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Journal Cover Journal of Cellular Physiology
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   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  [1604 journals]   [SJR: 1.608]   [H-I: 118]
  • PDE5 inhibitors enhance Celecoxib killing in multiple tumor types
    • Authors: Laurence Booth; Jane L. Roberts, Nichola Cruickshanks, Seyedmehrad Tavallai, Timothy Webb, Peter Samuel, Adam Conley, Brittany Binion, Harold F. Young, Andrew Poklepovic, Sarah Spiegel, Paul Dent
      Pages: n/a - n/a
      Abstract: The present studies determined whether clinically relevant phosphodiesterase 5 (PDE5) inhibitors interacted with a clinically relevant NSAID, celecoxib, to kill tumor cells. Celecoxib and PDE5 inhibitors interacted in a greater than additive fashion to kill multiple tumor cell types. Celecoxib and sildenafil killed ex vivo primary human glioma cells as well as their associated activated microglia. Knock down of PDE5 recapitulated the effects of PDE5 inhibitor treatment; the nitric oxide synthase inhibitor L‐NAME suppressed drug combination toxicity. The effects of celecoxib were COX2 independent. Over‐expression of c‐FLIP‐s or knock down of CD95/FADD significantly reduced killing by the drug combination. CD95 activation was dependent on nitric oxide and ceramide signaling. CD95 signaling activated the JNK pathway and inhibition of JNK suppressed cell killing. The drug combination inactivated mTOR and increased the levels of autophagy and knock down of Beclin1 or ATG5 strongly suppressed killing by the drug combination. The drug combination caused an ER stress response; knock down of IRE1α/XBP1 enhanced killing whereas knock down of eIF2α/ATF4/CHOP suppressed killing. Sildenafil and celecoxib treatment suppressed the growth of mammary tumors in vivo. Collectively our data demonstrate that clinically achievable concentrations of celecoxib and sildenafil have the potential to be a new therapeutic approach for cancer. J. Cell. Physiol. © 2014 Wiley Periodicals, Inc.
      PubDate: 2014-10-09T12:42:59.57235-05:0
      DOI: 10.1002/jcp.24843
  • Permanent culture of macrophages at physiological oxygen attenuates the
           antioxidant and immunomodulatory properties of dimethyl fumarate
    • Authors: Benjamin Haas; Sandra Chrusciel, Sarah Fayad‐Kobeissi, Jean‐Luc Dubois‐Randé, Francisco Azuaje, Jorge Boczkowski, Roberto Motterlini, Roberta Foresti
      Pages: n/a - n/a
      Abstract: We hypothesized that O2 tension influences the redox state and the immunomodulatory responses of inflammatory cells to dimethyl fumarate (DMF), an activator of the nuclear factor Nrf2 that controls antioxidant genes expression. This concept was investigated in macrophages permanently cultured at either physiological (5% O2) or atmospheric (20% O2) oxygen levels and then treated with DMF or challenged with lipopolysaccharide (LPS) to induce inflammation. RAW 264.7 macrophages cultured at 20% O2 exhibited a pro‐oxidant phenotype, reflected by a lower content of reduced glutathione, higher oxidized glutathione and increased production of reactive oxygen species when compared to macrophages continuously grown at 5% O2. At 20% O2, DMF induced a stronger antioxidant response compared to 5% O2 as evidenced by a higher expression of heme oxygenase‐1, NAD(P)H:quinone oxydoreductase‐1 and superoxide dismutase‐2. After challenge of macrophages with LPS, several pro‐inflammatory (iNOS, TNF‐α, MMP‐2, MMP‐9), anti‐inflammatory (arginase‐1, IL‐10) and pro‐angiogenic (VEGF‐A) mediators were evaluated in the presence or absence of DMF. All markers, with few interesting exceptions, were significantly reduced at 5% O2. This study brings new insights on the effects of O2 in the cellular adaptation to oxidative and inflammatory stimuli and highlights the importance of characterizing the effects of chemicals and drugs at physiologically relevant O2 tension. Our results demonstrate that the common practice of culturing cells at atmospheric O2 drives the endogenous cellular environment towards an oxidative stress phenotype, affecting inflammation and the expression of antioxidant pathways by exogenous modulators. J. Cell. Physiol. © 2014 Wiley Periodicals, Inc.
      PubDate: 2014-10-09T12:42:44.639233-05:
      DOI: 10.1002/jcp.24844
  • Cyclooxygenase‐2 Regulates NLRP3 Inflammasome‐Derived
           IL‐1β Production
    • Authors: Kuo‐Feng Hua; Ju‐Ching Chou, Shuk‐Man Ka, Yu‐Ling Tasi, Ann Chen, Shih‐Hsiung Wu, Hsiao‐Wen Chiu, Wei‐Ting Wong, Yih‐Fuh Wang, Change‐Ling Tsai, Chen‐Lung Ho, Cheng‐Hsiu Lin
      Pages: n/a - n/a
      Abstract: The NLR family, pyrin domain‐containing 3 (NLRP3) inflammasome is a reactive oxygen species‐sensitive multiprotein complex that regulates IL‐1β maturation via caspase‐1. It also plays an important role in the pathogenesis of inflammation‐related disease. Cyclooxygenase‐2 (COX‐2) is induced by inflammatory stimuli and contributes to the pathogenesis of inflammation‐related diseases. However, there is currently little known about the relationship between COX‐2 and the NLRP3 inflammasome. Here, we describe a novel role for COX‐2 in regulating the activation of the NLRP3 inflammasome. NLRP3 inflammasome‐derived IL‐1β secretion and pyroptosis in macrophages were reduced by pharmaceutical inhibition or genetic knockdown of COX‐2. COX‐2 catalyzes the synthesis of prostaglandin E2 and increases IL‐1β secretion. Conversely, pharmaceutical inhibition or genetic knockdown of prostaglandin E2 receptor 3 reduced IL‐1β secretion. The underlying mechanisms for the COX‐2‐mediated increase in NLRP3 inflammasome activation were determined to be the following: (1) enhancement of lipopolysaccharide‐induced proIL‐1β and NLRP3 expression by increasing NF‐κB activation and (2) enhancement of the caspase‐1 activation by increasing damaged mitochondria, mitochondrial reactive oxygen species production and release of mitochondrial DNA into cytosol. Furthermore, inhibition of COX‐2 in mice in vivo with celecoxib reduced serum levels of IL‐1β and caspase‐1 activity in the spleen and liver in response to lipopolysaccharide (LPS) challenge. These findings provide new insights into how COX‐2 regulates the activation of the NLRP3 inflammasome and suggest that it may be a new potential therapeutic target in NLRP3 inflammasome‐related diseases. J. Cell. Physiol. © 2014 Wiley Periodicals, Inc.
      PubDate: 2014-10-07T14:19:29.251235-05:
      DOI: 10.1002/jcp.24815
  • Effect of rMnSOD on survival signaling in pediatric high risk T‐cell
           acute lymphoblastic leukaemia
    • Authors: A Pica; A Di Santi, V D'angelo, A Iannotta, M Ramaglia, M Di Martino, Ml Pollio, A Schiattarella, A Borrelli, A Mancini, P Indolfi, F. Casale
      Pages: n/a - n/a
      Abstract: Manganese superoxide dismutase (MnSOD) is a mitochondrial enzyme that defends against oxidative damage due to reactive oxygen species (ROS). A new isoform of MnSOD with cytotoxic activity was recently discovered in liposarcoma cells. Here, we tested the effectiveness of a recombinant form of this isoform (rMnSOD) on leukemic T cells, Jurkat cells, and normal lymphocytes. Our results confirm that leukemic T cells can internalize rMnSOD and that rMnSOD causes apoptosis of 99% of leukemic cells without showing toxic effects on healthy cells. Using light and electron microscopy, we determined that an rMnSOD concentration of 0.067 μM most effective on apoptosis induction. Western blot analysis showed that treatment with 0.067 μM rMnSOD resulted in high expression of the pro‐apoptotic protein Bax and low expression of the anti‐apoptotic protein Bcl‐2 in leukemia cells. Concerning signal transduction pathway no influence was observed after treatment except for Jurkat cells showing a slightly decreased expression of ERK phosphorylation. These results suggest that rMnSOD may be an effective and non‐toxic treatment option for T‐cell leukemia. J. Cell. Physiol. © 2014 Wiley Periodicals, Inc.
