulky adducts. Approximately 50% of patients with CRC benefit from treatment with oxaliplatin. It is possible that genetic variants in biological pathways involved in drug transportation, drug metabolism, DNA damage repair, and cell cycle modulation might affect the activity, or efficacy of oxaliplatin. Because oxaliplatin resistance may be related to these genetic variants and may therefore be an important reason for treatment failure, we have summarized the genetic variations that have been reported to be predictive markers of the response to oxaliplatin based therapy in patients with advanced CRC. This article is protected by copyright. All rights reserved
      PubDate: 2017-04-19T08:24:07.710373-05:
      DOI: 10.1002/jcp.25966
  • Agarose Hydrogel Induced MCF-7 and BMG-1 Cell Line Progressive 3D and 3D
           Revert Cultures
    • Authors: Aishwarya Subramaniyan; Maddaly Ravi
      Abstract: 3D culture systems have enhanced the utility of cancer cell lines as they are considered closer to the in vivo systems. A variety of changes are induced in cells cultured in 3D systems; an apparent and striking feature being the spontaneous acquisition of distinct morphological entities. 3D reverts (3DRs) can be obtained by introducing 3D aggregates in scaffold/matrix-free culture units. It could be seen that the two cell lines used in this study exhibited differences in 3DR structures, though both were cultured on agarose hydrogels. Also, differences in 3DR formation, growth and survival were different. While 3D aggregates of several cell lines have been reported for a variety of studies, there are no studies that describe or utilize 3DRs. 3DRs can provide insights into complex events that can occur in cancer cells; especially as material to study metastasis, migration and invasion. This article is protected by copyright. All rights reserved
      PubDate: 2017-04-19T08:23:59.446-05:00
      DOI: 10.1002/jcp.25965
  • Hematopoietic Stem/Progenitor Cell Differentiation towards Myeloid Lineage
           is Modulated by LIGHT/LIGHT Receptor Signaling
    • Authors: Weikai Chen; Xin Lv, Changlong Liu, Ruoping Chen, Jianhe Liu, Haiyan Dai, Gang-Ming Zou
      Abstract: The cytokine LT-related inducible ligand that competes for glycoprotein D binding to herpesvirus entry mediator on T cells (LIGHT) is a member of the tumor necrosis factor (TNF) superfamily. It is expressed primarily on activated T lymphocytes, and detectable on monocytes, granulocytes, and immune dendritic cells. It mainly plays a role in immune regulation including T cell activation and dendritic cell maturation. We recently reported its role as an inducer in embryonic stem cell differentiation, but its role in regulation of adult stem cell has not been defined. In the present study, we examined the expression of LIGHT receptor in Lin-c-kit+Sca-1+ hematopoietic stem/progenitor cells (HSC/HPCs). We found that HSC express HVEM, a LIGHT receptor, on its surface. We further identified the role of LIGHT in promoting myeloid differentiation of HSCs driven by granulocyte-monocyte colony stimulating factor (GM-CSF). Further studies showed that LIGHT enhances both GM-CSF and GM-CSF receptor (GM-CSFR) expression in HSCs. LIGHT stimulation increases PU.1 expression in HSC/HPCs. In vivo administration of LIGHT increases the colony-forming unit-granulocyte/monocyte (CFU-GM) colony formation and plasma GM-CSF level. Altogether, the data suggest LIGHT promote myeloid differentiation of HSC/HPCs. This article is protected by copyright. All rights reserved
      PubDate: 2017-04-19T05:57:38.439815-05:
      DOI: 10.1002/jcp.25967
  • Isorhapontigenin induced cell growth inhibition and apoptosis by targeting
           EGFR-related pathways in prostate cancer
    • Authors: Cuicui Zhu; Qingyi Zhu, Zhaomeng Wu, Yingying Yin, Dan Kang, Shan Lu, Ping Liu
      Abstract: Isorhapontigenin (ISO), a naturally phytopolyphenol compound existing in Chinese herb, apples and various vegetables, has attracted extensive interest in recent years for its diverse pharmacological characteristics. Increasing evidences reveal that ISO can inhibit cancer cell growth by induced apoptosis, however the molecular mechanisms is not fully understood. In this study, we found for the first time that ISO apparently induced cell growth inhibition and apoptosis by targeting EGFR and its downstream signal pathways in prostate cancer (PCa) cells both in vitro and in vivo, whereas no obviously effect on normal prostate cells. From the results, we found that ISO competitively targeted EGFR with EGF and inhibited EGFR auto-phosphorylation, and then decreased the levels of p-Erk1/2, p-PI3K and p-AKT, and further induced down-regulation of p-FOXO1 and promoted FOXO1 nuclear translocation; and finally resulted in a significantly up-regulation of Bim/p21/27/Bax/cleaved Caspase-3/cleaved PARP-1 and a markedly down-regulation of Sp1/Bcl-2/XIAP/Cyclin D1. Moreover, our experimental data demonstrated that treatment of ISO decreased protein level of AR via both inhibiting the expression of AR gene and promoting the ubiquitination/degradation of AR proteins in proteasome. In vivo, we also found that ISO inhibited the growth of subcutaneous xenotransplanted tumor in nude mice by inducing PCa cell growth inhibition and apoptosis. Taken together, all findings here clearly implicated that EGFR-related signal pathways, including EGFR-PI3K-Akt and EGFR-Erk1/2 pathways, were involved in ISO-induced cell growth inhibition and apoptosis in PCa cells, providing a more solid theoretical basis for the application of ISO to treat patients with prostate cancer in clinic. This article is protected by copyright. All rights reserved
      PubDate: 2017-04-19T05:54:27.290005-05:
      DOI: 10.1002/jcp.25968
  • Establishment of a new conditionally immortalized human skeletal muscle
           microvascular endothelial cell line
    • Authors: Hironori Sano; Yasuteru Sano, Eri Ishiguchi, Fumitaka Shimizu, Masatoshi Omoto, Toshihiko Maeda, Hideaki Nishihara, Yukio Takeshita, Shiori Takahashi, Mariko Oishi, Takashi Kanda
      Abstract: In skeletal muscle, the capillaries have tight junctions (TJs) that are structurally similar to those in the blood-brain barrier (BBB) and blood-nerve barrier (BNB). Although many findings have been clarified in the territory of BBB and BNB, few have so far examined the TJs of capillaries in the skeletal muscle. In addition, no in vitro human skeletal muscle microvasculature models have been reported thus far. We newly established a new human skeletal muscle microvascular endothelial cell (HSMMEC) line. HSMMECs were isolated from human skeletal muscle and were infected with retroviruses harboring temperature-sensitive SV40 T antigen and telomerase genes. This cell line, termed TSM15, showed a spindle fiber-shaped morphology, an immunoreactivity to anti-factor VIII and anti-VE-cadherin antibodies, and a temperature-sensitive growth. TSM15 cells grew stably for more than 40 passages when they were cultured at 33°C, thereby retaining their spindle fiber-shaped morphology and contact inhibition at confluence. The cells expressed tight junctional molecules such as claudin-5, occludin, and zonula occludens-1, as well as transporters such as a glucose transporter 1. The transendothelial electrical resistance of TSM15 was as high as those of the human brain microvascular endothelial cell line. This novel cell line might facilitate the analyses of the pathophysiology of inflammatory myopathy, such as dermatomyositis, and can improve our understanding of the physiological and biochemical properties of the microvasculature in human skeletal muscle.A new conditionally immortalized human skeletal muscle microvascular endothelial cell line was established. This cell line, termed TSM15, expressed tight junctional molecules as well as transporters and has a barrier integrity relevant to that of a human brain microvascular endothelial cell line.
      PubDate: 2017-04-18T10:50:54.304015-05:
      DOI: 10.1002/jcp.25772
  • Purification of small molecule-induced cardiomyocytes from human induced
           pluripotent stem cells using a reporter system
    • Authors: Geun Hye Hwang; So Mi Park, Ho Jae Han, Joong Sun Kim, Seung Pil Yun, Jung Min Ryu, Ho Jin Lee, Woochul Chang, Su-Jin Lee, Jeong-Hee Choi, Jin-Sung Choi, Min Young Lee
      Abstract: In order to realize the practical use of human pluripotent stem cell (hPSC)-derived cardiomyocytes for the purpose of clinical use or cardiovascular research, the generation of large numbers of highly purified cardiomyocytes should be achieved. Here, we show an efficient method for cardiac differentiation of human induced pluripotent stem cells (hiPSCs) in chemically defined conditions and purification of hiPSC-derived cardiomyocytes using a reporter system. Regulation of the Wnt/β-catenin signaling pathway is implicated in the induction of the cardiac differentiation of hPSCs. We increased cardiac differentiation efficiency of hiPSCs in chemically defined conditions through combined treatment with XAV939, a tankyrase inhibitor and IWP2, a porcupine inhibitor and optimized concentrations. Although cardiac differentiation efficiency was high (>80%), it was difficult to suppress differentiation into non-cardiac cells, Therefore, we applied a lentiviral reporter system, wherein green fluorescence protein (GFP) and Zeocin-resistant gene are driven by promoter activation of a gene (TNNT2) encoding cardiac troponin T (cTnT), a cardiac-specific protein, to exclude non-cardiomyocytes from differentiated cell populations. We transduced this reporter construct into differentiated cells using a lentiviral vector and then obtained highly purified hiPSC-derived cardiomyocytes by treatment with the lowest effective dose of Zeocin. We significantly increased transgenic efficiency through manipulation of the cells in which the differentiated cells were simultaneously infected with virus and re-plated after single-cell dissociation. Purified cells specifically expressed GFP, cTnT, displayed typical properties of cardiomyocytes. This study provides an efficient strategy for obtaining large quantities of highly purified hPSC-derived cardiomyocytes for application in regenerative medicine and biomedical research.Combined treatment with XAV939 and IWP2 enhances hiPSC cardiac differentiation. Large amount of highly purified hiPSC-derived cardiomyocytes are obtained using a reporter system. Purified cells represent functional cardiomyocytes that possess electrophysiological properties and form cardiac junctions.
      PubDate: 2017-04-18T10:50:52.135818-05:
      DOI: 10.1002/jcp.25783
  • Evaluation the ethno-pharmacological studies in Iran during
           2004–2016: A systematic review
    • Authors: Zahra Sadeghi; Maryam Akaberi, Alireza Sobhkhizi, Amirhossein Sahebkar, Seyed Ahmad Emami
      Abstract: Although Iran has a deep history in herbal medicine and great heritage of ancient medical scholars, few efforts have been made to evaluate ethnopharmacological aspects of medicinal plants in this country. In the present study, the authors have reviewed all important literature about the ethnopharmacological investigations on medicinal plants used in the last decade in Iran. All provinces of Iran were categorized according to a phytogeographical division. Information was collected through bibliographic investigations from scientific journals and books. Afterward, the data were analyzed through the construction of specific ecological regions of the country. Fifty-five references reporting medicinal plants in five ecological zones were retrieved. The Irano-Turanian subregion has produced the greatest number of publications in this field among others (47%). Results illustrate that the most reported botanical families were Lamiaceae and Asteraceae (28.57% and 27.73%, respectively). Among various illnesses reported for these plants, gastrointestinal (30.15%), respiratory problems (14.28%), diabetes (11.11%), and cold/flu (11.11%) were the most cited. The most frequently cited medicinal uses were attributed to decoction and infusion preparations. Iran has a rich history of knowledge about phytotherapy and has also a diverse geographical regions, and a plant flora that is a good candidate for drug discovery. Documentation of indigenous knowledge about herbal medicine used by Iranian tribes is vital for the future development of herbal drugs. Ethnopharmacological studies of Iranian folk medicine with quantitative analytical techniques are warranted to find drug candidates, and also to preserve the precious knowledge of the Iranian folk medicine.This is the first ethno-pharmacological survery of medicinal plants used in the treatment of different diseases in Iran.
      PubDate: 2017-04-18T10:50:43.254221-05:
      DOI: 10.1002/jcp.25803
  • Optimized method for isolating highly purified and functional porcine
           aortic endothelial and smooth muscle cells
    • Authors: Farideh Beigi; Mitalben Patel, Marco A. Morales-Garza, Caitlin Winebrenner, Andrea S. Gobin, Eric Chau, Luiz C. Sampaio, Doris A. Taylor
      Abstract: Numerous protocols exist for isolating aortic endothelial and smooth muscle cells from small animals. However, establishing a protocol for isolating pure cell populations from large animal vessels that are more elastic has been challenging. We developed a simple sequential enzymatic approach to isolate highly purified populations of porcine aortic endothelial and smooth muscle cells. The lumen of a porcine aorta was filled with 25 U/ml dispase solution and incubated at 37°C to dissociate the endothelial cells. The smooth muscle cells were isolated by mincing the tunica media of the treated aorta and incubating the pieces in 0.2% and then 0.1% collagenase type I solution. The isolated endothelial cells stained positive for von Willebrand factor, and 97.2% of them expressed CD31. Early and late passage endothelial cells had a population doubling time of 38 hr and maintained a capacity to take up DiI-Ac-LDL and form tubes in Matrigel®. The isolated smooth muscle cells stained highly positive for alpha-smooth muscle actin, and an impurities assessment showed that only 1.8% were endothelial cells. Population doubling time for the smooth muscle cells was ∼70 hr at passages 3 and 7; and the cells positively responded to endothelin-1, as shown by a 66% increase in the intracellular calcium level. This simple protocol allows for the isolation of highly pure populations of endothelial and smooth muscle cells from porcine aorta that can survive continued passage in culture without losing functionality or becoming overgrown by fibroblasts.Isolating pure populations of endothelial and smooth muscle cells from large elastic mammalian vessels remains a challenge in vascular biology. A sequential enzymatic approach to isolate highly purified populations of porcine aortic endothelial and smooth muscle cells has been established. This simple protocol allows for the isolation of highly pure populations of endothelial and smooth muscle cells from porcine aorta that can survive continued passage in culture without losing functionality or becoming overgrown by fibroblasts.
      PubDate: 2017-04-18T10:46:06.50926-05:0
      DOI: 10.1002/jcp.25764
  • GM13133 is a negative regulator in mouse white adipocytes differentiation
           and drives the characteristics of brown adipocytes
    • Authors: LiangHui You; YaHui Zhou, XianWei Cui, XingYun Wang, YaZhou Sun, Yao Gao, Xing Wang, Juan Wen, Kaipeng Xie, RanRan Tang, ChenBo Ji, XiRong Guo
      Abstract: Obesity is tightly associated with the disturbance of white adipose tissue storing excess energy. Thermogenic adipocytes (brown and beige) exert a critical role of oxidizing nutrients at the high rates through non-shivering thermogenesis. The recruitment of brown characteristics in white adipocytes, termed browning, has been considered as a promising strategy for treating obesity and associated metabolic complications. Recently, long noncoding RNAs play a crucial role in regulating tissue development and participating in disease pathogenesis, yet their effects on the conversion of white into brown-like adipocytes and thermogenic function were not totally understood. Here, we identified a mouse brown adipose specific expressed lncRNA, termed GM13133. Moreover, a considerable amount of GM13133 is expressed in adipocytes and actively modulated by cold, β3-adrenergic agonist and cAMP stimuli, implying a potential role in the conversion from white to brown adipocytes. Overexpression of GM13133 did not affect the proliferation of mouse white pre-adipocytes, but inhibited white adipocyte differentiation by decreasing lipid accumulation. The forced expression of GM13133 also significantly drove the conversion of white into brown-like adipocytes with the enhanced mitochondrial biogenesis and the induced expression of brown adipocytes specific markers. A global mRNA analysis further indicated the possible regulatory role of cAMP signaling pathway in GM13133 mediated white-to-brown adipocytes conversion. Our results identified a lncRNA-mediated modulation in primary mouse white adipocyte differentiation and indicate the functional significance of GM13133 in promoting browning of white adipocytes and maintenance of thermogenesis, further providing a potential strategy to treating obesity.Our results provide identified a lncRNA-mediated modulation in primary mouse white adipocyte differentiation and further indicate the functional significance of GM13133 in promoting browning of white adipocytes and maintenance of thermogenesis, further providing a potential strategy to treating obesity.
      PubDate: 2017-04-18T10:20:36.14665-05:0
      DOI: 10.1002/jcp.25878
  • Activation of ClC-3 chloride channel by 17β-estradiol relies on the
           estrogen receptor α expression in breast cancer
    • Authors: Haifeng Yang; Lianshun Ma, Yawei Wang, Wanhong Zuo, Bingxue Li, Yaping Yang, Yehui Chen, Lixin Chen, Liwei Wang, Linyan Zhu
      Abstract: Although extensively studied, the mechanisms by which estrogen promotes breast cancer growth remain to be fully elucidated. Tamoxifen, an antiestrogen to treat ERα+ breast cancer, is also a high-affinity blocker of the chloride channels. In this study, we explored the involvement of the chloride channels in the action of estrogen in breast cancer. We found that 17β-estradiol (17β-E2) concentration-dependently activated the chloride currents in ERα+ breast cancer MCF-7 cells. Extracellular hypertonic challenge and chloride channel blockers, NPPB and DIDS inhibited the 17β-E2-activated chloride currents. Decreased the ClC-3 protein expression caused the depletion of the 17β-E2-activated chloride currents. 17β-E2-activated chloride currents which relied on the ERα expression were demonstrated by the following evidences. Firstly, 17β-E2-activated chloride currents could not be observed in ERα- breast cancer MDA-MB-231 cells. Secondly, ER antagonists, tamoxifen and ICI 182,780, and downregulation of ERα expression inhibited or abolished the 17β-E2-activated chloride currents. Thirdly, ERα expression was induced in MDA-MB-231 cells by ESR1 gene transfection, and then 17β-E2-activated chloride currents could be observed. In MCF-7 cells, ERα and ClC-3 mainly located in nucleus and translocated to cell plasma and membrane with respect to co-localization following treatment of 17β-E2. Downregulation of ERα expression could decrease the expression of ClC-3 protein. Conversely, downregulation of ClC-3 expression did not influence the ERα expression. Taken together, our findings demonstrated that ClC-3 is a potential target of 17β-E2 and modulates by the ERα in breast cancer cell. Pharmacological modulation of ClC-3 may provide a deep understanding in antiestrogen treatment of breast cancer patients. This article is protected by copyright. All rights reserved
      PubDate: 2017-04-17T09:25:38.767203-05:
      DOI: 10.1002/jcp.25963
  • Osteogenic differentiation Potential of Mesenchymal Stem Cells cultured on
           Nanofibrous Scaffold Improved in the Presence of Pulsed Electromagnetic
    • Authors: Monireh Arjmand; Abdolreza Ardeshirylajimi, Hossein Maghsoudi, Esmaeel Azadian
      Abstract: Nowadays, tissue engineering by using stem cells in combination with scaffolds and bioactive molecules has made significant contributions to the regeneration of damaged bone tissues. Since the usage of bioactive molecules including, growth factors to induce differentiation is safety limited in clinical applications, and it has also been previously observed that extremely low frequency pulsed electromagnetic fields (PEMF) can be effective in the enhancement of proliferation rate and osteogenic differentiation of stem cells, the aim of this study was investigating the osteoinductive potential of PEMF in combination with Poly(caprolactone) (PCL) nanofibrous scaffold. To achieve this aim, Adipose-derived mesenchymal stem cells (ADSCs) isolated and characterized and then osteogenic differentiation of them was investigated after culturing on the surface of PCL scaffold under treatments of PEMF, PEMF plus osteogenic medium (OM) and OM. Analysis of common osteogenic markers such as Alizarin red staining, ALP activity, calcium content and four important bone-related genes in days of 7, 14 and 21 confirmed that the effects of PEMF on the osteogenic differentiation of ADSCs are very similar to the effects of osteogenic medium. Thus, regarding the immunological concerns about the application of bioactive molecules for tissue engineering, PEMF could be a good alternative for osteogenic medium. Although, results were showed a synergetic effect for simultaneous application of PEMF and PCL scaffold in the osteogenesis process of ADSCs. Taking together, ADSCs-seeded PCL nanofibrous scaffold in combination with PEMF could be a great option for use in bone tissue engineering applications. This article is protected by copyright. All rights reserved
      PubDate: 2017-04-17T09:25:33.294085-05:
      DOI: 10.1002/jcp.25962
  • Therapeutic potential of novel formulated forms of curcumin in the
           treatment of breast cancer by the targeting of cellular and physiological
           dysregulated pathways
    • Authors: Amir Tajbakhsh; Malihe Hasanzadeh, Mehdi Rezaee, Mostafa Khedri, Majid Khazaei, Soodabeh Shahid Sales, Gordon A Ferns, Seyed Mahdi Hassanian, Amir Avan
      Abstract: Breast cancer is among the most important causes of cancer related death in women. There is a need for novel agents for targeting key signaling pathways to either improve the efficacy of the current therapy, or reduce toxicity. There is some evidence that curcumin may have antitumor activity in breast cancer. Several clinical trials have investigated its activity in patients with breast cancer, including a recent trial in breast cancer patients receiving radiotherapy, in whom it was shown that curcumin reduced the severity of radiation dermatitis, although it is associated with low bioavailability. Several approaches have been developed to increase its absorption rate (e.g., nano crystals, liposomes, polymers and micelles) and co-delivery of curcumin with adjuvants as well as different conjugation to enhance its bioavailability. In particular, micro-emulsions is an option for transdermal curcumin delivery, which has been reported to increase its absorption. Lipid-based nano-micelles is another approach to enhance curcumin absorption via gastrointestinal tract, while polymer-based nano-formulations (e.g., poly D, L-lactic-co-glycolic [PLGA]) allows the release of curcumin at a sustained level. This review summarizes the current data of the therapeutic potential of novel formulations of curcumin with particular emphasis on recent preclinical and clinical studies in the treatment of breast cancer. This article is protected by copyright. All rights reserved
      PubDate: 2017-04-17T09:25:28.861918-05:
      DOI: 10.1002/jcp.25961
  • Characterization of Runx2 Phosphorylation Sites Required for
           TGF-β1-Mediated stimulation of Matrix Metalloproteinase-13 Expression in
           Osteoblastic cells
    • Authors: B. Arumugam; M. Vairamani, N. C. Partridge, N. Selvamurugan
      Abstract: Transforming growth factor-beta1 (TGF-β1), a highly abundant growth factor in skeletal tissues, stimulates matrix metalloproteinase-13 (MMP-13) expression in osteoblastic cells. MMP-13 plays a critical role in bone remodeling. Runx2, a bone transcription factor, is required for TGF-β1-mediated stimulation of MMP-13 expression in osteoblastic cells. In this study, the molecular mechanism responsible for TGF-β1-stimulation of MMP-13 expression via Runx2 in osteoblastic cells was elucidated. TGF-β1 stimulated the phosphorylation of Runx2 at serine amino acids, and ERK inhibition blocked this effect in rat (UMR106-01) and human (MG-63) osteoblastic cells. Pretreatment with okadaic acid, a serine-threonine phosphatase inhibitor, increased Runx2 serine phosphorylation in osteoblastic cells. When cells were pretreated with an ERK inhibitor, TGF-β1-mediated stimulation of MMP-13 mRNA expression decreased. Nano-ESI/LC/MS analysis identified that TGF-β1 stimulates Runx2 phosphorylation at three serine amino acids. Transient transfection of mouse mesenchymal stem cells (C3H10T1/2) with Runx2 serine mutant constructs decreased TGF-β1-mediated Runx2 serine phosphorylation. A luciferase reporter assay identified that TGF-β1 stimulated MMP-13 promoter activity in these cells only in the presence of the wild Runx2 construct, and not with mutant Runx2. Thus, TGF-β1 stimulates the phosphorylation of Runx2 at three serine amino acids, and this event is required for MMP-13 expression in osteoblastic cells. Hence, this study contributes to the knowledge of events governing bone remodeling and bone-related diseases. This article is protected by copyright. All rights reserved
      PubDate: 2017-04-17T09:15:43.677904-05:
      DOI: 10.1002/jcp.25964
  • The Endoplasmic Reticulum-Associated Protein, OS-9, Behaves as a Lectin in
           Targeting the Immature Calcium-sensing Receptor
    • Authors: Bryan K Ward; Sarah L Rea, Aaron L Magno, Bernadette Pedersen, Suzanne J Brown, Shelby Mullin, Ajanthy Arulpragasam, Evan Ingley, Arthur D Conigrave, Thomas Ratajczak
      Abstract: The mechanisms responsible for the processing and quality control of the calcium-sensing receptor (CaSR) in the endoplasmic reticulum (ER) are largely unknown. In a yeast two-hybrid screen of the CaSR C-terminal tail (residues 865 -1078), we identified osteosarcoma-9 (OS-9) protein as a binding partner. OS-9 is an ER-resident lectin that targets misfolded glycoproteins to the ER-associated degradation (ERAD) pathway through recognition of specific N-glycans by its mannose-6-phosphate receptor homology (MRH) domain. We show by confocal microscopy that the CaSR and OS-9 co-localize in the ER in COS-1 cells. In immunoprecipitation studies with co-expressed OS-9 and CaSR, OS-9 specifically bound the immature form of wild type CaSR in the ER. OS-9 also bound the immature forms of a CaSR C-terminal deletion mutant and a C677A mutant that remains trapped in the ER, although binding to neither mutant was favoured over wild type receptor. OS-9 binding to immature CaSR required the MRH domain of OS-9 indicating that OS-9 acts as a lectin most likely to target misfolded CaSR to ERAD. Our results also identify two distinct binding interactions between OS-9 and the CaSR, one involving both C-terminal domains of the two proteins and the other involving both N-terminal domains. This suggests the possibility of more than one functional interaction between OS-9 and the CaSR. When we investigated the functional consequences of altered OS-9 expression, neither knockdown nor overexpression of OS-9 was found to have a significant effect on CaSR cell surface expression or CaSR-mediated ERK1/2 phosphorylation. This article is protected by copyright. All rights reserved
      PubDate: 2017-04-17T01:40:44.582414-05:
      DOI: 10.1002/jcp.25957
  • Spermicidal efficacy of VRP, a synthetic cationic antimicrobial peptide,
           inducing apoptosis and membrane disruption
    • Authors: Prasanta Ghosh; Arpita Bhoumik, Sudipta Saha, Sandipan Mukherjee, Sarfuddin Azmi, Jimut Kanti Ghosh, Sandhya Rekha Dungdung
      Abstract: Presently available contraceptives are mostly hormonal or detergent in nature with numerous side effects like irritation, lesion, inflammation in vagina, alteration of body homeostasis etc. Antimicrobial peptides with spermicidal activity but without adverse effects may be suitable alternatives. In the present study spermicidal activity of a cationic antimicrobial peptide VRP on human spermatozoa has been elucidated. Progressive forward motility of human spermatozoa was instantly stopped after 100 µM VRP treatment and at 350 µM all kinds of sperm motility ceased within 20 seconds as assessed by the Sander-Cramer assay. The spermicidal effect was confirmed by eosin-nigrosin assay and HOS test. VRP treatment (100µM) in human spermatozoa induced both the intrinsic and extrinsic pathways of apoptosis. TUNEL assay showed VRP treatment significantly disrupted the DNA integrity and changed the mitochondrial membrane permeability as evident from MPTP assay. AFM and SEM results depicted ultra structural changes including disruption of the acrosomal cap and plasma membrane of the head and midpiece region after treatment with 350 µM VRP. MTT assay showed after treatments with 100 µM and 350 µM of VRP for 24 hrs, a substantial amount of Lactobacillus acidophilus (about 90% and 75% respectively) remained viable. Hence, VRP being a small synthetic peptide with antimicrobial and spermicidal activity but tolerable to normal vaginal microflora, may be a suitable target for elucidating its contraceptive potentiality. This article is protected by copyright. All rights reserved
      PubDate: 2017-04-14T07:50:58.649528-05:
      DOI: 10.1002/jcp.25958
  • Platelet-derived microparticles regulates thrombin generation via
           phophatidylserine in abdominal sepsis
    • Authors: Yongzhi Wang; Su Zhang, Lingtao Luo, Eva Norström, Oscar Ö Braun, Matthias Mörgelin, Henrik Thorlacius
      Abstract: Sepsis is associated with dysfunctional coagulation. Recent data suggest that platelets play a role in sepsis by promoting neutrophil accumulation. Herein, we show that cecal ligation and puncture (CLP) triggered systemic inflammation, which is characterized by formation of IL-6 and CXC chemokines as well as neutrophil accumulation in the lung. Platelet depletion decreased neutrophil accumulation, IL-6 and CXC chemokines formation in septic lungs. Depletion of platelets increased peak thrombin formation and total thrombin generation (TG) in plasma from septic animals. CLP elevated circulating levels of platelet-derived microparticles (PMPs). In vitro generated PMPs were a potent inducer of TG. Interestingly, in vitro wild-type recombinant annexin V abolished PMP-induced thrombin formation whereas a mutant annexin V protein, which does not bind to phosphatidylserine (PS), had no effect. Administration of wild-type, but not mutant annexin V, significantly inhibited thrombin formation in septic animals. Moreover, CLP-induced formation of thrombin-antithrombin complexes were reduced in platelet-depleted mice and in animals pretreated with annexin V. PMP-induced TG attenuated in FXII- and FVII-deficient plasma. These findings suggest that sepsis-induced TG is dependent on platelets. Moreover, PMPs formed in sepsis are a potent inducer of TG via PS exposure and activation of both the intrinsic and extrinsic pathway of coagulation. In conclusion, these observations suggest that PMPs and PS play an important role in dysfunctional coagulation in abdominal sepsis. This article is protected by copyright. All rights reserved
      PubDate: 2017-04-14T07:50:35.251231-05:
      DOI: 10.1002/jcp.25959
  • Leucine elicits myotube hypertrophy and enhances maximal contractile force
           in tissue engineered skeletal muscle in vitro
    • Authors: N.R.W Martin; M.C Turner, R Farrington, D.J Player, M.P Lewis
      Abstract: The amino acid leucine is thought to be important for skeletal muscle growth by virtue of its ability to acutely activate mTORC1 and enhance muscle protein synthesis, yet little data exist regarding its impact on skeletal muscle size and its ability to produce force. We utilised a tissue engineering approach in order to test whether supplementing culture medium with leucine could enhance mTORC1 signalling, myotube growth and muscle function. Phosphorylation of the mTORC1 target proteins 4EBP-1 and rpS6 and myotube hypertrophy appeared to occur in a dose dependent manner, with 5 and 20mM of leucine inducing similar effects, which were greater than those seen with 1mM. Maximal contractile force was also elevated with leucine supplementation; however although this did not appear to be enhanced with increasing leucine doses, this effect was completely ablated by co-incubation with the mTOR inhibitor rapamycin, showing that the augmented force production in the presence of leucine was mTOR sensitive. Finally, by using electrical stimulation to induce chronic (24 hours) contraction of engineered skeletal muscle constructs, we were able to show that the effects of leucine and muscle contraction are additive, since the two stimuli had cumulative effects on maximal contractile force production. These results extend our current knowledge of the efficacy of leucine as an anabolic nutritional aid showing for the first time that leucine supplementation may augment skeletal muscle functional capacity, and furthermore validates the use of engineered skeletal muscle for highly-controlled investigations into nutritional regulation of muscle physiology. This article is protected by copyright. All rights reserved
      PubDate: 2017-04-14T07:50:34.066915-05:
      DOI: 10.1002/jcp.25960
  • Metformin attenuates albumin-induced alterations in renal tubular cells in
    • Authors: Soumaya Allouch; Shankar Munusamy
      Abstract: Proteinuria (albuminuria) plays a crucial role in the etiology of chronic kidney disease (CKD) via alteration of multiple signaling pathways and cellular process in renal cells. The objectives of this study are to investigate the effects of activation of the energy-sensing molecule AMP-activated kinase (AMPK) in renal cells using metformin on endoplasmic reticulum (ER) stress, AKT, mTOR, epithelial-to-mesenchymal transition (EMT), autophagy, and apoptosis that are thought to mediate renal cell injury during proteinuria, and to dissect the AMPK- and non-AMPK mediated effects of metformin using an in vitro model of albumin-induced renal cell injury. Rat renal proximal tubular (NRK-52E) cells were exposed to 10 and 15 mg/ml of albumin for 72 h in the presence of 1 mM Metformin and/or 0.5 µM compound C, and assessed for alterations in the aforementioned pathways. Metformin treatment restored AMPK phosphorylation and augmented autophagy in renal cells exposed to albumin. In addition, metformin treatment attenuated the albumin-induced phosphorylation of AKT and the downstream targets of mTOR, and prevented albumin-mediated inductions of EMT marker (α-SMA), pro-apoptotic ER stress marker CHOP, and apoptotic caspases -12 and -3 in renal cells. Blockade of metformin-induced AMPK activation with compound C blunted the ER defense response and autophagy but had no effect on the markers of EMT and apoptosis in our model. Our studies suggest that metformin protects renal cells against proteinuric cytotoxicity via suppression of AKT and mTOR activation, inhibition of EMT and apoptosis, and augmentation of autophagy and ER defense response through AMPK-independent and AMPK-dependent mechanisms, respectively.Proteinuria (albuminuria) plays a crucial role in the etiology of chronic kidney disease via alteration of multiple signaling pathways in renal cells. The current study using an in vitro model of albumin-induced cytotoxicity suggests that metformin protects renal cells against proteinuric cytotoxicity via suppression of AKT and mTOR activation, inhibition of epithelial-to-mesenchymal transition (EMT) and apoptosis, and augmentation of autophagy and ER defense response through AMPK-independent and AMPK-dependent mechanisms respectively.