      PubDate: 2014-10-07T11:36:06.767904-05:
      DOI: 10.1002/jcp.24840
  • Prostaglandin Dependent Control of an Endogenous Estrogen Receptor Agonist
           by Osteoblasts
    • Authors: Thomas L. McCarthy; Michael Centrella
      Pages: n/a - n/a
      Abstract: Estrogen receptor (ER) activation has complex effects on bone cells, and loss of circulating estradiol adversely affects skeletal status in women. Hormone replacement therapy effectively circumvents bone loss after menopause, but enhances disease risk in other tissues. Here we show that prostaglandin E2 (PGE2) augments the activity of an osteoblast‐derived selective ER modulator, ObSERM. The stimulatory effect of PGE2 is replicated in part by either the PG receptor EP3 agonist 17‐phenyl trinor PGE2 or by the PG receptor FP agonist PGF2α. Whereas activation of the various PG receptors induces multiple downstream signals, the response to PGE2 was mimicked by activators of protein kinase C, and suppressed by inhibition of protein kinase C but not by inhibition of protein kinase A. Moreover, inhibition of nitric oxide synthesis and activation of the PTH and Wnt pathways increases ObSERM activity. Our studies therefore reveal that ObSERM activity is controlled in distinct ways and revise our understanding of ER activation within bone by agents or events associated with PG expression. They also predict ways to sustain or improve bone formation, fracture repair, and surgical healing without adding the risk of disease in other tissues where ER activation also has important biological functions. J. Cell. Physiol. © 2014 Wiley Periodicals, Inc.
      PubDate: 2014-10-07T11:35:38.523439-05:
      DOI: 10.1002/jcp.24842
  • DNA‐PKcs deficiency inhibits glioblastoma cell‐derived
           angiogenesis after ionizing radiation
    • Authors: Yang Liu; Luwei Zhang, Yuanyuan Liu, Chao sun, Hong Zhang, Guoying Miao, Cui Xiao Di, Xin Zhou, Rong Zhou, Zhenhua Wang
      Pages: n/a - n/a
      Abstract: DNA‐dependent protein kinase catalytic subunit (DNA‐PKcs) plays a critical role in non‐homologous end‐joining repair of DNA double‐strand breaks (DSB) induced by ionizing radiation (IR). Little is known, however, regarding the relationship between DNA‐PKcs and IR‐induced angiogenesis; thus, in this study we aimed to further elucidate this relationship. Our findings revealed that lack of DNA‐PKcs expression or activity sensitized glioma cells to radiation due to the defective DNA DSB repairs and inhibition of phosphorylated AktSer473. Moreover, DNA‐PKcs deficiency apparently mitigated IR‐induced migration, invasion and tube formation of human microvascular endothelial cell (HMEC‐1) in conditioned media derived from irradiated DNA‐PKcs mutant M059J glioma cells or M059K glioma cells that have inhibited DNA‐PKcs kinase activity due to the specific inhibitor NU7026 or siRNA knockdown. Moreover, IR‐elevated vascular endothelial growth factor (VEGF) secretion was abrogated by DNA‐PKcs suppression. Supplemental VEGF antibody to irradiated‐conditioned media was negated enhanced cell motility with a concomitant decrease in phosphorylation of the FAK Try925 and SrcTry416. Furthermore, DNA‐PKcs suppression was markedly abrogated in IR‐induced transcription factor hypoxia inducible factor‐1α (HIF‐1α) accumulation, which is related to activation of VEGF transcription. These findings, taken together, demonstrate that depletion of DNA‐PKcs in glioblastoma cells at least partly suppressed IR‐inflicted migration, invasion and tube formation of HMEC‐1 cells, which may be associated with the reduced HIF‐1α level and VEGF secretion. Inhibition of DNA‐PKcs may be a promising therapeutic approach to enhance radio‐therapeutic efficacy for glioblastoma by hindering its angiogenesis. J. Cell. Physiol. © 2014 Wiley Periodicals, Inc.
      PubDate: 2014-10-07T11:34:26.23108-05:0
      DOI: 10.1002/jcp.24841
  • Role of Methionine Adenosyltransferase α2 and β Phosphorylation
           and Stabilization in Human Hepatic Stellate Cell
    • Authors: Komal Ramani; Shant Donoyan, Maria Lauda Tomasi, Sunhee Park
      Pages: n/a - n/a
      Abstract: Myofibroblastic trans‐differentiation of hepatic stellate cells (HSCs) is an essential event in the development of liver fibrogenesis. These changes involve modulation of key regulators of the genome and the proteome. Methionine adenosyltransferases (MAT) catalyze the biosynthesis of the methyl donor, S‐adenosylmethionine (SAMe) from methionine. We have previously shown that two MAT genes, MAT2A and MAT2B (encoding MATα2 and MATβ proteins respectively), are required for HSC activation and loss of MAT2A transcriptional control favors its up‐regulation during trans‐differentiation. Hence MAT genes are intrinsically linked to the HSC machinery during activation. In the current study, we have identified for the first time, post‐translational modifications in the MATα2 and MATβ proteins that stabilize them and favor human HSC trans‐differentiation. Culture‐activation of human HSCs induced the MATα2 and MATβ proteins. Using mass spectrometry, we identified phosphorylation sites in MATα2 and MATβ predicted to be phosphorylated by mitogen‐activated protein kinase (MAPK) family members [ERK1/2, V‐Raf Murine Sarcoma Viral Oncogene Homolog B1 (B‐Raf), MEK]. Phosphorylation of both proteins was enhanced during HSC activation. Blocking MEK activation lowered the phosphorylation and stability of MAT proteins without influencing their mRNA levels. Silencing ERK1/2 or B‐Raf lowered the phosphorylation and stability of MATβ but not MATα2. Reversal of the activated human HSC cell line, LX2 to quiescence lowered phosphorylation and destabilized MAT proteins. Mutagenesis of MATα2 and MATβ phospho‐sites destabilized them and prevented HSC trans‐differentiation. The data reveal that phosphorylation of MAT proteins during HSC activation stabilizes them thereby positively regulating trans‐differentiation. J. Cell. Physiol. © 2014 Wiley Periodicals, Inc.
      PubDate: 2014-10-07T09:36:32.892568-05:
      DOI: 10.1002/jcp.24839
  • Methods and Procedures in Adipose Stem Cells: State of the Art and
           Perspective For Translation Medicine
    • Authors: G.F. Nicoletti; F. De Francesco, F. D'Andrea, G.A. Ferraro
      Pages: n/a - n/a
      Abstract: Stem cells have potential in the retrieval and repair of injured tissue and renovation of organ function. To date, several studies have been carried out to elucidate how differentiation of stem cells can be used in regenerative medicine applications. Adipose tissue is an abundant and accessible source of stem cell, useful for regenerative therapeutic use. Adipose stem cells (ASCs) are favorable for future translational research and can be applied in many clinical settings. Adipose tissue repair has been recently adopted in clinical trials to prove that ASCs can be successfully used in patients. Variability in cell culture procedures (isolation, characterization and differentiation) may have an influence on the experimental outcome. In this report, we consider the selection mechanisms of ASCs using flow cytometry, cell culture, freezing/thawing, cell cycle evaluation, histochemistry/immunofluorescence and differentiation of ASCs. Both researchers and regulatory institutions should consider a new policy for GMP procedures and protocols, paying special attention to stem cell bio‐physiology, to facilitate more clinically oriented studies. ASCs show angiogenic properties, with prospects of repairing tissue damaged by radiotherapy, as well as possessing the ability to heal chronic wounds. They can also be useful in surgical practice. We focus on the potential clinical application of ASCs that are currently available regarding translational medicine and the methods and procedures for their isolation, differentiation and characterization. J. Cell. Physiol. © 2014 Wiley Periodicals, Inc.