      PubDate: 2017-04-13T12:07:37.793726-05:
      DOI: 10.1002/jcp.25838
  • Complex Osteoclastogenic Inductive Effects of Nicotine Over Hydroxyapatite
    • Authors: J. Costa-Rodrigues; I. Rocha, M.H. Fernandes
      Abstract: Cigarette smoke is associated to pathological weakening of bone tissue, being considered an important playmaker in conditions such as osteoporosis and periodontal bone loss. In addition, it is also associated with an increased risk of failure in bone regeneration strategies. The present work aimed to characterize the effects of nicotine on human osteoclastogenesis over a hydroxyapatite substrate. Osteoclast precursors were maintained in the absence or presence of the osteoclastogenesis enhancers M-CSF and RANKL, and were further treated with nicotine levels representative of the concentrations observed in the plasma and saliva of smokers. It was observed that nicotine at low concentrations elicit an increase in osteoclast differentiation, but only in the presence of M-CSF and RANKL it was also able to significantly increase the resorbing ability of osteoclasts. A slight downregulation of NFkB pathway and an increase in the production of TNF-α and, particularly PGE2, were involved in the observed effects of nicotine. At high concentrations, nicotine revealed cytotoxic effects, causing a decrease in cell density. In conclusion, nicotine at levels found in the plasma of the smokers, has the ability to act directly on osteoclast precursors, inducing its osteoclastogenic differentiation. The stimulatory behavior appears to be dependent on the stage of osteoclastic differentiation of the precursor cells, which means, in the absence of M-CSF and RANKL, it only favors the initial stages of osteoclast differentiation, while in the presence of the growth factors, a significant increase in their resorbing ability is also achieved. This article is protected by copyright. All rights reserved
      PubDate: 2017-04-13T11:35:23.039957-05:
      DOI: 10.1002/jcp.25956
  • Leptin accelerates the pathogenesis of heterotopic ossification in rat
           tendon tissues via mTORC1 signaling
    • Authors: Huaji Jiang; Yuhui Chen, Guorong Chen, Xinggui Tian, Jiajun Tang, Lei Luo, Minjun Huang, Bin Yan, Xiang Ao, Wen Zhou, Liping Wang, Xiaochun Bai, Zhongmin Zhang, Liang Wang, Cory J. Xian
      Abstract: Leptin, an adipocyte-derived cytokine associated with bone metabolism, is believed to play a critical role in the pathogenesis of heterotopic ossification (HO). The effect and underlying action mechanism of leptin were investigated on osteogenic differentiation of tendon-derived stem cells (TDSCs) in vitro and the HO formation in rat tendons. Isolated rat TDSCs were treated with various concentrations of leptin in the presence or absence of mTORC1 signaling specific inhibitor rapamycin in vitro. A rat model with Achilles tenotomy was employed to evaluate the effect of leptin on HO formation together with or without rapamycin treatment. In vitro studies with TDSCs showed that leptin increased the expression of osteogenic biomarkers (alkaline phosphatase, runt-related transcription factor 2, osterix, osteocalcin) and enhanced mineralization of TDSCs via activating the mTORC1 signal pathway (as indicated by phosphorylation of p70 ribosomal S6 kinase 1 and p70 ribosomal S6). However, mTORC1 signaling blockade with rapamycin treatment suppressed leptin-induced osteogenic differentiation and mineralization. In vivo studies showed that leptin promoted HO formation in the Achilles tendon after tenotomy, and rapamycin treatment blocked leptin-induced HO formation. In conclusion, leptin can promote TDSC osteogenic differentiation and heterotopic bone formation via mTORC1 signaling in both in vitro and in vivo models, which provides a new potential therapeutic target for HO prevention. This article is protected by copyright. All rights reserved
      PubDate: 2017-04-13T11:34:53.476177-05:
      DOI: 10.1002/jcp.25955
  • The therapeutic potential of targeting the BRAF in patients with
           colorectal cancer
    • Authors: Afsane Bahrami; AmirReza Hesari, Majid Khazaei, Seyed Mahdi Hassanian, Gordon Ferns, Amir Avan
      Abstract: Colorectal cancer is among the most lethal malignancies globally. BRAF is a member of the RAS/RAF/MEK/ERK signaling pathway. Its constitutive activation can result in increased cellular growth, development, invasion, and resistance to therapy. A mutation of the BRAF gene is present in 5-10% of metastatic colorectal cancers. BRAF mutations have been found to predict a lack of benefit to anti-EGFR therapy in metastatic CRC. Furthermore, CRC containing the BRAF V600E mutation display an innate resistance to BRAF inhibitors. The mechanisms of cell resistance can be explained at least in part by ERK dependent and ERK in-dependent pathway. Clinical trials evaluating the combinations of BRAF, PI3K, EGFR, and/or MEK inhibitors have revealed promising activity in BRAF mutant containing CRCs. There may be some benefit from future studies that focus on improving the efficacy of combined therapy in CRC with respect to the sustained effects. The aim of current review is to give an overview about the current status and prospective regarding the therapeutic potential of targeting BRAF mutant colorectal cancer. This article is protected by copyright. All rights reserved
      PubDate: 2017-04-13T11:28:44.42886-05:0
      DOI: 10.1002/jcp.25952
  • A highly efficient method for extracting next-generation sequencing
           quality RNA from adipose tissue of recalcitrant animal species
    • Authors: Davinder Sharma; Naresh Golla, Dheer Singh, Suneel Kumar Onteru
      Abstract: The next-generation sequencing (NGS) based RNA sequencing (RNA-Seq) and transcriptome profiling offers an opportunity to unveil complex evolutionary processes. Successful RNA-Seq and transcriptome profiling requires a large amount of high-quality RNA. However, NGS-quality RNA isolation is extremely difficult from recalcitrant adipose tissue (AT) with high lipid content and low cell numbers. Further, the amount and biochemical composition of AT lipid varies depending upon the animal species which can pose different degree of resistance to RNA extraction. Currently available approaches may work effectively in one species but can be almost unproductive in another species. Herein, we report a two step protocol for the extraction of NGS quality RNA from AT across a broad range of animal species. This article is protected by copyright. All rights reserved
      PubDate: 2017-04-13T11:28:33.730824-05:
      DOI: 10.1002/jcp.25951
  • Anti-tumor effects of crocetin and related molecular targets
    • Authors: Maliheh Moradzadeh; Hamid Reza Sadeghnia, Alijan Tabarraei, Amirhossein Sahebkar
      Abstract: Natural products have gained a wide popularity as chemopreventive and anti-cancer agents owing to their multi-mechanistic mode of action, availability and synergism with several conventional chemotherapeutic agents. Crocetin is a carotenoid compound isolated from the stigma of Crocus sativus L. (saffron). Crocetin has shown promising effects as an anti-tumor agent in animal models and cell culture systems. Crocetin retards the growth of cancer cells via inhibiting nucleic acid synthesis, enhancing anti-oxidative system, and inducing apoptosis and differentiation pathways. The present review outlines natural sources of crocetin, and its pharmacokinetic and pharmacological properties relevant to the prevention and treatment of cancer. Also, we discuss molecular targets underlying the putative anti-tumor effects of crocetin. This article is protected by copyright. All rights reserved
      PubDate: 2017-04-13T11:28:18.959964-05:
      DOI: 10.1002/jcp.25953
  • Acyl-CoA synthetase short-chain family member 2 (ACSS2) is regulated by
           SREBP-1 and plays a role in fatty acid synthesis in caprine mammary
           epithelial cells
    • Authors: Huifen Xu; Jun Luo, Gongzhen Ma, Xueying Zhang, Dawei Yao, Ming Li, Juan J. Loor
      Abstract: Sterol regulatory element binding protein 1 (SREBP-1) is well-known as the master regulator of lipogenesis in rodents. Acyl-CoA synthetase short-chain family member 2 (ACSS2) plays a key role in lipogenesis by synthesizing acetyl-CoA from acetate for lipogenesis. ATP citrate lyase (ACLY) catalyzes the conversion of citrate and coenzyme A to acetyl-CoA, hence, it is also important for lipogenesis. Although ACSS2 function in cancer cells has been elucidated, its essentiality in ruminant mammary lipogenesis is unknown. Furthermore, ACSS2 gene promoter and its regulatory mechanisms have not known. Expression of ACSS2 was high in lipid synthesizing tissues, and its expression increased during lactation compared with non-lactating period. Simultaneous knockdown of both ACSS2 and ACLY by siRNA in primary goat mammary epithelial cells decreased (P 
      PubDate: 2017-04-13T11:25:05.299581-05:
      DOI: 10.1002/jcp.25954
  • Cytoskeletal remodeling and regulation of cell fate in the hypertensive
           neonatal pulmonary artery in response to stress
    • Authors: Dina Johar
      Abstract: Neonatal pulmonary hypertension (PHN) is a lethal progressive disease that occurs in prenatal circulatory transition. Mechanical wall strain caused by cardiac pulsation integrates with hypoxia to generate rapidly progressive myocyte cytoskeleton disassembly and failure to exert force generation. The physiological responses to such an interaction have not been investigated. The persistent phenotype does not respond to traditional vasodilator therapy; hence, there is a need for new treatment strategies to improve the morbidity and mortality outcomes. We reviewed the current research methods, models and markers of persistent PHN relevant to oxidative and nitrosative stress as well as cell fate commitment, with an emphasis on apoptosis and proliferation. We surveyed potential investigations into the role of senescence in neonatal PHN cell fate decision programming during vasodilator treatment and suggested putative drug targets to improve clinical outcomes. We identified important signaling intermediates of senescence and cell cycle entry regulation in hypertensive pulmonary arterial tissues. This article is protected by copyright. All rights reserved
      PubDate: 2017-04-13T10:57:35.948814-05:
      DOI: 10.1002/jcp.25950
  • Pin1, the Master Orchestrator of Bone Cell Differentiation
    • Authors: Rabia Islam; Won-Joon Yoon, Hyun-Mo Ryoo
      Abstract: Pin1 is an enzyme that specifically recognizes the peptide bond between phosphorylated serine or threonine (pS/pT-P) and proline. This recognition causes a conformational change of its substrate, which further regulates downstream signaling. Pin1−/− mice show developmental bone defects and reduced mineralization. Pin1 targets RUNX2 (Runt-Related Transcription Factor 2), SMAD1/5, and β-catenin in the FGF, BMP, and WNT pathways, respectively. Pin1 has multiple roles in the crosstalk between different anabolic bone signaling pathways. For example, it controls different aspects of osteoblastogenesis and increases the transcriptional activity of Runx2, both directly and indirectly. Pin1 also influences osteoclastogenesis at different stages by targeting PU.1 (Purine-rich nucleic acid binding protein 1), C-FOS, and DC-STAMP. The phenotype of Pin1−/− mice has led to the recent identification of multiple roles of Pin1 in different molecular pathways in bone cells. These roles suggest that Pin1 can be utilized as an efficient drug target in congenital and acquired bone diseases. J. Cell. Physiol. 9999: 1–9, 2016. © 2016 The
      Authors . Journal of Cellular Physiology Published by Wiley Periodicals Inc.Pin1 is a peptidyl prolyl cis–trans isomerase that isomerizes only phosphorylated Ser/Thr-Pro motifs. Upon binding, Pin1 mediates structural modifications resulting in dramatic changes in substrate activity. In this review, we have summarized the known roles of Pin1 in molecular regulation of bone cell differentiation. Pin1 binds to Runx2, Smad1/5, and β-catenin, which act downstream of the FGF, BMP, and Wnt signaling pathways, respectively. Binding of Pin1 increases the stability and transcriptional activity of Runx2 and Smad1/5. Pin1 also inhibits the nuclear export of β-catenin and increases the level of transcriptionally active nuclear β-catenin.
      PubDate: 2017-04-12T14:14:53.463962-05:
      DOI: 10.1002/jcp.25442
  • Serotonin transporter protects the placental cells against apoptosis in
           caspase 3-independent pathway
    • Authors: Coedy Hadden; Tariq Fahmi, Anthonya Cooper, Alena V. Savenka, Vladimir V. Lupashin, Drucilla J. Roberts, Luc Maroteaux, Sylvie Hauguel-de Mouzon, Fusun Kilic
      Abstract: Serotonin (5-HT) and its specific transporter, SERT play important roles in pregnancy. Using placentas dissected from 18d gestational SERT-knock out (KO), peripheral 5-HT (TPH1)-KO, and wild-type (WT) mice, we explored the role of 5-HT and SERT in placental functions in detail. An abnormal thick band of fibrosis and necrosis under the giant cell layer in SERT-KO placentas appeared only moderately in TPH1-KO and minimally present in WT placentas. The majority of the changes were located at the junctional zone of the placentas in SERT. The etiology of these findings was tested with TUNEL assays. The placentas from SERT-KO and TPH1-KO showed 49- and 8-fold increase in TUNEL-positive cells without a concurrent change in the DNA repair or cell proliferation compared to WT placentas. While the proliferation rate in the embryos of TPH1-KO mice was 16-fold lower than the rate in gestational age matched embryos of WT or SERT-KO mice. These findings highlight an important role of continuous 5-HT signaling on trophoblast cell viability. SERT may contribute to protecting trophoblast cells against cell death via terminating the 5-HT signaling which changes cell death ratio in trophoblast as well as proliferation rate in embryos. However, the cell death in SERT-KO placentas is in caspase 3-independent pathway.
      PubDate: 2017-04-12T04:10:51.84766-05:0
      DOI: 10.1002/jcp.25812
  • Homocysteine as a Pathological Biomarker for Bone Disease
    • Authors: Jyotirmaya Behera; Jyoti Bala, Mohammed Nuru, Suresh C. Tyagi, Neetu Tyagi
      Abstract: In the last few decades, perturbation in methyl-group and homocysteine (Hcy) balance have emerged as independent risk factors in a number of pathological conditions including neurodegenerative disease, cardiovascular dysfunction, cancer development, autoimmune disease, and kidney disease. Recent studies report Hcy to be a newly recognized risk factor for osteoporosis. Elevated Hcy levels are known to modulate osteoclastgenesis by causing detrimental effects on bone via oxidative stress induced metalloproteinase-mediated extracellular matrix degradation and decrease in bone blood flow. Evidence from previous studies also suggests that the decreased chondrocytes mediated bone mineralization in chick limb-bud mesenchymal cells and during the gestational period of ossification in rat model. However, Hcy imbalance and its role in bone loss, regression in vascular invasion, and osteoporosis, are not clearly understood. More investigations are required to explore the complex interplay between Hcy imbalance and onset of bone disease progression. This article reviews the current body of knowledge on regulation of Hcy mediated oxidative stress and its role in bone remodeling, vascular blood flow and progression of bone disease. J. Cell. Physiol. 9999: 1–6, 2016. © 2016 Wiley Periodicals, Inc.This article reviews the current body of knowledge on regulation of Hcy-mediated oxidative stress and its role in bone remodeling, vascular blood flow, and progression of bone disease.
      PubDate: 2017-04-12T04:10:32.287337-05:
      DOI: 10.1002/jcp.25693
  • Dendritic Cell-Specific Transmembrane Protein (DC-STAMP) Regulates
           Osteoclast Differentiation via the Ca2+/NFATc1 Axis
    • Authors: Ya-Hui Chiu; Edward Schwarz, Dongge Li, Yuexin Xu, Tzong-Ren Sheu, Jinbo Li, Karen L. de Mesy Bentley, Changyong Feng, Baoli Wang, Jhih-Cheng Wang, Liz Albertorio-Saez, Ronald Wood, Minsoo Kim, Wensheng Wang, Christopher T. Ritchlin
      Abstract: DC-STAMP is a multi-pass transmembrane protein essential for cell–cell fusion between osteoclast precursors during osteoclast (OC) development. DC-STAMP−/− mice have mild osteopetrosis and form mononuclear cells with limited resorption capacity. The identification of an Immunoreceptor Tyrosine-based Inhibitory Motif (ITIM) on the cytoplasmic tail of DC-STAMP suggested a potential signaling function. The absence of a known DC-STAMP ligand, however, has hindered the elucidation of downstream signaling pathways. To address this problem, we engineered a light-activatable DC-STAMP chimeric molecule in which light exposure mimics ligand engagement that can be traced by downstream Ca2+ signaling. Deletion of the cytoplasmic ITIM resulted in a significant elevation in the amplitude and duration of intracellular Ca2+ flux. Decreased NFATc1 expression in DC-STAMP−/− cells was restored by DC-STAMP over-expression. Multiple biological phenotypes including cell–cell fusion, bone erosion, cell mobility, DC-STAMP cell surface distribution, and NFATc1 nuclear translocation were altered by deletion of the ITIM and adjacent amino acids. In contrast, mutations on each of the tyrosine residues surrounding the ITIM showed no effect on DC-STAMP function. Collectively, our results suggest that the ITIM on DC-STAMP is a functional motif that regulates osteoclast differentiation through the NFATc1/Ca2+ axis. J. Cell. Physiol. 9999: 1–12, 2016. © 2016 Wiley Periodicals, Inc.
      PubDate: 2017-04-12T03:46:10.033247-05:
      DOI: 10.1002/jcp.25638
  • The Role of Exercise and TFAM in Preventing Skeletal Muscle Atrophy
    • Authors: Nicholas T. Theilen; George H. Kunkel, Suresh C. Tyagi
      Abstract: Skeletal muscle atrophy is the consequence of protein degradation exceeding protein synthesis. This arises for a multitude of reasons including the unloading of muscle during microgravity, post-surgery bedrest, immobilization of a limb after injury, and overall disuse of the musculature. The development of therapies prior to skeletal muscle atrophy settings to diminish protein degradation is scarce. Mitochondrial dysfunction is associated with skeletal muscle atrophy and contributes to the induction of protein degradation and cell apoptosis through increased levels of ROS observed with the loss of organelle function. ROS binds mtDNA, leading to its degradation and decreasing functionality. Mitochondrial transcription factor A (TFAM) will bind and coat mtDNA, protecting it from ROS and degradation while increasing mitochondrial function. Exercise stimulates cell signaling pathways that converge on and increase PGC-1α, a well-known activator of the transcription of TFAM and mitochondrial biogenesis. Therefore, in the present review we are proposing, separately, exercise and TFAM treatments prior to atrophic settings (muscle unloading or disuse) alleviate skeletal muscle atrophy through enhanced mitochondrial adaptations and function. Additionally, we hypothesize the combination of exercise and TFAM leads to a synergistic effect in targeting mitochondrial function to prevent skeletal muscle atrophy. J. Cell. Physiol. 9999: 1–11, 2017. © 2016 The
      Authors . Journal of Cellular Physiology Published by © 2016 Wiley Periodicals, Inc.Skeletal muscle atrophy via unloading and disuse of the musculature is the consequence of protein degradation exceeding protein synthesis. This state is correlated with increased markers of mitochondrial dysfunction. We hypothesize targeting increased mitochondrial function through exercise and mitochondrial transcription factor A treatments will diminish skeletal muscle atrophy after a period of muscle unloading and disuse.
      PubDate: 2017-04-12T03:45:30.354215-05:
      DOI: 10.1002/jcp.25737
  • Dysbiosis and Disease: Many Unknown Ends, Is It Time to Formulate
           Guidelines for Dysbiosis Research?
    • Authors: Sudhakar Veeranki; Suresh C. Tyagi
      Abstract: Dysbiosis has been implicated in modulation of disease and treatment outcome. It also has been linked to the reproducibility concerns. However, research community needs guidelines on animal models and dysbiosis research to tackle the complexities associated with it. There is a necessity for multi-disciplinary collaborative approach in setting up certain guidelines to hasten the dysbiosis research in a hassle-free manner. J. Cell. Physiol. 9999: 1–2, 2016. © 2016 Wiley Periodicals, Inc.Dysbiosis has been implicated in modulation of disease and treatment outcome. It also has been linked to the reproducibility concerns. However, research community needs guidelines on animal models and dysbiosis research to tackle the complexities associated with it. There is a necessity for multi-disciplinary collaborative approach in setting up certain guidelines to hasten the dysbiosis research in a hassle-free manner.
      PubDate: 2017-04-12T03:45:26.938277-05:
      DOI: 10.1002/jcp.25719
  • Promoting Cancer Control in Africa With “Ubuntu”: A Report of the
           African Organization for Research and Training in Africa (AORTIC) 10th
           Conference, 2015 in Marrakech, Morocco
    • Authors: Kenneth O. Simbiri; Christopher K. Williams, Marcella Macaluso, Antonio Giordano
      Abstract: The objectives of the African Organization for Research and Training in Cancer (AORTIC), includes bringing products of decades of advances in cancer research to African populations through local and international collaboration. The consistent and huge growth in participation in the conferences and the diversity of the nations is a witness to the success of the organization thus far. The theme for the Tenth AORTIC International Conference on Cancer in Africa in Morocco in 2015 was “Road map to Cancer Control in Africa” and topics of discussion of paramount importance for low- and middle-income African countries included childhood cancers such as BL, cancers of the cervix, breast, and prostate; cancers associated with HIV-infection such as cervical, vulvar, and anal; as well as cancer care challenges associated with palliative care. The role of environmental factors that underlie some epigenetic changes in some of the cancers was emphasized. Oral and poster presentations from various parts of the continent indicate the growth of basic and translational science of cancer in the region, with studies revealing regional diversity in the frequencies of the triple-negative breast cancer, cervical cancer, prostate cancer, HCC, and Burkitt's lymphoma. There was a sign that Africa is trying to keep pace with the paradigm shift and focusing on translational medicine. This was shown by suggestions for application of genome-wide association studies, new generation sequencing, as well as the evaluation of single nucleotide polymorphisms that may be responsible for variable susceptibility in some of the prevalent cancers in people of African descent. J. Cell. Physiol. 9999: 1–9, 2016. © 2016 Wiley Periodicals, Inc.
      PubDate: 2017-04-12T03:45:25.755658-05:
      DOI: 10.1002/jcp.25671
  • Human White Adipocytes Convert Into “Rainbow” Adipocytes In
    • Authors: Giulia Maurizi; Antonella Poloni, Domenico Mattiucci, Spartaco Santi, Angela Maurizi, Valerio Izzi, Angelica Giuliani, Stefania Mancini, Maria Cristina Zingaretti, Jessica Perugini, Ilenia Severi, Massimo Falconi, Marco Vivarelli, Maria Rita Rippo, Silvia Corvera, Antonio Giordano, Pietro Leoni, Saverio Cinti
      Abstract: White adipocytes are plastic cells able to reversibly transdifferentiate into brown adipocytes and into epithelial glandular cells under physiologic stimuli in vivo. These plastic properties could be used in future for regenerative medicine, but are incompletely explored in their details. Here, we focused on plastic properties of human mature adipocytes (MA) combining gene expression profile through microarray analysis with morphologic data obtained by electron and time lapse microscopy. Primary MA showed the classic morphology and gene expression profile of functional mature adipocytes. Notably, despite their committed status, MA expressed high levels of reprogramming genes. MA from ceiling cultures underwent transdifferentiation toward fibroblast-like cells with a well-differentiated morphology and maintaining stem cell gene signatures. The main morphologic aspect of the transdifferentiation process was the secretion of large lipid droplets and the development of organelles necessary for exocrine secretion further supported the liposecretion process. Of note, electron microscope findings suggesting liposecretion phenomena were found also in explants of human fat and rarely in vivo in fat biopsies from obese patients. In conclusion, both MA and post-liposecretion adipocytes show a well-differentiated phenotype with stem cell properties in line with the extraordinary plasticity of adipocytes in vivo. J. Cell. Physiol. 9999: 1–13, 2016. © 2016 Wiley Periodicals, Inc.Plastic properties of human mature adipocytes (MA) have been explored combining gene expression profile made by microarray analysis with morphologic data obtained by electron and time lapse microscopy. Mature adipocytes from ceiling cultures underwent transdifferentiation toward fibroblast-like cells with a well-differentiated morphology and maintaining stem cell gene signatures.
      PubDate: 2017-04-12T03:42:35.986628-05:
      DOI: 10.1002/jcp.25743
  • Contributions of 3D Cell Cultures for Cancer Research
    • Authors: Maddaly Ravi; Aarthi Ramesh, Aishwarya Pattabhi
      Abstract: Cancer cell lines have contributed immensely in understanding the complex physiology of cancers. They are excellent material for studies as they offer homogenous samples without individual variations and can be utilised with ease and flexibility. Also, the number of assays and end-points one can study is almost limitless; with the advantage of improvising, modifying or altering several variables and methods. Literally, a new dimension to cancer research has been achieved by the advent of 3Dimensional (3D) cell culture techniques. This approach increased many folds the ways in which cancer cell lines can be utilised for understanding complex cancer biology. 3D cell culture techniques are now the preferred way of using cancer cell lines to bridge the gap between the ‘absolute in vitro’ and ‘true in vivo’. The aspects of cancer biology that 3D cell culture systems have contributed include morphology, microenvironment, gene and protein expression, invasion/migration/metastasis, angiogenesis, tumour metabolism and drug discovery, testing chemotherapeutic agents, adaptive responses and cancer stem cells. We present here, a comprehensive review on the applications of 3D cell culture systems for these aspects of cancers. J. Cell. Physiol. 9999: 1–19, 2016. © 2016 Wiley Periodicals, Inc.3D cell cultures are gaining ground for several applications, of which cancer research is an important area. Several aspects of cancer physiology have been unraveled using these cultures. Also, 3D cell cultures provide realistic material for drug discovery and biomarker discovery.
      PubDate: 2017-04-12T03:30:48.581371-05:
      DOI: 10.1002/jcp.25664
  • MAP Kinase-Dependent RUNX2 Phosphorylation Is Necessary for Epigenetic
           Modification of Chromatin During Osteoblast Differentiation
    • Authors: Yan Li; Chunxi Ge, Renny T. Franceschi
      Abstract: RUNX2, an essential transcription factor for osteoblast differentiation and bone formation is activated by ERK/MAP kinase-dependent phosphorylation. However, relationship between these early events and specific epigenetic modifications of chromatin during osteoblast differentiation have not been previously examined. Here, we explore these relationships using chromatin immunoprecipitation (ChIP) to detect chromatin modifications in RUNX2-binding regions of Bglap2 and Ibsp. Growth of MC3T3-E1c4 preosteoblast cells in differentiation conditions rapidly induced Bglap2 and lbsp mRNAs. For both genes, osteogenic stimulation increased chromatin-bound P-ERK, P-RUNX2, p300, and RNA polymerase II as well as histone H3K9 and H4K5 acetylation. The level of H3K4 di-methylation, another gene activation-associated histone mark, also increased. In contrast, levels of the gene repressive marks, H3K9 mono-, di-, and tri-methylation in the same regions were reduced. Inhibition of MAP kinase signaling blocked differentiation-dependent chromatin modifications and Bglap2 and Ibsp expression. To evaluate the role of RUNX2 phosphorylation in these responses, RUNX2-deficient C3H10T1/2 cells were transduced with adenovirus encoding wild type or phosphorylation site mutant RUNX2 (RUNX2 S301A/S319A). Wild type RUNX2, but not the non-phosphorylated mutant, increased H3K9 and H4K5 acetylation as well as chromatin-associated P-ERK, p300, and polymerase II. Thus, RUNX2 phosphorylation is necessary for subsequent epigenetic changes required for osteoblast gene expression. Taken together, this study reveals a molecular mechanism through which osteogenic genes are controlled by a MAPK and P-RUNX2-dependent process involving epigenetic modifications of specific promoter regions. J. Cell. Physiol. 9999: 1–9, 2016. © 2016 Wiley Periodicals, Inc.The RUNX2 transcription factor requires MAP kinase-dependent phosphorylation to become transcriptionally active. Here, we show that phosphorylation is required for transcription-related changes in histone acetylation and methylation and recruitment of cofactors such as P300/CBP and RNA polymerase II to promoter regions of bone-related genes.
      PubDate: 2017-04-10T08:30:54.649747-05:
      DOI: 10.1002/jcp.25517
  • Live Cell Imaging: Assessing the Phototoxicity of 488 and 546 nm Light
           and Methods to Alleviate it
    • Authors: Stephen Douthwright; Greenfield Sluder
      Abstract: In live cell imaging of fluorescent proteins, phototoxicity of the excitation light can be problematical. Cell death is obvious, but reduced cell viability can make the interpretation of observations error prone. We characterized the phototoxic consequences of 488 and 546 nm light on untransformed human cells and tested methods that have or could be used to alleviate photodamage. Unlabeled RPE1 cells were given single 0.5–2.5 min irradiations in early G1 from a mercury arc lamp on a fluorescence microscope. Four hundred eighty-eight nanometer light produced a dose-dependent decrease in the percentage of cells that progressed to mitosis, slowing of the cell cycle for some of those entering mitosis, and a ∼12% incidence of cell death for the highest dose. For 546 nm light we found a 10–15% reduction in the percentage of cells entering mitosis, no strong dose dependency, and a ∼2% incidence of cell death for the longest irradiations. For cells expressing GFP-centrin1 or mCherry-centrin1, fewer entered mitosis for each dose than unlabeled cells. For constant total dose 488 nm light irradiations of unlabeled cells, reducing the intensity 10-fold or spreading the exposures out as a series of 10 sec pulses at 1 min intervals produced a minor and not consistent improvement in the percentage of cells entering mitosis. Reducing oxidative processes, by culturing at ∼3% oxygen or adding the reducing agent Trolox noticeably increased the fraction of cells entering mitosis. Thus, for long-term imaging there can be value to using RFP constructs and for GFP-tagged proteins reducing oxidative processes. J. Cell. Physiol. 9999: 1–8, 2016. © 2016 Wiley Periodicals, Inc.We characterized the phototoxic consequences of 488 and 546 nm light on untransformed human cells. Four hundred eighty-eight nanometer light produced a dose-dependent decrease in the percentage of cells that progressed to mitosis, and slowing of the cell cycle for some of those entering mitosis and for 546 nm light, we found a 10–15% reduction in the percentage of cells entering mitosis, but no strong dose dependency. Reducing oxidative processes, by culturing at ∼3% oxygen or adding the reducing agent Trolox noticeably increased the fraction of cells entering mitosis.
      PubDate: 2017-04-10T08:30:45.4095-05:00
      DOI: 10.1002/jcp.25588
  • Use of a Collagen Membrane to Enhance the Survival of Primary Intestinal
           Epithelial Cells
    • Authors: Fiorella Di Claudio; Cecilia I. Muglia, Paola L. Smaldini, María Lucía Orsini Delgado, Fernando M. Trejo, J. Raúl Grigera, Guillermo H. Docena
      Abstract: Intestinal epithelial cell culture is important for biological, functional, and immunological studies. Since enterocytes have a short in vivo life span due to anoikis, we aimed to establish a novel and reproducible method to prolong the survival of mouse and human cells. Cells were isolated following a standard procedure, and cultured on ordered-cow's collagen membranes. A prolonged cell life span was achieved; cells covered the complete surface of bio-membranes and showed a classical enterocyte morphology with high expression of enzymes supporting the possibility of cryopreservation. Apoptosis was dramatically reduced and cultured enterocytes expressed cytokeratin and LGR5 (low frequency). Cells exposed to LPS or flagellin showed the induction of TLR4 and TLR5 expression and a functional phenotype upon exposure to the probiotic Bifidobacterium bifidum or the pathogenic Clostridium difficile. The secretion of the homeostatic (IL-25 and TSLP), inhibitory (IL-10 and TGF-β), or pro-inflammatory mediators (IL-1β and TNF) were induced. In conclusion, this novel protocol using cow's collagen-ordered membrane provides a simple and reproducible method to maintain intestinal epithelial cells functional for cell-microorganism interaction studies and stem cell expansion. J. Cell. Physiol. 9999: 1–8, 2016. © 2016 Wiley Periodicals, Inc.We have developed a method for culturing primary intestinal epithelial cells useful for the study of their interaction with microbes.
      PubDate: 2017-04-10T08:26:20.026321-05:
      DOI: 10.1002/jcp.25594
  • Neurotransmitters: The Critical Modulators Regulating Gut–Brain Axis
    • Authors: Rahul Mittal; Luca H. Debs, Amit P. Patel, Desiree Nguyen, Kunal Patel, Gregory O'Connor, M'hamed Grati, Jeenu Mittal, Denise Yan, Adrien A. Eshraghi, Sapna K. Deo, Sylvia Daunert, Xue Zhong Liu
      Abstract: Neurotransmitters, including catecholamines and serotonin, play a crucial role in maintaining homeostasis in the human body. Studies on these neurotransmitters mainly revolved around their role in the “fight or flight” response, transmitting signals across a chemical synapse and modulating blood flow throughout the body. However, recent research has demonstrated that neurotransmitters can play a significant role in the gastrointestinal (GI) physiology. Norepinephrine (NE), epinephrine (E), dopamine (DA), and serotonin have recently been a topic of interest because of their roles in the gut physiology and their potential roles in GI and central nervous system pathophysiology. These neurotransmitters are able to regulate and control not only blood flow, but also affect gut motility, nutrient absorption, GI innate immune system, and the microbiome. Furthermore, in pathological states, such as inflammatory bowel disease (IBD) and Parkinson's disease, the levels of these neurotransmitters are dysregulated, therefore causing a variety of GI symptoms. Research in this field has shown that exogenous manipulation of catecholamine serum concentrations can help in decreasing symptomology and/or disease progression. In this review article, we discuss the current state-of-the-art research and literature regarding the role of neurotransmitters in regulation of normal GI physiology, their impact on several disease processes, and novel work focused on the use of exogenous hormones and/or psychotropic medications to improve disease symptomology. J. Cell. Physiol. 9999: 1–14, 2016. © 2016 Wiley Periodicals, Inc.Neurotransmitters affect microbiota in the gut: Neurotransmitters, including serotonin, alter the microbiota in the gut that can modulate the production of cytokines and bacterial by-products, leading to either healthy or diseased state.
      PubDate: 2017-04-10T08:26:17.781595-05:
      DOI: 10.1002/jcp.25518
  • Anti-Tumor Effects of Cardiac Glycosides on Human Lung Cancer Cells and
           Lung Tumorspheres
    • Authors: Vivek Kaushik; Juan Sebastian Yakisich, Neelam Azad, Yogesh Kulkarni, Rajkumar Venkatadri, Clayton Wright, Yon Rojanasakul, Anand Krishnan V. Iyer
      Abstract: Lung cancer is a leading cause of cancer-related death in the United States. Although several drugs have been developed that target individual biomarkers, their success has been limited due to intrinsic or acquired resistance for the specific targets of such drugs. A more effective approach is to target multiple pathways that dictate cancer progression. Cardiac glycosides demonstrate such multimodal effects on cancer cell survival, and our aim was to evaluate the effect of two naturally occurring monosaccaridic cardiac glycosides—Convallatoxin and Peruvoside on lung cancer cells. Although both drugs had significant anti-proliferative effects on H460 and Calu-3 lung cancer cells, Convallatoxin demonstrated twofold higher activity as compared to Peruvoside using both viability and colony forming assays, suggesting a role for the aglycone region in dictating drug potency. The tumor suppressor p53 was found to be important for action of both drugs—p53-underexpressing cells were less sensitive as compared to p53-positive H460 cells. Further, assessment of p53-underexpressing H460 cells showed that drugs were able to arrest cells in the G0/G1 phase of the cell cycle in a dose-dependent manner. Both drugs significantly inhibited migration and invasion of cancer cells and decreased the viability of floating tumorspheres. An assessment of intracellular pathways indicated that both drugs were able to modulate proteins that are involved in apoptosis, autophagy, cell cycle, proliferation, and EMT. Our data suggest, a promising role for cardiac glycosides in lung cancer treatment, and provides impetus for further investigation of the anti-cancer potential of this class of drugs. J. Cell. Physiol. 9999: 1–11, 2016. © 2016 Wiley Periodicals, Inc.The cardiac glycosides convallatoxin and peruvoside exert potent anti-cancer effects in lung cancer cells. A significant inhibition of cell migration, invasion, and lung tumorsphere formation was observed in response to both drugs.
      PubDate: 2017-04-10T08:26:10.364208-05:
      DOI: 10.1002/jcp.25611
  • TRIM33 is essential for osteoblast proliferation and differentiation via
           BMP pathway
    • Authors: Jia Guo; Wei Qin, Quan Xing, Manman Gao, Fuxin Wei, Zhi Song, Lingling Chen, Ying Lin, Xianling Gao, Zhengmei Lin
      Abstract: Tripartite motif containing 33 (TRIM33) functions both as a positive and negative regulator of the TGF-β/BMP pathway in tumors; however, its effect and mechanism during osteoblast proliferation and differentiation, which involves the TGF-β/BMP pathway is not defined. In this study, we used mouse C3H10T1/2 mesenchymal stem cell line and MC3T3-E1 preosteoblasts to investigate the role of TRIM33 during this process. The results demonstrated that the expression of TRIM33 increased during the differentiation. Moreover, the overexpression or knockdown of TRIM33 resulted in both an augmentation or decrease in osteoblast differentiation, which were measured by the expression of alkaline phosphatase (ALP) at the mRNA level, both Runt-related transcription factor 2 (Runx2) and osteocalcin (OCN) at the protein level, and the formation of mineral modules. To further demonstrate the mechanism of TRIM33 in this process, we found that TRIM33 could positively mediate the BMP pathway by forming TRIM33-Smad1/5 complex. This interaction between TRIM33 and Smad1/5 triggered the phosphorylation of Smad1/5. In addition, the essential role of TRIM33 in osteoblast proliferation was determined in this study by CellCounting Kit (CCK) −8 and cell cycle assays. In summary, we establish the function of TRIM33 as a positive regulator of osteoblast differentiation in BMP pathway, which mediates its effect through its interaction with and activation of Smad1/5. In addition, the results clearly demonstrate that TRIM33 is necessary for osteoblast proliferation by regulating cell cycle. These results suggest that TRIM33 can be a positive target of osteoblast proliferation and differentiation through BMP pathway.We establish the function of TRIM33 as a positive regulator of osteoblast differentiation in BMP pathway, which mediates its effect through its interaction with and activation of Smad1/5. In addition, the results clearly demonstrate that TRIM33 is necessary for osteoblast proliferation by regulating cell cycle. These results suggest that TRIM33 can be a positive target of osteoblast proliferation and differentiation through BMP pathway.