      PubDate: 2014-10-07T09:34:51.035164-05:
      DOI: 10.1002/jcp.24837
  • Pregnane X Receptor as the “Sensor and Effector” in Regulating
    • Authors: Xi Ma; Jingshu Chen, Yanan Tian
      Pages: n/a - n/a
      Abstract: The pregnane X receptor (PXR, NR1I2) is a ligand‐activated nuclear receptor which plays an essential role in organism's metabolic detoxification system by sensing the presence of xenobiotics and triggering detoxification responses. In addition to its role in xenobiotic metabolism, PXR has pleiotropic functions in regulating immune/inflammatory responses, cell proliferation, bile acid/cholesterol metabolism, glucose and lipid metabolism, steroid/endocrine homeostasis and bone metabolism. Recent research suggests that the PXR is required for maintaining healthy commensalism between microbiota and gut. Interestingly, the metabolites such as indole derivatives from commensal microbes serve as the ligands for the PXR in intestinal epithelium forming an intricate mutualistic interaction between host and microbiota. PXR‐regulated gene responses are controlled at epigenetic level by chromatin modifications, DNA methylation and noncoding RNA. Developmental alterations of the epigenome by exposure to the xenobiotics or diseases may produce persistent changes in PXR‐regulated physiological responses. These new areas of research promise to vastly increase our understanding of PXR‐regulated responses. In this review we highlight recent results on the epigenetic mechanisms for the PXR‐regulated gene expression and discuss the physiological significance of these findings. J. Cell. Physiol. © 2014 Wiley Periodicals, Inc.
      PubDate: 2014-10-07T09:33:13.495063-05:
      DOI: 10.1002/jcp.24838
  • Gastroprotective Effects of L‐Lysine Salification of Ketoprofen in
           Ethanol‐Injured Gastric Mucosa
    • Authors: A. Cimini; L. Brandolini, R. Gentile, L. Cristiano, P. Menghini, A. Fidoamore, A. Antonosante, E. Benedetti, A. Giordano, M. Allegretti
      Pages: n/a - n/a
      Abstract: Ketoprofen L‐lysine salt (KLS), a NSAID, is widely used for its analgesic efficacy and tolerability. L‐lysine salification was reported to increase the solubility and the gastric absorption and tolerance of Ketoprofen. Since the management of NSAIDs gastrotoxicity still represents a major limitation in prolonged therapies, mainly when gastric lesions are present, this study investigated the gastro‐protective activity of L‐lysine by using a well‐established model of gastric mucosa injury, the ethanol‐gastric injury model. Several evidences show that the damaging action of ethanol could be attributed to the increase of ROS, which plays a key role in the increase of lipid peroxidation products, including malonyldialdehyde and 4‐hydroxy‐2‐nonenal. With the aim to unravel the mechanism of L‐lysine gastroprotection, cellular MDA levels and 4‐HNE protein adducts as markers of lipid peroxidation and a panel of key endogenous gastro‐protective proteins were assayed. The data obtained indicate a gastroprotective effect of L‐lysine on gastric mucosa integrity. J. Cell. Physiol. © 2014 Wiley Periodicals, Inc.
      PubDate: 2014-10-06T22:53:18.120733-05:
      DOI: 10.1002/jcp.24809
  • Amyloid Precursor Protein (APP) Affects Global Protein Synthesis in
           Dividing Human Cells
    • Authors: Anna Sobol; Paola Galluzzo, Shuang Liang, Brittany Rambo, Sylvia Skucha, Megan J. Weber, Sara Alani, Maurizio Bocchetta
      Pages: n/a - n/a
      Abstract: Hypoxic non‐small cell lung cancer (NSCLC) is dependent on Notch‐1 signaling for survival. Targeting Notch‐1 by means of γ‐secretase inhibitors (GSI) proved effective in killing hypoxic NSCLC. Post‐mortem analysis of GSI‐treated, NSCLC‐burdened mice suggested enhanced phosphorylation of 4E‐BP1 at threonines 37/46 in hypoxic tumor tissues. In vitro dissection of this phenomenon revealed that Amyloid Precursor Protein (APP) inhibition was responsible for a non‐canonical 4E‐BP1 phosphorylation pattern rearrangement‐a process, in part, mediated by APP regulation of the pseudophosphatase Styx. Upon APP depletion we observed modifications of eIF‐4F composition indicating increased recruitment of eIF‐4A to the mRNA cap. This phenomenon was supported by the observation that cells with depleted APP were partially resistant to silvestrol, an antibiotic that interferes with eIF‐4A assembly into eIF‐4F complexes. APP downregulation in dividing human cells increased the rate of global protein synthesis, both cap‐ and IRES‐dependent. Such an increase seemed independent of mTOR inhibition. After administration of Torin‐1, APP downregulation and Mechanistic Target of Rapamycin Complex 1 (mTORC‐1) inhibition affected 4E‐BP1 phosphorylation and global protein synthesis in opposite fashions. Additional investigations indicated that APP operates independently of mTORC‐1. Key phenomena described in this study were reversed by overexpression of the APP C‐terminal domain. The presented data suggest that APP may be a novel regulator of protein synthesis in dividing human cells, both cancerous and primary. Furthermore, APP appears to affect translation initiation using mechanisms seemingly dissimilar to mTORC‐1 regulation of cap‐dependent protein synthesis. J. Cell. Physiol. © 2014 Wiley Periodicals, Inc.
      PubDate: 2014-10-06T01:57:08.048627-05:
      DOI: 10.1002/jcp.24835
  • Emerging Concept in DNA Methylation: Role of Transcription Factors in
           Shaping DNA Methylation Patterns
    • Authors: Claire Marchal; Benoit Miotto
      Pages: n/a - n/a
      Abstract: DNA methylation in mammals is a key epigenetic modification essential to normal genome regulation and development. DNA methylation patterns are established during early embryonic development, and subsequently maintained during cell divisions. Yet, discrete site‐specific de novo DNA methylation or DNA demethylation events play a fundamental role in a number of physiological and pathological contexts, leading to critical changes in the transcriptional status of genes such as differentiation, tumor suppressor or imprinted genes. How the DNA methylation machinery targets specific regions of the genome during early embryogenesis and in adult tissues remains poorly understood. Here, we report advances being made in the field with a particular emphasis on the implication of transcription factors in establishing and in editing DNA methylation profiles. J. Cell. Physiol. © 2014 Wiley Periodicals, Inc.
      PubDate: 2014-10-06T01:56:48.790137-05:
      DOI: 10.1002/jcp.24836
  • Editor's Choice
    • PubDate: 2014-09-29T19:20:45.629226-05:
      DOI: 10.1002/jcp.24829
  • Highlights: Volume 230, Number 1
    • PubDate: 2014-09-29T19:20:45.545979-05:
      DOI: 10.1002/jcp.24740
  • Journal of Cellular Physiology: Volume 230, Number 1, January 2015
    • Abstract: Cover: Fluorescence images of H9 human embryonic stem cells (hESCs) cultured in pluripotency media for 0 days; top, DAPI nuclear staining; middle/bottom, immunofluorescence using with primary antibodies for cytoplasmic and nuclear survivin, respectively, detected with Alexa‐fluor conjugated secondary antibodies. See article by Kapinas et al. on pages 63–70.
      PubDate: 2014-09-29T19:20:44.997738-05:
      DOI: 10.1002/jcp.24738
  • Table of Contents: Volume 230, Number 1
    • PubDate: 2014-09-29T19:20:37.146126-05:
      DOI: 10.1002/jcp.24739
  • Could lncRNAs be the Missing Links in Control of Mesenchymal Stem Cell
    • Authors: Coralee E. Tye; Jonathan A.R. Gordon, Lori A. Martin‐Buley, Janet L. Stein, Jane B. Lian, Gary S. Stein
      Abstract: Long suspected, recently recognized, and increasingly studied, non protein‐coding RNAs (ncRNAs) are emerging as key drivers of biological control and pathology. Since their discovery in 1993, microRNAs (miRNAs) have been the subject of intense research focus and investigations have revealed striking findings, establishing that these molecules can exert a shocking level of biological control in numerous tissues. More recently, long ncRNAs (lncRNAs), the lesser‐studied siblings of miRNA, have been suggested to have a similar robust role in developmental and adult tissue regulation. Mesenchymal stem cells (MSCs) are an important source of multipotent cells for normal and therapeutic tissue repair. Much is known about the critical role of miRNAs in biogenesis and differentiation of MSCs however; recent studies have suggested lncRNAs may play an equally important role in the regulation of these cells. Here we highlight the role of lncRNAs in the regulation of mesenchymal stem cell lineages including adipocytes, chondrocytes, myoblasts, and osteoblasts. In addition, the potential for these noncoding RNAs to be used as biomarkers for disease or therapeutic targets is also discussed. J. Cell. Physiol. © 2014 Wiley Periodicals, Inc.