      PubDate: 2017-04-10T08:25:45.251448-05:
      DOI: 10.1002/jcp.25769
  • New Roles of Lkb1 in Regulating Adipose Tissue Development and
    • Authors: Ziye Xu; Jiaqi Liu, Tizhong Shan
      Abstract: Adipose tissues regulate energy metabolism and reproduction. There are three types of adipocytes (brown, white, and beige adipocytes) in mammals. White adipocytes store energy and are closely associated with obesity and other metabolic diseases. The beige and brown adipocytes have numerous mitochondria and high levels of UCP1 that dissipates lipid to generate heat and defend against obesity. The global epidemic of obesity and its associated metabolic diseases urge an imperative need for understating the regulation of adipogenesis. Liver kinase B1 (Lkb1), also called STK11, is a master kinase of the AMPK subfamily and plays crucial roles in regulating glucose and energy homeostasis in various metabolic tissues. In this review, we focus on the regulatory roles of Lkb1 in regulating preadipocyte differentiation, adipose tissue development, and thermogenesis. J. Cell. Physiol. 9999: 1–3, 2016. © 2016 Wiley Periodicals, Inc.(i) Lkb1 regulates adipocyte progenitors differentiation; (ii) Adipocyte-specific deletion of Lkb1 promotes brown adipose tissue development and browning of white adipose tissues; (iii) Lkb1 affects Ucp1 expression and thermogenesis.
      PubDate: 2017-04-10T08:21:41.418218-05:
      DOI: 10.1002/jcp.25643
  • Arabinogalactan Proteins From Baobab and Acacia Seeds Influence Innate
           Immunity of Human Keratinocytes In Vitro
    • Authors: Abderrakib Zahid; Julie Despres, Magalie Benard, Eric Nguema-Ona, Jerome Leprince, David Vaudry, Christophe Rihouey, Maité Vicré-Gibouin, Azeddine Driouich, Marie-Laure Follet-Gueye
      Abstract: Plant derived arabinogalactan proteins (AGP) were repeatedly confirmed as immunologically as well as dermatologically active compounds. However, little is currently known regarding their potential activity toward skin innate immunity. Here, we extracted and purified AGP from acacia (Acacia senegal) and baobab (Adansonia digitata) seeds to investigate their biological effects on the HaCaT keratinocyte cell line in an in vitro system. While AGP from both sources did not exhibit any cytotoxic effect, AGP from acacia seeds enhanced cell viability. Moreover, real-time quantitative reverse transcription-polymerase chain reaction (qRT-PCR) analysis showed that AGP extracted from both species induced a substantial overexpression of hBD-2, TLR-5, and IL1-α genes. These data suggest that plant AGP, already known to control plant defensive processes, could also modulate skin innate immune responses. J. Cell. Physiol. 9999: 1–11, 2016. © 2016 Wiley Periodicals, Inc.Real-time quantitative reverse transcription-polymerase chain reaction (qRT-PCR) analysis showed that AGP extracted from acacia or baobab seeds induced a substantial overexpression of hBD-2, TLR-5, and IL1-α genes. These data suggest that plant AGP, already known to control plant defensive processes, could also modulate skin innate immune responses.
      PubDate: 2017-04-10T08:21:26.504739-05:
      DOI: 10.1002/jcp.25646
  • Regulation of chondrocyte functions by transient receptor potential cation
           channel V6 in osteoarthritis
    • Authors: Tengfei Song; Jun Ma, Lei Guo, Peng Yang, Xuhui Zhou, Tianwen Ye
      Abstract: Transient receptor potential vanilloid (TRPV) channels function to maintain the dynamic balance of calcium signaling and calcium metabolism in bones. The goal of this study was to determine the potential role of TRPV6 in regulation of chondrocytes. The level of TRPV6 expression was analyzed by western blot in articular cartilage derived from the knee joints of osteoarthritis (OA) rat models and OA patients. Bone structure and osteoarthritic changes in the knee joints of TRPV6 knockout mice were examined using micro-computed and histological analysis at the age of 6 and 12 months old. Furthermore, to investigate the effects of TRPV6 on chondrocyte extracellular matrix secretion, the release of matrix degrading enzymes, cell proliferation, and apoptosis, we decreased and increased TRPV6 expression in chondrocytes with lentiviral constructs encoding shRNA targeting TRPV6 and encoding TRPV6, respectively. The results showed that the level of TRPV6 expression in an OA rat model was markedly down-regulated. TRPV6 knockout mice showed severe osteoarthritis changes, including cartilage fibrillation, eburnation, and loss of proteoglycans. In addition, deficiency of TRPV6 clearly affected chondrocyte function, such as extracellular matrix secretion, the release of matrix degrading enzymes, cell proliferation, and apoptosis. Taken together, our results implicated that TRPV6 channel, as a chondro-protective factor, was involved in the pathogenesis of OA.The level of TRPV6 expression in an OA rat model was markedly down-regulated. TRPV6 knockout mice showed severe osteoarthritic changes. Deficiency of TRPV6 clearly affected chondrocyte function.
      PubDate: 2017-04-10T08:21:22.239837-05:
      DOI: 10.1002/jcp.25770
  • Irx3 and Bmp2 regulate mouse mesenchymal cell chondrogenic differentiation
           in both a Sox9-dependent and -independent manner
    • Authors: Yoshihiro Tamamura; Kenichi Katsube, Hisashi Mera, Maki Itokazu, Shigeyuki Wakitani
      Abstract: Sox9, a master regulator of cartilage development, controls the cell fate decision to differentiate from mesenchymal to chondrogenic cells. In addition, Sox9 regulates the proliferation and differentiation of chondrocytes, as well as the production of cartilage-specific proteoglycans. The existence of Sox9-independent mechanisms in cartilage development remains to be determined. Here, we attempted to identify genes involved in such putative mechanisms via microarray analysis using a mouse chondrogenic cell line, N1511. We first focused on transcription factors that exhibited upregulated expression following Bmp2 treatment, which was not altered by subsequent treatment with Sox9 siRNA. Among these, we selected positive regulators for chondrogenesis and identified Iroquois-related homeobox 3 (Irx3) as one of the candidate genes. Irx3 expression gradually increased with chondrocyte terminal differentiation in a reciprocal manner to Sox9 expression, and promoted the chondrogenic differentiation of mesenchymal cells upon Bmp2 treatment. Furthermore, Irx3 partially rescued impaired chondrogenesis by upregulating the expression of epiphycan and lumican under reduced Sox9 expression. Finally, Irx3 was shown to act in concert with Bmp2 signaling to activate the p38 MAPK pathway, which in turn stimulated Sox9 expression, as well as the expression of epiphycan and lumican in a Sox9-independent manner. These results indicate that Irx3 represents a novel chondrogenic factor of mesenchymal cells, acts synergistically with Bmp2-mediated signaling, and regulates chondrogenesis independent of the transcriptional machinery associated with Sox9-mediated regulation.Irx3 cooperates with Bmp2 to regulate the expression of type II collagen and aggrecan in a Sox9-dependent manner, whereas that of epiphycan and lumican in a Sox9-independent manner in C3H10T1/2 cells.
      PubDate: 2017-04-10T08:21:02.381908-05:
      DOI: 10.1002/jcp.25776
  • Inhibitory effect of blue light emitting diode on migration and invasion
           of cancer cells
    • Authors: Phil-Sun Oh; Hyun-Soo Kim, Eun-Mi Kim, Hyosook Hwang, Hyang Hwa Ryu, SeokTae Lim, Myung-Hee Sohn, Hwan-Jeong Jeong
      Abstract: The aim of this study was to determine the effects and molecular mechanism of blue light emitting diode (LED) in tumor cells. A migration and invasion assay for the metastatic behavior of mouse colon cancer CT-26 and human fibrosarcoma HT-1080 cells was performed. Cancer cell migration-related proteins were identified by obtaining a 2-dimensional gel electrophoresis (2-DE) in total cellular protein profile of blue LED-irradiated cancer cells, followed by matrix-assisted laser desorption/ionization-time of flight (MALDI-TOF) analysis of proteins. Protein levels were examined by immunoblotting. Irradiation with blue LED inhibited CT-26 and HT-1080 cell migration and invasion. The anti-metastatic effects of blue LED irradiation were associated with inhibition of matrix metalloproteinase (MMP)-2 and MMP-9 expression. P38 MAPK phosphorylation was increased in blue LED-irradiated CT-26 and HT-1080 cells, but was inhibited after pretreatment with SB203580, a specific inhibitor of p38 MAPK. Inhibition of p38 MAPK phosphorylation by SB203580 treatment increased number of migratory cancer cells in CT-26 and HT-1080 cells, indicating that blue LED irradiation inhibited cancer cell migration via phosphorylation of p38 MAPK. Additionally blue LED irradiation of mice injected with CT-26 cells expressing luciferase decreased early stage lung metastasis compared to untreated control mice. These results indicate that blue LED irradiation inhibits cancer cell migration and invasion in vitro and in vivo.
      PubDate: 2017-04-10T08:20:43.277853-05:
      DOI: 10.1002/jcp.25805
  • Nuclear Localization of Diacylglycerol Kinase Alpha in K562 Cells Is
           Involved in Cell Cycle Progression
    • Authors: Alessandro Poli; Roberta Fiume, Gianluca Baldanzi, Daniela Capello, Stefano Ratti, Marco Gesi, Lucia Manzoli, Andrea Graziani, Pann-Ghill Suh, Lucio Cocco, Matilde Y. Follo
      Abstract: Phosphatidylinositol (PI) signaling is an essential regulator of cell motility and proliferation. A portion of PI metabolism and signaling takes place in the nuclear compartment of eukaryotic cells, where an array of kinases and phosphatases localize and modulate PI. Among these, Diacylglycerol Kinases (DGKs) are a class of phosphotransferases that phosphorylate diacylglycerol and induce the synthesis of phosphatidic acid. Nuclear DGKalpha modulates cell cycle progression, and its activity or expression can lead to changes in the phosphorylated status of the Retinoblastoma protein, thus, impairing G1/S transition and, subsequently, inducing cell cycle arrest, which is often uncoupled with apoptosis or autophagy induction. Here we report for the first time not only that the DGKalpha isoform is highly expressed in the nuclei of human erythroleukemia cell line K562, but also that its nuclear activity drives K562 cells through the G1/S transition during cell cycle progression. J. Cell. Physiol. 9999: 1–8, 2016. © 2016 Wiley Periodicals, Inc.
      PubDate: 2017-04-10T08:15:55.422872-05:
      DOI: 10.1002/jcp.25642
  • Bone Shaft Revascularization After Marrow Ablation Is Dramatically
           Accelerated in BSP-/- Mice, Along With Faster Hematopoietic Recolonization
    • Authors: Wafa Bouleftour; Renata Neves Granito, Arnaud Vanden-Bossche, Odile Sabido, Bernard Roche, Mireille Thomas, Marie Thérèse Linossier, Jane E. Aubin, Marie-Hélène Lafage-Proust, Laurence Vico, Luc Malaval
      Abstract: The bone organ integrates the activity of bone tissue, bone marrow, and blood vessels and the factors ensuring this coordination remain ill defined. Bone sialoprotein (BSP) is with osteopontin (OPN) a member of the small integrin binding ligand N-linked glycoprotein (SIBLING) family, involved in bone formation, hematopoiesis and angiogenesis. In rodents, bone marrow ablation induces a rapid formation of medullary bone which peaks by ∼8 days (d8) and is blunted in BSP-/- mice. We investigated the coordinate hematopoietic and vascular recolonization of the bone shaft after marrow ablation of 2 month old BSP+/+ and BSP-/- mice. At d3, the ablated area in BSP-/- femurs showed higher vessel density (×4) and vascular volume (×7) than BSP+/+. Vessel numbers in the shaft of ablated BSP+/+ mice reached BSP-/- values only by d8, but with a vascular volume which was twice the value in BSP-/-, reflecting smaller vessel size in ablated mutants. At d6, a much higher number of Lin− (×3) as well as LSK (Lin− IL-7Rα− Sca-1hi c-Kithi, ×2) and hematopoietic stem cells (HSC: Flt3− LSK, ×2) were counted in BSP-/- marrow, indicating a faster recolonization. However, the proportion of LSK and HSC within the Lin− was lower in BSP-/- and more differentiated stages were more abundant, as also observed in unablated bone, suggesting that hematopoietic differentiation is favored in the absence of BSP. Interestingly, unablated BSP-/- femur marrow also contains more blood vessels than BSP+/+, and in both intact and ablated shafts expression of VEGF and OPN are higher, and DMP1 lower in the mutants. In conclusion, bone marrow ablation in BSP-/- mice is followed by a faster vascular and hematopoietic recolonization, along with lower medullary bone formation. Thus, lack of BSP affects the interplay between hematopoiesis, angiogenesis, and osteogenesis, maybe in part through higher expression of VEGF and the angiogenic SIBLING, OPN. J. Cell. Physiol. 9999: 1–10, 2016. © 2016 Wiley Periodicals, Inc.In mice with a knockout of the SIBLING gene, Bone Sialoprotein (BSP-/-) vascular and hematopoietic recolonization are faster than in wild type, despite lower medullary bone formation. This goes with enhanced expression of osteopontin and VEGF, and downregulation of Dentin Matrix Protein 1, another SIBLING shown to inhibit angiogenesis. Lack of BSP thus affects the interplay between hematopoiesis, angiogenesis, and osteogenesis, in part through the mediation of pro- and antiangiogenic factors.
      PubDate: 2017-04-10T08:15:49.54356-05:0
      DOI: 10.1002/jcp.25630
  • Testosterone Rescues the De-Differentiation of Smooth Muscle Cells Through
           Serum Response Factor/Myocardin
    • Authors: Carolina Leimgruber; Amado A. Quintar, Nahuel Peinetti, María V. Scalerandi, Juan P. Nicola, Joseph M. Miano, Cristina A. Maldonado
      Abstract: Prostatic smooth muscle cells (pSMCs) differentiation is a key factor for prostatic homeostasis, with androgens exerting multiple effects on these cells. Here, we demonstrated that the myodifferentiator complex Srf/Myocd is up-regulated by testosterone in a dose-dependent manner in primary cultures of rat pSMCs, which was associated to the increase in Acta2, Cnn1, and Lmod1 expressions. Blocking Srf or Myocd by siRNAs inhibited the myodifferentiator effect of testosterone. While LPS led to a dedifferentiated phenotype in pSMCs, characterized by down-regulation of Srf/Myocd and smooth muscle cell (SMC)-restricted genes, endotoxin treatment on Myocd-overexpressing cells did not result in phenotypic alterations. Testosterone at a physiological dose was able to restore the muscular phenotype by normalizing Srf/Myocd expression in inflammation-induced dedifferentiated pSMCs. Moreover, the androgen reestablished the proliferation rate and IL-6 secretion increased by LPS. These results provide novel evidence regarding the myodifferentiating role of testosterone on SMCs by modulating Srf/Myocd. Thus, androgens preserve prostatic SMC phenotype, which is essential to maintain the normal structure and function of the prostate. J. Cell. Physiol. 9999: 1–12, 2016. © 2016 Wiley Periodicals, Inc.Androgens favor a contractile phenotype on smooth muscle cells that relies on the Myocd/SRF myodifferentiator complex, with this role being critical for cellular homeostasis and pathophysiology.
      PubDate: 2017-04-10T08:15:47.086012-05:
      DOI: 10.1002/jcp.25679
  • Sensitivity of TP53-Mutated Cancer Cells to the Phytoestrogen Genistein Is
           Associated With Direct Inhibition of Plk1 Activity
    • Authors: Sol-Bi Shin; Sang-Uk Woo, Young-Won Chin, Young-Joo Jang, Hyungshin Yim
      Abstract: Polo-like kinase 1 (Plk1), a conserved Ser/Thr mitotic kinase, has been identified as a promising target for anticancer drug development because its overexpression is correlated with malignancy. Here, we found that genistein, an isoflavone, inhibits Plk1 kinase activity directly. Previously the mitotic disturbance phenomenon induced by treatment with genistein was not fully explained by its inhibitory effect on EGFR. In kinase profiling assays, it showed selectivity relative to a panel of kinases, including EGFR. Treatment with genistein induced cell death in a concentration-dependent manner in cancer cells from diverse tissue origins, but not in non-transformed cells such as hTERT-RPE or MCF10A cells. We also observed that genistein tended to be more selective against cancer cells with mutations in the TP53 gene. TP53-depeleted LNCaP and NCI-H460 cells using shRNA targeting human TP53 were more sensitive to cell death by treatment of genistein. Furthermore, genistein induced mitotic arrest by inhibiting Plk1 activity and, consequently, led to mitotic catastrophe and apoptosis. These data suggest that genistein may be a promising anticancer drug candidate due to its inhibitory activity against Plk1 as well as EGFR and effectiveness toward cancer cells, especially those with p53-mutation. J. Cell. Physiol. 9999: 1–11, 2016. © 2016 Wiley Periodicals, Inc.The inhibitory activity of genistein on the Plk1 kinase was relatively selective to its activity against a panel of other kinases, including EGFR. Genistein had a cytotoxic effect in human cancer cells from diverse tissue origins, but not in non-transformed cells. The sensitivity of cancer cells containing mutations in the TP53 gene to genistein tended to be higher when compared with that of cells with wild-type TP53. Since around 50% of solid tumors have mutant TP53, the development of genistein as an anticancer drug would be valuable for treatment of such tumors.
      PubDate: 2017-04-10T08:15:41.562342-05:
      DOI: 10.1002/jcp.25680
  • Epithelial–mesenchymal transition (EMT): A biological process in the
           development, stem cell differentiation, and tumorigenesis
    • Authors: Tong Chen; Yanan You, Hua Jiang, Zack Z. Wang
      Abstract: The lineage transition between epithelium and mesenchyme is a process known as epithelial–mesenchymal transition (EMT), by which polarized epithelial cells lose their adhesion property and obtain mesenchymal cell phenotypes. EMT is a biological process that is often involved in embryogenesis and diseases, such as cancer invasion and metastasis. The EMT and the reverse process, mesenchymal–epithelial transition (MET), also play important roles in stem cell differentiation and de-differentiation (or reprogramming). In this review, we will discuss current research progress of EMT in embryonic development, cellular differentiation and reprogramming, and cancer progression, all of which are representative models for researches of stem cell biology in normal and in diseases. Understanding of EMT and MET may help to identify specific markers to distinguish normal stem cells from cancer stem cells in future.In this review, we discuss current research progress of EMT in embryonic development, cellular differentiation and reprogramming, and cancer progression, all of which are representative models for researches of stem cell biology in normals and in diseases.
      PubDate: 2017-04-10T08:15:29.511501-05:
      DOI: 10.1002/jcp.25797
  • 4th European Seminars in Virology on Oncogenic and Oncolytic Viruses, in
           Bertinoro (Bologna), Italy
    • Authors: Alberto Reale; Lorenzo Messa, Adriana Vitiello, Arianna Loregian, Giorgio Palù
      Abstract: The 4th European Seminars in Virology (EuSeV), which was focused on oncogenic and oncolytic viruses, was held in Bertinoro (Bologna), Italy, from June 10 to 12, 2016. This article summarizes the plenary lectures and aims to illustrate the main topics discussed at 4th EuSeV, which brought together knowledge and expertise in the field of oncogenic and oncolytic viruses from all over the world. The meeting was divided in two parts, “Mechanisms of Viral Oncogenesis” and “Viral Oncolysis and Immunotherapy,” which were both focused on dissecting the complex and multi-factorial interplay between cancer and human viruses and on exploring new anti-cancer strategies. J. Cell. Physiol. 9999: 1–8, 2016. © 2016 Wiley Periodicals, Inc.The 4th European Seminars in Virology (EuSeV), which was focused on oncogenic and oncolytic viruses, was held in Bertinoro (Bologna), Italy, from June 10 to 12, 2016. This article summarizes the plenary lectures and aims to illustrate the main topics discussed at 4th EuSeV, which brought together knowledge and expertise in the field of oncogenic and oncolytic viruses from all over the world. The meeting was divided in two parts, “Mechanisms of Viral Oncogenesis” and “Viral Oncolysis and Immunotherapy,” which were both focused on dissecting the complex and multi-factorial interplay between cancer and human viruses and on exploring new anti-cancer strategies.
      PubDate: 2017-04-10T08:12:26.323088-05:
      DOI: 10.1002/jcp.25692
  • A critical role for adiponectin-mediated development of endometrial
    • Authors: Whasun Lim; Myung Jin Choi, Hyocheol Bae, Fuller W. Bazer, Gwonhwa Song
      Abstract: Adiponectin is one of the adipokines in the collagen superfamily. It is secreted primarily by white adipocytes and influences reproductive processes including ovarian and uterine functions. Adiponectin regulates energy homeostasis, insulin sensitivity, and is anti-inflammatory in various tissues. Its receptors (ADIPOR1 and ADIPOR2) are widely expressed in mammalian tissues, including porcine conceptuses and endometrial during the estrous cycle and peri-implantation period of pregnancy. However, regulatory effects of adiponectin on endometrial epithelial cells are unknown. Therefore, we investigated the effects of parity on expression of ADIPOR1 and ADIPOR2 and the effects of adiponectin in the porcine endometrium during early pregnancy. Results of this study revealed robust expression of ADIPOR1 and ADIPOR2 in uterine luminal (LE) and glandular (GE) epithelia during early pregnancy and expression decreased as with increasing parity. For porcine luminal epithelial (pLE) cells, adiponectin enhanced proliferation, and increased phosphorylation of AKT, P70S6K, S6, ERK1/2, JNK, P38, and P90RSK in a time-dependent manner. Moreover, the abundance of adiponectin-activated signaling molecules were suppressed by pharmacological inhibitors including wortmannin, U0126, SP600125, and SB203580, respectively, in pLE cells. Furthermore, inhibition of each targeted signal transduction molecule influenced proliferation of adiponectin-stimulated pLE cells. In addition, adiponectin inhibited tunicamycin-induced endoplasmic reticulum (ER)-stress through effects on ER stress regulated proteins in pLE cells. Collectively, these results suggest that adiponectin affects development of porcine uterine epithelia and reproductive performance through modulation of PI3K/AKT and MAPK cell signaling pathways.Adiponectin enhance uterine receptivity to implantation and placentation through PI3K/AKT and MAPK pathways for development of the porcine uterus and reproductive performance.
      PubDate: 2017-04-10T08:10:49.228743-05:
      DOI: 10.1002/jcp.25768
  • KCNQ1 variants associate with hypertension in type 2 diabetes and affect
           smooth muscle contractility in vitro
    • Authors: Kuo-Chin Huang; Te-Mao Li, Xiang Liu, Jin-Hua Chen, Wen-Kuei Chien, Yi-Tzone Shiao, Hsinyi Tsang, Ting-Hsu Lin, Chiu-Chu Liao, Shao-Mei Huang, Ju-Pi Li, Cheng-Wen Lin, Jung-Chun Lin, Chih-Chien Lin, Chih-Ho Lai, Chi-Fung Cheng, Wen-Miin Liang, Chien-Hui Hung, Ching-Chu Chen, Ying-Ju Lin, Fuu-Jen Tsai
      Abstract: KCNQ1 encodes a potassium voltage-gated channel and represents a susceptibility locus for type 2 diabetes mellitus (T2DM). Here, we explored the association between KCNQ1 polymorphisms and hypertension risk in individuals with T2DM, as well as the role of KCNQ1 in vascular smooth muscle cell contraction in vitro. To investigate the relationship between KCNQ1 and the risk of developing hypertension in patients with T2DM, we divided the T2DM cohort into hypertension (n = 452) and non-hypertension (n = 541) groups. The Mann–Whitney U test, chi-square test, and multivariate regression analyses were used to assess the clinical characteristics and genotypic frequencies. In vitro studies utilized the rat aortic smooth muscle A10 cell line. Patients in the hypertension group were significantly older at the time of enrollment and had higher levels of body mass index, waist-to-hip ratio, and triglyceride than those in the non-hypertension group. The KCNQ1 rs3864884 and rs12576239 genetic variants were associated with hypertension in T2DM. KCNQ1 expression was lower in the individuals with the CC versus the CT and TT genotypes. Smooth muscle cell contractility was inhibited by treatment with a KCNQ1 inhibitor. These results suggest that KCNQ1 might be associated with hypertension in individuals with T2DM.The KCNQ1 rs3864884 and rs12576239 genetic variants were associated with hypertension in T2DM. Smooth muscle cell contractility was inhibited by treatment with a KCNQ1 inhibitor. KCNQ1 might be associated with hypertension in individuals with T2DM.
      PubDate: 2017-04-10T08:06:52.00554-05:0
      DOI: 10.1002/jcp.25775
  • SirT3 and p53 Deacetylation in Aging and Cancer
    • Authors: Jijun Chen; Aiqin Wang, Qingqi Chen
      Abstract: The mammalian Sirtuins are a family of highly conserved nicotinamide adenine dinucleotide (NAD+)-dependent histone deacetylase. The mammalian Sirtuins family has identified seven different types of sirtuin. Sirtuins 3(SirT3) is localized to mitochondria, and is a multiple functional protein deacylase through deacetylating a variety of substrates. Cellular senescence represents a permanent withdraw from cell cycle in response to diverse stress; it is controlled by the p53 and retinoblastoma protein (RB) tumor suppressors, and constitutes a potent anticancer mechanisms. p53 can be deacetylated by a protein complex containing histone deacetylases (HDAC1). There is possibility that SirT3 may also deacetylate p53. We introduce the recent research on the SirT3-p53 interplay directly and indirectly, and discuss the significance of p53 deacetylation by SirT3 in the aging and cancer. J. Cell. Physiol. 9999: 1–4, 2016. © 2016 Wiley Periodicals, Inc.Cellular senescence is controlled by the p53 and retinoblastoma protein (RB) tumor suppressors, and constitutes a potent anticancer mechanisms. p53 may be deacetylated by Sirtuins 3.
      PubDate: 2017-04-10T08:06:30.349039-05:
      DOI: 10.1002/jcp.25669
  • Let-7i-Induced Atg4B Suppression Is Essential for Autophagy of Placental
           Trophoblast in Preeclampsia
    • Authors: Yinyan Xu; Xinyan Huang, Juan Xie, Yanni Chen, Jing Fu, Li Wang
      Abstract: Autophagy, identified as type II programmed cell death, has already been known to be involved in the pathophysiology of preeclampsia (PE), which is a gestational disease with high morbidity. The present study aims to investigate the functional role of let-7i, a miRNA, in trophoblastic autophagy. Placental tissue used in this study was collected from patients with severe preeclampsia (SPE) or normal pregnant women. A decreased level of let-7i was found in placenta of SPE. In addition, autophagic vacuoles were observed in SPE and the expression of microtubule associated protein 1 light chain 3 (LC3) II/I was elevated. In vitro, let-7i mimics suppressed the autophagic activities in human HTR-8/SVneo trophoblast cell line (HTR-8) and human placental choriocarcinoma cell line JEG-3, whereas let-7i inhibitor enhanced the activities. As a potential target of let-7i, autophagy-related 4B cysteine peptidase (Atg4B) had an increased expression level in SPE. As expected, the increased expression of Atg4B was negatively regulated by let-7i using dual luciferase reporter assay. Furthermore, these trophoblast-like cells transfected with the let-7i mimic or inhibitors resulted in a significant change of Atg4B in both mRNA and protein level. More importantly, Atg4B overexpression could partly reverse let-7i mimic-reduced LC3II/I levels; whereas Atg4B silencing partly attenuated let-7i inhibitor-induced the level of LC3II/I expression. Taken together, these findings suggest that let-7i is able to regulate autophagic activity via regulating Atg4B expression, which might contribute to the pathogenesis of PE. J. Cell. Physiol. 9999: 1–9, 2016. © 2016 Wiley Periodicals, Inc.This study provided insight into the role of let-7i in the pathophysiology of PE. Our study demonstrates for the first time that a novel let-7i-Atg4B-autophagy signaling pathway plays a key role in PE. This finding suggests that miRNAs are key molecules involved in autophagic activity, and might be regard as a new therapeutic target against PE.
      PubDate: 2017-04-10T08:05:39.959857-05:
      DOI: 10.1002/jcp.25661
  • Cre Recombinase Strains Used for the Study of Adipose Tissues and
           Adipocyte Progenitors
    • Authors: Jiaqi Liu; Ziye Xu, Weiche Wu, Yizhen Wang, Tizhong Shan
      Abstract: Adipose tissues play important roles in whole body energy homeostasis and lifer span. Understanding the mechanisms of controlling adipose tissues development is significant for providing useful information to treat the worldwide epidemic of obesity and its associated metabolic diseases. Several different Cre transgenes have been generated and used for determining the origin of adipose tissues and the function of individual gene in regulating adipose growth and development. Here, we mainly review and discuss the efficiency and specific of those Cre recombinase mouse strains used for the study of adipose tissues and adipocyte progenitors. J. Cell. Physiol. 9999: 1–6, 2016. © 2016 Wiley Periodicals, Inc.We mainly review and discuss the efficiency and specific of those Cre recombinase mouse strains used for determining the origin of adipose tissues and the function of individual gene in regulating adipose growth and development.
      PubDate: 2017-04-10T08:05:29.836113-05:
      DOI: 10.1002/jcp.25675
  • Overexpression of Large-Conductance Calcium-Activated Potassium Channels
           in Human Glioblastoma Stem-Like Cells and Their Role in Cell Migration
    • Authors: Paolo Rosa; Luigi Sforna, Silvia Carlomagno, Giorgio Mangino, Massimo Miscusi, Mauro Pessia, Fabio Franciolini, Antonella Calogero, Luigi Catacuzzeno
      Abstract: Glioblastomas (GBMs) are brain tumors characterized by diffuse invasion of cancer cells into the healthy brain parenchyma, and establishment of secondary foci. GBM cells abundantly express large-conductance, calcium-activated potassium (BK) channels that are thought to promote cell invasion. Recent evidence suggests that the GBM high invasive potential mainly originates from a pool of stem-like cells, but the expression and function of BK channels in this cell subpopulation have not been studied. We investigated the expression of BK channels in GBM stem-like cells using electrophysiological and immunochemical techniques, and assessed their involvement in the migratory process of this important cell subpopulation. In U87-MG cells, BK channel expression and function were markedly upregulated by growth conditions that enriched the culture in GBM stem-like cells (U87-NS). Cytofluorimetric analysis further confirmed the appearance of a cell subpopulation that co-expressed high levels of BK channels and CD133, as well as other stem cell markers. A similar association was also found in cells derived from freshly resected GBM biopsies. Finally, transwell migration tests showed that U87-NS cells migration was much more sensitive to BK channel block than U87-MG cells. Our data show that BK channels are highly expressed in GBM stem-like cells, and participate to their high migratory activity. J. Cell. Physiol. 9999: 1–11, 2016. © 2016 Wiley Periodicals, Inc.We investigated the expression of BK channels in GBM stem-like cells using electrophysiological and immunochemical techniques, and assessed their involvement in the migratory process of this important cell subpopulation. In U87-MG cells, BK channel expression and function were markedly upregulated in GBM stem-like cells expressing CD133, as well as other stem cell markers. In addition, transwell migration tests showed that GBM stem-like cell migration was very sensitive to BK channel block.
      PubDate: 2017-04-10T07:26:09.346707-05:
      DOI: 10.1002/jcp.25592
  • The Role of Calcium-Sensing Receptors in Endothelin-1-Dependent Effects on
           Adult Rat Ventricular Cardiomyocytes: Possible Contribution to Adaptive
           Myocardial Hypertrophy
    • Authors: Elena Dyukova; Rolf Schreckenberg, Christoph Arens, Guzel Sitdikova, Klaus-Dieter Schlüter
      Abstract: Nitric oxide (NO)-deficiency as it occurs during endothelial dysfunction activates the endothelin-1 (ET-1) system and increases the expression of receptor activity modifying protein (RAMP)-1 that acts as a chaperon for calcium-sensing receptors (CaR) that have recently been identified to improve cardiac function. Here, we hypothesized that ET-1 increases the cardiac expression of CaR and thereby induces an adaptive type of hypertrophy. Expressions of RAMP-1, endothelin receptors, and CaR were analyzed by RT-PCR in left ventricular tissues of L-NAME-treated rats. Effects of ET-1 on CaR expression and cell function (load free cell shortening) were analyzed in adult rat ventricular cardiomyocytes. siRNA directed against CaR and RAMP-1 was used to investigate a causal relationship. PD142893 and BQ788 were used to dissect the contribution of ETB1, ETB2, and ETA receptors. Non-specific NO synthase inhibition with L-Nitro arginine methyl ester (L-NAME) caused a cardiac upregulation of ETB receptors and CaR suggesting a paracrine effect of ET-1 on cardiomyocytes. Indeed, ET-1 induced the expression of CaR in cultured cardiomyocytes. Under these conditions, cardiomyocytes increased cell size (hypertrophy) but maintained normal function. Inhibition of ETA and ETB1 receptors led to ET-1-dependent reduction in cell shortening and attenuated up-regulation of CaR. Down-regulation of RAMP-1 reduced CaR responsiveness. In conclusion, ET-1 causes an adaptive type of hypertrophy by up-regulation of CaR in cardiomyocytes via ETA and/or ETB1 receptors. J. Cell. Physiol. 9999: 1–11, 2016. © 2016 Wiley Periodicals, Inc.Current manuscript is the first report indicating the ET-1-dependent upregulation of CaR receptor in cardiomyocytes and it shows that this upregulation allows cardiac cells to maintain proper function. Second new finding indicates involvement of ETA and /or ETB1 receptors in adaptive hypertrophy in cardiomyocytes, whereas ETB2 receptors activation leads to mal-adaptive effect. Finally, chaperone activity of RAMP1 is required for the functional coupling of CaR in cardiomyocytes.
      PubDate: 2017-04-10T07:26:03.913152-05:
      DOI: 10.1002/jcp.25612
  • Cardiotrophin-1 Regulates Adipokine Production in 3T3-L1 Adipocytes and
           Adipose Tissue From Obese Mice
    • Authors: Miguel López-Yoldi; Beatriz Marcos-Gomez, María Asunción Romero-Lozano, Neira Sáinz, Jesús Prieto, Jose Alfredo Martínez, Matilde Bustos, Maria J. Moreno-Aliaga
      Abstract: Cardiotrophin-1 (CT-1) belongs to the IL-6 family of cytokines. Previous studies of our group revealed that CT-1 is a key regulator of glucose and lipid metabolism. The aim of the present study was to analyze the in vitro and in vivo effects of CT-1 on the production of several adipokines involved in body weight regulation, nutrient metabolism, and inflammation. For this purpose, 3T3-L1 adipocytes were incubated with recombinant protein CT-1 (rCT-1) (1–40 ng/ml) for 1 and 18 h. Moreover, the acute effects of rCT-1 administration (0.2 mg/kg, i.v.) for 30 min and 3 h on adipokines levels were also evaluated in high-fat fed obese mice. In 3T3-L1 adipocytes, rCT-1 treatment downregulated the expression and secretion of leptin, resistin, and visfatin. However, rCT-1 significantly stimulated apelin mRNA and secretion. rCT-1 (18 h) also promoted the activation by phosphorylation of AKT, ERK 1/2, and STAT3. Interestingly, pre-treatment with the PI3K inhibitor LY294002 reversed the stimulatory effects of rCT-1 on apelin expression, suggesting that this pathway could be mediating the effects of rCT-1 on apelin production. In contrast, acute administration of rCT-1 (30 min and 3 h) to diet-induced obese mice downregulated leptin and resistin, without significantly modifying apelin or visfatin mRNA in adipose tissue. Furthermore, CT-1 null mice exhibited altered expression of adipokines in adipose tissue. The present study demonstrates that rCT-1 modulates the production of adipokines in vitro and in vivo, suggesting that the regulation of the secretory function of adipocytes could be involved in the metabolic actions of this cytokine. J. Cell. Physiol. 9999: 1–9, 2016. © 2016 Wiley Periodicals, Inc.The current study demonstrates that CT-1 is capable of regulating the secretory pattern of adipokines by adipocytes, decreasing the production of pro-inflammatory adipokines, such as leptin, resistin, and visfatin, while stimulating the secretion of apelin. These suggest that the beneficial actions of CT-1 on glucose and lipid metabolism could be also related to its ability to control adipokine secretion and therefore the cross-talk between adipose tissue and other key metabolic organs.
      PubDate: 2017-04-10T07:20:40.955197-05:
      DOI: 10.1002/jcp.25590
  • Anti-Atherosclerotic Effects of Vitamins D and E in Suppression of
    • Authors: Bahman Rashidi; Zahra Hoseini, Amirhossein Sahebkar, Hamed Mirzaei
      Abstract: Atherosclerosis is a progressive and multifactorial disease which occurs under the influence of various risk factors including endothelial dysfunction (ED), oxidative stress, and low-density lipoprotein (LDL) oxidation. In contract to the initial hypotheses on the usefulness of vitamin E supplementation for cardiovascular disease prevention, large outcome trials showed consumption of vitamin E has no obvious effect on cardiovascular disease and, in some cases, it may even increase the rate of mortality. This seemingly unexpected finding may be due to the opposite effects of vitamin E compounds. Vitamin E is a group of compounds which have different and even opposing effects, yet in most of the studies, the exact consumed component of vitamin E is not determined. It appears that the combined consumption of gamma-tocopherol, vitamin C, D, and tetrahydrobiopterin (BH4) may be extremely effective in both preventing atherogenesis and suppressing plaque development. In this regard, one of main issues is effect of vitamins E and D deficiency on microRNAs network in atherosclerosis. Various studies have indicated that miRNAs have key roles in atherosclerosis pathogenesis. The deficiency of vitamins E and D could provide a deregulation for miRNAs network and these events could lead to progression of atherosclerosis. Here, we highlighted a variety of mechanisms involve in the progression of atherosclerosis and effects of vitamins D and E on these mechanisms. Moreover, we summarized miRNAs involve in atherosclerosis and their regulation by vitamins E and D deficiency. J. Cell. Physiol. 9999: 1–9, 2016. © 2016 Wiley Periodicals, Inc.
      PubDate: 2017-04-10T07:15:57.272442-05:
      DOI: 10.1002/jcp.25738
  • The Sabotaging Role of Myeloid Cells in Anti-Angiogenic Therapy:
           Coordination of Angiogenesis and Immune Suppression by Hypoxia
    • Authors: Chunyan Li; Tao Liu, Alexandr V. Bazhin, Yuhui Yang
      Abstract: Tumor angiogenesis has become a promising target for anti-tumor therapy. Unfortunately, the somewhat inevitable occurrence of resistance has limited the efficacy of anti-angiogenic therapy. In addition to their well-established role in immune suppression, bone marrow-derived myeloid cells actively contribute to tumor angiogenesis. More importantly, myeloid cells constitute one of the major mechanisms of resistance to angiogenesis inhibition. As the most pervasive feature in tumor microenvironment, hypoxia is able to initiate both pro-angiogenic and immunosuppressive capacities of myeloid cells. Tumor adapts to hypoxic stress primarily through signaling mediated by hypoxic inducible factors (HIFs) and consequently utilizes hypoxia to its own advantage. In this regard, hypoxia orchestrates both angiogenesis and immune evasion to support tumor growth. In this article, we will review available information on the sabotaging role of myeloid cells in anti-angiogenic therapy. We will also discuss how hypoxia coordinates the dual-role cellular and molecular participants in microenvironment to maximize the efficiency of angiogenesis and immunosuppression to promote tumor progression. J. Cell. Physiol. 9999: 1–11, 2016. © 2016 Wiley Periodicals, Inc.The signaling mediated by hypoxia-inducible factors (HIFs) provides tumor cells and stromal cells with adaptation to the hypoxic microenvironment. In this regards, hypoxia and hypoxia-driven signaling orchestrate the cellular and molecular participants in angiogenesis and immune suppression, maximizing the efficiency of both programs.
      PubDate: 2017-04-10T07:10:43.351525-05:
      DOI: 10.1002/jcp.25726
  • C-Met as a potential target for the treatment of gastrointestinal cancer:
           Current status and future perspectives
    • Authors: Afsane Bahrami; Soodabeh Shahidsales, Majid Khazaei, Majid Ghayour-Mobarhan, Mina Maftouh, Seyed Mahdi Hassanian, Amir Avan
      Abstract: Aberrant activation of the HGF/c-Met signalling pathways is shown to be related with cell proliferation, progression, metastasis, and worse prognosis in several tumor types, including gastrointestinal cancers, suggesting its value as a stimulating-target for cancer-therapy. Several approaches have been developed for targeting HGF and/or c-Met, and one of them, crizotinib (dual c-Met/ALK inhibitor), is recently been approved by FDA for lung-cancers with ALK-rearrangement. The main aim of current review is to give an overview on the role of c-Met/HGF pathway in gastrointestinal cancer, in preclinical and clinical trials. Although several important matters is still remained to be elucidated on the molecular pathways underlying the antitumor effects of this therapy in gastrointestinal-cancers. Further investigations are warranted to recognize the main determinants of the activity of c-Met inhibitors, for parallel targeting signalling pathway associated/activated via MET/HGF pathway or in response to the cell resistance to anti-c-Met agents. Additionally, identification of patients that might benefit from therapy could help to increase the selectivity and efficacy of the therapy.
      PubDate: 2017-04-10T07:07:21.969917-05:
      DOI: 10.1002/jcp.25794
  • Isolation, Characterization, and Establishment of Spontaneously
           Immortalized Cell Line HRPE-2S With Stem Cell Properties.
    • Authors: Hoda Shams Najafabadi; Zahra-soheila Soheili, Shahram Samiei, Hamid Ahmadieh, Ehsan Ranaei Pirmardan, Maryam Masoumi
      Abstract: The retinal pigment epithelium is a monolayer of highly specialized pigmented cells located between the neural retina and the Bruch's membrane of the choroid. RPE cells play a crucial role in the maintenance and function of the underlying photoreceptors. This study introduces a spontaneously arising human retinal pigment epithelial cell line, HRPE-2S, which was isolated from primary RPE cell culture of 2 days old male donor. We characterized morphology and functional properties of the new cell line. The immortalized cell line was maintained in culture for more than 70 passages and 240 divisions. The average doubling time of the cells was approximately 22 h and got freezed at 26th passage. The cell line expressed RPE-specific markers RPE65 and cell junction protein ZO1 as an epithelial cell marker. It also expressed CHX10, PAX6, Nestin, SOX2 as stem and retinal progenitor cell markers. Ki67 as a marker of cell proliferation was expressed in all HRPE-2S cells. It represented typical epithelial cobblestone morphology and did not phenotypically change through several passages. Stem cell-like aggregations (neurospheres) were observed in SEM microscopy. The cells represented high mitotic index. They could be viable under hypoxic conditions and serum deprivation. According to functional studies, the cell line exhibited stem cell-like behaviors with particular emphasis on its self-renewal capacity. LDH isoenzymes expression pattern confirmed the same cellular source for both of the HRPE-2S cells and primary RPE cells. Characteristics of HRPE-2S cells promise it as an in vitro model for RPE stem cell-based researches. J. Cell. Physiol. 9999: 1–15, 2016. © 2016 Wiley Periodicals, Inc.This research introduced a novel spontaneously immortalized retinal pigment epithelial cell line, HRPE-2S, as a tool for researches in RPE stem cell based biology.
      PubDate: 2017-04-10T07:01:16.534238-05:
      DOI: 10.1002/jcp.25729
  • Stimulatory actions of a novel thiourea derivative on large-conductance,
           calcium-activated potassium channels
    • Authors: Sheng-Nan Wu; Jyh-Haur Chern, Santai Shen, Hwei-Hisen Chen, Ying-Ting Hsu, Chih-Chin Lee, Ming-Huan Chan, Ming-Chi Lai, Feng-Shiun Shie
      Abstract: In this study, we examine whether an anti-inflammatory thiourea derivative, compound #326, actions on ion channels. The effects of compound #326 on Ca2+-activated K+ channels were evaluated by patch-clamp recordings obtained in cell-attached, inside-out or whole-cell configuration. In pituitary GH3 cells, compound #326 increased the amplitude of Ca2+-activated K+ currents (IK(Ca)) with an EC50 value of 11.6 μM, which was reversed by verruculogen, but not tolbutamide or TRAM-34. Under inside-out configuration, a bath application of compound #326 raised the probability of large-conductance Ca2+-activated K+ (BKCa) channels. The activation curve of BKCa channels was shifted to less depolarised potential with no modification of the gating charge of the curve; consequently, the difference of free energy was reduced in the presence of this compound. Compound #326-stimulated activity of BKCa channels is explained by a shortening of mean closed time, despite its inability to alter single-channel conductance. Neither delayed-rectifier nor erg-mediated K+ currents was modified. Compound #326 decreased the peak amplitude of voltage-gated Na+ current with no clear change in the overall current–voltage relationship of this current. In HEK293T cells expressing α-hSlo, compound #326 enhanced BKCa channels effectively. Intriguingly, the inhibitory actions of compound #326 on interleukin 1β in lipopolysaccharide-activated microglia were significantly reversed by verruculogen, whereas BKCa channel inhibitors suppressed the expressions of inducible nitric oxide synthase. The BKCa channels could be an important target for compound #326 if similar in vivo results occur, and the multi-functionality of BKCa channels in modulating microglial immunity merit further investigation.Compound #326-stimulated activity of BKCa channels is explained by a shortening of mean closed time, despite its inability to alter single-channel conductance. Intriguingly, the inhibitory actions of compound #326 on interleukin 1β in lipopolysaccharide-activated microglia were significantly reversed by verruculogen, whereas BKCa channel inhibitors suppressed the expressions of inducible nitric oxide synthase.
      PubDate: 2017-04-10T07:00:50.629408-05:
      DOI: 10.1002/jcp.25788
  • TNF-α promotes osteoclastogenesis through JNK signaling-dependent
           induction of Semaphorin3D expression in estrogen-deficiency induced
    • Authors: Chenglin Sang; Jiefeng Zhang, Yongxian Zhang, Fangjing Chen, Xuecheng Cao, Lei Guo
      Abstract: Tumor necrosis factor α (TNF-α)-induced osteoclast formation have been demonstrated to play an important role in the pathogenesis of estrogen deficiency-mediated bone loss, but the exact mechanisms by which TNF-α enhanced osteoclast differentiation were not fully elucidated. The class III semaphorins members were critical to regulate bone homeostasis. Here, we identified a novel mechanism whereby TNF-α increasing Semaphorin3D expression contributes to estrogen deficiency-induced osteoporosis. In this study, we found that Semaphorin3D expression was upregulated by TNF-α during the process of RANKL-induced osteoclast differentiation. Inhibition of Semaphorin3D in pre-osteoclasts could attenuate the stimulatory effects of TNF-α on osteoclast proliferation and differentiation. Mechanistically, blocking of the Jun N-terminal kinase (JNK) signaling markedly rescued TNF-α-induced Semaphorin3D expression, suggesting that JNK signaling was involved in the regulation of Semaphorin3D expression by TNF-α. In addition, silencing of Semaphorin3D in vivo could alleviate estrogen deficiency-induced osteoporosis. Our results revealed a novel function for Semaphorin3D and suggested that increased Semaphorin3D may contribute to enhanced bone loss by increased TNF-α in estrogen deficiency-induced osteoporosis. Thus, Semaphorin3D may provide a potential therapeutic target for the treatment of estrogen-deficiency induced osteoporosis.We found that TNF-a could increase Sema3D expression in RANKL-induced osteoclast formation. Upregulation of Sema3D expression was required for TNF-a-mediated osteoclast proliferation and differentiation. JNK signaling was involved in the regulation of Sema3D expression by TNF-a.
      PubDate: 2017-04-10T06:56:16.414109-05:
      DOI: 10.1002/jcp.25784
  • Lkb1 regulation of skeletal muscle development, metabolism and muscle
           progenitor cell homeostasis
    • Authors: Tizhong Shan; Ziye Xu, Jiaqi Liu, Weiche Wu, Yizhen Wang
      Abstract: Liver kinase B1 (Lkb1), also named as Serine/Threonine protein kinase 11 (STK11), is a serine/threonine kinase that plays crucial roles in various cellular processes including cell survival, cell division, cellular polarity, cell growth, cell differentiation, and cell metabolism. In metabolic tissues, Lkb1 regulates glucose homeostasis and energy metabolism through phosphorylating and activating the AMPK subfamily proteins. In skeletal muscle, Lkb1 affects muscle development and postnatal growth, lipid and fatty acid oxidation, glucose metabolism, and insulin sensitivity. Recently, the regulatory roles of Lkb1 in regulating division, self-renew, proliferation, and differentiation of skeletal muscle progenitor cells have been reported. In this review, we discuss the roles of Lkb1 in regulating skeletal muscle progenitor cell homeostasis and skeletal muscle development and metabolism.In skeletal muscle, Lkb1 affects muscle development and postnatal growth, lipid, and fatty acid oxidation, glucose metabolism and insulin sensitivity. In skeletal muscle progenitor cells, Lkb1 regulates cell division, self-renew, proliferation, and differentiation. Here, we review the role of Lkb1 in skeletal muscle development, metabolism and muscle progenitor cell homeostasis.
      PubDate: 2017-04-10T06:55:39.58097-05:0
      DOI: 10.1002/jcp.25786
  • Chick derived induced pluripotent stem cells by the poly-cistronic
           transposon with enhanced transcriptional activity
    • Authors: Masafumi Katayama; Takashi Hirayama, Tetsuya Tani, Katsuhiko Nishimori, Manabu Onuma, Tomokazu Fukuda
      Abstract: Induced pluripotent stem (iPS) cell technology lead terminally differentiated cells into the pluripotent stem cells through the expression of defined reprogramming factors. Although iPS cells have been established in a number of mammalian species, including mouse, human, and monkey, studies on iPS cells in avian species are still very limited. To establish chick iPS cells, six factors were used within the poly-cistronic reprogramming vector (PB-R6F), containing M3O (MyoD derived transactivation domain fused with Oct3/4), Sox2, Klf4, c-Myc, Lin28, and Nanog. The PB-R6F derived iPS cells were alkaline-phosphatase and SSEA-1 positive, which are markers of pluripotency. Elevated levels of endogenous Oct3/4 and Nanog genes were detected in the established iPS cells, suggesting the activation of the FGF signaling pathway is critical for the pluripotent status. Histological analysis of teratoma revealed that the established chick iPS cells have differentiation ability into three-germ-layer derived tissues. This is the first report of establishment of avian derived iPS cells with a single poly-cistronic transposon based expression system. The establishment of avian derived iPS cells could contribute to the genetic conservation and modification of avian species. This article is protected by copyright. All rights reserved
      PubDate: 2017-04-07T08:06:50.802967-05:
      DOI: 10.1002/jcp.25947
  • APN/CD13 Is Over-expressed by Psoriatic Fibroblasts and Is Modulated by
           CGRP and IL-4 But not by Retinoic Acid Treatment
    • Authors: Pascale Gerbaud; Jean Guibourdenche, Rafika Jarray, Marc Conti, Patricia Palmic, Stéphanie Leclerc-Mercier, Julie Bruneau, Olivier Hermine, Yves Lepelletier, Françoise Raynaud
      Abstract: Psoriasis vulgaris is a common skin inflammatory disease characterized by recurrent flare episodes associated with scaly well-demarcated skin plaques. Skin biopsies from psoriatic patients with high PASI score (22.67 ± 8.67) and from HD were used to study APN/CD13. APN/CD13 is over-expressed in LP and nLP compare to HD skins and fibroblasts. This over-expression is positively correlated with specific enzymatic activity enhancement. However, discrepancies between APN/CD13 expression in LP and nLP prompt us to focus our study on APN/CD13 modulation. Calcitonin Gene Related Peptide (CGRP), a neuropeptide, positively modulated expression and activity of APN/CD13. CGRP consistently induced IL4 secretion, which is also involved in the increase of APN/CD13 expression and activity, which is significantly reversed using IL-4 blocking antibody. Surprisingly, retinoic acid altered the APN/CD13 enzymatic activity only in nLP fibroblasts without modification of APN/CD13 expression. APN/CD13 is over-expressed on psoriatic fibroblasts and exerted high level of activity compare to HD fibroblasts. Taken together, several factors such as CGRP and IL-4 acted on positive regulation of APN/CD13 expression and activity. This study highlighted the interest of APN/CD13 as a new potential target, which should be investigated in psoriasis. This article is protected by copyright. All rights reserved
      PubDate: 2017-04-07T05:37:00.604059-05:
      DOI: 10.1002/jcp.25941
  • Stimulation of Oral Fibroblast Chemokine Receptors Identifies CCR3 and
           CCR4 as Potential Wound Healing Targets
    • Authors: Jeroen Kees Buskermolen; Sanne Roffel, Susan Gibbs
      Abstract: The focus of this study was to determine which chemokine receptors are present on oral fibroblasts and whether these receptors influence proliferation, migration and/or the release of wound healing mediators. This information may provide insight into the superior wound healing characteristics of the oral mucosa. The gingiva fibroblasts expressed 12 different chemokine receptors (CCR3, CCR4, CCR6, CCR9, CCR10, CXCR1, CXCR2, CXCR4, CXCR5, CXCR7, CX3CR1 and XCR1), as analyzed by flow cytometry. Fourteen corresponding chemokines (CCL5, CCL15, CCL20, CCL22, CCL25, CCL27, CCL28, CXCL1, CXCL8, CXCL11, CXCL12, CXCL13, CX3CL1 and XCL1) were used to study the activation of these receptors on gingiva fibroblasts. Twelve of these fourteen chemokines stimulated gingiva fibroblast migration (all except for CXCL8 and CXCL12). Five of the chemokines stimulated proliferation (CCL5/CCR3, CCL15/CCR3, CCL22/CCR4, CCL28/CCR3/CCR10 and XCL1/XCR1). Furthermore, CCL28/CCR3/CCR10 and CCL22/CCR4 stimulation increased IL-6 secretion and CCL28/CCR3/CCR10 together with CCL27/CCR10 upregulated HGF secretion. Moreover TIMP-1 secretion was reduced by CCL15/CCR3. In conclusion, this in-vitro study identifies chemokine receptor-ligand pairs which may be used in future targeted wound healing strategies. In particular, we identified the chemokine receptors CCR3 and CCR4, and the mucosa specific chemokine CCL28, as having an predominant role in oral wound healing by increasing human gingiva fibroblast proliferation, migration and the secretion of IL-6 and HGF and reducing the secretion of TIMP-1. This article is protected by copyright. All rights reserved
      PubDate: 2017-04-07T05:36:55.242489-05:
      DOI: 10.1002/jcp.25946
  • HSF1 phosphorylation by ERK/GSK3 suppresses RNF126 to sustain IGF-IIR
           expression for hypertension-induced cardiomyocyte hypertrophy
    • Authors: Chih-Yang Huang; Fa-Lun Lee, Shu-Fen Peng, Kuan-Ho Lin, Ray-Jade Chen, Tsung-Jung Ho, Fu-Jen Tsai, V. Vijaya Padma, Wei-Wen Kuo, Chih-Yang Huang
      Abstract: Hypertension-induced cardiac hypertrophy and apoptosis are major characteristics of early-stage heart failure (HF). Inhibition of extracellular signal-regulated kinases (ERK) efficaciously suppressed angiotensin II (ANG II)-induced cardiomyocyte hypertrophy and apoptosis by blocking insulin-like growth factor II receptor (IGF-IIR) signaling. However, the detailed mechanism by which ANG II induces ERK-mediated IGF-IIR signaling remains elusive. Here, we found that ANG II activated ERK to upregulate IGF-IIR expression via the angiotensin II type I receptor (AT1R). ERK activation subsequently phosphorylates HSF1 at serine 307, leading to a secondary phosphorylation by glycogen synthase kinase III (GSK3) at serine 303. Moreover, we found that ANG II mediated ERK/GSK3-induced IGF-IIR protein stability by downregulating the E3 ubiquitin ligase of IGF-IIR RING finger protein CXXVI (RNF126). The expression of RNF126 decreased following ANG II-induced HSF1S303 phosphorylation, resulting in IGF-IIR protein stability and increased cardiomyocyte injury. Inhibition of GSK3 significantly alleviated ANG II-induced cardiac hypertrophy in vivo and in vitro. Taken together, these results suggest that HSF1 phosphorylation stabilizes IGF-IIR protein stability by downregulating RNF126 during cardiac hypertrophy. ANG II activates ERK/GSK3 to phosphorylate HSF1, resulting in RNF126 degradation, which stabilizes IGF-IIR protein expression and eventually results in cardiac hypertrophy. HSF1 could be a valuable therapeutic target for cardiac diseases among hypertensive patients. This article is protected by copyright. All rights reserved
      PubDate: 2017-04-06T10:30:45.112758-05:
      DOI: 10.1002/jcp.25945
  • Role of adenosine signaling in the pathogenesis of breast cancer
    • Authors: Amirhossein Bahreyni; Seyed Sattar Samani, Farzad Rahmani, Reihane Behnam-Rassouli, Majid Khazaei, Mikhail Ryzhikov, Mohammad Reza Parizadeh, Amir Avan, Seyed Mahdi Hassanian
      Abstract: The plasma level of adenosine increases under ischemic and inflamed conditions in tumor microenvironment. Adenosine elicits a range of signaling pathways in tumors, resulting in either inhibition or enhancement of tumor growth depending upon different subtypes of adenosine receptors activation and type of cancer. Metabolism of adenosine-5′-triphosphate (ATP) and its derivatives including adenosine is dysregulated in the breast tumor microenvironment, supporting the role of this metabolite in the pathogenesis of breast cancer. Adenosine regulates inflammation, apoptosis, cell proliferation and metastasis in breast cancer cells. This review summarizes the role of adenosine in the pathogenesis of breast cancer for a better understanding and hence a better management of this disease. This article is protected by copyright. All rights reserved
      PubDate: 2017-04-06T10:30:39.100635-05:
      DOI: 10.1002/jcp.25944
  • Polydatin Impairs Mitochondria Fitness and Ameliorates Podocyte Injury by
           Suppressing Drp1 Expression
    • Authors: Ni Zheng; Tao Liang, Xiaohui Xu, Zelin Zhao, Jiangang Liu, Zhao Song, Weikang Huo, Hongchao Xu, Qiujing Wang, Xin Li
      Abstract: Polydatin (PD), a resveratrol glycoside, has been shown to protect renal function in diabetic nephropathy (DN), but the underlying molecular mechanism remains unclear. This study demonstrates that PD stabilize the mitochondrial morphology and attenuate mitochondrial malfunction in both KKAy mice and in hyperglycemia (HG)-induced MPC5 cells. We use western blot analysis to demonstrate that PD reversed podocyte apoptosis induced by HG via suppressing dynamin-related protein 1 (Drp1). This effect may depend on the ability of PD to inhibit the generation of cellular reactive oxygen species (ROS). In conclusion, we demonstrate that PD may be therapeutically useful in DN, and that, podocyte apoptosis induced by HG can be reversed by PD through suppressing Drp1 expression. This article is protected by copyright. All rights reserved
      PubDate: 2017-04-06T10:28:29.565786-05:
      DOI: 10.1002/jcp.25943
  • TWEAK promotes migration and invasion in MEFs through a mechanism
           dependent on ERKs activation and Fibulin 3 down-regulation
    • Authors: Celia Sequera; Ana Vázquez-Carballo, María Arechederra, Sonia Fernández-Veledo, Almudena Porras
      Abstract: TWEAK regulates multiple physio-pathological processes in fibroblasts such as fibrosis. It also induces migration and invasion in tumors and it can activate p38 MAPK in various cell types. Moreover, p38α MAPK promotes migration and invasion in several cancer cells types and in mouse embryonic fibroblasts (MEFs). However, it remains unknown if TWEAK could promote migration in fibroblasts and whether p38α MAPK might play a role. Our results reveal that TWEAK activates ERKs, Akt and p38α/β MAPKs and reduces secreted Fibulin 3 in MEFs. TWEAK also increases migration and invasion in wt and p38α deficient MEFs, which indicates that p38α MAPK is not required to mediate these effects. In contrast, ERKs inhibition significantly decreases TWEAK-induced migration and Fibulin 3 knock-down mimics TWEAK effect. These results indicate that both ERKs activation and Fibulin 3 down-regulation would contribute to mediate TWEAK pro-migratory effect. In fact, the additional regulation of ERKs and/or p38β as a consequence of Fibulin 3 decrease might be also involved in the pro-migratory effect of TWEAK in MEFs. In conclusion, our studies uncover novel mechanisms by which TWEAK would favor tissue repair by promoting fibroblasts migration.This article is protected by copyright. All rights reserved
      PubDate: 2017-04-06T09:41:07.497554-05:
      DOI: 10.1002/jcp.25942
  • Crosstalk of autophagy and apoptosis: Involvement of the dual role of
           autophagy under ER stress
    • Authors: Shuling Song; Jin Tan, Yuyang Miao, Mengmeng Li, Qiang Zhang
      Abstract: Endoplasmic reticulum (ER) stress is a common cellular stress response that is triggered by a variety of conditions that disturb cellular homeostasis, and induces cell apoptosis. Autophagy, an important and evolutionarily conserved mechanism for maintaining cellular homeostasis, is closely related to the apoptosis induced by ER stress. There are common upstream signaling pathways between autophagy and apoptosis induced by ER stress, including PERK/ATF4, IRE1α, ATF6, and Ca2+. Autophagy can not only block the induction of apoptosis by inhibiting the activation of apoptosis-associated caspase which could reduce cellular injury, but also help to induce apoptosis. In addition, the activation of apoptosis-related proteins can also inhibit autophagy by degrading autophagy-related proteins, such as Beclin-1, Atg4D, Atg3, and Atg5. Although the interactions of different autophagy- and apoptosis-related proteins, and also common upstream signaling pathways have been found, the potential regulatory mechanisms have not been clearly understood. In this review, we summarize the dual role of autophagy, and the interplay and potential regulatory mechanisms between autophagy and apoptosis under ER stress condition.The dura role of autophagy, pro-survival, and -death, may be dependent on the extent of ER stress, and it implements by the regulation of apoptosis. Autophagy can not only block the induction of apoptosis by inhibiting the activation of apoptosis-associated caspase which could reduce cellular injury, but also help to induce apoptosis. In addition, the activation of apoptosis-related proteins can also inhibit autophagy by degrading autophagy-related proteins, such as Beclin-1, Atg4D, Atg3, and Atg5.
      PubDate: 2017-03-31T02:45:23.567359-05:
      DOI: 10.1002/jcp.25785
  • Targeting stroma in pancreatic cancer: Promises and failures of targeted
    • Authors: Afsane Bahrami; Majid Khazaei, Fariba Bagherieh, Majid Ghayour-Mobarhan, Mina Maftouh, Seyed Mahdi Hassanian, Amir Avan
      Abstract: Desmoplasia or abundant fibrotic stroma is a typical property of most malignancies, which has a great effect on tumorigenesis, angiogenesis, and resistance to therapy. The activated stroma cells comprises several cell types including endothelial cells, nerve cells, inflammatory/macrophages cells, stellate cells, and extracellular matrix. In other word, the interactions of cancer-stroma modulate tumorigenesis, therapy resistance, and poor delivery of drugs. Therefore, targeting the tumor stroma in combination with conventional chemotherapeutic agents could provide a promising approach in the treatment of pancreatic cancer. This review summarizes the current knowledge about pancreatic stellate cells, targeting stroma compartments with particular emphasis on preclinical, and clinical trials on targeting of stroma as an option in pancreatic cancer treatment.
      PubDate: 2017-03-31T02:41:10.662556-05:
      DOI: 10.1002/jcp.25798
  • Effects of Carbocysteine and Beclomethasone on Histone
           Acetylation/Deacetylation Processes in Cigarette Smoke Exposed Bronchial
           Epithelial Cells
    • Authors: Elisabetta Pace; Serena Di Vincenzo, Maria Ferraro, Liboria Siena, Giuseppina Chiappara, Paola Dino, Patrizio Vitulo, Alessandro Bertani, Federico Saibene, Luigi Lanata, Mark Gjomarkaj
      Abstract: Histone deacetylase expression/activity may control inflammation, cell senescence, and responses to corticosteroids. Cigarette smoke exposure, increasing oxidative stress, may negatively affect deacetylase expression/activity. The effects of cigarette smoke extracts (CSE), carbocysteine, and beclomethasone dipropionate on chromatin remodeling processes in human bronchial epithelial cells are largely unknown. The present study was aimed to assess the effects of cigarette smoke, carbocysteine, and beclomethasone dipropionate on histone deacetylase 3 (HDAC3) expression/activity, N-CoR (nuclear receptor corepressor) expression, histone acetyltransferases (HAT) (p300/CBP) expression, p-CREB and IL-1 m-RNA expression, neutrophil chemotaxis. Increased p-CREB expression was observed in the bronchial epithelium of smokers. CSE increased p-CREB expression and decreased HDAC3 expression and activity and N-CoR m-RNA and protein expression. At the same time, CSE increased the expression of the HAT, p300/CBP. All these events increased acetylation processes within the cells and were associated to increased IL-1 m-RNA expression and neutrophil chemotaxis. The incubation of CSE exposed cells with carbocysteine and beclomethasone counteracted the effects of cigarette smoke on HDAC3 and N-CoR but not on p300/CBP. The increased deacetylation processes due to carbocysteine and beclomethasone dipropionate incubation is associated to reduced p-CREB, IL-1 m-RNA expression, neutrophil chemotaxis. These findings suggest a new role of combination therapy with carbocysteine and beclomethasone dipropionate in restoring deacetylation processes compromised by cigarette smoke exposure. J. Cell. Physiol. 9999: 1–9, 2016. © 2016 Wiley Periodicals, Inc.Cigarette smoke exposure, increasing oxidative stress, may negatively affect deacetylase expression/activity, thus, supporting increased pro-inflammatory responses.The findings here provided suggest a new role of combination therapy with carbocysteine and beclomethasone dipropionate in restoring deacetylation processes compromised by cigarette smoke exposure.
      PubDate: 2017-03-31T02:35:37.888802-05:
      DOI: 10.1002/jcp.25710
  • Acetoacetate induces hepatocytes apoptosis by the ROS-mediated MAPKs
           pathway in ketotic cows
    • Authors: Xiliang Du; Zhen Shi, Zhicheng Peng, Chenxu Zhao, Yuming Zhang, Zhe Wang, Xiaobing Li, Guowen Liu, Xinwei Li
      Abstract: Dairy cows with ketosis are characterized by oxidative stress, hepatic damage, and hyperketonemia. Acetoacetate (AA) is the main component of ketone bodies in ketotic cows, and is associated with the above pathological process. However, the potential mechanism was not illuminated. Therefore, the aim of this study was to investigate the mechanism of AA-induced hepatic oxidative damage in ketotic cows. Compared with healthy cows, ketotic cows exhibited severe oxidative stress and hepatic damage. Moreover, the extent of hepatic damage and oxidative stress had a positive relationship with the AA levels. In vitro, AA treatment increased reactive oxygen species (ROS) content and further induced oxidative stress and apoptosis of bovine hepatocytes. In this process, AA treatment increased the phosphorylation levels of JNK and p38MAPK and decreased the phosphorylation level of ERK, which could increase p53 and inhibit nuclear factor E2-related factor 2 (Nrf2) expression, nuclear localization, and DNA-binding affinity, thereby inducing the overexpression of pro-apoptotic molecules Bax, Caspase 3, Caspase 9, PARP and inhibition of anti-apoptotic molecule Bcl-2. Antioxidant N-acetylcysteine (NAC) treatment or interference of MAPKs pathway could attenuate the hepatocytes apoptosis induced by AA. Collectively, these results indicate that AA triggers hepatocytes apoptosis via the ROS-mediated MAPKs pathway in ketotic cows.Excess acetoacetate induced imbalance of oxidation and antioxidation, resulting in overproduction of reactive oxygen species (ROS) in the mitochondria. High concentration of ROS increased the phosphorylation levels of p38MAPK and JNK and decreased the phosphorylation levels of ERK1/2, which could activate p53 and inhibit Nrf2 expression, nuclear localization, and DNA-binding affinity. Consequently, the expression of pro-apoptotic molecules Bax, Casepase9, Casepase3, and PARP was significantly upregulated and anti-apoptotic molecules Bcl-2 were markedly downregulated, resulting in hepatocytes apoptosis in ketotic cows.
      PubDate: 2017-03-31T02:35:33.101056-05:
      DOI: 10.1002/jcp.25773
  • Pyk2 inhibition promotes contractile differentiation in arterial smooth
    • Authors: Mario Grossi; Anirban Bhattachariya, Ina Nordström, Karolina M. Turczyńska, Daniel Svensson, Sebastian Albinsson, Bengt-Olof Nilsson, Per Hellstrand
      Abstract: Modulation from contractile to synthetic phenotype of vascular smooth muscle cells is a central process in disorders involving compromised integrity of the vascular wall. Phenotype modulation has been shown to include transition from voltage-dependent toward voltage-independent regulation of the intracellular calcium level, and inhibition of non-voltage dependent calcium influx contributes to maintenance of the contractile phenotype. One possible mediator of calcium-dependent signaling is the FAK-family non-receptor protein kinase Pyk2, which is activated by a number of stimuli in a calcium-dependent manner. We used the Pyk2 inhibitor PF-4594755 and Pyk2 siRNA to investigate the role of Pyk2 in phenotype modulation in rat carotid artery smooth muscle cells and in cultured intact arteries. Pyk2 inhibition promoted the expression of smooth muscle markers at the mRNA and protein levels under stimulation by FBS or PDGF-BB and counteracted phenotype shift in cultured intact carotid arteries and balloon injury ex vivo. During long-term (24–96 hr) treatment with PF-4594755, smooth muscle markers increased before cell proliferation was inhibited, correlating with decreased KLF4 expression and differing from effects of MEK inhibition. The Pyk2 inhibitor reduced Orai1 and preserved SERCA2a expression in carotid artery segments in organ culture, and eliminated the inhibitory effect of PDGF stimulation on L-type calcium channel and large-conductance calcium-activated potassium channel expression in carotid cells. Basal intracellular calcium level, calcium wave activity, and store-operated calcium influx were reduced after Pyk2 inhibition of growth-stimulated cells. Pyk2 inhibition may provide an interesting approach for preserving vascular smooth muscle differentiation under pathophysiological conditions.The Pyk2 inhibitor PF-4594755 potently inhibits Pyk2 phosphorylation of vascular smooth muscle cells stimulated by FBS or PDGF-BB. This is accompanied by increased expression of smooth muscle marker proteins without concomitant effects on cell proliferation. Pyk2 inhibition upregulates SERCA2a expression and may preserve the contractile phenotype by lowering basal intracellular calcium and promoting voltage-dependent control of calcium influx over the plasma membrane.