      PubDate: 2014-09-25T00:21:51.876637-05:
      DOI: 10.1002/jcp.24834
  • LIGHT is a crucial mediator of airway remodeling
    • Authors: Jen‐Yu Hung; Shyh‐Ren Chiang, Ming‐Ju Tsai, Ying‐Ming Tsai, Inn‐Wen Chong, Jiunn‐Min Shieh, Ya‐Ling Hsu
      Abstract: Chronic inflammatory airway diseases like asthma and chronic obstructive pulmonary disease are major health problems globally. Airway epithelial cells play important role in airway remodeling, which is a critical process in the pathogenesis of diseases. This study aimed to demonstrate that LIGHT, an inflammatory factor secreted by T cells after allergen exposure, is responsible for promoting airway remodeling. LIGHT increased primary human bronchial epithelial cells (HBECs) undergoing epithelial‐mesenchymal transition (EMT) and expressing MMP‐9. The induction of EMT was associated with increased NF‐κB activation and p300/NF‐κB association. The interaction of NF‐κB with p300 facilitated NF‐κB acetylation, which in turn, was bound to the promoter of ZEB1, resulting in E‐cadherin downregulation. LIGHT also stimulated HBECs to produce numerous cytokines/chemokines that could worsen airway inflammation. Furthermore, LIGHT enhanced HBECs to secrete activin A, which increased bronchial smooth muscle cell (BSMC) migration. In contrast, depletion of activin A decreased such migration. The findings suggest a new molecular determinant of LIGHT‐mediated pathogenic changes in HBECs and that the LIGHT‐related vicious cycle involving HBECs and BSMCs may be a potential target for the treatment of chronic inflammation airway diseases with airway remodeling. J. Cell. Physiol. © 2014 Wiley Periodicals, Inc.
      PubDate: 2014-09-24T00:46:50.134114-05:
      DOI: 10.1002/jcp.24832
  • Lkb1 deletion promotes ectopic lipid accumulation in muscle progenitor
           cells and mature muscles
    • Authors: Tizhong Shan; Pengpeng Zhang, Pengpeng Bi, Shihuan Kuang
      Abstract: Excessive intramyocellular triglycerides (muscle lipids) are associated with reduced contractile function, insulin resistance and Type 2 diabetes, but what governs lipid accumulation in muscle is unclear. Here we report a role of Lkb1 in regulating lipid metabolism in muscle stem cells and their descendent mature muscles. We used MyodCre and Lkb1flox/flox mice to specifically delete Lkb1 in myogenic cells including stem and differentiated cells, and examined the lipid accumulation and gene expression of myoblasts cultured from muscle stem cells (satellite cells). Genetic deletion of Lkb1 in myogenic progenitors led to elevated expression of lipogenic genes and ectopic lipid accumulation in proliferating myoblasts. Interestingly, the Lkb1‐deficient myoblasts differentiated into adipocyte‐like cells upon adipogenic induction. However, these adipocyte‐like cells maintained myogenic gene expression with reduced ability to form myotubes efficiently. Activation of AMPK by AICAR prevented ectopic lipid formation in the Lkb1 null myoblasts. Notably, Lkb1‐deficient muscles accumulated excessive lipids in vivo in response to high‐fat diet feeding. These results demonstrate that Lkb1 acts through AMPK to limit lipid deposition in muscle stem cells and their derivative mature muscles, and point to the possibility of controlling muscle lipid content using AMPK activating drugs. J. Cell. Physiol. © 2014 Wiley Periodicals, Inc.
      PubDate: 2014-09-24T00:44:13.870582-05:
      DOI: 10.1002/jcp.24831
  • UCP2‐related mitochondrial pathway participates in oroxylin
           A‐induced apoptosis in human colon cancer cells
    • Authors: Chen Qiao; Libin Wei, Qinsheng Dai, Yuxin Zhou, Qian Yin, Zhiyu Li, Yuanming Xiao, Qinglong Guo, Na Lu
      Abstract: Oroxylin A is a flavonoid extracted from the root of Scutellaria baicalensis Georgi. Our previous research demonstrated that oroxylin A have various anti‐tumor effects including apoptosis, cell cycle arrest, drug‐resistant reversion and others. This paper explores the mechanism how oroxylin A induce apoptosis by regulating uncoupling protein 2 (UCP2) in human colon cancer cells. We found that the inhibition of UCP2 by UCP2 siRNA significantly increased the sensitivity of cells to drugs, reactive oxygen species (ROS) generation and the opening of mitochondrial permeability transition pore (MPTP) of CaCo‐2 cells. We also found that UCP2 inhibition could lead to ROS‐mediated MPTP activation. Furthermore, we demonstrated that oroxylin A triggered MPTP‐dependent pro‐apoptotic protein release from mitochondria to matrix and then induced apoptotic cascade by inhibiting UCP2. Intriguingly, the inhibition of UCP2 by oroxylin A was able to block Bcl‐2 translocation to the mitochondria, keeping MPTP at open‐state. In conclusion, we have demonstrate that UCP2 play a key role in mitochondrial apoptotic pathway; UCP2's inhibition by oroxylin A triggers the MPTP opening, and promotes the apoptosis in CaCo‐2 cells. J. Cell. Physiol. © 2014 Wiley Periodicals, Inc.
      PubDate: 2014-09-24T00:44:07.232297-05:
      DOI: 10.1002/jcp.24833
  • Alpha‐synuclein as a Pathological Link between Chronic Traumatic
           Brain Injury and Parkinson's disease
    • Authors: Sandra A. Acosta; Naoki Tajiri, Ike de la Pena, Marina Bastawrous, Paul R. Sanberg, Yuji Kaneko, Cesar V. Borlongan
      Abstract: The long‐term consequences of traumatic brain injury (TBI) are closely associated with the development of histopathological deficits. Notably, TBI may predispose long‐term survivors to age‐related neurodegenerative diseases, such as Parkinson's disease (PD), which is characterized by a gradual degeneration of the nigrostriatal dopaminergic neurons. However, preclinical studies on the pathophysiological changes in substantia nigra (SN) after chronic TBI are lacking. In the present in vivo study, we examined the pathological link between PD‐associated dopaminergic neuronal loss and chronic TBI. Sixty days post TBI, rats were euthanized and brain tissues harvested. Immunostaining was performed using tyrosine hydroxylase (TH), an enzyme required for the synthesis of dopamine in neurons, á‐synuclein, a presynaptic protein that plays a role in synaptic vesicle recycling, and major histocompatibility complex II (MHCII), a protein found in antigen presenting cells such as inflammatory microglia cells, all key players in PD pathology. Unbiased stereology analyses revealed significant decrease of TH‐positive expression in the surviving dopaminergic neurons of the SN pars compacta (SNpc) relative to sham control. In parallel, increased á‐synuclein accumulation was detected in the ipsilateral SN compared to the contralateral SN in TBI animals or sham control. In addition, exacerbation of MHCII+ cells was recognized in the SN and cerebral peduncle ipsilateral to injury relative to contralateral side and sham control. These results suggest á‐synuclein as a pathological link between chronic effects of TBI and PD symptoms as evidenced by significant overexpression and abnormal accumulation of á‐synuclein in inflammation‐infiltrated SN of rats exposed to chronic TBI. J. Cell. Physiol. © 2014 Wiley Periodicals, Inc.