      PubDate: 2017-03-31T02:25:35.8512-05:00
      DOI: 10.1002/jcp.25760
  • Biological and pharmacological evaluation of dimethoxycurcumin: A
           metabolically stable curcumin analogue with a promising therapeutic
    • Authors: Manouchehr Teymouri; Nastaran Barati, Matteo Pirro, Amirhosein Sahebkar
      Abstract: Dimethoxycurcumin (DiMC) is a synthetic analog of curcumin with superior inter-related pro-oxidant and anti-cancer activity, and metabolic stability. Numerous studies have shown that DiMC reserves the biologically beneficial features, including anti-inflammatory, anti-carcinogenic, and cytoprotective properties, almost to the same extent as curcumin exhibits. DiMC lacks the phenolic-OH groups as opposed to curcumin, dimethoxycurcumin, and bis-demethoxycurcumin that all vary in the number of methoxy groups per molecule, and has drawn the attentions of researchers who attempted to discover the structure-activity relationship (SAR) of curcumin. In this regard, tetrahydrocurcumin (THC), the reduced and biologically inert metabolite of curcumin, denotes the significance of the conjugated α,β diketone moiety for the curcumin activity. DiMC exerts unique molecular activities compared to curcumin, including induction of androgen receptor (AR) degradation and suppression of the transcription factor activator protein-1 (AP-1). The enhanced AR degradation on DiMC treatment suggests it as a novel anticancer agent against resistant tumors with androgenic etiology. Further, DiMC might be a potential treatment for acne vulgaris. DiMC induces epigenetic alteration more effectively than curcumin, although both showed no direct DNA hypomethylating activity. Given the metabolic stability, nanoparticulation of DiMC is more promising for in vivo effectiveness. However, studies in this regard are still in its infancy. In the current review, we portray the various molecular and biological functions of DiMC reported so far. Whenever possible, the efficiency is compared with curcumin and the reasons for DiMC being more metabolically stable are elaborated. We also provide future perspective investigations with respect to varying DiMC-nanoparticles.This review, being the first of its kind, is a comprehensive evaluation of the molecular, cellular, and clinical properties of dimethoxycurcumin as a promising analog of curcumin. We have discussed various molecular and biological functions of DiMC reported so far. Whenever possible, the efficiency compared with curcumin and the reasons for DiMC being more metabolically stable than curcumin has been explained. We have also provided future perspective investigations with respect to varying DiMC-nanoparticles.
      PubDate: 2017-03-31T02:25:31.602396-05:
      DOI: 10.1002/jcp.25749
  • Cancer Cell Plasticity: Rapid Reversal of Chemosensitivity and Expression
           of Stemness Markers in Lung and Breast Cancer Tumorspheres
    • Authors: Juan Sebastian Yakisich; Neelam Azad, Vivek Kaushik, Anand Krishnan V. Iyer
      Abstract: In cancer cells, the reversible nature of the stemness status in terms of chemoresistance has been poorly characterized. In this study, we have simulated one cycle of environmental conditions to study such reversibility by first generating floating tumorspheres (FTs) from lung and breast cancer cells by culturing them in serum-free media without the addition of any external mitogenic stimulation, and subsequently (after 2 weeks) re-incubating them back in serum-containing media to simulate routine culture conditions (RCCs). We found that cancer cells are extremely plastic: cells grown under RCCs become multidrug-resistant when grown as FTs, but upon re-incubation under RCCs quickly re-attach and lose the acquired resistance. These phenotypic changes are accompanied by concomitant changes in the expression of key proteins associated with multiple pathways important for chemoresistance, survival, and stemness maintenance. Therefore, our strategy provides an excellent experimental model to study environmental factors that modulate the plasticity of cancer cells. J. Cell. Physiol. 9999: 1–7, 2016. © 2016 Wiley Periodicals, Inc.In this report, we have developed a protocol to grow lung and breast cancer cells as floating tumorspheres (FTs) in the absence of any external mitogenic stimulation. By using routine culture conditions (RCCs), FTs, and revertant cells (FTs cell that reattach when incubated back under RCCs), we simulated one cycle of environmental conditions in order to study the plasticity of cancer cells. We found that cancer cells are extremely plastic: cells grown under RCCs become multidrug-resistant, when grown as FTs but upon re-incubation under RCCs quickly re-attach and lose the acquired resistance. These phenotypic changes are accompanied by concomitant changes in the expression of key proteins associated with multiple pathways important for chemoresistance, survival, and stemness maintenance.
      PubDate: 2017-03-31T02:25:24.598579-05:
      DOI: 10.1002/jcp.25725
  • Challenging of AS160/TBC1D4 Alters Intracellular Lipid milieu in L6
           Myotubes Incubated With Palmitate
    • Authors: Agnieszka Mikłosz; Bartłomiej Łukaszuk, Małgorzata Żendzian-Piotrowska, Justyna Brańska-Januszewska, Halina Ostrowska, Adrian Chabowski
      Abstract: The Akt substrate of 160 kDa (AS160) is a key regulator of GLUT4 translocation from intracellular depots to the plasma membrane in myocytes. Likely, AS160 also controls LCFAs transport, which requires relocation of fatty acid transporters. The aim of the present study was to determine the impact of AS160 knockdown on lipid milieu in L6 myotubes incubated with palmitate (PA). Therefore, we compared two different settings, namely: 1) AS160 knockdown prior to palmitate incubation (pre-PA-silencing, AS160−/PA); 2) palmitate incubation with subsequent AS160 knockdown (post-PA-silencing, PA/AS160−). The efficiency of AS160 silencing was checked at mRNA and protein levels. The expression and localization of FA transporters were determined using Western Blot and immunofluorescence analyses. Intracellular lipid content (FFA, DAG, TAG, and PL) and FA composition were estimated by GLC, whereas basal palmitate uptake was analyzed by means of scintigraphy. Both groups with silenced AS160 were characterized by a greater expression of FA transporters (FAT/CD36, FATP-1, 4) which had contributed to an increased FA cellular influx. Accordingly, we observed that post-PA-silencing of AS160 resulted in a marked decrement in DAG, TAG, and PL contents, but increased FFA content (PA/AS160− vs. PA). The opposite effect was observed in the group with pre-PA-silencing of AS160 in which AS160 knockdown did not affect the lipid pools (AS160−/PA vs. PA). Our results indicate that post-PA-silencing of AS160 has a capacity to decrease the lipotoxic effect(s) of PA by decreasing the content of lipids (DAG and PL) that promote insulin resistance in myotubes. J. Cell. Physiol. 9999: 1–14, 2017. © 2016 The
      Authors . Journal of Cellular Physiology Published by Wiley Periodicals Inc.AS160/TBC1D4 is a key regulator of GLUT4 translocation to the plasma membrane in myocytes, moreover, it role in the control of protein mediated fatty acids transport is also likely. The current study indicates that AS160/TBC1D4 may influence not only FA transporters content, and therefore their influx, but also their storage and oxidation. However, the effect differs depending on the events sequence, i.e., AS160 knock-down prior to or after palmitate incubation.
      PubDate: 2017-03-31T02:23:29.244944-05:
      DOI: 10.1002/jcp.25632
  • TNT-Induced Phagocytosis: Tunneling Nanotubes Mediate the Transfer of
           Pro-Phagocytic Signals From Apoptotic to Viable Cells
    • Authors: Margarethe Bittins; Xiang Wang
      Abstract: The exposure of phosphatidylserine (PS) on the surface membrane of apoptotic cells triggers the recruitment of phagocytic receptors and subsequently results in uptake by phagocytes. Here we describe how apoptotic cells can use intercellular membrane nanotubes to transfer exposed PS to neighboring viable cells, and thus deposit an “eat-me” tag on the viable cells. Tunneling nanotubes (TNTs) connected UV-treated apoptotic rat pheochromocytoma PC12 cells with neighboring untreated cells. These TNTs were composed of PS-exposed plasma membrane and facilitated the transfer of the membrane from apoptotic to viable cells. Other pro-phagocytic signals, such as oxidized phospholipids and calreticulin, were also transferred to viable cells. In addition, anti-phagocytic signal CD47 presenting on the plasma membrane of viable cells was masked by the transferred PS-membrane. Confocal imaging revealed an increase of phagocytosis of viable PC12 cells by murine RAW264.7 macrophages when the viable PC12 cells were cocultured with UV-treated PC12 cells. Treatment with 50 nM cytochalasin D would abolish TNTs and correspondingly inhibit this phagocytosis of the viable cells. Our study indicates that exposed-PS membrane is delivered from apoptotic to viable cells through TNTs. This transferred membrane may act as a pro-phagocytic signal for macrophages to induce phagocytosis of viable cells in a situation where they are in the vicinity of apoptotic cells. J. Cell. Physiol. 9999: 1–9, 2017. © 2016 The
      Authors . Journal of Cellular Physiology Published by Wiley Periodicals Inc.Tunneling nanotubes (TNTs) connected UV-treated apoptotic PC12 cells with neighboring untreated cells. These phosphatidylserine (PS)-positive TNTs facilitated the transfer of pro-phagocytic signals from apoptotic to viable cells. Notably, the phagocytosis of viable PC12 cells by macrophages increased when apoptotic cells were present, but only under conditions that allowed for the formation of TNTs. Our findings suggest that TNTs are a means of transferring “eat-me” signals from dying to living cells, and thus induce phagocytosis of previously healthy cells.
      PubDate: 2017-03-31T02:23:26.392479-05:
      DOI: 10.1002/jcp.25584
  • Maintenance of Bone Homeostasis by DLL1-Mediated Notch Signaling
    • Authors: Yukari Muguruma; Katsuto Hozumi, Hiroyuki Warita, Takashi Yahata, Tomoko Uno, Mamoru Ito, Kiyoshi Ando
      Abstract: Adult bone mass is maintained through a balance of the activities of osteoblasts and osteoclasts. Although Notch signaling has been shown to maintain bone homeostasis by controlling the commitment, differentiation, and function of cells in both the osteoblast and osteoclast lineages, the precise mechanisms by which Notch performs such diverse and complex roles in bone physiology remain unclear. By using a transgenic approach that modified the expression of delta-like 1 (DLL1) or Jagged1 (JAG1) in an osteoblast-specific manner, we investigated the ligand-specific effects of Notch signaling in bone homeostasis. This study demonstrated for the first time that the proper regulation of DLL1 expression, but not JAG1 expression, in osteoblasts is essential for the maintenance of bone remodeling. DLL1-induced Notch signaling was responsible for the expansion of the bone-forming cell pool by promoting the proliferation of committed but immature osteoblasts. However, DLL1-Notch signaling inhibited further differentiation of the expanded osteoblasts to become fully matured functional osteoblasts, thereby substantially decreasing bone formation. Osteoblast-specific expression of DLL1 did not alter the intrinsic differentiation ability of cells of the osteoclast lineage. However, maturational arrest of osteoblasts caused by the DLL1 transgene impaired the maturation and function of osteoclasts due to a failed osteoblast-osteoclast coupling, resulting in severe suppression of bone metabolic turnover. Taken together, DLL1-mediated Notch signaling is critical for proper bone remodeling as it regulates the differentiation and function of both osteoblasts and osteoclasts. Our study elucidates the importance of ligand-specific activation of Notch signaling in the maintenance of bone homeostasis. J. Cell. Physiol. 9999: 1–12, 2017. © 2016 The
      Authors . Journal of Cellular Physiology Published by Wiley Periodicals Inc.The proper regulation of DLL1 expression, but not JAG1 expression, in osteoblasts is essential for the maintenance of bone remodeling. DLL1-Notch signaling directly controls the physiological expansion and differentiation of osteoblasts and indirectly affects the maturation and function of osteoclasts, thereby ensuring the homeostasis of bone mass.
      PubDate: 2017-03-31T02:23:23.179938-05:
      DOI: 10.1002/jcp.25647
  • Reduced primary cilia length and altered Arl13b expression are associated
           with deregulated chondrocyte Hedgehog signaling in alkaptonuria
    • Authors: Stephen D. Thorpe; Silvia Gambassi, Clare L. Thompson, Charmilie Chandrakumar, Annalisa Santucci, Martin M. Knight
      Abstract: Alkaptonuria (AKU) is a rare inherited disease resulting from a deficiency of the enzyme homogentisate 1,2-dioxygenase which leads to the accumulation of homogentisic acid (HGA). AKU is characterized by severe cartilage degeneration, similar to that observed in osteoarthritis. Previous studies suggest that AKU is associated with alterations in cytoskeletal organization which could modulate primary cilia structure/function. This study investigated whether AKU is associated with changes in chondrocyte primary cilia and associated Hedgehog signaling which mediates cartilage degradation in osteoarthritis. Human articular chondrocytes were obtained from healthy and AKU donors. Additionally, healthy chondrocytes were treated with HGA to replicate AKU pathology (+HGA). Diseased cells exhibited shorter cilia with length reductions of 36% and 16% in AKU and +HGA chondrocytes respectively, when compared to healthy controls. Both AKU and +HGA chondrocytes demonstrated disruption of the usual cilia length regulation by actin contractility. Furthermore, the proportion of cilia with axoneme breaks and bulbous tips was increased in AKU chondrocytes consistent with defective regulation of ciliary trafficking. Distribution of the Hedgehog-related protein Arl13b along the ciliary axoneme was altered such that its localization was increased at the distal tip in AKU and +HGA chondrocytes. These changes in cilia structure/trafficking in AKU and +HGA chondrocytes were associated with a complete inability to activate Hedgehog signaling in response to exogenous ligand. Thus, we suggest that altered responsiveness to Hedgehog, as a consequence of cilia dysfunction, may be a contributing factor in the development of arthropathy highlighting the cilium as a novel target in AKU.This study investigates whether the rare inherited disease alkaptonuria (AKU) is associated with changes in chondrocyte primary cilia and associated Hedgehog signaling, which is known to mediate cartilage degradation in osteoarthritis. AKU results in alterations to primary cilia structure which are associated with ciliary trafficking defects with the potential to impact a range of fundamental signaling pathways. In particular, AKU chondrocytes demonstrated a complete inability to activate Hedgehog signaling in response to endogenous ligand; which we propose may be the result of the observed dysfunction of cilia structure and trafficking. Thus, we suggest that altered responsiveness to Hedgehog, as a consequence of cilia dysfunction, may be a contributing factor in the development of arthropathy highlighting the cilium as a novel target in AKU.
      PubDate: 2017-03-31T02:20:25.756423-05:
      DOI: 10.1002/jcp.25839
  • Staphylococcal protein A promotes osteoclastogenesis through MAPK
           signaling during bone infection
    • Authors: Yuan Wang; Xin Liu, Ce Dou, Zhen Cao, Chuan Liu, Shiwu Dong, Jun Fei
      Abstract: Bone infection is a common and serious complication in the orthopedics field, which often leads to excessive bone destruction and non-union. Osteoclast is the only type of cells which have the function of bone resorption. Its over activation is closely related to excessive bone loss. Staphylococcus aureus (S. aureus) is a major pathogen causing bone infection, which can produce a large number of strong pathogenic substances staphylococcal protein A (SPA). However, few studies were reported about the effects of SPA on osteoclastogenesis. In our study, we observed that S. aureus activated osteoclasts and promoted bone loss in bone infection specimens. Then, we investigated the effects of SPA on RANKL-induced osteoclastogenesis in vitro, the results revealed that SPA promoted osteoclastic differentiation and fusion, and enhanced osteoclastic bone resorption. In addition, we also showed that SPA upregulated the expression of NFATc1 and c-FOS through the activation of MAPK signaling to promote osteoclastogenesis. Our findings might help us better understand the pathogenic role of S. aureus in bone infection and develop new therapeutic strategies for infectious bone diseases.Staphylococcal protein A (SPA) is one of the most important pathogenic components of Staphylococcus aureus (S. aureus). Here we reported that S. aureus caused bone loss in bone infection specimens. In vitro study showed that SPA promoted osteoclastic differentiation, fusion and bone resorption activity by upregulating NFATc1 and c-FOS through MAPK signaling.
      PubDate: 2017-03-31T02:15:32.243969-05:
      DOI: 10.1002/jcp.25774
  • Reduced Glucose-Induced Insulin Secretion in Low-protein-fed Rats Is
           Associated With Altered Pancreatic Islets Redox Status
    • Authors: Ana Paula G. Cappelli; Claudio C. Zoppi, Leonardo R. Silveira, Thiago M. Batista, Flávia M. Paula, Priscilla M. R. da Silva, Alex Rafacho, Helena C. Barbosa-Sampaio, Antonio C. Boschero, Everardo M. Carneiro
      Abstract: In the present study, we investigated the relationship between early life protein malnutrition-induced redox imbalance, and reduced glucose-stimulated insulin secretion. After weaning, male Wistar rats were submitted to a normal-protein-diet (17%-protein, NP) or to a low-protein-diet (6%-protein, LP) for sixty days. Pancreatic islets were isolated and hydrogen peroxide (H2O2), oxidized (GSSG) and reduced (GSH) glutathione content, CuZn-superoxide dismutase (SOD1), glutathione peroxidase (GPx1) and catalase (CAT) gene expression, as well as enzymatic antioxidant activities were quantified. Islets that were pre-incubated with H2O2 and/or N-acetylcysteine, were subsequently incubated with glucose for insulin secretion measurement. Protein malnutrition increased CAT mRNA content by 100%. LP group SOD1 and CAT activities were 50% increased and reduced, respectively. H2O2 production was more than 50% increased whereas GSH/GSSG ratio was near 60% lower in LP group. Insulin secretion was, in most conditions, approximately 50% lower in LP rat islets. When islets were pre-incubated with H2O2 (100 µM), and incubated with glucose (33 mM), LP rats showed significant decrease of insulin secretion. This effect was attenuated when LP islets were exposed to N-acetylcysteine. This article is protected by copyright. All rights reserved
      PubDate: 2017-03-30T07:35:46.528717-05:
      DOI: 10.1002/jcp.25908
  • End Stage Renal Disease-induced Hypercalcemia May Promote Aortic Valve
           Calcification via Annexin VI Enrichment of Valve Interstitial Cell
           Derived-Matrix Vesicles
    • Authors: Lin Cui; Nabil A Rashdan, Dongxing Zhu, Elspeth Milne, Paul Ajuh, Gillian Milne, Miep Helfrich, Kelvin Lim, Sai Prasad, Daniel Lerman, Alex Vesey, Marc Dweck, WS Jenkins, David Newby, Colin Farquharson, Vicky E Macrae
      Abstract: Patients with end-stage renal disease (ESRD) have elevated circulating calcium (Ca) and phosphate (Pi), and exhibit accelerated progression of calcific aortic valve disease (CAVD). We hypothesised that matrix vesicles (MVs) initiate the calcification process in CAVD.Ca induced rat valve interstitial cells (VICs) calcification at 4.5 mM (16.4 fold; P 
      PubDate: 2017-03-30T07:01:12.82548-05:0
      DOI: 10.1002/jcp.25935
  • Stem Cells Applications in Bone and Tooth Repair and Regeneration: New
           Insights, Tools and Hopes
    • Authors: Eiman Abdel Meguid; Yuehai Ke, Junfeng Ji, Ahmed HK El-Hashash
      Abstract: The exploration of stem and progenitor cells holds promise for advancing our understanding of the biology of tissue repair and regeneration mechanisms after injury. This will also help in the future use of stem cell therapy for the development of regenerative medicine approaches for the treatment of different tissue-specie defects or disorders such as bone, cartilages and tooth defects or disorders. Bone is a specialized connective tissue, with mineralized extracellular components that provide bones with both strength and rigidity, and thus enable bones to function in body mechanical supports and necessary locomotion process. New insights have been added to the use of different types of stem cells in bone and tooth defects over the last few years. In this concise review, we briefly describe bone structure as well as summarize recent research progress and accumulated information regarding the osteogenic differentiation of stem cells, as well as stem cell contributions to bone repair/regeneration, bone defects or disorders and both restoration and regeneration of bones and cartilages. We also discuss advances in the osteogenic differentiation and bone regeneration of dental and periodontal stem cells as well as in stem cell contributions to dentine regeneration and tooth engineering. This article is protected by copyright. All rights reserved
      PubDate: 2017-03-30T07:01:01.27359-05:0
      DOI: 10.1002/jcp.25940
  • Connexin43 Intercellular Communication Drives the Early Differentiation of
           Human Bone Marrow Stromal Cells into Osteoblasts
    • Authors: Julie Talbot; Régis Brion, Audrey Lamora, Mathilde Mullard, Sarah Morice, Dominique Heymann, Franck Verrecchia
      Abstract: Although it has been demonstrated that human bone marrow stromal cells (hBMSCs) express the ubiquitous connexin43 (Cx43) and form functional gap junctions, their role in the early differentiation of hBMSCs into osteoblasts remains poorly documented. Using in vitro assays, we show that Cx43 expression and gap junctional intercellular communication (GJIC) are increased during the differentiation of hBMSCs into osteoblasts, both at the protein and mRNA levels. Two independent procedures to reduce GJIC, a pharmacological approach with GJIC inhibitors (18α-glycyrrhetinic acid and Gap27 peptide) and a molecular approach using small interfering RNA against Cx43, demonstrated that the presence of Cx43 and functional junctional channels are essential to the ability of hBMSCs to differentiate into osteoblasts in vitro. In addition, a reduced GJIC decreases the expression of Runx2, the major transcription factor implicated in the control of osteoblast commitment and early differentiation of hBMSCs into osteoblasts, suggesting that GJIC mediated by Cx43 is implicated in this process. Together our results demonstrate that GJIC mediated by the Cx43 channels plays a central role throughout the differentiation of hBMSC into osteoblasts, from the early stages to the process of mineralization. This article is protected by copyright. All rights reserved
      PubDate: 2017-03-30T07:00:27.158567-05:
      DOI: 10.1002/jcp.25938
  • Sphingolipids Metabolism in the Salivary Glands of Rats With Obesity and
           Streptozotocin Induced Diabetes
    • Authors: Marta Garbowska; Bartłomiej Łukaszuk, Agnieszka Mikłosz, Igor Wróblewski, Krzysztof Kurek, Lucyna Ostrowska, Adrian Chabowski, Małgorzata Żendzian-Piotrowska, Anna Zalewska
      Abstract: BackgroundsDiabetes is considered a major public health problem affecting millions of individuals worldwide. Remarkably, scientific reports regarding salivary glands sphingolipid metabolism in diabetes are virtually non-existent. This is odd given the well-established link between the both in other tissues (e.g. skeletal muscles, liver) and the key role of these glands in oral health preservation. The aim of this paper is to examine sphingolipids metabolism in the salivary glands in (pre)diabetes (evoked by high fat diet feeding or streptozotocin).MethodsWistar rats were allocated into three groups: control, HFD- or STZ-diabetes. The content of major sphingolipid classes in the parotid (PSG) and submandibular (SMSG) glands was assessed via chromatography. Additionally, Western Blott analyses were employed for the evaluation of key sphingolipid signaling pathway enzyme levels.ResultsNo changes in ceramide content in the PSG were found, whereas an increase in ceramide concentration for SMSG of the STZ group was observed. This was accompanied by an elevation in SPT1 level. Probably also sphingomyelin hydrolysis was increased in the SMSG of the STZ-diabetic rats, since we observed a significant drop in the amount of SM.ConclusionPSG and SMSG respond differently to (pre)diabetes, with clearer pattern presented by the later gland. An activation of sphingomyelin signaling pathway was observed in the course of STZ-diabetes, i.e. metabolic condition with rapid onset/progression. Whereas, chronic HFD lead to an inhibition of sphingomyelin signaling pathway in the salivary glands (manifested in an inhibition of ceramide de novo synthesis and accumulation of S1P). This article is protected by copyright. All rights reserved
      PubDate: 2017-03-30T07:00:08.158859-05:
      DOI: 10.1002/jcp.25939
  • Roles of Neurotrophins in Skeletal Tissue Formation and Healing
    • Authors: Yu-Wen Su; Xin-Fu Zhou, Bruce K. Foster, Brian L. Grills, Jiake Xu, Cory J. Xian
      Abstract: Neurotrophins and their receptors are key molecules that are known to be critical in regulating nervous system development and maintenance and have been recognized to be also involved in regulating tissue formation and healing in skeletal tissues. Studies have shown that neurotrophins and their receptors are widely expressed in skeletal tissues, implicated in chondrogenesis, osteoblastogenesis and osteoclastogenesis, and are also involved in regulating tissue formation and healing events in skeletal tissue. Increased mRNA expression for neurotrophins NGF, BDNF, NT-3 and NT-4 and their Trk receptors has been observed in injured bone tissues, and NT-3 and its receptor, TrkC, have been identified to have the highest induction at the injury site in a drill-hole injury repair model in both bone and the growth plate. In addition, NT-3 has also recently been shown to be both an osteogenic and angiogenic factor, and this neurotrophin can also enhance expression of the key osteogenic factor, BMP-2, as well as the major angiogenic factor, VEGF, to promote bone formation, vascularization, and healing of the injury site. Further studies, however, are needed to investigate if different neurotrophins have differential roles in skeletal repair, and if NT-3 can be a potential target of intervention for promoting bone fracture healing. This article is protected by copyright. All rights reserved
      PubDate: 2017-03-30T07:00:04.252342-05:
      DOI: 10.1002/jcp.25936
  • Cholinergic and Dopaminergic Neuronal Differentiation of Human Adipose
           Tissue Derived Mesenchymal Stem Cells
    • Authors: HE Marei; A El-Gamal, A Althani, N Afifi, A Abd-Elmaksoud, A Farag, C Cenciarelli, Thomas Caceci, Anwarul Hasan
      Abstract: Mesenchymal stem cells (MSCs) are multipotent cells that can differentiate into various cell types such as cartilage, bone, and fat cells. Recent studies have shown that induction of MSCs in vitro by growth factors including epidermal growth factor (EGF) and fibroblast growth factor (FGF2) causes them to differentiate into neural like cells. These cultures also express ChAT, a cholinergic marker; and TH, a dopaminergic marker for neural cells. To establish a protocol with maximum differentiation potential, we examined MSCs under three experimental culture conditions using neural induction media containing FGF2, EGF, BMP-9, retinoic acid and heparin. Adipose-derived MSCs were extracted and expanded in vitro for 3 passages after reaching >80% confluency, for a total duration of 9 days. Cells were then characterized by flow cytometry for CD markers as CD44 positive and CD45 negative. MSCs were then treated with neural induction media and were characterized by morphological changes and Q-PCR. Differentiated MSCs expressed markers for immature and mature neurons; β Tubulin III (TUBB3) and MAP2, respectively showing the neural potential of these cells to differentiate into functional neurons. Improved protocols for MSCs induction will facilitate and ensure the reproducibility and standard production of MSCs for therapeutic applications in neurodegenerative diseases. This article is protected by copyright. All rights reserved
      PubDate: 2017-03-30T07:00:00.347246-05:
      DOI: 10.1002/jcp.25937
  • Cover Image, Volume 232, Number 7, July 2017
    • Authors: Erik H. Koritzinsky; Jonathan M. Street, Robert A. Star, Peter S.T. Yuen
      Abstract: Cover: The cover image, by Peter S.T. Yuen et al. is based on the From the Bench Paper Quantification of Exosomes,
      DOI : 10.1002/jcp.25387.