      PubDate: 2014-09-24T00:43:55.605155-05:
      DOI: 10.1002/jcp.24830
  • Genome‐wide transcriptional and functional analysis of endoglin
           isoforms in the human promonocytic cell line U937
    • Authors: Francisco J Blanco; Luisa Ojeda‐Fernandez, Mikel Aristorena, Eunate Gallardo‐Vara, Alberto Benguria, Ana Dopazo, Carmen Langa, Luisa M Botella, Carmelo Bernabeu
      Abstract: Endoglin is an auxiliary cell surface receptor for TGF‐β family members. Two different alternatively spliced isoforms, long (L)‐endoglin and short (S)‐endoglin, have been reported. S‐endoglin and L‐endoglin proteins vary from each other in their cytoplasmic tails that contain 14 and 47 amino acids, respectively. A critical role for endoglin in vascular development has primarily been studied in endothelial cells. In addition, endoglin expression is upregulated during monocyte‐to‐macrophage differentiation, however, little is known about its role in this myeloid context. To investigate the function of endoglin in monocytes, stable transfectants expressing the two endoglin isoforms in the promonocytic human cell line U937 were generated. The differential gene expression fingerprinting of these endoglin transfectants using DNA microarrays and further bioinformatics analysis showed a clear alteration in essential biological functions, mainly those related to “Cellular Movement”, including cell adhesion and transmigration. Interestingly, these cellular functions are highly dependent on adhesion molecules, including integrins α1 (CD49a, ITGA1 gene), αL (CD11a, ITGAL gene), αM (CD11b, ITGAM gene) and β2 (CD18, ITGB2 gene) and the chemokine receptor CCR2 (CD192, CCR2 gene), which are downregulated in endoglin transfectants. Moreover, activin A (INHBA gene), a TGF‐β superfamily member involved in macrophage polarization, was distinctly affected in each endoglin transfectant, and may contribute to the regulated expression of integrins. These data were confirmed by quantitative PCR, flow cytometry and functional tests. Taken together, these results provide new insight into endoglin function in monocytes. J. Cell. Physiol. © 2014 Wiley Periodicals, Inc.
      PubDate: 2014-09-12T05:48:01.205103-05:
      DOI: 10.1002/jcp.24827
  • Novel Hedgehog Agonists Promote Osteoblast Differentiation in Mesenchymal
           Stem Cells
    • Authors: Takashi Nakamura; Masahiro Naruse, Yuta Chiba, Toshihisa Komori, Keiichi Sasaki, Masahiro Iwamoto, Satoshi Fukumoto
      Abstract: Hedgehog (Hh) family members are involved in multiple cellular processes including proliferation, migration, differentiation, and cell fate determination. Recently, the novel Hh agonists Hh‐Ag 1.3 and 1.7 were identified in a high‐throughput screening of small molecule compounds that activate the expression of Gli1, a target of Hh signaling. This study demonstrates that Hh‐Ag 1.3 and 1.7 strongly activate the expression of endogenous Gli1 and promote osteoblast differentiation in the mesenchymal stem cell line C3H10T1/2. Both compounds stimulated alkaline phosphatase activity in a dose‐dependent manner, and induced osteoblast marker gene expression in C3H10T1/2 cells, which indicated that they had acquired an osteoblast identity. Of the markers, the expression of osterix/Sp7, a downstream target of runt‐related transcription factor (Runx)2, was induced by Hh‐Ag 1.7, which also rescued the osteoblast differentiation defect ofRD‐127, a mesenchymal cell line from Runx2‐deficient mice. Hh‐Ags also activated canonical Wnt signaling and synergized with low doses of BMP‐2 to enhance osteoblastic potential. Thus, Hh‐Ag 1.7 could be useful for bone healing in individuals with abnormalities in osteogenesis, such as osteoporosis patients and the elderly, and can contribute to the development of novel therapeutics for the treatment of bone fractures and defects. J. Cell. Physiol. © 2014 Wiley Periodicals, Inc.
      PubDate: 2014-09-12T05:47:21.000731-05:
      DOI: 10.1002/jcp.24823
  • MicroRNA‐302a Stimulates Osteoblastic Differentiation by Repressing
           COUP‐TFII Expression
    • Authors: In‐Hong Kang; Byung‐Chul Jeong, Sung‐Woong Hur, Hyuck Choi, Seung‐Ho Choi, Je‐Hwang Ryu, Yun‐Chan Hwang, Jeong‐Tae Koh
      Abstract: Chicken ovalbumin upstream promoter transcription factor II (COUP‐TFII) is a potent transcription factor that represses osteoblast differentiation and bone formation. Previously we observed that stimuli for osteoblast differentiation, such as bone morphogenetic protein 2 (BMP2), inhibits COUP‐TFII expression. This study was undertaken to identify BMP2‐regulated and COUP‐TFII‐targeting microRNAs (miRNAs), and to explore their regulatory roles in osteoblast differentiation. Based on in silico analysis, 12 miRNAs were selected and their expression in BMP2‐treated MC3T3‐E1 cells was examined. BMP2 induced miR‐302a expression in dose‐ and time‐dependent manners with the decrease in COUP‐TFII expression. Runx2, a BMP2‐downstream transcription factor, specifically regulated miR‐302a expression and its promoter activity. A computer‐based prediction algorithm led to the identification of two miR‐302a binding sites on the 3'‐untranslational region of COUP‐TFII mRNA (S1: 620‐626 bp, S2: 1016‐1022 bp), and a luciferase assay showed that miR‐302a directly targeted S1 and S2. Transfection of miR‐302a precursor significantly enhanced expression of osteogenic marker genes with decreasing COUP‐TFII mRNA and protein level, alkaline phosphatase activity and matrix mineralization. On the other hand, inhibition of miR‐302a significantly attenuated BMP2‐induced osteoblast specific gene expression, alkaline phosphatase activity, and matrix mineralization with increasing COUP‐TFII mRNA and protein level. These results indicate that miR‐302a is induced by osteogenic stimuli and promotes osteoblast differentiation by targeting COUP‐TFII. MiR‐302a could be a positive regulator for osteoblast differentiation. J. Cell. Physiol. © 2014 Wiley Periodicals, Inc.
      PubDate: 2014-09-12T05:45:31.80351-05:0
      DOI: 10.1002/jcp.24822
  • miR‐203 is a Direct Transcriptional Target of E2F1 and Causes G1
           Arrest in Esophageal Cancer Cells
    • Authors: Kun Zhang; Limeng Dai, Bo Zhang, Xueqing Xu, Jiazhong Shi, Liyuan Fu, Xuedan Chen, Juan Li, Yun Bai
      Abstract: miR‐203 act as tumor repressor by inhibiting cell proliferation and is repressed in a variety of human tumors, although the molecular mechanisms responsible have not been elucidated. Here, we reveal that miR‐203 is regulated by E2F1, an important transcription factor that can induce cell proliferation by controlling cell cycle progression. We found that miR‐203 expression was induced by cisplatin, which also induced E2F1 protein accumulation in esophageal squamous cell carcinoma (ESCC) cell lines. miR‐203 expression was elevated upon activation of ectopic E2F1, whereas this induction was abolished when the E2F1 gene was silenced. Moreover, with luciferase reporter assays and chromatin immunoprecipitation (ChIP) assays, we demonstrated that E2F1 transactivates miR‐203 by directly binding to the miR‐203 gene promoter. In addition, we found that miR‐203 inhibited cell proliferation by inducing G1/S cell cycle arrest, but not apoptosis, in ESCC cell lines. Finally, we observed that miR‐203 negatively inhibited the expression of CDK6, subsequently decreasing E2F1 expression possibly through Rb phosphorylation. Taken together, our data show that cancer‐related miR‐203 is a novel transcriptional target of E2F1 and that it regulates cell cycle arrest by participating in a feedback loop with E2F1. J. Cell. Physiol. © 2014 Wiley Periodicals, Inc.
      PubDate: 2014-09-12T05:43:18.257391-05:
      DOI: 10.1002/jcp.24821
  • FoxK2 is required for cellular proliferation and survival
    • Authors: Lars P. van der Heide; Patrick J. E. C. Wijchers, Lars von Oerthel, J. Peter H. Burbach, Marco F. M. Hoekman, Marten P. Smidt
      Abstract: FoxK2 is a forkhead transcription factor expressed ubiquitously in the developing murine central nervous system. Here we investigated the role of FoxK2 in vitro and focused on proliferation and cellular survival. Knockdown of FoxK2 results in a decrease in BrdU incorporation and H3 phosphorylation, suggesting attenuation of proliferation. In the absence of growth factors FoxK2 knockdown results in a dramatic increase in caspase 3 activity and propidium iodide positive cells, indicative of cell death. Additionally, knockdown of FoxK2 results in an increase in the mRNA of Gadd45α, Gadd45γ, as well as an increase in the phosphorylation of the mTOR dependent kinase p70S6K. Rapamycin treatment completely blocked the increase in p70S6K and synergistically potentiated the decrease in H3 phosphorylation upon FoxK2 knockdown. To gain more insight into the pro‐apoptotic effects upon FoxK2 knockdown we screened for changes in Bcl2 genes. Upon FoxK2 knockdown both Puma and Noxa were significantly upregulated. Both genes were not inhibited by rapamycin treatment, instead rapamycin increased Noxa mRNA. FoxK2 requirement in cellular survival is further emphasized by the fact that resistance to TGFβ‐induced cell death was greatly diminished after FoxK2 knockdown. Overall our data suggest FoxK2 is required for proliferation and survival, that mTOR is part of a feedback loop partly compensating for FoxK2 loss, possibly by upregulating Gadd45s, whereas cell death upon FoxK2 loss is induced in a Bcl2 dependent manner via Puma and Noxa. J. Cell. Physiol. © 2014 Wiley Periodicals, Inc.