      PubDate: 2017-03-28T07:35:07.258023-05:
  • Individual-specific Variation in the Respiratory Activities of HMECs and
           Their Bioenergetic Response to IGF1 and TNFα
    • Authors: Sallie S. Schneider; Elizabeth M. Henchey, Nazneen Sultana, Stephanie M. Morin, D. Joseph Jerry, Grace Makari-Judson, Giovanna M. Crisi, Richard B. Arenas, Melissa Johnson, Holly S. Mason, Nagendra Yadava
      Abstract: Metabolic reprograming is a hallmark of cancer cells. However, the roles of pre-existing differences in normal cells metabolism toward cancer risk is not known. In order to assess pre- existing variations in normal cell metabolism, we have quantified the inter-individual variation in oxidative metabolism of normal primary human mammary epithelial cells (HMECs). We then assessed their response to selected cytokines such as insulin growth factor 1 (IGF1) and tumor necrosis factor alpha (TNFα), which are associated with breast cancer risk. Specifically, we compared the oxidative metabolism of HMECs obtained from women with breast cancer and without cancer. Our data show considerable inter-individual variation in respiratory activities of HMECs from different women. A bioenergetic parameter called pyruvate-stimulated respiration (PySR) was identified as a key distinguishing feature of HMECs from women with breast cancerand without cancer. Samples showing PySR over 20% of basal respiration rate were considered PySR + ve and the rest as PySR-ve. By this criterion, HMECs from tumor-affected breasts (AB) and non-tumor affected breasts (NAB) of cancer patients were mostly PySR-ve (88% and 89%, respectively), while HMECs from non-cancer patients were mostly PySR + ve (57%). This suggests that PySR-ve/ + ve phenotypes are individual-specific and are not caused by field effects due to the presence of tumor. The effects of IGF1 and TNFα treatments on HMECs revealed that both suppressed respiration and extracellular acidification. In addition, IGF1 altered PySR-ve/ + ve phenotypes. These results reveal individual-specific differences in pyruvate metabolism of normal breast epithelial cells and its association with breast cancer risk. This article is protected by copyright. All rights reserved
      PubDate: 2017-03-28T07:15:51.276264-05:
      DOI: 10.1002/jcp.25932
  • Acute Ethanol Exposure-induced Autophagy-mediated Cardiac Injury via
           Activation of the ROS-JNK-Bcl-2 Pathway
    • Authors: Zhongxin Zhu; Yewei Huang, Lingchun Lv, Youli Tao, Minglong Shao, Congcong Zhao, Mei Xue, Jia Sun, Chao Niu, Yang Wang, Sunam Kim, Weitao Cong, Wei Mao, Litai Jin
      Abstract: Binge drinking is associated with increased cardiac autophagy, and often triggers heart injury. Given the essential role of autophagy in various cardiac diseases, this study was designed to investigate the role of autophagy in ethanol-induced cardiac injury and the underlying mechanism. Our study showed that ethanol exposure enhanced the levels of LC3-II and LC3-II positive puncta and promoted cardiomyocyte apoptosis in vivo and in vitro. In addition, we found that ethanol induced autophagy and cardiac injury largely via the sequential triggering of reactive oxygen species (ROS) accumulation, activation of c-Jun NH2-terminal kinase (JNK), phosphorylation of Bcl-2, and dissociation of the Beclin 1/Bcl-2 complex. By contrast, inhibition of ethanol-induced autophagic flux with pharmacologic agents in the hearts of mice and cultured cells significantly alleviated ethanol-induced cardiomyocyte apoptosis and heart injury. Elimination of ROS with the antioxidant N-acetyl cysteine (NAC) or inhibition of JNK with the JNK inhibitor SP600125 reduced ethanol-induced autophagy and subsequent autophagy-mediated apoptosis. Moreover, metallothionein (MT), which can scavenge reactive oxygen and nitrogen species, also attenuated ethanol-induced autophagy and cell apoptosis in MT-TG mice. In conclusion, our findings suggest that acute ethanol exposure induced autophagy-mediated heart toxicity and injury mainly through the ROS-JNK-Bcl-2 signaling pathway. This article is protected by copyright. All rights reserved
      PubDate: 2017-03-28T07:15:49.967808-05:
      DOI: 10.1002/jcp.25934
  • Global Chondrocyte Gene Expression after a Single Anabolic Loading Period:
           Time Evolution and Re-inducibility of Mechano-responses
    • Authors: Simone Scholtes; Elisabeth Krämer, Melanie Weisser, Wolfgang Roth, Reto Luginbühl, Tobias Grossner, Wiltrud Richter
      Abstract: ObjectiveA genome-wide identification of mechano-regulated genes and candidate pathways in human chondrocytes subjected to a single anabolic loading episode and characterization of time evolution and re-inducibility of the response.DesignOsteochondral constructs consisting of a chondrocyte-seeded collagen-scaffold connected to β-tricalcium-phosphate were pre-cultured for 35 days and subjected to dynamic compression (25% strain, 1 Hz, 9 × 10 minutes over 3h) before microarray-profiling was performed. Proteoglycan synthesis was determined by 35S-sulfate-incorporation over 24 hours. Protein alterations were determined by Western blotting.ResultsCell viability and hardness of constructs were unaltered by dynamic compression while proteoglycan synthesis was significantly stimulated (1.45-fold, p = 0.016). Among 115 significantly regulated genes, 114 were up-regulated, 48 of them > two-fold. AP-1-relevant transcription factors FOSB and FOS strongly increased in line with elevated ERK1/2-phosphorylation and rising MAP3K4 expression. Expression of proteoglycan-synthesizing enzymes CHSY1 and GALNT4 was load-responsive as were factors associated with the MAPK-, TGF-β-, calcium-, retinoic-acid-, Wnt- and Notch-signaling pathway which were significantly altered. SOX9, BMP4 and BMP6 levels rose significantly also after multiple loading episodes at daily intervals even at the 14th cycle with no indication for desensitation. Canonical pSmad2/3 and pSmad1/5/9-signalling showed no consistent regulation.ConclusionThis study associates novel genes with mechanoregulation in chondrocytes, raising SOX9 protein levels with anabolic loading and suggests that more pathways than so far anticipated apparently work together in a complex network of stimulators and feedback-regulators. Knowledge on time evolution of mechanosensitive indicators responding to anabolic loading is crucial to maximize cartilage matrix-deposition for the generation of high-level cartilage replacement tissue. This article is protected by copyright. All rights reserved
      PubDate: 2017-03-28T06:55:27.141559-05:
      DOI: 10.1002/jcp.25933
  • The Role of Runx2 in Facilitating Autophagy in Metastatic Breast Cancer
    • Authors: Manish Tandon; Ahmad H. Othman, Vivek Ashok, Gary S. Stein, Jitesh Pratap
      Abstract: Breast cancer metastases cause significant patient mortality. During metastases, cancer cells use autophagy, a catabolic process to recycle nutrients via lysosomal degradation, to overcome nutritional stress for their survival. The Runt-related transcription factor, Runx2, promotes cell survival under metabolic stress and regulates breast cancer progression and bone metastases. Here, we identify that Runx2 enhances autophagy in metastatic breast cancer cells. We defined Runx2 function in cellular autophagy by monitoring microtubule-associated protein light chain (LC3B-II) levels, an autophagy-specific marker. The electron and confocal microscopic analyses were utilized to identify alterations in autophagic vesicles. The Runx2 knockdown cells accumulate LC3B-II protein and autophagic vesicles due to reduced turnover. Interestingly, Runx2 promotes autophagy by enhancing trafficking of LC3B vesicles. Our mechanistic studies revealed that Runx2 promotes autophagy by increasing acetylation of α-tubulin sub-units of microtubules. Inhibiting autophagy decreased cell adhesion and survival of Runx2 knockdown cells. Furthermore, analysis of LC3B protein in clinical breast cancer specimens and tumor xenografts revealed significant association between high Runx2 and low LC3B protein levels. Our studies reveal a novel regulatory mechanism of autophagy via Runx2 and provide molecular insights into the role of autophagy in metastatic cancer cells. This article is protected by copyright. All rights reserved
      PubDate: 2017-03-27T05:41:18.712795-05:
      DOI: 10.1002/jcp.25916
  • NLRP3 Inflammasome: Its Regulation and Involvement in Atherosclerosis
    • Authors: Zahra Hoseini; Fatemeh Sepahvand, Bahman Rashidi, Amirhossein Sahebkar, Aria Masoudifar, Hamed Mirzaei
      Abstract: Inflammasomes are intracellular complexes involved in the innate immunity that convert proIL-1β and proIL-18 to mature forms and initiate pyroptosis via cleaving procaspase-1. The most well-known inflammasome is NLRP3. Several studies have indicated a decisive and important role of NLRP3 inflammasome, IL-1β, IL-18, and pyroptosis in atherosclerosis. Modern hypotheses introduce atherosclerosis as an inflammatory/lipid-based disease and NLRP3 inflammasome has been considered as a link between lipid metabolism and inflammation because crystalline cholesterol and oxidized low-density lipoprotein (oxLDL) (two abundant components in atherosclerotic plaques) activate NLRP3 inflammasome. In addition, oxidative stress, mitochondrial dysfunction, endoplasmic reticulum (ER) stress, and lysosome rupture, which are implicated in inflammasome activation, have been discussed as important events in atherosclerosis. In spite of these clues, some studies have reported that NLRP3 inflammasome has no significant effect in atherogenesis. Our review reveals that some molecules such as JNK-1 and ASK-1 (upstream regulators of inflammasome activation) can reduce atherosclerosis through inducing apoptosis in macrophages. Notably, NLRP3 inflammasome can also cause apoptosis in macrophages, suggesting that NLRP3 inflammasome may mediate JNK-induced apoptosis, and the apoptotic function of NLRP3 inflammasome may be a reason for the conflicting results reported. The present review shows that the role of NLRP3 in atherogenesis can be significant. Here, the molecular pathways of NLRP3 inflammasome activation and the implications of this activation in atherosclerosis are explained. This article is protected by copyright. All rights reserved
      PubDate: 2017-03-27T05:12:57.347885-05:
      DOI: 10.1002/jcp.25930
  • The Diversified Function and Potential Therapy of Ectopic Olfactory
           Receptors in Non-olfactory Tissues
    • Authors: Zhe Chen; Hong Zhao, Nian Fu, Linxi Chen
      Abstract: Olfactory receptors (ORs) are mainly distributed in olfactory neurons and play a key role in detecting volatile odorants, eventually resulting in the production of smell perception. Recently, it is also reported that ORs are expressed in non-olfactory tissues including heart, lung, sperm, skin and cancerous tissues. Interestingly, ectopic ORs are associated with the development of diseases in non-olfactory tissues. For instance, ectopic ORs initiate the hypoxic ventilatory responses and maintain the oxygen homeostasis of breathing in the carotid body when oxygen levels decline. Ectopic ORs induce glucose homeostasis in diabetes. Ectopic ORs regulate systemic blood pressure by increasing renin secretion and vasodilation. Ectopic ORs participate in the process of tumor cell proliferation, apoptosis, metastasis and invasiveness. Ectopic ORs accelerate the occurrence of obesity, angiogenesis and wound-healing processes. Ectopic ORs affect fetal hemoglobin levels in sickle cell anemia and thalassemia. Finally, we also elaborate some ligands targeting for ORs. Obviously, the diversified function and related signal pathway of ectopic ORs may play a potential therapeutic target in non-olfactory tissues. Thus, this review focuses on the latest research results about the diversified function and therapeutic potential of ectopic ORs in non-olfactory tissues. This article is protected by copyright. All rights reserved
      PubDate: 2017-03-24T06:40:33.749115-05:
      DOI: 10.1002/jcp.25929
  • Fasting Up-regulates Ferroportin 1 Expression via a Ghrelin/GHSR/MAPK
           Signaling Pathway
    • Authors: Qian-Qian Lou; Yu-Fu Zhou, Mesona Yung-Jin Chen, Li Liu, Juan Ma, Meng-Wan Zhang, Fa-Li Zhang, Ya Ke, Zhong-Ming Qian
      Abstract: The significant positive correlation between ghrelin and iron and hepcidin levels in the plasma of children with iron deficiency anemia prompted us to hypothesize that ghrelin may affect iron metabolism. Here, we investigated the effects of fasting or ghrelin on the expression of hepcidin, ferroportin 1 (Fpn1), transferrin receptor 1 (TfR1), ferritin light chain (Ft-L) proteins and ghrelin, and also hormone secretagogue receptor 1 alpha (GHSR1α) and ghrelin O-acyltransferase (GOAT) mRNAs in the spleen and/or macrophage. We demonstrated that fasting induces a significant increase in the expression of ghrelin, GHSR1α, GOAT and hepcidin mRNAs, as well as Ft-L and Fpn1 but not TfR1 proteins in the spleens of mice in vivo. Similar to the effects of fasting on the spleen, ghrelin induced a significant increase in the expression of Ft-L and Fpn1 but not TfR1 proteins in macrophages in vitro. In addition, ghrelin was found to induce a significant enhancement in phosphorylation of ERK as well as translocation of pERK from the cytosol to nuclei. Furthermore, the increased pERK and Fpn1 induced by ghrelin was demonstrated to be preventable by pre-treatment with either GHSR1α antagonist or pERK inhibitor. Our findings support the hypothesis that fasting up-regulates Fpn1 expression, probably via a ghrelin/GHSR/MAPK signaling pathway. This article is protected by copyright. All rights reserved
      PubDate: 2017-03-24T06:40:31.342975-05:
      DOI: 10.1002/jcp.25931
  • miR-124 and miR-9 Mediated Downregulation of HDAC5 Promotes Neurite
           Development Through Activating MEF2C- GPM6A Pathway
    • Authors: Xi Gu; Congcong Fu, Lifang Lin, Shuhu Liu, Xiaohong Su, Aili Li, Qiaoqi Wu, Chunhong Jia, Peidong Zhang, Lu Chen, Xinhong Zhu, Xuemin Wang
      Abstract: The class IIa histone deacetylases (HDACs) play important roles in the central nervous system during diverse biological processes such as synaptic plasticity, axon regeneration, cell apoptosis, and neural differentiation. Although it is known that HDAC5 regulates neuronal differentiation, neither the physiological function nor the regulation of HDAC5 in neuronal differentiation is clear. Here, we identify HDAC5 as an inhibitor of neurite elongation and show that HDAC5 is regulated by the brain enriched microRNA miR-124 and miR-9. We discover that HDAC5 inhibits neurite extension both in differentiated P19 cells and primary neurons. We also show that the neuronal membrane glycoprotein GPM6A (M6a) is a direct target gene of HDAC5 regulated transcriptional factor MEF2C. HDAC5 inhibits neurite elongation, acting at least partially via a MEF2C/M6a signaling pathway. We also confirmed the miR-124/miR-9 regulated HDAC5-MEF2C-M6a pathway regulates neurite development in primary neurons. Thus, HDAC5 emerges as a cellular conductor of MEF2C and M6a activity and is regulated by miR-124 and miR-9 to control neurite development. This article is protected by copyright. All rights reserved
      PubDate: 2017-03-23T06:15:41.705303-05:
      DOI: 10.1002/jcp.25927
  • Dual-specificity Tyrosine Phosphorylation-regulated Kinase 2 Regulates
           Osteoclast Fusion in a Cell Heterotypic Manner
    • Authors: Gali Guterman-Ram; Milena Pesic, Ayelet Orenbuch, Tal Czeiger, Anastasia Aflalo, Noam Levaot
      Abstract: Monocyte fusion into osteoclasts, bone resorbing cells, plays a key role in bone remodeling and homeostasis; therefore, aberrant cell fusion may be involved in a variety of debilitating bone diseases. Research in the last decade has led to the discovery of genes that regulate osteoclast fusion, but the basic molecular and cellular regulatory mechanisms underlying the fusion process are not completely understood. Here, we reveal a role for Dyrk2 in osteoclast fusion. We demonstrate that Dyrk2 down regulation promotes osteoclast fusion, whereas its overexpression inhibits fusion. Moreover, Dyrk2 also promotes the fusion of foreign-body giant cells, indicating that Dyrk2 plays a more general role in cell fusion. In an earlier study, we showed that fusion is a cell heterotypic process initiated by fusion-founder cells that fuse to fusion-follower cells, the latter of which are unable to initiate fusion. Here, we show that Dyrk2 limits the expansion of multinucleated founder cells through the suppression of the fusion competency of follower cells. This article is protected by copyright. All rights reserved
      PubDate: 2017-03-23T06:15:39.506116-05:
      DOI: 10.1002/jcp.25922
  • Involvement of Testicular DAAM1 Expression in Zinc Protection Against
           Cadmium-induced Male Rat Reproductive Toxicity
    • Authors: Marouane Chemek; Massimo Venditti, Sana Boughamoura, Safa Ben Mimouna, Imed Messaoudi, Sergio Minucci
      Abstract: In order to verify the effects of exposure to Cd and Zn on testicular DAAM1 gene and protein expression and also to ascertain their involvement in the protective role of Zn in prevent the testicular toxicity Cd-induced in male offspring rats at adult age after gestational and lactational exposure, male offspring rats, from mothers receiving either tap water, Cd, Zn or Cd + Zn during gestation and lactation periods, were scarified on postnatal days (PND) 70. The reproductive organ (testis, epididymis and vesicle seminal) were collected, weighed and analysed. The results showed that exposure to Cd in utero and through lactation decreased the relative reproductive organ weight, altered the testicular histology at the interstitial and tubular levels, and causing a significant reduction in the daily sperm production (DSP) per testis and per gram of testis, and other then altering the epididymal sperm quality. Furthemore, both mRNA and protein expression of rat testicular DAAM1 were also inhibited in Cd-treated group. Zn supply has completely corrected the most of these toxic effects. Our results imply that Zn could prevent Cd-induced testicular toxicity and sperm quality alteration in adult male rat after gestational and lactational exposure, probably via the restoration of the testicular DAAM1 expression inhibited by Cd. This article is protected by copyright. All rights reserved
      PubDate: 2017-03-23T03:50:58.697216-05:
      DOI: 10.1002/jcp.25923
  • Retinoic Acid Receptor-related Orphan Receptor RORα Regulates
           Differentiation and Survival of Keratinocytes During Hypoxia
    • Authors: Hongyu Li; Longjian Zhou, Jun Dai
      Abstract: Low O2 pressures present in the microenvironment of epidermis control keratinocyte differentiation and epidermal barrier function through hypoxia inducible factors (HIFs) dependent gene expression. This study focuses on investigating relations of the retinoic acid receptor-related orphan receptor alpha (RORα) to HIF-1α in keratinocytes under hypoxic conditions. The expression level of RORα is significantly elevated under hypoxia in both human and murine keratinocytes. Gene silencing of RORA attenuates hypoxia-stimulated expression of genes related to late differentiation and epidermal barrier function, and leads to an enhanced apoptotic response. While the hypoxic induction of RORα is dependent on HIF-1α, RORα is in turn critical for nuclear accumulation of HIF-1α and activation of HIF transcriptional activity. These results collectively suggest that RORα functions as an important mediator of HIF-1α activities in regulating keratinocyte differentiation/survival and epidermal barrier function during the oxygen sensing stage. This article is protected by copyright. All rights reserved
      PubDate: 2017-03-23T03:50:27.051248-05:
      DOI: 10.1002/jcp.25924
  • Primary Cilium Alterations and Expression Changes of Patched1 Proteins in
           Niemann-pick Type C Disease
    • Authors: Patrizia Formichi; Carla Battisti, Maria Margherita De Santi, Raffaella Guazzo, Sergio Antonio Tripodi, Elena Radi, Benedetta Rossi, Ermelinda Tarquini, Antonio Federico
      Abstract: Niemann-Pick type C disease (NPC) is a disorder characterized by abnormal intracellular accumulation of unesterified cholesterol and glycolipids. Two distinct disease-causing genes have been isolated, NPC1 and NPC2. The NPC1 protein is involved in the sorting and recycling of cholesterol and glycosphingolipids in the late endosomal/lysosomal system. It has extensive homology with the Patched 1 (Ptc1) receptor, a transmembrane protein localized in the primary cilium and involved in the Hedgehog signaling (Shh) pathway.We assessed the presence of NPC1 and Ptc1 proteins and evaluated the relative distribution and morphology of primary cilia in fibroblasts from five NPC1 patients and controls, and in normal fibroblasts treated with 3-ß-[2-(diethylamino)ethoxy]androst-5-en-17-one (U18666A), a cholesterol transport-inhibiting drug that is widely used to mimic NPC. Immunofluorescence and western blot analyses showed a significant decrease in expression of NPC1 and Ptc1 in NPC1 fibroblasts, while they were normally expressed in U18666A-treated fibroblasts. Moreover, fibroblasts from NPC1 patients and U18666A-treated cells showed a lower percentage distribution of primary cilia and a significant reduction in median cilia length with respect to controls. These are the first results demonstrating altered cytoplasmic expression of Ptc1 and reduced number and length of primary cilia, where Ptc1 is located, in fibroblasts from NPC1 patients. We suggest that the alterations in Ptc1 expression in cells from NPC1 patients are closely related to NPC1 expression deficit, while the primary cilia alterations observed in NPC1 and U18666A-treated fibroblasts may represent a secondary event derived from a defective metabolic pathway. This article is protected by copyright. All rights reserved
      PubDate: 2017-03-23T03:46:12.925397-05:
      DOI: 10.1002/jcp.25926
  • Mechanism of Prolactin Inhibition of miR-135b via Methylation in Goat
           Mammary Epithelial Cells
    • Authors: Zhi Chen; Jun Luo, ChangHui Zhang, Yue Ma, Shuang Sun, Tianyin Zhang, Juan J. Loor
      Abstract: Prolactin is an important endocrine activator of lactogenesis. This study investigated the function and mechanism of miR-135b in the enhancement of lactation by prolactin in goat mammary epithelial tissue. We utilized S-Poly (T) sequencing to evaluate changes in gene regulation in the goat mammary gland after incubation with 2.5 ug/ml prolactin and 2.5 ug/ml IGF-1 by examining highly expressed miRNAs during early-lactation and late-lactation. The results illustrated that miR-135b is highly expressed in the goat mammary gland during early-lactation and late-lactation, and also after treatment with 2.5 ug/ml prolactin and 2.5 ug/ml IGF-1. We used Q-RT PCR, Western Blot, immunofluorescence, and luciferase reporter assay analysis, and found that PRL was significantly down-regulated in response to the expression of miR-135b in a manner that was functionally related to TAG synthesis via the large tumor suppressor 2 gene (Lats2), an important regulator of adipocyte proliferation via Hippo Signaling. Furthermore, using bisulfite-sequencing PCR (BSP), Q-PCR and Western Blot we discovered an increase in expression of DNMT I (DNA methyl transferase I) in goat mammary epithelial cells with the 2.5 ug/ml PRL incubation, which led to DNA methylation of the CpG island upstream of miR-135b and inhibited the transcription and expression of miR-135b. This article is protected by copyright. All rights reserved
      PubDate: 2017-03-23T03:46:09.289111-05:
      DOI: 10.1002/jcp.25925
  • Autophagy Maintains the Integrity of Endothelial Barrier in LPS-induced
           Lung Injury
    • Authors: Dan Zhang; Jian Zhou, Le Chi Ye, Jing Li, Zhenzhou Wu, Yuping Li, Chichi Li
      Abstract: Understanding the role and underlying regulation mechanism of autophagy in lipopolysaccharide-induced lung injury (LPS-LI) may provide potentially new pharmacological targets for treatment of acute lung injury. The aim of this study was to investigate the functional significance of autophagy in LPS-LI. The autophagy of human pulmonary microvascular endothelial cells (HPMVECs) and mice was inhibited before they were challenged with LPS. In vitro, permeability, vitality and the lactate dehydrogenase (LDH) release rate of the cells were detected, the zonula occluden-1 (ZO-1) expression and the stress fiber formation were determined. In vivo, the lung injury was assessed. We found LPS caused high permeability and increased LDH release rate, lowered viability of the cells, inhibited the ZO-1 expression and induced stress fiber formation, these effects were further aggravated by prohibiting the level of autophagy. Consistently, in in vivo experiments, LPS induced serious lung injury, which was reflected as edema, leukocyte infiltration and hemorrhage in lung tissue, and the high concentration of pro-inflammation cytokines tumor necrosis factor (TNF)-α and interleukin (IL)-1β in bronchoalveolar lavage fluid (BALF). Inhibiting autophagy further exacerbated LPS-LI. It appears that autophagy played a protective role in LPS-LI in part through restricting the injury of lung microvascular barrier. This article is protected by copyright. All rights reserved
      PubDate: 2017-03-22T05:01:27.054343-05:
      DOI: 10.1002/jcp.25928
  • Gax Suppresses Chemerin/CMKLR1-induced Preadipocyte Biofunctions Through
           the Inhibition of Akt/mTOR and ERK Signaling Pathways
    • Authors: Yunqi Jiang; Ping Liu, Wenlin Jiao, Juan Meng, Jinbo Feng
      Abstract: Adipose tissue is closely associated with angiogenesis and vascular remodeling. Chemerin is involved in inflammatory reaction and vascular dysfunction. However, the mechanisms of chemerin participating in vascular remodeling and whether Growth arrest–specific homeobox (Gax) can effectively intervene it remain obscured. Here, 3T3-F442A preadipocytes were cultured, injected into athymic mice to model fat pads, and treated respectively with Ad-chemerin, Ad-Gax, or specific inhibitors in vitro and in vivo. MTT, flow cytometry, Western blotting, and imunohisto(cyto)-chemistry analyses showed that chemerin enhanced the expression of FABP4 and VEGF, activated Akt/mTOR and ERK pathways, increased the cell percent of S phase, decreased the percent of G0-G1 phase and apoptotic cells, and augmented neovascular density in fat pads. Inversely, Gax suppressed the expression of these adipogenic and vasifactive markers and these signaling proteins, decreased the percent of S phase cells, and increased those of G0-G1 phase and apoptotic cells, and reduced the neovascular density. Our results indicate that chemerin-CMKLR1 activates Akt/mTOR and ERK pathways and facilitates preadipocyte proliferation, adipogenesis, and angiogenesis. Contrarily, Gax weakens the effect of chemerin on preadipocyte biofunctions. This article is protected by copyright. All rights reserved
      PubDate: 2017-03-21T23:01:57.895306-05:
      DOI: 10.1002/jcp.25918
  • All-trans Retinoic Acid Ameliorates Hepatic Stellate Cell Activation via
           Suppression of Thioredoxin Interacting Protein Expression
    • Authors: Hiroki Shimizu; Toshiaki Tsubota, Keita Kanki, Goshi Shiota
      Abstract: Activation of hepatic stellate cells (HSCs) is the effector factor of hepatic fibrosis and hepatocellular carcinoma (HCC) development. Accumulating evidence suggests that retinoic acids (RAs), derivatives of vitamin A, contribute to prevention of liver fibrosis and carcinogenesis, however, regulatory mechanisms of RAs still remain exclusive. To elucidate RA signaling pathway, we previously performed a genome-wide screening of RA-responsive genes by in silico analysis of RA-response elements, and identified 26 RA-responsive genes. We found that thioredoxin interacting protein (TXNIP), which inhibits antioxidant activity of thioredoxin (TRX), was downregulated by all-trans retinoic acid (ATRA). In the present study, we demonstrate that ATRA ameliorates activation of HSCs through TXNIP suppression. HSC activation was attenuated by TXNIP downregulation, whereas potentiated by TXNIP upregulation, indicating that TXNIP plays a crucial role in activation of HSCs. Notably, we showed that TXNIP-mediated HSC activation was suppressed by antioxidant N-acetylcysteine. In addition, ATRA treatment or downregulation of TXNIP clearly declined oxidative stress levels in activated HSCs. These data suggest that ATRA plays a key role in inhibition of HSC activation via suppressing TXNIP expression, which reduces oxidative stress levels. This article is protected by copyright. All rights reserved
      PubDate: 2017-03-21T07:27:39.140761-05:
      DOI: 10.1002/jcp.25921
  • TGF-β Inhibits Osteogenesis by Upregulating the Expression of Ubiquitin
           Ligase SMURF1 via MAPK-ERK Signaling
    • Authors: Xuewu Sun; Ziang Xie, Yan Ma, Xin Pan, Jiying Wang, Zhijun Chen, Peihua Shi
      Abstract: High incidence of osteoporotic fractures emphasizes the necessity of developing effective measures to promote osteogenesis. In our study, we investigated a possible role of MAPK-ERK signaling in the TGF-β-mediated osteoblastic differentiation. Our results indicated that TGF-β activated the MAPK-ERK pathway and inhibited osteogenesis in mesenchymal pluripotent cell line, C3H10T1/2, and preosteoblastic cell line, MC3T3 cells. And the downregulation of MAPK-ERK signaling using pharmacological inhibitor U0126 and RNA interference rescued osteoblast differentiation suppressed by TGF-β, which was confirmed by Alkaline phosphatase (ALP) staining and alizarrn red staining and the enhanced expression of osteogenesic markers. Western blotting analysis indicated that TGF-β induced protein expression of E3 ubiquitin-protein ligase SMURF1, which contributed to the degradation of RUNX2 and SMAD1 as evidenced by SMURF1 inhibition using RNA interference and proteasome inhibitor MG132. Moreover, we observed that the expression of SMURF1 was decreased, while that of SMAD1 and RUNX2 increased by MAPK-ERK inhibitor U0126 in TGF-β-treated differentiating preosteoblasts, suggesting that MAPK-ERK regulated the transcription of osteogenesis-related genes. Furthermore, a synergistic effect between U0126 and bone morphogenic protein (BMP)-2 on osteoblast differentiation and bone formation was observed both in cell cultures and experimental animals. In conclusion, our results revealed that TGF-β inhibited osteoblastic differentiation by inducing the MAPK-ERK pathway which upregulated the expression of ubiquitin ligase SMURF1 and resulted in reduced presence of osteogenic proteins. In addition, the potentiation of BMP-2 on osteogenic activity by ERK1/2 inhibitor U0126 suggests that it may have potential clinical utility for promoting osteogenesis in bone fracture repair. This article is protected by copyright. All rights reserved
      PubDate: 2017-03-21T07:27:31.075954-05:
      DOI: 10.1002/jcp.25920
  • Regulation Role of CRTC3 in Skeletal Muscle and Adipose Tissue
    • Authors: Jiaqi Liu; Ziye Xu, Weiche Wu, Yizhen Wang, Tizhong Shan
      Abstract: The cyclic adenosine monophosphate (cAMP) - protein kinase A (PKA) signaling pathway plays important role in regulating energy homeostasis. Many of the effects of the cAMP-PKA signaling is mediated through the cAMP responsive element binding protein (CREB) and its coactivator CREB-regulated transcription coactivators (CRTCs). CRTC3 is a member of CRTCs family proteins and plays important roles in glucose and energy metabolism. Previous studies shown that global knockout of CRTC3 enhances oxygen consumption and energy expenditure and subsequently protects the knockout animal against obesity. In skeletal muscle, CRTC3 affects lipid and glycogen metabolism and mitochondrial biogenesis. In white adipocytes, CRTC3 regulates GLUT4 expression and glucose uptake. More recently, the localization and function of CRTC3 in brown fat have been reported. In this review, we mainly discuss the regulatory role of CRTC3 in skeletal muscle and adipose tissues. This article is protected by copyright. All rights reserved
      PubDate: 2017-03-21T07:21:11.234825-05:
      DOI: 10.1002/jcp.25917
  • Transplantation of Mesenchymal Stem Cells Overexpressing IL10 Attenuates
           Cardiac Impairments in Rats with Myocardial Infarction
    • Authors: Xin Meng; Jianping Li, Ming Yu, Jian Yang, Minjuan Zheng, Jinzhou Zhang, Chao Sun, Hongliang Liang, Liwen Liu
      Abstract: Mesenchymal stem cell (MSC) has been well known to exert therapeutic potential for patients with myocardial infarction (MI). In addition, interleukin-10 (IL10) could attenuate MI through suppressing inflammation. Thus, the combination of MSC implantation with IL10 delivery may extend health benefits to ameliorate cardiac injury after MI. Here we established overexpression of IL10 in bone marrow-derived MSC through adenoviral transduction. Cell viability, apoptosis and IL10 secretion under ischemic challenge in vitro were examined. In addition, MSC was transplanted into the injured hearts in a rat model of MI. Four weeks after the MI induction, myocardial infarction, cardiac functions, apoptotic cells and inflammation cytokines were assessed. In response to in vitro oxygen-glucose deprivation (OGD), IL10 overexpression in MSC (Ad.IL10-MSC) enhanced cell viability, decreased apoptosis and increased IL10 secretion. Consistently, the implantation of Ad.IL10-MSCs into MI animals resulted in more reductions in myocardial infarct size, cardiac impairment and cell apoptosis, compared to the individual treatments of either MSC or IL10 administration. Moreover, the attenuation of both systemic and local inflammations was most prominent for Ad.IL10-MSC treatment. IL10 overexpression and MSC may exert a synergistic anti-inflammatory effect to alleviate cardiac injury after MI. This article is protected by copyright. All rights reserved
      PubDate: 2017-03-21T07:20:58.420555-05:
      DOI: 10.1002/jcp.25919
  • Identification and Characterization of Site-specific N-glycosylation in
           the Potassium Channel Kv3.1b
    • Authors: Paul Christian Vicente; Jin Young Kim, Jeong-Ju Ha, Min-Young Song, Hyun-Kyung Lee, Dong-Hyun Kim, Jin-Sung Choi, Kang-Sik Park
      Abstract: The potassium ion channel Kv3.1b is a member of a family of voltage-gated ion channels that are glycosylated in their mature form. In the present study, we demonstrate the impact of N-glycosylation at specific asparagine residues on the trafficking of the Kv3.1b protein. Large quantities of asparagine 229 (N229)-glycosylated Kv3.1b reached the plasma membrane, whereas N220-glycosylated and unglycosylated Kv3.1b were mainly retained in the endoplasmic reticulum (ER). These ER-retained Kv3.1b proteins were susceptible to degradation when co-expressed with calnexin, whereas Kv3.1b pools located at the plasma membrane were resistant. Mass spectrometry analysis revealed a complex type Hex3HexNAc4Fuc1 glycan as the major glycan component of the N229-glycosylated Kv3.1b protein, as opposed to a high-mannose type Man8GlcNAc2 glycan for N220-glycosylated Kv3.1b. Taken together, these results suggest that trafficking-dependent roles of the Kv3.1b potassium channel are dependent on N229 site-specific glycosylation and N-glycan structure, and operate through a mechanism whereby specific N-glycan structures regulate cell surface expression. This article is protected by copyright. All rights reserved
      PubDate: 2017-03-21T07:20:43.960399-05:
      DOI: 10.1002/jcp.25915
  • HDAC1 and HDAC3 Underlie Dynamic H3K9 Acetylation During Embryonic
           Neurogenesis and in Schizophrenia-like Animals
    • Authors: Josef Večeřa; Eva Bártová, Jana Krejčí, Soňa Legartová, Denisa Komůrková, Jana Rudá-Kučerová, Tibor Štark, Eva Dražanová, Tomáš Kašpárek, Alexandra Šulcová, Frank J. Dekker, Wiktor Szymanski, Christian Seiser, Georg Weitzer, Raphael Mechoulam, Vincenzo Micale, Stanislav Kozubek
      Abstract: Although histone acetylation is one of the most widely studied epigenetic modifications, there is still a lack of information regarding how the acetylome is regulated during brain development and pathophysiological processes. We demonstrate that the embryonic brain (E15) is characterized by an increase in H3K9 acetylation as well as decreases in the levels of HDAC1 and HDAC3. Moreover, experimental induction of H3K9 hyperacetylation led to the overexpression of NCAM in the embryonic cortex and depletion of Sox2 in the subventricular ependyma, which mimicked the differentiation processes. Inducing differentiation in HDAC1-deficient mouse ESCs resulted in early H3K9 deacetylation, Sox2 downregulation and enhanced astrogliogenesis, whereas neuro-differentiation was almost suppressed. Neuro-differentiation of (wt) ESCs was characterized by H3K9 hyperacetylation that was associated with HDAC1 and HDAC3 depletion. Conversely, the hippocampi of schizophrenia-like animals showed H3K9 deacetylation that was regulated by an increase in both HDAC1 and HDAC3. The hippocampi of schizophrenia-like brains that were treated with the cannabinoid receptor-1 inverse antagonist AM251 expressed H3K9ac at the level observed in normal brains. Together, the results indicate that co-regulation of H3K9ac by HDAC1 and HDAC3 is important to both embryonic brain development and neuro-differentiation as well as the pathophysiology of a schizophrenia-like phenotype. This article is protected by copyright. All rights reserved
      PubDate: 2017-03-16T07:06:05.26055-05:0
      DOI: 10.1002/jcp.25914
  • Rapamycin Attenuates BAFF-extended Proliferation and Survival via
           Disruption of mTORC1/2 Signaling in Normal and Neoplastic B-lymphoid Cells
    • Authors: Qingyu Zeng; Shanshan Qin, Hai Zhang, Beibei Liu, Jiamin Qin, Xiaoxue Wang, Ruijie Zhang, Chunxiao Liu, Xiaoqing Dong, Shuangquan Zhang, Shile Huang, Long Chen
      Abstract: B cell activating factor from the TNF family (BAFF) stimulates B-cell proliferation and survival, but excessive BAFF promotes the development of aggressive B cells leading to malignant and autoimmune diseases. Recently, we have reported that rapamycin, a macrocyclic lactone, attenuates human soluble BAFF (hsBAFF)-stimulated B-cell proliferation/survival by suppressing mTOR-mediated PP2A-Erk1/2 signaling pathway. Here, we show that the inhibitory effect of rapamycin on hsBAFF-promoted B cell proliferation/survival is also related to blocking hsBAFF-stimulated phosphorylation of Akt, S6K1 and 4E-BP1, as well as expression of survivin in normal and B-lymphoid (Raji and Daudi) cells. It appeared that both mTORC1 and mTORC2 were involved in the inhibitory activity of rapamycin, as silencing raptor or rictor enhanced rapamycin's suppression of hsBAFF-induced survivin expression and proliferation/viability in B cells. Also, PP242, an mTORC1/2 kinase inhibitor, repressed survivin expression and cell proliferation/viability more potently than rapamycin (mTORC1 inhibitor) in B cells in response to hsBAFF. Of interest, ectopic expression of constitutively active Akt (myr-Akt) or constitutively active S6K1 (S6K1-ca), or downregulation of 4E-BP1 conferred resistance to rapamycin's attenuation of hsBAFF-induced survivin expression and B-cell proliferation/viability, whereas overexpression of dominant negative Akt (dn-Akt) or constitutively hypophosphorylated 4E-BP1 (4EBP1-5A), or downregulation of S6K1, or co-treatment with Akt inhibitor potentiated the inhibitory effects of rapamycin. The findings indicate that rapamycin attenuates excessive hsBAFF-induced cell proliferation/survival via blocking mTORC1/2 signaling in normal and neoplastic B-lymphoid cells. Our data underscore that rapamycin may be a potential agent for preventing excessive BAFF-evoked aggressive B-cell malignancies and autoimmune diseases. This article is protected by copyright. All rights reserved
      PubDate: 2017-03-16T07:00:45.056107-05:
      DOI: 10.1002/jcp.25913
  • Knockout of ATG5 Leads to Malignant Cell Transformation and Resistance to
           Src Family Kinase Inhibitor PP2
    • Authors: Sung-Hee Hwang; Byeal-I Han, Michael Lee
      Abstract: Autophagy can either promote or inhibit cell death in different cellular contexts. In this study, we investigated the role of autophagy in ATG5 knockout (KO) cell line established using CRISPR/Cas9 system. In ATG5 KO cells, RT-PCR and immunoblot of LC3 confirmed the functional gene knockout. We found that knockout of ATG5 significantly increased proliferation of NIH 3T3 cells. In particular, autophagy deficiency enhanced susceptibility to cellular transformation as determined by an in vitro clonogenic survival assay and a soft agar colony formation assay. We also found that ATG5 KO cells had a greater migration ability as compared to wild-type (WT) cells. Moreover, ATG5 KO cells were more resistant to treatment with a Src family tyrosine kinase inhibitor (PP2) than WT cells were. Cyto-ID Green autophagy assay revealed that PP2 failed to induce autophagy in ATG5 KO cells. PP2 treatment decreased the percentage of cells in the S and G2/M phases among WT cells but had no effect on cell cycle distribution of ATG5 KO cells, which showed a high percentage of cells in the S and G2/M phases. Additionally, the proportion of apoptotic cells significantly decreased after treatment of ATG5 KO cells with PP2 in comparison with WT cells. We found that expression levels of p53 were much higher in ATG5 KO cells. The ATG5 KO seems to lead to compensatory upregulation of the p53 protein because of a decreased apoptosis rate. Taken together, our results suggest that autophagy deficiency can lead to malignant cell transformation and resistance to PP2. This article is protected by copyright. All rights reserved
      PubDate: 2017-03-15T06:55:54.137758-05:
      DOI: 10.1002/jcp.25912
  • Three-dimensional Culture of Buffalo Granulosa Cells in Hanging Drop
           Mimics the Preovulatory Follicle Stage
    • Authors: Monica Yadav; Himanshu Agrawal, Mamta Pandey, Dheer Singh, Suneel Kumar Onteru
      Abstract: Granulosa cell (GC) culture models mimicking the intrafollicular environment are limited. Such models have a great potential in reproductive toxicity studies. The buffalo, a monovulatory species like humans, could be a better model than polyovulatory rodents. Therefore, we targeted the development and characterization of three-dimensional (3D) culture systems for buffalo GCs. The GCs from small ovarian follicles (SF) maintained the CYP19 gene expression for 144 h in a 2D culture system. Hence, GCs from SF were cultured directly in 3D using hanging drop and Poly-{(2-hydroxyethyl methacrylate)} (polyHEMA) methods in the DMEM media containing 1ng/mL FSH and 10 ng/mL IGF-1 for 144 h. The expression profile of nine GC-specific transcripts; CYP19, TNFAIP6, AMH, PTI, NR4A1, FSHR, RUNX, LHR and COX2/PTGS2; revealed that 3D-spheroids developed in hanging drop method maintained the GC phenotype of preovulatory follicles. Therefore, hanging drop method is a best method for culturing GCs to mimic the intrafollicular environment. This article is protected by copyright. All rights reserved
      PubDate: 2017-03-15T06:55:50.136304-05:
      DOI: 10.1002/jcp.25909
  • Autophagy Mediates Cytotoxicity of Human Colorectal Cancer Cells Treated
           with Garcinielliptone FC
    • Authors: Shen-Jeu Won; Cheng-Hsin Yen, Ting-Yu Lin, Ya-Fen Jiang-Shieh, Chun-Nan Lin, Jyun-Ti Chen, Chun-Li Su
      Abstract: The tautomeric pair of garcinielliptone FC (GFC) is a novel tautomeric pair of polyprenyl benzophenonoid isolated from the pericarps of Garcinia subelliptica Merr. (G. subelliptica, Clusiaceae), a tree with abundant sources of polyphenols. Our previous report demonstrated that GFC induced apoptosis on various types of human cancer cell lines including chemoresistant human colorectal cancer HT-29 cells. In the present study, we observed that many autophagy-related genes in GFC-treated HT-29 cells were up- and down-regulated using a cDNA microarray containing oncogenes and kinase genes. GFC-induced autophagy of HT-29 cells was confirmed by observing the formation of acidic vesicular organelles, LC3 puncta, and double-membrane autophagic vesicles using flow cytometry, confocal microscopy, and transmission electron microscopy, respectively. Inhibition of AKT/mTOR/P70S6K signaling as well as formation of Atg5-Atg12 and PI3K/Beclin-1 complexes were observed using Western blot. Administration of autophagy inhibitor (3-methyladenine and shRNA Atg5) and apoptosis inhibitor Z-VAD showed that the GFC-induced autophagy was cytotoxic form and GFC-induced apoptosis enhanced GFC-induced autophagy. Our data suggest the involvement of autophagy and apoptosis in GFC-induced anticancer mechanisms of human colorectal cancer. This article is protected by copyright. All rights reserved
      PubDate: 2017-03-15T06:55:39.862725-05:
      DOI: 10.1002/jcp.25910
  • Quantification of Exosomes
    • Authors: Erik H. Koritzinsky; Jonathan M. Street, Robert A. Star, Peter S.T. Yuen
      Abstract: Exosomes are released by cells as self-contained vesicles with an intact lipid bilayer that encapsulates a small portion of the parent cell. Exosomes have been studied widely as information-rich sources of potential biomarkers that can reveal cellular physiology. We suggest that quantification is essential to understand basic biological relationships between exosomes and their parent cells and hence the underlying interpretation of exosome signals. The number of methods for quantifying exosomes has expanded as interest in exosomes has increased. However, a consensus on proper quantification has not developed, making each study difficult to compare to another. Overcoming this ad hoc approach will require widely available standards that have been adequately characterized, and multiple comparative studies across platforms. We outline the current status of these technical approaches and our view of how they can become more coherent. J. Cell. Physiol. 9999: 1–4, 2016. © 2016 Wiley Periodicals, Inc.As exosomes are released from cells, they represent not only a rich source of biomarkers, but also a potential source of insight into parent cell physiology. Quantitation of exosomes is essential for progress in both of these areas. Several methods have been developed to quantitate exosomes, and we summarize the current status and future directions toward standardization of these methods.
      PubDate: 2017-03-01T07:02:02.139595-05:
      DOI: 10.1002/jcp.25387
  • Differential Expression of Estrogen Receptor Variants in Response to
           Inflammation Signals in Human Airway Smooth Muscle
    • Authors: Bharathi Aravamudan; Katelyn J. Goorhouse, Ghanashyam Unnikrishnan, Michael A. Thompson, Christina M. Pabelick, John R. Hawse, Y. S. Prakash, Venkatachalem Sathish
      Abstract: The prevalence of asthma is higher in pre-pubescent and aging males, and in post-pubertal females, strongly indicating that sex steroids (especially estrogen) may be an important modulator in lung disease. We recently demonstrated that airway smooth muscle (ASM) expresses both alpha and beta forms of the estrogen receptor (ERα and ERβ) in males and females, and that these receptors regulate intracellular [Ca2+] and ASM contractility. Although both ERα and ERβ have multiple splice variants, it is unclear if and how the expression of these variants is modulated under conditions such as chronic inflammation/asthma. In order to test the hypothesis that the differential expression of ERα and ERβ variants contributes to the pathogenesis of asthma, we profiled the expression of various ERα and ERβ genes in asthmatic and inflamed (TNFα- or IL-13-treated) ASM. Gene expression was assessed at both the mRNA and protein levels in asthmatic ASM cells or non-asthmatic cells treated with TNFα (20 ng/ml) or IL-13 (50 ng/ml). We observed marked variation in the expression of ER isoforms in response to inflammatory stimuli, and in non-asthmatic versus asthmatic ASM. Changes in protein levels of ERα and ERβ corresponded with the observed differential mRNA patterns. Pharmacological studies implicate cytosolic (p42/44 MAPK and PI3 K) and nuclear (NFκB, STAT6, and AP-1) signaling pathways as putative mechanisms that mediate and/or regulate effects of inflammation on ER expression. We conclude that variations in ASM ER expression profiles occur with inflammation and that ER variants could contribute to estrogen signaling in airway diseases such as asthma. J. Cell. Physiol. 9999: 1–7, 2016. © 2016 Wiley Periodicals, Inc.
      PubDate: 2017-02-24T09:10:37.512884-05:
      DOI: 10.1002/jcp.25674
  • Cytoplasmic Localization of RUNX3 via Histone Deacetylase-Mediated SRC
           Expression in Oxidative-Stressed Colon Cancer Cells
    • Authors: Kyoung Ah Kang; Mei Jing Piao, Yea Seong Ryu, Young Hee Maeng, Jin Won Hyun
      Abstract: Runt domain transcription factor 3 (RUNX3) is a transcription factor that functions as a tumor suppressor. RUNX3 is frequently inactivated by epigenetic silencing or its protein mislocalization (cytoplasmic localization) in many cancer types. This study investigated whether oxidative stress induces redistribution of RUNX3 from the nucleus to the cytoplasm. The cytoplasmic localization of RUNX3 was associated with oxidative stress-induced RUNX3 phosphorylation at tyrosine residues via SRC activation. Moreover, oxidative stress increased expression of histone deacetylases (HDACs). RUNX3 phosphorylation and SRC expression induced by oxidative stress were inhibited by knockdown of HDAC1, restoring the nuclear localization of RUNX3 under oxidative stress. In conclusion, these results demonstrate that HDAC1- and SRC-mediated phosphorylation of RUNX3 induced by oxidative stress is associated with the cytoplasmic localization of RUNX3 and can lead to RUNX3 inactivation and carcinogenesis. J. Cell. Physiol. 9999: 1–8, 2016. © 2016 Wiley Periodicals, Inc.RUNX3 cytoplasmic localization induced by oxidative stress in colon cancer cells. Oxidative stress induces HDAC1 expression and then activates SRC. Activated SRC induces expression of phospho-RUNX3. Phospho-RUNX3 is retained in the cytoplasm, leading to degradation of cytoplasmic RUNX3 by the proteasome system and finally resulting carcinogenesis.
      PubDate: 2017-02-21T05:31:21.597574-05:
      DOI: 10.1002/jcp.25746
  • Regulation of mitotic spindle orientation during epidermal stratification
    • Authors: Wei Xie; Jun Zhou
      Abstract: The epidermis is a stratified epithelium that serves as a barrier to infection from environmental pathogens and prevents water loss. Epidermal stratification is tightly controlled during embryogenesis. Progenitor cells in the developing epidermis undergo both symmetric and asymmetric cell divisions to balance the growth of the skin surface area against the generation of differentiated cell layers. Therefore, understanding the relationship between oriented divisions of progenitor cells and the development and stratification of the epidermis is of paramount importance in the field of skin biology and pathology. We provide here an integrated view of recent studies implicating that improper orientation of the mitotic spindle contributes to disorders associated with abnormal epidermal stratification and suggesting that spindle orientation could serve as a potential therapeutic target in skin diseases.We provide here an integrated view of recent studies implicating that improper orientation of the mitotic spindle contributes to disorders associated with abnormal epidermal stratification. We also suggest that spindle orientation could serve as a potential therapeutic target in skin diseases.
      PubDate: 2017-02-21T05:15:49.12907-05:0
      DOI: 10.1002/jcp.25750
  • Na+/H+ exchanger NHE1 and NHE2 have opposite effects on migration velocity
           in rat gastric surface cells
    • Authors: Anja Paehler vor der Nolte; Giriprakash Chodisetti, Zhenglin Yuan, Florian Busch, Brigitte Riederer, Min Luo, Yan Yu, Manoj B. Menon, Andreas Schneider, Renata Stripecke, Katerina Nikolovska, Sunil Yeruva, Ursula Seidler
      Abstract: Following superficial injury, neighbouring gastric epithelial cells close the wound by rapid cell migration, a process called epithelial restitution. Na+/H+ exchange (NHE) inhibitors interfere with restitution, but the role of the different NHE isoforms expressed in gastric pit cells has remained elusive. The role of the basolaterally expressed NHE1 (Slc9a1) and the presumably apically expressed NHE2 (Slc9a2) in epithelial restitution was investigated in the nontransformed rat gastric surface cell line RGM1. Migration velocity was assessed by loading the cells with the fluorescent dye DiR and following closure of an experimental wound over time. Since RGM1 cells expressed very low NHE2 mRNA and have low transport activity, NHE2 was introduced by lentiviral gene transfer. In medium with pH 7.4, RGM1 cells displayed slow wound healing even in the absence of growth factors and independently of NHE activity. Growth factors accelerated wound healing in a partly NHE1-dependent fashion. Preincubation with acidic pH 7.1 stimulated restitution in a NHE1-dependent fashion. When pH 7.1 was maintained during the restitution period, migratory speed was reduced to ∼10% of the speed at pH 7,4, and the residual restitution was further inhibited by NHE1 inhibition. Lentiviral NHE2 expression increased the steady-state pHi and reduced the restitution velocity after low pH preincubation, which was reversible by pharmacological NHE2 inhibition. The results demonstrate that in RGM1 cells, migratory velocity is increased by NHE1 activation, while NHE2 activity inhibit this process. A differential activation of NHE1 and NHE2 may therefore, play a role in the initiation and completion of the epithelial restitution process.Genetic deletion of the Na+/H+ exchanger NHE2 leads to compromised wound healing in the gastrointestinal tract of mice. We studied the roles of NHE1 and NHE2 in gastrointestinal wound healing in vitro, and while NHE1 activity supported wound healing velocity, NHE2/GFP-overexpression dramatically reduced wound healing velocity compared to GFP expression alone. This effect was abolished by a NHE2- but not a NHE1 inhibitory concentration of HOE642. Thus, NHE1 and NHE2 may be important for the initiation and the resolution of gastrointestinal wound healing.
      PubDate: 2017-02-21T05:05:58.470701-05:
      DOI: 10.1002/jcp.25758
  • Osteogenic differentiation of human gingival mesenchymal stem cells by
           Aristolochia bracteolata supplementation through enhanced Runx2 expression
    • Authors: Dinesh Murugan Girija; Suresh Y Ranga Rao, Mangathayaru Kalachaveedu, Rajasekaran Subbarayan
      Abstract: The phenotypic characteristics of human gingival derived mesenchymal stem cells (HGMSCs) on induction with total methanol extract of Aristolochia bracteolata have been evaluated. HGMSCs were cultured in control and two different induction medium: Control medium (basal medium), OM1 (Standard induction medium), and OM2 (100 μg/ml of A. bracteolata). Osteogenic differentiation of the cultured cells was assessed by studying the calcium deposition and osteoblastic gene expression. OM2 medium showed an enhanced osteogenic differentiation potential than OM1 as measured by increased calcium deposition and elevated expression of Runx2, osteopontin, osteonectin, osteocalcin, Collagen type I, and ALP levels in comparison with OM1 differentiated cells. We conclude that at 100 μg/ml A. bracteolata has induced HGMSC differentiation into osteogenic lineage consequent to enhanced Runx2 expression and related osteogenic genes.The phenotypic characteristics of human gingival derived mesenchymal stem cells (HGMSCs) on induction with total methanol extract of Aristolochia bracteolata have been evaluated. A. bracteolata has induced HGMSC differentiation into osteogenic lineage consequent to enhanced Runx2 expression and related osteogenic genes.
      PubDate: 2017-02-21T04:55:23.652165-05:
      DOI: 10.1002/jcp.25835
  • Subclinical alteration of the cervical–vaginal microbiome in women
           with idiopathic infertility
    • Authors: Giuseppina Campisciano; Fiorella Florian, Angela D'Eustacchio, David Stanković, Giuseppe Ricci, Francesco De Seta, Manola Comar
      Abstract: Biomarkers have a wide application in research and clinic, they help to choose the correct treatment for diseases. Recent studies, addressing the vaginal microbiome using next generation sequencing (NGS), reported the involvement of bacterial species in infertility. We compared the vaginal microbiome of idiopathic infertile women with that of healthy, including bacterial vaginosis affected women and non-idiopathic infertile women, to identify bacterial species suitable as biomarkers. Information on microorganisms was obtained from the V3-16S rDNA sequencing of cervical–vaginal fluids of 96 women using the Ion Torrent platform. Data were processed with QIIME and classified against the Vaginal 16S rDNA Reference Database. The analysis revealed a significant beta-diversity variation (p 
      PubDate: 2017-02-16T10:35:27.562047-05:
      DOI: 10.1002/jcp.25806
  • Mesenchyme Homeobox 2 Enhances Migration of Endothelial Colony Forming
           Cells Exposed to Intrauterine Diabetes Mellitus
    • Authors: Cassandra R. Gohn; Emily K. Blue, BreAnn M. Sheehan, Kaela M. Varberg, Laura S. Haneline
      Abstract: Diabetes mellitus (DM) during pregnancy has long-lasting implications for the fetus, including cardiovascular morbidity. Previously, we showed that endothelial colony forming cells (ECFCs) from DM human pregnancies have decreased vasculogenic potential. Here, we evaluate whether the molecular mechanism responsible for this phenotype involves the transcription factor, Mesenchyme Homeobox 2 (MEOX2). In human umbilical vein endothelial cells, MEOX2 upregulates cyclin-dependent kinase inhibitor expression, resulting in increased senescence and decreased proliferation. We hypothesized that dysregulated MEOX2 expression in neonatal ECFCs from DM pregnancies decreases network formation through increased senescence and altered cell cycle progression. Our studies show that nuclear MEOX2 is increased in ECFCs from DM pregnancies. To determine if MEOX2 is sufficient and/or required to induce impaired network formation, MEOX2 was overexpressed and depleted in ECFCs from control and DM pregnancies, respectively. Surprisingly, MEOX2 overexpression in control ECFCs resulted in increased network formation, altered cell cycle progression, and increased senescence. In contrast, MEOX2 knockdown in ECFCs from DM pregnancies led to decreased network formation, while cell cycle progression and senescence were unaffected. Importantly, migration studies demonstrated that MEOX2 overexpression increased migration, while MEOX2 knockdown decreased migration. Taken together, these data suggest that altered migration may be mediating the impaired vasculogenesis of ECFCs from DM pregnancies. While initially believed to be maladaptive, these data suggest that MEOX2 may serve a protective role, enabling increased vessel formation despite exposure to a DM intrauterine environment. J. Cell. Physiol. 9999: 1–8, 2016. © 2016 Wiley Periodicals, Inc.Endothelial colony forming cells (ECFCs) from diabetic human pregnancies have decreased vasculogenic potential both in vitro and in vivo. The goal of this study was to evaluate whether the molecular mechanism responsible for this phenotype involves the transcription factor, Mesenchyme Homeobox 2 (MEOX2). While initially believed to be maladaptive, these data suggest that MEOX2 may serve a protective role, enabling increased vessel formation despite exposure to a DM intrauterine environment.
      PubDate: 2017-02-16T10:25:26.580862-05:
      DOI: 10.1002/jcp.25734
  • The Role of Cardiac Myosin Binding Protein C3 in Hypertrophic
           Cardiomyopathy-Progress and Novel Therapeutic Opportunities
    • Authors: Iman A. Mohamed; Navaneethakrishnan T. Krishnamoorthy, Gheyath K. Nasrallah, Sahar I. Da'as
      Abstract: Hypertrophic cardiomyopathy (HCM) is a common autosomal dominant genetic cardiovascular disorder marked by genetic and phenotypic heterogeneity. Mutations in the gene encodes the cardiac myosin-binding protein C, cMYBPC3 is amongst the various sarcomeric genes that are associated with HCM. These mutations produce mutated mRNAs and truncated cMyBP-C proteins. In this review, we will discuss the implications and molecular mechanisms involved in MYBPC3 different mutations. Further, we will highlight the novel targets that can be developed into potential therapeutics for the treatment of HMC. J. Cell. Physiol. 9999: 1–10, 2016. © 2016 Wiley Periodicals, Inc.This review discusses the implications and the and molecular mechanisms of MYBPC3 different mutations. Further, we will highlight the novel targets that can be developed into potential therapeutics for the treatment of Hypertrophic cardiomyopathy.
      PubDate: 2017-02-16T10:15:27.995734-05:
      DOI: 10.1002/jcp.25639
  • Identification of MSX1 and DCLK1 as mRNA Biomarkers for Colorectal Cancer
           Detection Through DNA Methylation Information
    • Authors: Ai-Jun Sun; Hai-Bo Gao, Gao Liu, Heng-Fa Ge, Zun-Ping Ke, Sen Li
      Abstract: Colorectal cancer is the second most deadly malignancy in the United States. However, the currently screening options had their limitation. Novel biomarkers for colorectal cancer detections are necessary to reduce the mortality. The clinical information, mRNA expression levels and DNA methylation information of colorectal cancer were downloaded from TCGA. The patients were separated into training group and testing group based on their platforms for DNA methylation. Beta values of DNA methylation from tumor tissues and normal tissues were utilized to figure out the position that were differentially methylated. The expression levels of mRNA of thirteen genes, whose CpG islands were differentially methylated, were extracted from the RNA-Seq results from TCGA. The probabilities whether the mRNA was differentially expressed between tumor and normal samples were calculated using Student's t-test. Logistic regression and decision tree were built for cancer detection and their performances were evaluated by the area under the curve (AUC). Twenty-four genomic locations were differentially methylated, which could be mapped to eleven genes. Nine out of eleven genes had differentially expressed mRNA levels, which were used to build the model for cancer detection. The final detection models consisting of mRNA expression levels of these nine genes had great performances on both training group and testing group. The model that constructed in this study suggested MSX1 and DCLK1 might be used in colorectal cancer detection or as target of cancer therapies. J. Cell. Physiol. 9999: 1–6, 2016. © 2016 Wiley Periodicals, Inc.Nine out of eleven genes had differentially expressed mRNA levels, which were used to build the model for cancer detection. The final detection models consisting of mRNA expression levels of these nine genes had great performances on both training group and testing group. The model that constructed in this study suggested MSX1 and DCLK1 might be used in colorectal cancer detection or as target of cancer therapies.
      PubDate: 2017-02-16T10:10:26.023949-05:
      DOI: 10.1002/jcp.25733
  • MicroRNA-125a-5p Is a Downstream Effector of Sorafenib in Its
           Antiproliferative Activity Toward Human Hepatocellular Carcinoma Cells
    • Authors: Nicoletta Potenza; Nicola Mosca, Silvia Zappavigna, Filomena Castiello, Marta Panella, Carmela Ferri, Daniela Vanacore, Antonio Giordano, Paola Stiuso, Michele Caraglia, Aniello Russo
      Abstract: Sorafenib is an antitumor drug for treatment of advanced hepatocellular carcinoma (HCC). It acts as a multikinase inhibitor suppressing cell proliferation and angiogenesis. Human microRNA-125a-5p (miR-125a) is endowed with similar activities and is frequently downregulated in HCC. Looking for a potential microRNA-based mechanism of action of the drug, we found that sorafenib increases cellular expression of miR-125a in cultured HuH-7 and HepG2 HCC cells. Upregulation of the microRNA inhibited cell proliferation by suppression of sirtuin-7, a NAD(+)-dependent deacetylase, and p21/p27-dependent cell cycle arrest in G1. Later, recruitment of miR-125a in the antiproliferative activity of sorafenib was inquired by modulating its expression in combination with the drug treatment. This analysis showed that intracellular delivery of miR-125a had no additive effect on the antiproliferative activity of sorafenib, whereas a miR-125a inhibitor could counteract it. Finally, evaluation of other oncogenic targets of miR-125a revealed its ability to interfere with the expression of matrix metalloproteinase-11, Zbtb7a proto-oncogene, and c-Raf, possibly contributing to the antiproliferative activity of the drug. J. Cell. Physiol. 9999: 1–7, 2016. © 2016 Wiley Periodicals, Inc.Upregulation of the microRNA inhibited cell proliferation by suppression of sirtuin-7, a NAD(+)-dependent deacetylase, and p21/p27-dependent cell cycle arrest in G1. Intracellular delivery of miR-125a had no additive effect on the antiproliferative activity of sorafenib, whereas a miR-125a inhibitor could counteract it. miR-125a interfered with the expression of matrix metalloproteinase-11, Zbtb7a proto-oncogene, and c-Raf, possibly contributing to the antiproliferative activity of the drug.
      PubDate: 2017-02-16T10:05:25.695604-05:
      DOI: 10.1002/jcp.25744
  • Proteomics Applications in Dental Derived Stem Cells
    • Authors: Jie Li; Weidong Tian, Jinlin Song
      Abstract: At present, the existence of a variety of dental derived stem cells has been documented. These cells displayed promising clinical application potential not only for teeth and its surrounding tissue regeneration, but also for other tissues, such as nerve and bone regeneration. Proteomics is an unbiased, global informatics tool that provides information on all protein expression levels as well as post-translational modification in cells or tissues and is applicable to dental derived stem cells research. Over the last decade, considerable progress has been made to study the global proteome, secrotome, and membrane proteome of dental derived stem cells. Here, we present an overview of the proteomics studies in the context of stem cell research. Particular attention is given to dental derived stem cell types as well as current challenges and opportunities. J. Cell. Physiol. 9999: 1–9, 2016. © 2016 Wiley Periodicals, Inc.(1) Dental derived stem cells displayed promising clinical application potential not only for teeth and its surrounding tissue regeneration, but also for other tissues regeneration. (2) Here, we present an overview of the proteomics studies in the context of dental derived stem cell research. (3) Particular attention is given to dental derived stem cell types as well as current challenges and opportunities.
      PubDate: 2017-02-16T09:40:25.198627-05:
      DOI: 10.1002/jcp.25667
  • Protective Role of GPER Agonist G-1 on Cardiotoxicity Induced by
    • Authors: Ernestina M. De Francesco; Carmine Rocca, Francesco Scavello, Daniela Amelio, Teresa Pasqua, Damiano C. Rigiracciolo, Andrea Scarpelli, Silvia Avino, Francesca Cirillo, Nicola Amodio, Maria C. Cerra, Marcello Maggiolini, Tommaso Angelone
      Abstract: The use of Doxorubicin (Dox), a frontline drug for many cancers, is often complicated by dose-limiting cardiotoxicity in approximately 20% of patients. The G-protein estrogen receptor GPER/GPR30 mediates estrogen action as the cardioprotection under certain stressful conditions. For instance, GPER activation by the selective agonist G-1 reduced myocardial inflammation, improved immunosuppression, triggered pro-survival signaling cascades, improved myocardial mechanical performance, and reduced infarct size after ischemia/reperfusion (I/R) injury. Hence, we evaluated whether ligand-activated GPER may exert cardioprotection in male rats chronically treated with Dox. 1 week of G-1 (50 μg/kg/day) intraperitoneal administration mitigated Dox (3 mg/kg/day) adverse effects, as revealed by reduced TNF-α, IL-1β, LDH, and ROS levels. Western blotting analysis of cardiac homogenates indicated that G-1 prevents the increase in p-c-jun, BAX, CTGF, iNOS, and COX2 expression induced by Dox. Moreover, the activation of GPER rescued the inhibitory action elicited by Dox on the expression of BCL2, pERK, and pAKT. TUNEL assay indicated that GPER activation may also attenuate the cardiomyocyte apoptosis upon Dox exposure. Using ex vivo Langendorff perfused heart technique, we also found an increased systolic recovery and a reduction of both infarct size and LDH levels in rats treated with G-1 in combination with Dox respect to animals treated with Dox alone. Accordingly, the beneficial effects induced by G-1 were abrogated in the presence of the GPER selective antagonist G15. These data suggest that GPER activation mitigates Dox-induced cardiotoxicity, thus proposing GPER as a novel pharmacological target to limit the detrimental cardiac effects of Dox treatment. J. Cell. Physiol. 9999: 1–10 , 2016. © 2016 Wiley Periodicals, Inc.1. The potential beneficial effects of GPER activation in Doxorubicin (Dox)-induced cardiotoxicity was evaluated. 2. In male rat heart, the treatment with the selective GPER ligand G-1 prevented the increase in TNF-α, IL-1β, and LDH plasma levels induced by Dox. In Langendorff perfused rat heart, G-1 also increased the systolic recovery at the end of ischemia and reduced infarct size and LDH levels triggered by Dox. Moreover, G-1 attenuated the Dox-induced myocyte apoptosis, as well as the inhibition of pro-survival signaling cascades like ERK1/2 and AKT. Next, G-1 counteracted the down-regulation of BCL2 as well as the up-regulation of apoptotic, inflammatory and fibrotic targets, such as BAX, iNOS, COX2, and CTGF induced by Dox. 3. Our study paves the way for further analyzing the role of GPER as pharmacological target in novel therapeutic strategies aimed to prevent the cardiotoxicity exerted by Dox.
      PubDate: 2017-02-16T09:20:30.356957-05:
      DOI: 10.1002/jcp.25585
  • Postnatal Hyperplasic Effects of ActRIIB Blockade in a Severely Dystrophic
    • Authors: Cory Nielsen; Ross M. Potter, Christopher Borowy, Kimberly Jacinto, Ravi Kumar, C. George Carlson
      Abstract: The efficacy of two ActRIIB ligand-trapping agents (RAP-031 and RAP-435) in treating muscular dystrophy was examined by determining their morphological effects on the severely dystrophic triangularis sterni (TS) muscle of the mdx mouse, a model for Duchenne muscular dystrophy. These agents trap all endogenous ligands to the ActRIIB receptor and thereby block myostatin signaling in a highly selective manner. Short-term (1 month) and long-term (3 months) in vivo treatment of 1-month-old mdx mice increased myonuclei and fiber cross section (FCS) density but did not alter individual fiber size. Vehicle-treated mdx mice exhibited age-dependent increases in myonuclei and FCS density, and age-dependent reductions in centronucleation that were each enhanced by treatment with RAP-435. Distributions of FCS area (FCSA) in the mdx TS were 90% identical to those from untreated age-matched nondystrophic mice and were unaltered by the substantial fiber hyperplasia observed with age and RAP-435 treatment. These results were inconsistent with injury-induced fiber regeneration which produces altered FCSA distributions characterized by a distinct class of smaller regenerated fibers. Nondystrophic mice exhibited a constant postnatal density of fiber cross sections and myonuclei, and RAP-435 treatment of nondystrophic mice increased TS mean FCSA but had no effects on myonuclei or FCS density. These results demonstrating a continual postnatal proliferation and fusion of satellite cells and a response to myostatin blockade characteristic of developing prenatal muscle suggest that the lack of dystrophin directly results in unrestrained postnatal satellite cell activation that is not necessarily dependent upon prior fiber degeneration. J. Cell. Physiol. 9999: 1–20, 2016. © 2016 Wiley Periodicals, Inc.We examined the efficacy of two ActRIIB ligand-trapping agents in altering the morphology of a severely dystrophic muscle in the mdx mouse. The results indicate that severely dystrophic muscle exhibits monotonic postnatal increases in myonuclei and fiber density that are not associated with alterations in the distribution of fiber cross-sectional areas. Long-term treatment with an ActRIIB ligand-trapping agent substantially enhanced this postnatal hyperplasia and reduced centronucleation. These results demonstrating a continual postnatal proliferation and fusion of satellite cells and a response to myostatin blockade characteristic of developing prenatal muscle suggest that the lack of dystrophin directly results in unrestrained postnatal satellite cell activation that is not necessarily dependent upon prior fiber degeneration.
      PubDate: 2017-02-16T09:15:56.755686-05:
      DOI: 10.1002/jcp.25694
  • Role of Wnt5a in the Pathogenesis of Inflammatory Diseases
    • Authors: Mehran Pashirzad; Mojtaba Shafiee, Farzad Rahmani, Reihane Behnam-Rassouli, Fatemeh Hoseinkhani, Mikhail Ryzhikov, Maryam Moradi Binabaj, Mohammad Reza Parizadeh, Amir Avan, Seyed Mahdi Hassanian
      Abstract: Wnt5a initiates pro-inflammatory responses through activation of non-canonical Wnt signaling pathway. Pro-inflammatory functions of Wnt5a trigger pro-inflammatory signaling cascades and increase secretion of pro-inflammatory cytokines and chemokines. Wnt5a as a potent signaling molecule is strongly implicated in a number of diseases including cancer, diabetes, metabolic disorders, and of special interest in this review, inflammatory diseases. This review summarizes the role of Wnt5a in the pathogenesis of inflammatory diseases including atherosclerosis, rheumatoid arthritis, psoriasis vulgaris and sepsis, promoting greater understanding, and clinical management of these diseases. J. Cell. Physiol. 9999: 1–6, 2017. © 2016 Wiley Periodicals, Inc.Wnt5a as a potent signaling molecule is strongly implicated in a number of diseases including cancer, diabetes, metabolic disorders, and of special interest in this review, inflammatory diseases. This review summarizes the role of Wnt5a in the pathogenesis of inflammatory diseases including atherosclerosis, rheumatoid arthritis, psoriasis vulgaris and sepsis, promoting greater understanding and clinical management of these diseases.
      PubDate: 2017-02-16T09:10:27.206473-05:
      DOI: 10.1002/jcp.25687
  • Involvement of CX3CL1 in the Migration of Osteoclast Precursors Across
           Osteoblast Layer Stimulated by Interleukin-1ß
    • Authors: Tsuyoshi Matsuura; Shizuko Ichinose, Masako Akiyama, Yuki Kasahara, Noriko Tachikawa, Ken-ichi Nakahama
      Abstract: The trigger for bone remodeling is bone resorption by osteoclasts. Osteoclast differentiation only occurs on the old bone, which needs to be repaired under physiological conditions. However, uncontrolled bone resorption is often observed in pro-inflammatory bone diseases, such as rheumatoid arthritis. Mature osteoclasts are multinuclear cells that differentiate from monocyte/macrophage lineage cells by cell fusion. Although Osteoclast precursors should migrate across osteoblast layer to reach bone matrix before maturation, the underlying mechanisms have not yet been elucidated in detail. We herein found that osteoclast precursors utilize two routes to migrate across osteoblast layer by confocal- and electro-microscopic observations. The osteoclast supporting activity of osteoblasts inversely correlated with osteoblast density and was positively related to the number of osteoclast precursors under the osteoblast layer. Osteoclast differentiation was induced by IL-1ß, but not by PGE2 in high-density osteoblasts. Osteoblasts and osteoclast precursors expressed CX3CL1 and CX3CR1, respectively, and the expression of CX3CL1 increased in response to interleukin-1ß. An anti-CX3CL1-neutralizing antibody inhibited the migration of osteoclast precursors and osteoclast differentiation. These results strongly suggest the involvement of CX3CL1 in the migration of osteoclast precursors and osteoclastogenesis, and will contribute to the development of new therapies for bone diseases. J. Cell. Physiol. 9999: 1–7, 2016. © 2016 Wiley Periodicals, Inc.Transcellular and paracellular migration of osteoclast precursors across osteoblast layer were observed. CX3CL1 was involved in the migration of osteoclast precursors and osteoclastogenesis.
      PubDate: 2017-02-03T10:00:30.140846-05:
      DOI: 10.1002/jcp.25577
  • DNA Methylation Profiling in Chondrocyte Dedifferentiation In Vitro
    • Authors: Li Duan; Yujie Liang, Bin Ma, Daming Wang, Wei Liu, Jianghong Huang, Jianyi Xiong, Liangquan Peng, Jielin Chen, Weimin Zhu, Daping Wang
      Abstract: DNA methylation has emerged as a crucial regulator of chondrocyte dedifferentiation, which severely compromises the outcome of autologous chondrocyte implantation (ACI) treatment for cartilage defects. However, the full-scale DNA methylation profiling in chondrocyte dedifferentiation remains to be determined. Here, we performed a genome-wide DNA methylation profiling of dedifferentiated chondrocytes in monolayer culture and chondrocytes treated with DNA methylation inhibitor 5-azacytidine (5-AzaC). This research revealed that the general methylation level of CpG was increased while the COL-1A1 promoter methylation level was decreased during the chondrocyte dedifferentiation. 5-AzaC could reduce general methylation levels and reverse the chondrocyte dedifferentiation. Surprisingly, the DNA methylation level of COL-1A1 promoter was increased after 5-AzaC treatment. The COL-1A1 expression level was increased while that of SOX-9 was decreased during the chondrocyte dedifferentiation. 5-AzaC treatment up-regulated the SOX-9 expression while down-regulated the COL-1A1 promoter activity and gene expression. Taken together, these results suggested that differential regulation of the DNA methylation level of cartilage-specific genes might contribute to the chondrocyte dedifferentiation. Thus, the epigenetic manipulation of these genes could be a potential strategy to counteract the chondrocyte dedifferentiation accompanying in vitro propagation. J. Cell. Physiol. 9999: 1–9, 2016. © 2016 Wiley Periodicals, Inc.Differential regulation of the DNA methylation level of cartilage-specific genes might contribute the chondrocyte dedifferentiation. Thus, the epigenetic manipulation of these genes could be a potential strategy to counteract the chondrocyte dedifferentiation accompanying in vitro propagation.
      PubDate: 2017-01-31T07:47:23.34891-05:0
      DOI: 10.1002/jcp.25486
  • Glioblastoma Multiforme and Adult Neurogenesis in the
           Ventricular-Subventricular Zone: A Review
    • Authors: Claudia Capdevila; Lucía Rodríguez Vázquez, Joaquín Martí
      Abstract: Brain cancers account for
      PubDate: 2017-01-31T07:47:08.85698-05:0
      DOI: 10.1002/jcp.25502
  • Cytoskeleton Aberrations in Alkaptonuric Chondrocytes
    • Authors: Michela Geminiani; Silvia Gambassi, Lia Millucci, Pietro Lupetti, Giulia Collodel, Lucia Mazzi, Bruno Frediani, Daniela Braconi, Barbara Marzocchi, Marcella Laschi, Giulia Bernardini, Annalisa Santucci
      Abstract: Alkaptonuria (AKU) is an ultra-rare autosomal genetic disorder caused by a defect in the activity of the enzyme homogentisate 1,2-dioxygenase (HGD) that leads to the accumulation of homogentisic acid (HGA) and its oxidized product, benzoquinone acetic acid (BQA), in the connective tissues causing a pigmentation called “ochronosis.” The consequent progressive formation of ochronotic aggregates generate a severe condition of oxidative stress and inflammation in all the affected areas. Experimental evidences have also proved the presence of serum amyloid A (SAA) in several AKU tissues and it allowed classifying AKU as a secondary amyloidosis. Although AKU is a multisystemic disease, the most affected system is the osteoarticular one and articular cartilage is the most damaged tissue. In this work, we have analyzed for the first time the cytoskeleton of AKU chondrocytes by means of immunofluorescence staining. We have shown the presence of SAA within AKU chondrocytes and finally we have demonstrated the co-localization of SAA with three cytoskeletal proteins: actin, vimentin, and β-tubulin. Furthermore, in order to observe the ultrastructural features of AKU chondrocytes we have performed TEM analysis, focusing on the Golgi apparatus structure and, to demonstrate that pigmented areas in AKU cartilage are correspondent to areas of oxidation, 4-HNE presence has been evaluated by means of immunofluorescence. J. Cell. Physiol. 9999: 1–11, 2016. © 2016 Wiley Periodicals, Inc.The cytoskeleton of alkaptonuric chondrocytes is severely alterated. Serum amyloid A is demonstrated to be present within alkaptonuric chondrocytes and to co-localize with the three main cytoskeletal proteins: actin, vimentin, and β-tubulin. These findings lead to hypothesize a role of SAA in cytoskeleton alterations of alkaptonuric chondrocytes.
      PubDate: 2017-01-31T07:29:41.564847-05:
      DOI: 10.1002/jcp.25500
  • Table of Contents, Editor's Choice, Highlights
    • First page: 1579
      PubDate: 2017-03-28T07:35:12.196675-05:
      DOI: 10.1002/jcp.25892
  • Condensin Regulation of Genome Architecture
    • Authors: Vibhuti Rana; Giovanni Bosco
      First page: 1617
      Abstract: Condensin complexes exist across all domains of life and are central to the structure and organization of chromatin. As architectural proteins, condensins control chromatin compaction during interphase and mitosis. Condensin activity has been well studied in mitosis but have recently emerged as important regulators of genome organization and gene expression during interphase. Here, we focus our discussion on recent findings on the molecular mechanism and how condensins are used to shape chromosomes during interphase. These findings suggest condensin activity during interphase is required for proper chromosome organization. J. Cell. Physiol. 232: 1617–1625, 2017. © 2016 Wiley Periodicals, Inc.This article reviews recent works exploring condensin molecular mechanism and activity. We specifically explore activity during interphase and how condensins are regulated.