      PubDate: 2014-09-12T05:42:38.06738-05:0
      DOI: 10.1002/jcp.24828
  • Involvement of intracellular signalling in the IL‐1β inhibitory
           effect on fructose intestinal absorption
    • Authors: M Jesús Rodríguez‐Yoldi; Sonia Gascón, Cristina Barranquero, Alberto García‐Barrios, Jesús Osada
      Abstract: A variety of bacteria and their excreted/secreted products having direct effects on epithelial ion transport and permeability and the release of cytokines during bacterial infection may impact directly on epithelial function. Interleukin‐1β (IL‐1β) is a pleiotropic cytokine that affects the intestinal absorption of nutrients. The aim of this work was to study the intracellular signalling pathways involved in the inhibitory effect of IL‐1β on D‐fructose intestinal transport in rabbit jejunum and Caco‐2 cells. The results show that the cytokine inhibitory effect was completely reversed in presence of proteasome or PKC selective inhibitors in IL‐1β treated rabbits. In addition, the activation of PI3K abolished the IL‐1β effect. Likewise, these results were confirmed in Caco‐2 cells. In addition, p‐PI3K expression was increased by IL‐1β‐treatment whereas the expression of p‐PKCα was not significantly affected. In summary, the results suggest that IL‐1β could regulate the activation of pPKCα 73, pPI3K 55 and NF‐kB proteins. These events could exert an inhibitory effect on fructose intestinal absorption by a modification of GLUT5 insertion to brush‐border membrane and/or the functional transporter activity. This effect is independent of hormonal milieu and nervous stimuli. J. Cell. Physiol. © 2014 Wiley Periodicals, Inc.
      PubDate: 2014-09-12T05:42:20.411333-05:
      DOI: 10.1002/jcp.24820
  • The dysfunction of the trabecular meshwork during glaucoma course
    • Authors: Sergio Claudio Saccà; Alessandra Pulliero, Alberto Izzotti
      Abstract: Primary open angle glaucoma is a multi‐tissue disease that targets, in an ascending order, the trabecular meshwork, the optic nerve head, the lateral geniculate nuclei and the visual cortex. Oxidative stress and vascular damage play major roles in triggering apoptotic cell loss in these tissues. Molecular alterations occurring in the ocular anterior chamber during the early course of glaucoma trigger this cell loss. These molecular events are mainly of endogenous origin and related to the long‐term accumulation of oxidative damages arising from mitochondrial failure and endothelial dysfunction. This situation results in decreased antioxidant defences in aqueous humour and apoptosis activation in trabecular meshwork cells as triggered by severe mitochondrial damage altering tissue function and integrity. The presence of neural proteins in glaucomatous aqueous humour indicate that a molecular interconnection exists between the anterior and the posterior chamber tissues. Trabecular meshwork and lamina cribrosa share a common neuro‐ectodermal embryological, which contribute to explain the interconnection between anterior and the posterior chamber during glaucoma pathogenesis. During glaucoma, proteins deriving from the damage occurring in endothelial trabecular meshwork cells are released into aqueous humour. Accordingly, aqueous humour composition is characterised in glaucomatous patients by the presence of proteins deriving from apoptosis activation, mitochondrial damage, loss of intercellular connections, antioxidant decrease. Many questions remain unanswered, but molecular events illuminate TM damage and indicate that trabecular cells protection plays a role in the treatment and prevention of glaucoma. J. Cell. Physiol. © 2014 Wiley Periodicals, Inc.
      PubDate: 2014-09-12T05:41:42.077463-05:
      DOI: 10.1002/jcp.24826
  • Flt3 ligand induces monocyte proliferation and enhances the function of
           monocyte‐derived dendritic cells in vitro
    • Authors: Sung‐Whan Kim; Seong‐Mi Choi, Yee Shin Choo, Il‐Kwon Kim, Byeung‐Wook Song, Han‐Soo Kim
      Abstract: Flt3 ligand (FL), a potent hematopoietic cytokine, plays an important role in development and activation of dendritic cells (DCs) and natural killer cells (NK). Although some post‐receptor signaling events of FL have been characterized, the role of FL on Flt3 expressing human peripheral blood monocyte is unclear. In the current study, we examined the role of FL on cell survival and growth of peripheral blood monocytes and function of monocyte‐derived DCs. FL promoted monocyte proliferation in a dose‐dependent manner and prevented spontaneous apoptosis. FL induced ERK phosphorylation and a specific ERK inhibitor completely abrogated FL‐mediated cellular growth, while p38 MAPK, JNK and AKT were relatively unaffected. Addition of FL to GM‐CSF and IL‐4 during DCs generation from monocytes increased the yield of DCs through induction of cell proliferation. DCs generated in the presence of FL expressed more costimulatory molecules on their surfaces and stimulated allogeneic T cell proliferation in MLR to a higher magnitude. Furthermore, FL partially antagonized IL‐10‐mediated inhibition on DCs function. Further characterization of FL actions may provide new and important information for immunotherapeutic approaches utilizing DCs. J. Cell. Physiol. © 2014 Wiley Periodicals, Inc.
      PubDate: 2014-09-12T05:41:13.443492-05:
      DOI: 10.1002/jcp.24824
  • Justicidin A‐Induced Autophagy Flux Enhances Apoptosis of Human
           Colorectal Cancer Cells via Class III PI3K and Atg5 Pathway
    • Authors: Shen‐Jeu Won; Cheng‐Hsin Yen, Hsiao‐Sheng Liu, Shan‐Ying Wu, Sheng‐Hui Lan, Ya‐Fen Jiang‐Shieh, Chun‐Nan Lin, Chun‐Li Su
      Abstract: Our previous reports showed that justicidin A (JA), a novel and pure arylnaphthalide lignan isolated from Justicia procumbens, induces apoptosis of human colorectal cancer cells and hepatocellular carcinoma cells, leading to the suppression of both tumor cell growth in NOD‐SCID mice. Here, we reveal that JA induces autophagy in human colorectal cancer HT‐29 cells by conversion of autophagic marker LC3‐I to LC3‐II. Furthermore, LC3 puncta and autophagic vesicle formation, and SQSTM1/p62 suppression were observed. Administration of autophagy inhibitor (bafilomycin A1 and chloroquine) and transfection of a tandem fluorescent‐tagged LC3 (mRFP‐GFP) reporter plasmid (ptfLC3) demonstrated that JA induces autophagy flux in HT‐29 cells. Expression of LC3, SQSTM1, Beclin 1, and nuclear DNA double‐strand breaks (representing apoptosis) were also detected in the tumor tissue of HT‐29 cells transplanted into NOD‐SCID mice orally administrated with JA. In addition, the expression of autophagy signaling pathway‐related molecules p‐PDK1, p‐mTOR, p‐p70S6k/p‐RPS6KB2 was decreased, whereas that of class III PI3K, Beclin 1, Atg5‐Atg12, and mitochondrial BNIP3 was increased in response to JA. Pre‐treatment of the cells with class III PI3K inhibitor 3‐methyladenine or Atg5 shRNA attenuated JA‐induced LC3‐II expression and LC3 puncta formation, indicating the involvement of class III PI3K and Atg5. A novel mechanism was demonstrated in the anticancer compound JA; pre‐treatment with 3‐methyladenine or Atg5 shRNA blocked JA‐induced suppression in cell growth and colony formation, respectively, via inhibition of apoptosis. In contrast, administration of apoptosis inhibitor Z‐VAD did not affect JA‐induced autophagy. Our data suggest the chemotherapeutic potential of JA for treatment of human colorectal cancer. J. Cell. Physiol. © 2014 Wiley Periodicals, Inc.
      PubDate: 2014-09-12T05:28:03.183994-05:
      DOI: 10.1002/jcp.24825
  • p27 and Leukemia: Cell cycle and beyond
    • Authors: Anita Roy; Subrata Banerjee
      Abstract: Cell division is the foundation to development and the regulation of cell cycle progression is therefore of paramount importance to the living organisms. Primary control of cell cycle is achieved by an array of cyclins and cyclin dependent kinases (CDKs). The functions of these cyclin‐CDK complexes are again regulated by a host of cyclin dependent kinase inhibitors (CDKI). Till date CDKIs are broadly classified into two groups – INK4 family (p15, p16, p18 and p19) and the cip/kip family (p21, p27 and p57). Collectively these CDKIs regulate the progression from G1 to S phase of cell cycle. This review summarizes the functions of p27 while highlighting its emerging roles in leukemia. J. Cell. Physiol. © 2014 Wiley Periodicals, Inc.
      PubDate: 2014-09-10T00:28:21.380767-05:
      DOI: 10.1002/jcp.24819
  • Hesperetin induces apoptosis in breast carcinoma by triggering
           accumulation of ROS and activation of ASK1/JNK pathway
    • Authors: Shreyasi Palit; Susanta Kar, Gunjan Sharma, Pijush K. Das
      Abstract: Hesperetin, a flavanone glycoside predominantly found in citrus fruits, exhibits a wide array of biological properties. In the present study hesperetin exhibited a significant cytotoxic effect in human breast carcinoma MCF‐7 cells in a concentration‐ and time‐dependent manner without affecting normal (HMEC) as well as immortalized normal mammary epithelial cells (MCF‐10A). The cytotoxic effect of hesperetin was due to the induction of apoptosis as evident from the phosphatidyl‐serine externalization, DNA fragmentation, caspase‐7 activation and PARP cleavage. Apoptosis was associated with caspase‐9 activation, mitochondrial membrane potential loss, release of cytochrome c and increase in Bax:Bcl‐2 ratio. Pre‐treatment with caspase‐9 specific inhibitor (Z‐LEHD‐fmk) markedly attenuated apoptosis suggesting an involvement of intrinsic mitochondrial apoptotic cascade. Further, DCFDA flow‐cytometric analysis revealed triggering of ROS in a time‐dependent manner. Pre‐treatment with ROS scavenger N‐acetylcysteine (NAC) and glutathione markedly abrogated hesperetin‐mediated apoptosis whereas carbonyl cyanide m‐chlorophenylhydrazone (CCCP) pretreatment along with DHR123‐based flow‐cytometry indicated the generation of cytosolic ROS. Profiling of MAPKs revealed activation of JNK upon hesperetin treatment which was abrogated upon NAC pretreatment. Additionally, inhibition of JNK by SP600125 significantly reversed hesperetinmediated apoptosis. The activation of JNK was associated with the activation of ASK1. Silencing of ASK1 resulted in significant attenuation of JNK activation as well as reversed the hesperetin‐mediated apoptosis suggesting that hesperetin‐mediated apoptosis of MCF‐7 cells involves accumulation of ROS and activation of ASK1/JNK pathway. In addition, hesperetin also induced apoptosis in triple negative breast cancer MDA‐MB‐231 cells via intrinsic pathway via activation of caspase ‐9 and ‐3 and increase in Bax:Bcl‐2 ratio. J. Cell. Physiol. © 2014 Wiley Periodicals, Inc.
      PubDate: 2014-09-10T00:16:32.838662-05:
      DOI: 10.1002/jcp.24818
  • The GPER agonist G‐1 induces mitotic arrest and apoptosis in human
           vascular smooth muscle cells independent of GPER
    • Authors: Yu Gui; Zhan Shi, Zeng Yong Wang, Jing‐Jing Li, Can Xu, Rui Juan Tian, Xin Xing Song, Michael P. Walsh, Dong Li, Jie Gao, Xi‐Long Zheng
      Abstract: The G protein‐coupled estrogen receptor (GPER) has been implicated in the regulation of smooth muscle cell (SMC) proliferation. The GPER selective agonist G‐1 has been a useful tool for exploring the biological roles of GPER in a variety of experimental settings, including SMC proliferation. The present study, originally designed to investigate cellular and signaling mechanisms underlying the regulatory role of GPER in vascular SMC proliferation using G‐1, unexpectedly revealed off‐target effects of G‐1. G‐1 (1‐10 μM) inhibited bromodeoxyuridine (BrdU) incorporation of human SMCs and caused G2/M cell accumulation. G‐1 treatment also increased mitotic index concurrent with a decrease in phosphorylation of Cdk1 (Tyr 15) and an increase in phosphorylation of the mitotic checkpoint protein BuBR1. Furthermore, G‐1 caused microtubule disruption, mitotic spindle damage, and tubulin depolymerization. G‐1 induced cell apoptosis as indicated by the appearance of TUNEL‐positive and annexin V‐positive cells with enhanced cleavage of caspases 3 and 9. However, neither the GPER antagonist G‐15 nor the MAPK kinase inhibitor PD98059 prevented these G‐1 effects. Down‐regulation of GPER or p44/42 MAPK with siRNA transfection also did not affect the G‐1‐induced apoptosis. We conclude that G‐1 inhibits proliferation of SMCs through mechanisms involving mitotic arrest and apoptosis, independent of GPER and the MAPK pathway. J. Cell. Physiol. © 2014 Wiley Periodicals, Inc.
      PubDate: 2014-09-10T00:12:53.242319-05:
      DOI: 10.1002/jcp.24817
  • IL‐1β Promotes Malignant Transformation and Tumor
           Aggressiveness in Oral Cancer
    • Authors: Chia‐Huei Lee; Jeffrey Shu‐Ming Chang, Shih‐Han Syu, Thian‐Sze Wong, Jimmy Yu‐Wai Chan, Ya‐Chu Tang, Zhi‐Ping Yang, Wen‐Chan Yang, Chiung‐Tong Chen, Shao‐Chun Lu, Pei‐Hua Tang, Tzu‐Ching Yang, Pei‐Yi Chu, Jenn‐Ren Hsiao, Ko‐Jiunn Liu
      Abstract: Chronic inflammation, coupled with alcohol, betel quid, and cigarette consumption, is associated with oral squamous cell carcinoma (OSCC). Interleukin‐1 beta (IL‐1β) is a critical mediator of chronic inflammation and implicated in many cancers. In this study, we showed that increased pro‐IL‐1β expression was associated with the severity of oral malignant transformation in a mouse OSCC model induced by 4‐Nitroquinolin‐1‐oxide (4‐NQO) and arecoline, two carcinogens related to tobacco and betel quid, respectively. Using microarray and quantitative PCR assay, we showed that pro‐IL‐1β was upregulated in human OSCC tumors associated with tobacco and betel quid consumption. In a human OSCC cell line TW2.6, we demonstrated nicotine‐derived nitrosamine ketone (NNK) and arecoline stimulated IL‐1β secretion in an inflammasome‐dependent manner. IL‐1β treatment significantly increased the proliferation and dysregulated the Akt signaling pathways of dysplastic oral keratinocytes (DOKs). Using cytokine antibodies and inflammation cytometric bead arrays, we found that DOK and OSCC cells secreted high levels of IL‐6, IL‐8, and growth‐regulated oncogene‐α following IL‐1β stimulation. The conditioned medium of IL‐1β‐treated OSCC cells exerted significant proangiogenic effects. Crucially, IL‐1β increased the invasiveness of OSCC cells through the epithelial‐mesenchymal transition (EMT), characterized by downregulation of E‐cadherin, upregulation of Snail, Slug, and Vimentin, and alterations in morphology. These findings provide novel insights into the mechanism underlying OSCC tumorigenesis. Our study suggested that IL‐1β□ can be induced by tobacco and betel quid‐related carcinogens, and participate in the early and late stages of oral carcinogenesis by increasing the proliferation of dysplasia oral cells, stimulating oncogenic cytokines, and promoting aggressiveness of OSCC. J. Cell. Physiol. © 2014 Wiley Periodicals, Inc.