      PubDate: 2017-01-05T08:50:27.155548-05:
      DOI: 10.1002/jcp.25702
  • Exosomes: Nanoparticulate tools for RNA interference and drug delivery
    • Authors: Fahimeh Shahabipour; Nastaran Barati, Thomas P. Johnston, Giuseppe Derosa, Pamela Maffioli, Amirhossein Sahebkar
      First page: 1660
      Abstract: Exosomes are naturally occurring extracellular vesicles released by most mammalian cells in all body fluids. Exosomes are known as key mediators in cell-cell communication and facilitate the transfer of genetic and biochemical information between distant cells. Structurally, exosomes are composed of lipids, proteins, and also several types of RNAs which enable these vesicles to serve as important disease biomarkers. Moreover, exosomes have emerged as novel drug and gene delivery tools owing to their multiple advantages over conventional delivery systems. Recently, increasing attention has been focused on exosomes for the delivery of drugs, including therapeutic recombinant proteins, to various target tissues. Exosomes are also promising vehicles for the delivery of microRNAs and small interfering RNAs, which is usually hampered by rapid degradation of these RNAs, as well as inefficient tissue specificity of currently available delivery strategies. This review highlights the most recent accomplishments and trends in the use of exosomes for the delivery of drugs and therapeutic RNA molecules.Exosomes are naturally occurring extracellular vesicles released by most mammalian cells in all body fluids. Recently, increasing attention has been focused on exosomes for the delivery of drugs, including therapeutic recombinant proteins, to various target tissues. This review highlights the most recent accomplishments and trends in the use of exosomes for the delivery of drugs and therapeutic RNA molecules.
      PubDate: 2017-01-31T07:18:54.765898-05:
      DOI: 10.1002/jcp.25766
  • ­NOD Mice Having a Lyn Tyrosine Kinase Mutation Exhibit Abnormal
           Neutrophil Chemotaxis
    • Authors: Yue Wu; Michael Hannigan, Lijun Zhan, Joseph A. Madri, Chi-Kuang Huang
      First page: 1689
      Abstract: Neutrophils from NOD (Non-Obese Diabetic) mice exhibited reduced migration speed, decreased frequency of directional changes, and loss of directionality during chemotaxis (compared to wild-type [WT] C57BL/6 mice). Additionally, F-actin of chemotaxing NOD neutrophils failed to orient toward the chemoattractant gradient and NOD neutrophil adhesion was impaired. A point mutation near the autophosphorylation site of Lyn in NOD mice was identified. Point mutations of G to A (G1412 in LynA and G1199 in LynB) cause a change of amino acid E393 (glutamic acid) to K (lysine) in LynA (E393K) (E372 of LynB), affecting fMLP-induced tyrosine phosphorylation. These data indicate that the Lyn mutation in NOD neutrophils is likely responsible for dysregulation of neutrophil adhesion and directed migration, implying the role of Lyn in modulating diabetic patient's susceptibility to bacterial and fungal infections. J. Cell. Physiol. 232: 1689–1695, 2017. © 2016 Wiley Periodicals, Inc.A specific Lyn kinase mutation has been documented in NOD mice which affects neutrophil directed migration. This may thus affect response to infections in this murine model of diabetes.
      PubDate: 2017-03-27T07:40:28.147043-05:
      DOI: 10.1002/jcp.25583
  • HDAC1, HDAC4, and HDAC9 Bind to PC3/Tis21/Btg2 and Are Required for Its
           Inhibition of Cell Cycle Progression and Cyclin D1 Expression
    • Authors: Laura Micheli; Giorgio D'Andrea, Luca Leonardi, Felice Tirone
      First page: 1696
      Abstract: PC3/Tis21 is a transcriptional cofactor that inhibits proliferation in several cell types, including neural progenitors. Here, we report that PC3/Tis21 associates with HDAC1, HDAC4, and HDAC9 in vivo, in fibroblast cells. Furthermore, when HDAC1, HDAC4, or HDAC9 are silenced in fibroblasts or in a line of cerebellar progenitor cells, the ability of PC3/Tis21 to inhibit proliferation is significantly reduced. Overexpression of HDAC1, HDAC4, or HDAC9 in fibroblasts and in cerebellar precursor cells synergizes with PC3/Tis21 in inhibiting the expression of cyclin D1, a cyclin selectively inhibited by PC3/Tis21. Conversely, the depletion of HDAC1 or HDAC4 (but not HDAC9) in fibroblasts and in cerebellar precursor cells significantly impairs the ability of PC3/Tis21 to inhibit cyclin D1 expression. An analysis of HDAC4 deletion mutants shows that both the amino-terminal moiety and the catalytic domain of HDAC4 associate to PC3/Tis21, but neither alone is sufficient to potentiate the inhibition of cyclin D1 by PC3/Tis21. As a whole, our findings indicate that PC3/Tis21 inhibits cell proliferation in a way dependent on the presence of HDACs, in fibroblasts as well as in neural cells. Considering that several reports have demonstrated that HDACs can act as transcriptional corepressors on the cyclin D1 promoter, our data suggest that the association of PC3/Tis21 to HDACs is functional to recruit them to target genes, such as cyclin D1, for repression of their expression. J. Cell. Physiol. 232: 1696–1707, 2017. © 2016 Wiley Periodicals, Inc.By silencing HDAC1, HDAC4, and HDAC9 in fibroblast as well as in neural cells, we reveal that HDAC1, HDAC4, and HDAC9 are each required for the negative control of the cell cycle exerted by PC3/Tis21. Such effect is likely dependent on the ability of HDAC1, HDAC4, and HDAC9 to associate with PC3/Tis21, that we demonstrate here. Moreover, we find that HDAC1 and HDAC4—but not HDAC9—are required for the inhibition exerted by PC3/Tis21 of cyclin D1 expression.
      PubDate: 2017-01-27T06:45:37.185904-05:
      DOI: 10.1002/jcp.25467
  • Modulation of Chloride Currents in Human Lung Epithelial Cells Exposed to
           Exogenous Oxidative Stress
    • Authors: Rita Canella; Marta Martini, Roberta Borriello, Carlotta Cavicchio, Ximena M. Muresan, Mascia Benedusi, Franco Cervellati, Giuseppe Valacchi
      First page: 1817
      Abstract: Air pollution continues to be a major public health concern affecting 9 out of 10 individuals living in urban areas worldwide. Respiratory tract is the organ most exposed to gas pollution, and ozone has been shown to be one of the most noxious pollutants to which living organisms are exposed. In the present work, we have investigated the effects of 0.1 ppm of ozone on chloride currents in human lung epithelial cells (A549 line) and whether this effect could be modulated by vitamin E pre-treatment. Whole-cell patch clamp technique was applied to not excitable cells in order to obtain information about chloride currents behavior, important for epithelial lung cells homeostasis. Significant alteration of the I–V curve after ozone treatment was observed, with the appearance of a large outward rectifier component decreasing over time and returning to the basal state levels after 24 h. Statistical analysis indicated a modification of the amount of ions passing the membrane in the unit of time as a possible cause of this difference. RT-qPCR analysis showed an increase in ClC-2 and ORCC mRNA after ozone exposure. In addition, pre-treatment with vitamin E was able to suppress the outward rectifier component induced by ozone, bringing back the current values to the control level and preventing ozone induced chloride channels up regulation. Our data suggest that ozone exposure is able to modify chloride current density and the use of vitamin E can prevent the above-mentioned damage. J. Cell. Physiol. 232: 1817–1825, 2017. © 2016 Wiley Periodicals, Inc.Pollutant ozone is able to induce oxidative stress to lung epithelial cells and this affect chloride currents and the expression of ClC2 and ORCC channel genes levels. Vitamin E pre-treatment is able to rescue this noxious effect.
      PubDate: 2017-01-05T08:55:22.244087-05:
      DOI: 10.1002/jcp.25705
  • A Human 3D In Vitro Model to Assess the Relationship Between Osteoporosis
           and Dissemination to Bone of Breast Cancer Tumor Cells
    • Authors: Francesca Salamanna; Veronica Borsari, Silvia Brogini, Paola Torricelli, Simona Cepollaro, Matteo Cadossi, Milena Fini
      First page: 1826
      Abstract: Despite consistent improvements in diagnostic and therapeutic strategies for breast cancer, up to 40% of patients will develop bone metastases. To reduce the morbidity and complications related with bone metastases, it is imperative to reduce their etiological factors. Osteoporosis, being characterized by a sudden estrogen deficiency, may provide a favorable condition for bone metastasis. This work, using a humanized 3D in vitro model, aims at evaluating the relationship between osteoporosis and breast cancer-derived bone metastases. Bone tissue discarded from total hip replacement surgery of healthy and osteoporotic patients was cultured in a rolling apparatus system in hypoxic environment. Protein levels (i.e., vascular endothelial growth factor (VEGF), VEGF receptor 1, VEGF receptor 2, interleukin (IL)-6, IL-1β, IL-8 IL-10, tumor necrosis factor α (TNF-α), osteoprotegerin (OPG), receptor activator for nuclear factor KB ligand (RANKL)) and histological and immunohistochemical (i.e., cytokeratin 8 and 18) analyses showed a noticeable specificity of breast cancer cells for the colonization of osteoporotic bone. These data are the first to demonstrate that using humanized 3D in vitro systems, which individually model the pre- and postmenopausal bone microenvironment, it is possible to recognize major differences in tumor growth and colonization between healthy and osteoporotic status. Thus, this system might help to develop a shared system between basic and clinical sciences where a personalized diagnosis is associated to a therapeutic strategy designed for a single patient: a model able to achieve a translational research approach in the clinical setting, which may lead to the application and dissemination of personalized medicine. J. Cell. Physiol. 232: 1826–1834, 2017. © 2016 Wiley Periodicals, Inc.Humanized 3D in vitro model; relationship between osteoporosis and breast cancer-derived bone metastases; differences in tumor growth and colonization between healthy and osteoporotic status.
      PubDate: 2017-01-05T08:40:23.492705-05:
      DOI: 10.1002/jcp.25708
  • SIRT1-SIRT3 Axis Regulates Cellular Response to Oxidative Stress and
    • Authors: Ilaria Carnevale; Laura Pellegrini, Patrizia D'Aquila, Serena Saladini, Emanuela Lococo, Lucia Polletta, Enza Vernucci, Eleonora Foglio, Stefano Coppola, Luigi Sansone, Giuseppe Passarino, Dina Bellizzi, Matteo A. Russo, Massimo Fini, Marco Tafani
      First page: 1835
      Abstract: Sirtuins are conserved NAD+-dependent deacylases. SIRT1 is a nuclear and cytoplasmic sirtuin involved in the control of histones a transcription factors function. SIRT3 is a mitochondrial protein, which regulates mitochondrial function. Although, both SIRT1 and SIRT3 have been implicated in resistance to cellular stress, the link between these two sirtuins has not been studied so far. Here we aimed to unravel: i) the role of SIRT1-SIRT3 axis for cellular response to oxidative stress and DNA damage; ii) how mammalian cells modulate such SIRT1-SIRT3 axis and which mechanisms are involved. Therefore, we analyzed the response to different stress stimuli in WT or SIRT1-silenced cell lines. Our results demonstrate that SIRT1-silenced cells are more resistant to H2O2 and etoposide treatment showing decreased ROS accumulation, γ-H2AX phosphorylation, caspase-3 activation and PARP cleavage. Interestingly, we observed that SIRT1-silenced cells show an increased SIRT3 expression. To explore such a connection, we carried out luciferase assays on SIRT3 promoter demonstrating that SIRT1-silencing increases SIRT3 promoter activity and that such an effect depends on the presence of SP1 and ZF5 recognition sequences on SIRT3 promoter. Afterwards, we performed co-immunoprecipitation assays demonstrating that SIRT1 binds and deacetylates the transcription inhibitor ZF5 and that there is a decreased interaction between SP1 and ZF5 in SIRT1-silenced cells. Therefore, we speculate that acetylated ZF5 cannot bind and sequester SP1 that is free, then, to increase SIRT3 transcription. In conclusion, we demonstrate that cells with low SIRT1 levels can maintain their resistance and survival by increasing SIRT3 expression. J. Cell. Physiol. 232: 1835–1844, 2017. © 2016 Wiley Periodicals, Inc.Our article demonstrate that cells with low SIRT1 levels can maintain their resistance and survival to exogenous stresses by increasing SIRT3 expression.
      PubDate: 2017-01-06T10:15:44.033347-05:
      DOI: 10.1002/jcp.25711
  • Extracellular Hsp70 Enhances Mesoangioblast Migration via an Autocrine
           Signaling Pathway
    • Authors: Maria M. Barreca; Walter Spinello, Vincenzo Cavalieri, Giuseppina Turturici, Gabriella Sconzo, Punit Kaur, Rosaria Tinnirello, Alexzander A. A. Asea, Fabiana Geraci
      First page: 1845
      Abstract: Mouse mesoangioblasts are vessel-associated progenitor stem cells endowed with the ability of multipotent mesoderm differentiation. Therefore, they represent a promising tool in the regeneration of injured tissues. Several studies have demonstrated that homing of mesoangioblasts into blood and injured tissues are mainly controlled by cytokines/chemokines and other inflammatory factors. However, little is known about the molecular mechanisms regulating their ability to traverse the extracellular matrix (ECM). Here, we demonstrate that membrane vesicles released by mesoangioblasts contain Hsp70, and that the released Hsp70 is able to interact by an autocrine mechanism with Toll-like receptor 4 (TLR4) and CD91 to stimulate migration. We further demonstrate that Hsp70 has a positive role in regulating matrix metalloproteinase 2 (MMP2) and MMP9 expression and that MMP2 has a more pronounced effect on cell migration, as compared to MMP9. In addition, the analysis of the intracellular pathways implicated in Hsp70 regulated signal transduction showed the involvement of both PI3K/AKT and NF-κB. Taken together, our findings present a paradigm shift in our understanding of the molecular mechanisms that regulate mesoangioblast stem cells ability to traverse the extracellular matrix (ECM). J. Cell. Physiol. 232: 1845–1861, 2017. © 2016 Wiley Periodicals, Inc.Extracellular Hsp70 released through extracellular vesicles regulates mesoangioblast migration via the activation of NF-κB and PI3K/AKT pathways.
      PubDate: 2017-01-31T07:19:01.293202-05:
      DOI: 10.1002/jcp.25722
  • Calprotectin Induces IL-6 and MCP-1 Production via Toll-Like Receptor 4
           Signaling in Human Gingival Fibroblasts
    • Authors: Yasufumi Nishikawa; Yukari Kajiura, Jung Hwan Lew, Jun-ichi Kido, Toshihiko Nagata, Koji Naruishi
      First page: 1862
      Abstract: Calprotectin, a heterodimer of S100A8 and S100A9 molecules, is associated with inflammatory diseases such as inflammatory bowel disease. We have reported that calprotectin levels in gingival crevicular fluids of periodontitis patients are significantly higher than in healthy subjects. However, the functions of calprotectin in pathophysiology of periodontitis are still unknown. The aim of this study is to investigate the effects of calprotectin on the productivity of inflammatory cytokines in human gingival fibroblasts (HGFs). The HGFs cell line CRL-2014® (ATCC) were cultured, and total RNAs were collected to examine the expression of TLR2/4 and RAGE mRNA using RT-PCR. After the cells were treated with S100A8, S100A9, and calprotectin, supernatants were collected and the levels of IL-6 and MCP-1 were measured using ELISA methods. To examine the intracellular signals involved in calprotectin-induced cytokine production, several chemical inhibitors were used. Furthermore, after the siRNA-mediated TLR4 down-regulated cells were treated with S100A8, S100A9, and calprotectin, the levels of IL-6 and MCP-1 were also measured. HGFs showed greater expression of TLR4 mRNA, but not TLR2 and RAGE mRNA compared with human oral epithelial cells. Calprotectin increased significantly the production of MCP-1 and IL-6 in HGFs, and the cytokine productions were significantly suppressed in the cells treated with MAPKs, NF-κB, and TLR4 inhibitors. Furthermore, calprotectin-mediated MCP-1 and IL-6 production were significantly suppressed in TLR4 down-regulated cells. Taken together, calprotectin induces IL-6 and MCP-1 production in HGFs via TLR4 signaling that involves MAPK and NF-κB, resulting in the progression of periodontitis. J. Cell. Physiol. 232: 1862–1871, 2017. © 2016 Wiley Periodicals, Inc.
      PubDate: 2017-01-06T10:15:40.270932-05:
      DOI: 10.1002/jcp.25724
  • Calretinin Immunoreactivity in the Human Testis Throughout Fetal Life
    • Authors: Giovanna G. Altobelli; Francesca Pentimalli, Mariarosaria D'Armiento, Susan Van Noorden, Vincenzo Cimini
      First page: 1872
      Abstract: The main functions of the testis are sex hormone and sperm cell production. Steroidogenesis occurs in the Leydig interstitial cells and spermatogenesis in the seminiferous tubules. Male gonad morphogenesis is a finely orchestrated process, mainly coordinated by hormones, whose actions can significantly affect post-pubertal testicular function. Calcium is a key intracellular messenger, which regulates many signal transduction pathways, and is also implicated in steroidogenesis. Calcium homeostasis and signaling rely on many calcium-binding proteins including calretinin, of the “EF-hand” protein family. Calretinin is a highly conserved protein mainly expressed in the nervous system but also detected in rat and human adult and fetal testis as well as in pathological conditions. Calretinin expression in the fetal testis, however, has not been thoroughly analyzed probably owing to limited availability and paucity of tissues. Here, we examined by immunocytochemistry the expression of calretinin in human fetal testis specimens, obtained from natural and therapeutic abortions, at various developmental ages. We found that calretinin-immunoreactive Leydig cells were visible throughout the timeframe studied (14th–27th week). Immunoreactivity was also observed in Sertoli cells and in the germ cells of the immature seminiferous tubules. Overall our data indicate that calretinin expression parallels the decline in Leydig cell number, suggesting that its presence is indeed correlated to their steroidogenic activity. They also suggest that the intratubular positivity of calretinin could be linked to the ability of Sertoli cells to produce locally acting hormones contributing to the histodifferentiation of the male genital tract. J. Cell. Physiol. 232: 1872–1878, 2017. © 2016 Wiley Periodicals, Inc.Human fetal testis specimens, obtained upon natural and therapeutic abortions at various developmental ages (weeks 14–27), show calretinin expression in the Leydig interstitial cells, in the epididymis and in the immature seminiferous tubules. Calretinin expression parallels the decline in Leydig cell number. The presence of calretinin seems correlated to the Leydig cells steroidogenic activity, which is consistent with the role that calcium intracellular signaling plays in the steroidogenic process. Our findings, by adding calretinin to the list of players underlying fetal testis development, are particularly timely and prompt further assessment of calretinin role in this context both in normal and pathological conditions.
      PubDate: 2017-01-05T08:25:28.973764-05:
      DOI: 10.1002/jcp.25727
  • Fibroblast Growth Factor 21 (FGF21) Promotes Formation of Aerobic
           Myofibers via the FGF21-SIRT1-AMPK-PGC1α Pathway
    • Authors: Xinyi Liu; Yongliang Wang, Liming Hou, Yuanzhu Xiong, Shuhong Zhao
      First page: 1893
      Abstract: Fibroblast growth factor 21(FGF21) is a pivotal regulator of energy metabolism, which is currently being assessed as a potential drug target for the treatment of insulin-resistant conditions. However, the cellular mechanisms by which FGF21 affects myogenesis remain unclear. In this study, we explored the function of FGF21 in myogenesis both in vitro and in vivo. Our experiments showed for the first time that FGF21 promotes myoblast differentiation and serves as a switch of molecular transformation from anaerobic myofibers to aerobic myofibers via the FGF21-SIRT1-AMPK-PGC1α axis. Furthermore, we employed the Dual-Luciferase Reporter Assay System and Electrophoretic Mobility Shift Assay (EMSA) and demonstrated that MYOD, a major myogenic transcription factor, binds directly to the promoter region of Fgf21, leading to the activation of Fgf21 expression in mouse C2C12 myoblasts. Our study revealed a novel mechanism of myogenesis and muscle fiber transformation and indicated that FGF21 serves as a vital regulator of muscle development and important contributor to the pathogenesis of myopathy. J. Cell. Physiol. 232: 1893–1906, 2017. © 2016 Wiley Periodicals, Inc.(1) The mouse FGF21 gene expression is positively regulated by the transcription factor MyoD. (2) FGF21 promotes myogenic differentiation and facilitates the switch of the muscle fiber type from anaerobic myofibers to aerobic myofibers via stimulation the FGF21-SIRT1-AMPK-PGC1α axis. (3) The increased FGF21 protein induced by fasting is likely to change the expression of the PGC-1α protein from a physiological to an abnormally excessive level via the activation of AMPK signaling, which results in muscle atrophy.
      PubDate: 2017-01-05T08:30:37.504911-05:
      DOI: 10.1002/jcp.25735
  • Rabs, Membrane Dynamics, and Parkinson's Disease
    • Authors: Bor Luen Tang
      First page: 1626
      Abstract: Genes encoding cellular membrane trafficking components, namely RAB7L1 and RAB39B, are more recently recognized factors associated with Parkinson's disease (PD). Encoded by a gene within the PARK16 locus, RAB7L1 interacts with Leucine-rich repeat kinase 2 (LRRK2) to act in intracellular transport processes that are likely important for neuronal survival and function. LRRK2 also directly phosphorylates a number of other Rab proteins. On the other hand, nonsense and missense mutations of the X-chromosome localized RAB39B were shown to underlie X-linked intellectual disability (ID) in male patients with early-onset PD. The cellular or neuronal functions of RAB39B are not yet known with certainty, but it has recently been shown to play a role in glutamate receptor trafficking. Importantly, RAB39B is also functionally connected to components for autophagy regulation, which affects α-synuclein processing and clearance. In this review, we discuss the association of Rabs with PD pathology, and potential etiological mechanisms whereby defects or deficiencies in certain Rab proteins could lead to PD susceptibility. J. Cell. Physiol. 232: 1626–1633, 2017. © 2016 Wiley Periodicals, Inc.In this review, I outlined and discussed a few recently recognized membrane traffic components that impinge on Parkinson Disease pathology, with specific attention on how some classic PD factors interacts with Rab GTPases, and RAB7L1 and RAB39B, whose variance and mutations predisposes an individual to the disease.
      PubDate: 2016-12-20T08:30:41.441112-05:
      DOI: 10.1002/jcp.25713
  • Nestin Expressed by Pre-Existing Cardiomyocytes Recapitulated in Part an
           Embryonic Phenotype; Suppressive Role of p38 MAPK
    • Authors: Marc-Andre Meus; Vanessa Hertig, Louis Villeneuve, Jean-Francois Jasmin, Angelino Calderone
      First page: 1717
      Abstract: Nestin(+)-cardiomyocytes were identified in the ischemically damaged human/rodent heart, albeit the cellular source, and signaling events implicated in the appearance of the intermediate filament protein remained undefined. Expression of the enhanced green fluorescent protein (EGFP) driven by the second intron of the nestin gene identified a subpopulation of EGFP/nestin(+) cells that differentiated to a vascular phenotype in the peri-infarct/infarct region of post-MI mice albeit the transgene was not detected in nestin(+)-cardiomyocytes. α-MHC-driven expression of the reporter mCherry was detected in troponin-T(+)- and nestin(+)-cardiomyocytes in the peri-infarct/infarct region of post-MI mice. However, the cell cycle re-entry of nestin/mCherry(+)-cardiomyocytes was not observed. Nestin staining was identified in a paucity of neonatal rat ventricular cardiomyocytes (NNVM). Exposure to phorbol 12,13-dibutyrate (PDBu) induced NNVM hypertrophy but did not promote nestin expression or Brdu incorporation. PDBu treatment of NNVMs phosphorylated p38 MAPK and HSP27 and HSP27 phosphorylation was abrogated by the p38 MAPK inhibitor SB203580. PDBu/SB203580 co-treatment significantly increased the percentage of NNVMs that expressed nestin and incorporated Brdu. In the heart of embryonic 10.5 day mice, nestin immunoreactivity was observed in cycling troponin-T(+)-cardiomyocytes. Nestin was also detected in embryonic rat ventricular cardiomyocytes and depletion of the intermediate filament protein attenuated cell cycle re-entry. Thus, nestin expressed by pre-existing cardiomyocytes following ischemic damage recapitulated in part an embryonic trait and may provide the requisite phenotype to initiate cell cycle re-entry. However, the overt activation of the p38 MAPK pathway post-MI may in part limit the appearance and inhibit the cell cycle re-entry of nestin(+)-cardiomyocytes. J. Cell. Physiol. 232: 1717–1727, 2017. © 2016 Wiley Periodicals, Inc.Nestin was expressed by pre-existing cardiomyocytes following ischemic damage to the adult heart and recapitulated in part an embryonic phenotype. However, nestin-expressing cardiomyocytes were unable to re-enter the cell cycle following ischemic damage because of the suppressive effect of the p38 MAPK pathway.
      PubDate: 2016-07-27T10:50:36.765274-05:
      DOI: 10.1002/jcp.25496
  • Differences in Activation of HIV-1 Replication by Superinfection With
           HIV-1 and HIV-2 in U1 Cells
    • Authors: Xue Wang; Bing Sun, Christelle Mbondji, Santanu Biswas, Jiangqin Zhao, Indira Hewlett
      First page: 1746
      Abstract: Macrophages contribute to HIV-1 pathogenesis by forming a viral reservoir that serve as a viral source for the infection of CD4 T cells. The relationship between HIV-1 latent infection and superinfection in macrophages has not been well studied. Using susceptible U1 cells chronically infected with HIV-1, we studied the effects of HIV superinfection on latency and differences in superinfection with HIV-1 and HIV-2 in macrophages. We found that HIV-1 (MN) superinfection displayed increased HIV-1 replication in a time-dependent manner; while cells infected with HIV-2 (Rod) initially showed increased HIV-1 replication, followed by a decrease in HIV-1 RNA production. HIV-1 superinfection upregulated/activated NF-ĸB, NFAT, AP-1, SP-1, and MAPK Erk through expression/activation of molecules, CD4, CD3, TCRβ, Zap-70, PLCγ1, and PKCΘ in T cell receptor-related signaling pathways; while HIV-2 superinfection initially increased expression/activation of these molecules followed by decreased protein expression/activation. HIV superinfection initially downregulated HDAC1 and upregulated acetyl-histone H3 and histone H3 (K4), while HIV-2 superinfection demonstrated an increase in HDAC1 and a decrease in acetyl-histone H3 and histone H3 (K4) relative to HIV-1 superinfection. U1 cells superinfected with HIV-1 or HIV-2 showed differential expression of proteins, IL-2, PARP-1, YB-1, and LysRS. These findings indicate that superinfection with HIV-1 or HIV-2 has different effects on reactivation of HIV-1 replication. HIV-1 superinfection with high load of viral replication may result in high levels of cytotoxicity relative to HIV-2 superinfection. Cells infected with HIV-2 showed lower level of HIV-1 replication, suggesting that co-infection with HIV-2 may result in slower progression toward AIDS. J. Cell. Physiol. 232: 1746–1753, 2017. © 2016 Wiley Periodicals, Inc.
      PubDate: 2016-09-30T09:20:42.86365-05:0
      DOI: 10.1002/jcp.25614
  • Lgr4 Expression in Osteoblastic Cells Is Suppressed by Hydrogen Peroxide
    • Authors: Chantida Pawaputanon Na Mahasarakham; Yayoi Izu, Katsuhiko Nishimori, Yuichi Izumi, Masaki Noda, Yoichi Ezura
      First page: 1761
      Abstract: LGR4 is expressed in bone and has been shown to be involved in bone metabolism. Oxidative stress is one of the key issues in pathophysiology of osteoporosis. However, the link between Lgr4 and oxidative stress has not been known. Therefore, effects of hydrogen peroxide on Lgr4 expression in osteoblasts were examined. Hydrogen peroxide treatment suppressed the levels of Lgr4 mRNA expression in an osteoblastic cell line, MC3T3-E1. The suppressive effects were not obvious at 0.1 mM, while 1 mM hydrogen peroxide suppressed Lgr4 expression by more than 50%. Hydrogen peroxide treatment suppressed Lgr4 expression within 12 h and this suppression lasted at least up to 48 h. Hydrogen peroxide suppression of Lgr4 expression was still observed in the presence of a transcription inhibitor but was no longer observed in the presence of a protein synthesis inhibitor. Although Lgr4 expression in osteoblasts is enhanced by BMP2 treatment as reported before, hydrogen peroxide treatment suppressed Lgr4 even in the presence of BMP2. Finally, hydrogen peroxide suppressed Lgr4 expression in primary cultures of osteoblasts similarly to MC3T3-E1 cells. These date indicate that hydrogen peroxide suppresses Lgr4 expression in osteoblastic cells. J. Cell. Physiol. 232: 1761–1766, 2017. © 2016 Wiley Periodicals, Inc.
      PubDate: 2016-12-29T05:37:03.982426-05:
      DOI: 10.1002/jcp.25684
  • CD93 as a Potential Target in Neovascular Age-Related Macular Degeneration
    • Authors: Gian Marco Tosi; Elena Caldi, Barbara Parolini, Paolo Toti, Giovanni Neri, Federica Nardi, Claudio Traversi, Gabriele Cevenini, Davide Marigliani, Elisabetta Nuti, Tommaso Bacci, Federico Galvagni, Maurizio Orlandini
      First page: 1767
      Abstract: In patients with age-related macular degeneration (AMD), choroidal neovascularization is the major cause of severe visual loss. In these patients, the persistence of neovascular growth despite vascular endothelial growth factor-A blockage needs the discovery of new endothelial cell targets. The glycoprotein CD93, highly expressed in activated endothelial cells, has been recently involved in the regulation of the angiogenic process both as transmembrane and soluble protein. Choroidal neovascular membranes from patients affected by AMD were examined by immunofluorescence using anti-CD93 and anti-von Willebrand factor antibodies. Blood vessels within intraocular and extraocular neoplasias were used as controls for CD93 expression. All choroidal neovascular membranes displayed strong CD93 staining in the von Willebrand factor-positive endothelial cells, consistently with the analyses showing a high colocalization coefficient in the blood vessels. Intraocular and extraocular tumor vessels showed similar results, whereas the normal choroid displayed blood vessels with only faint CD93 staining. Additionally, the concentration of soluble CD93 was determined in the aqueous humor of patients affected by naïve neovascular AMD by enzyme-linked immunosorbent assays. Age-matched cataract patients served as controls. Soluble CD93 was significantly increased in the aqueous humor of naïve neovascular AMD patients and tended to decrease after treatment with an antiangiogenic drug. In conclusion, both transmembrane and soluble CD93 are overexpressed in patients with neovascular AMD, indicating that CD93 may represent a potential new antiangiogenic target in the treatment of choroidal neovascularization. J. Cell. Physiol. 232: 1767–1773, 2017. © 2016 Wiley Periodicals, Inc.In patients with age-related macular degeneration (AMD), the formation of new blood vessels in the choroid, which is the major cause of severe visual loss, persists even after therapy. In AMD patients, both in the endothelial cells of choroidal neovascular membranes and in the aqueous humor, we found high levels of the glycoprotein CD93, which has been recently involved in the enhancement of tumor vascular function. The presence of CD93 in AMD patients unveils a potential new target for choroidal neovascularization treatment.
      PubDate: 2016-11-30T10:30:45.761141-05:
      DOI: 10.1002/jcp.25689
  • TRPV4 Regulates Tight Junctions and Affects Differentiation in a Cell
           Culture Model of the Corneal Epithelium
    • Authors: Jacqueline Martínez-Rendón; Erika Sánchez-Guzmán, Angélica Rueda, James González, Rosario Gulias-Cañizo, Guillermo Aquino-Jarquín, Federico Castro-Muñozledo, Refugio García-Villegas
      First page: 1794
      Abstract: TRPV4 (transient receptor potential vanilloid 4) is a cation channel activated by hypotonicity, moderate heat, or shear stress. We describe the expression of TRPV4 during the differentiation of a corneal epithelial cell model, RCE1(5T5) cells. TRPV4 is a late differentiation feature that is concentrated in the apical membrane of the outmost cell layer of the stratified epithelia. Ca2+ imaging experiments showed that TRPV4 activation with GSK1016790A produced an influx of calcium that was blunted by the specific TRPV4 blocker RN-1734. We analyzed the involvement of TRPV4 in RCE1(5T5) epithelial differentiation by measuring the development of transepithelial electrical resistance (TER) as an indicator of the tight junction (TJ) assembly. We showed that TRPV4 activity was necessary to establish the TJ. In differentiated epithelia, activation of TRPV4 increases the TER and the accumulation of claudin-4 in cell–cell contacts. Epidermal Growth Factor (EGF) up-regulates the TER of corneal epithelial cultures, and we show here that TRPV4 activation mimicked this EGF effect. Conversely, TRPV4 inhibition or knock down by specific shRNA prevented the increase in TER. Moreover, TRPP2, an EGF-activated channel that forms heteromeric complexes with TRPV4, is also concentrated in the outmost cell layer of differentiated RCE1(5T5) sheets. This suggests that the EGF regulation of the TJ may involve a heterotetrameric TRPV4-TRPP2 channel. These results demonstrated TRPV4 activity was necessary for the correct establishment of TJ in corneal epithelia and as well as the regulation of both the barrier function of TJ and its ability to respond to EGF. J. Cell. Physiol. 232: 1794–1807, 2017. © 2016 Wiley Periodicals, Inc.TRPV4 channel activity is necessary for the correct establishment of tight Junctions (TJ) in corneal epithelia and as well as the regulation of both the barrier function of TJ and its ability to respond to EGF. Thus, TRPV4 function is necessary for the terminal differentiation of corneal epithelia.
      PubDate: 2016-11-30T10:06:26.658836-05:
      DOI: 10.1002/jcp.25698
  • Supernatants of Adipocytes From Obese Versus Normal Weight Women and
           Breast Cancer Cells: In Vitro Impact on Angiogenesis
    • Authors: Lauriane Bougaret; Laetitia Delort, Hermine Billard, Charlotte Lequeux, Nicolas Goncalves-Mendes, Ali Mojallal, Odile Damour, Marie-Paule Vasson, Florence Caldefie-Chezet
      First page: 1808
      Abstract: Breast cancer is correlated with a higher risk of metastasis in obese postmenopausal women. Adipokines, whose plasma concentrations are modulated in obese subjects and adipocytes surround mammary cells, suggesting that adipocyte secretome affect mammary tumorogenesis. We hypothesize that mature adipocyte secretions from obese women conditioned or not by breast neoplasic cells, increase changes on the angiogenesis stages. Supernatants of human mature adipocytes, differentiated from stem cells of either adipose tissue of normal weight (MA20) or obese (MA30) women or obtained from co-cultures between MA20 and MA30 and breast cancer cell line MCF-7, were collected. The impact of these supernatants was investigated on proliferation, migration, and tube formation by endothelial cells (HUVEC). MA20 and MA30 showed a preservation of their “metabolic memory” (increase of Leptin, ObR, VEGF, CYP19A1, and a decrease of Adiponectin expression in MA30 compared to MA20). Supernatants from obese-adipocytes increased HUVEC proliferation, migration, and sprouting like with supernatants obtained from co-cultures of MA/MCF-7 regardless the women's BMI. Additional analyses such as the use of neutralizing antibodies, analysis of supernatants (Milliplex®) and variations in gene expression (qRT-PCR), strongly suggest an implication of IL-6, or a synergistic action among adipokines, probably associated with that of VEGF or IL-6. As a conclusion, supernatants from co-cultures of MA30 and MCF-7 cells increase proliferation, migration, and sprouting of HUVEC cells. These results provide insights into the interaction between adipocytes and epithelial cancer cells, particularly in case of obesity. The identification of synergistic action of adipokines would therefore be a great interest in developing preventive strategies. J. Cell. Physiol. 232: 1808–1816, 2017. © 2016 Wiley Periodicals, Inc.We characterized the impact of adipokines from obese or normal weight women on different angiogenesis stages (proliferation, migration, and tube formation of endothelial cells) in order to determine the influence of obesity on this process and the impact of adipocytes on breast tumor cells.
      PubDate: 2016-12-20T08:30:45.692881-05:
      DOI: 10.1002/jcp.25701
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