      PubDate: 2014-09-10T00:11:17.930538-05:
      DOI: 10.1002/jcp.24816
  • Novel Insights into TRPM7 Function in Fibrotic Diseases: A Potential
           Therapeutic Target
    • Authors: Tao Xu; Bao‐ming Wu, Xiao‐ming Meng, Cheng Huang, Ming‐ming Ni, Jun Li
      Abstract: Transient receptor potential (TRP) channels are cellular sensors for a wide spectrum of physical and chemical stimuli. Activation of TRP channels changes the membrane potential, translocates important signaling ions crossing the cell membrane, alters enzymatic activity, and initiates endocytosis/exocytosis. Fibrosis is the leading cause of organ dysfunction in diseases, which is characterized by an imbalance in the turnover of extracellular matrix components. Accumulating evidence has demonstrated that TRPM7, a member of TRP channels superfamily, participates in the development and pathogenesis of fibrotic diseases, such as hepatic, pulmonary and cardiac fibrosis. In this review, we discuss the comprehensive role of TRPM7 in modulating profibrotic response and its potential as therapeutic target for fibrotic diseases. J. Cell. Physiol. © 2014 Wiley Periodicals, Inc.
      PubDate: 2014-09-09T19:18:08.739893-05:
      DOI: 10.1002/jcp.24801
  • Epithelial‐to‐Mesenchymal Transition Induced by
           TGF‐β1 Is Mediated by AP1‐Dependent EpCAM Expression in
           MCF‐7 Cells
    • Authors: Jiujiao Gao; Qiu Yan, Jiao Wang, Shuai Liu, Xuesong Yang
      Abstract: The epithelial‐to‐mesenchymal transition (EMT), a process involving the breakdown of cell‐cell junctions and loss of epithelial polarity, is closely related to cancer metastasis and invasion. The epithelial cell adhesion molecule (EpCAM) is a type I transmembrane protein expressed in the majority of normal epithelial tissues and overexpressed in the majority of human epithelial cancers including breast cancer. EpCAM plays an important role in cancer progression. We showed that EpCAM participated in TGF‐β1‐induced EMT. TGF‐β1 treatment of MCF‐7 breast cancer cells was shown to induce EpCAM expression, which promoted the EMT and cell migration. EpCAM overexpression further enhanced TGF‐β1‐induced EMT, and EpCAM knockdown inhibited TGF‐β1‐induced EMT. We further demonstrated that TGF‐β1 treatment induced the phosphorylation of JNK that was in turn responsible for the increased expression of Jun and Fos. This result suggests an important role of the JNK to AP‐1 signaling to EpCAM downstream of TGF‐β1 for the induction of EMT in the breast cancer cells. Collectively, our study highlights a novel function for EpCAM in TGF‐β1‐induced EMT process and suggests that targeting of EpCAM may be an attractive strategy to treat breast cancer. This study implicates the potential value of EpCAM as a molecular marker for breast cancer. J. Cell. Physiol. © 2014 Wiley Periodicals, Inc.
      PubDate: 2014-09-09T19:17:54.701705-05:
      DOI: 10.1002/jcp.24802
  • WW Domain of BAG3 is Required for the Induction of Autophagy in Glioma
    • Authors: Nana Merabova; Ilker Kudret Sariyer, A Sami Saribas, Tijana Knezevic, Jennifer Gordon, M. Caterina Turco, Alessandra Rosati, Michael Weaver, Jacques Landry, Kamel Khalili
      Abstract: Autophagy is an evolutionarily conserved, selective degradation pathway of cellular components that is important for cell homeostasis under healthy and pathologic conditions. Here we demonstrate that an increase in the level of BAG3 results in stimulation of autophagy in glioblastoma cells. BAG3 is a member of a co‐chaperone family of proteins that associates with Hsp70 through a conserved BAG domain positioned near the C‐terminus of the protein. Expression of BAG3 is induced by a variety of environmental changes that cause stress to cells. Our results show that BAG3 overexpression induces autophagy in glioma cells. Interestingly, inhibition of the proteasome caused an increase in BAG3 levels and induced autophagy. Further analysis using specific siRNA against BAG3 suggests that autophagic activation due to proteosomal inhibition is mediated by BAG3. Analyses of BAG3 domain mutants suggest that the WW domain of BAG3 is crucial for the induction of autophagy. BAG3 overexpression also increased the interaction between Bcl2 and Beclin‐1, instead of disrupting them, suggesting that BAG3 induced autophagy is Beclin‐1 independent. These observations reveal a novel role for the WW domain of BAG3 in the regulation of autophagy. J. Cell. Physiol. © 2014 Wiley Periodicals, Inc.
      PubDate: 2014-09-09T17:57:46.553115-05:
      DOI: 10.1002/jcp.24811
  • A Group of Novel HIF‐1α Inhibitors, Glyceollins, Blocks
           HIF‐1α Synthesis and Decreases Its Stability via Inhibition of
           the PI3K/AKT/mTOR Pathway and Hsp90 Binding
    • Authors: Sun Hee Lee; Jun‐Goo Jee, Kwanghyun Ryu, You Mie Lee
      Abstract: Glyceollins, a group of phytoalexins isolated from soybean, are known to exhibit anticancer, antiestrogenic, and antiangiogenic activities. However, whether glyceollins regulate tumor growth through regulation of hypoxia‐inducible factor (HIF)‐1α has not been investigated. We determined whether and how glyceollins regulate the synthesis and stability of HIF‐1α. Quantitative real‐time PCR revealed that glyceollins inhibited the expression of HIF‐1‐induced genes such as vascular endothelial growth factor (VEGF) in cancer cells. Enzyme‐linked immunosorbent assay and reporter luciferase assay showed that glyceollins decreased VEGF secretion and its promoter activity, respectively. Treatment of various cancer cells with 0.5‐100 µM glyceollins under hypoxic conditions reduced the expression of HIF‐1α. Glyceollins blocked translation of HIF‐1α by inhibiting the PI3K/AKT/mTOR pathway under hypoxic conditions. Glyceollins decreased the stability of HIF‐1α after treatment with cycloheximide, a protein synthesis inhibitor, and increased the ubiquitination of HIF‐1α after treatment with MG132, a proteasome inhibitor. Glyceollins blocked the interaction of Hsp90 with HIF‐1α, as shown by immunoprecipitation assay. Chemical binding of Hsp90 with glyceollins, as confirmed by computational docking analysis, was stronger than that with geldanamycin at the HSP90 ATP‐binding pocket. We found that glyceollins decreased microvessel density, as well as expression of phosphorylated AKT/mTOR and the Hsp90 client protein CDK4, in solid tumor tissues. Glyceollins potently inhibited HIF‐1α synthesis and decreased its stability by blocking the PI3K/AKT/mTOR pathway and HSP90 binding activity, respectively. These results may provide new perspectives into potential therapeutic application of glyceollins for the prevention and treatment of hypervascularized diseases and into the mechanism of their anticancer activity. J. Cell. Physiol. © 2014 Wiley Periodicals, Inc.
      PubDate: 2014-09-09T17:51:50.62946-05:0
      DOI: 10.1002/jcp.24813
  • Gene Expression Changes in Progression of Cervical Neoplasia Revealed by
           Microarray Analysis of Cervical Neoplastic Keratinocytes
    • Authors: John Charles Rotondo; Silvia Bosi, Cristian Bassi, Manuela Ferracin, Giovanni Lanza, Roberta Gafà, Eros Magri, Rita Selvatici, Stefania Torresani, Roberto Marci, Paola Garutti, Massimo Negrini, Mauro Tognon, Fernanda Martini
      Abstract: To evaluate the gene expression changes involved in neoplastic progression of cervical intraepithelial neoplasia. Using microarray analysis, large‐scale gene expression profile was carried out on HPV16‐CIN2, HPV16‐CIN3 and normal cervical keratinocytes derived from two HPV16‐CIN2, two HPV‐CIN3 lesions and two corresponding normal cervical tissues, respectively. Differentially expressed genes were analyzed in normal cervical keratinocytes compared with HPV16‐CIN2 keratinocytes and in HPV16‐CIN2 keratinocytes compared with HPV16‐CIN3 keratinocytes; 37 candidate genes with continuously increasing or decreasing expression during CIN progression were identified. One of these genes, phosphoglycerate dehydrogenase, was chosen for further characterization. Quantitative reverse transcription‐polymerase chain reaction and immunohistochemical analysis confirmed that expression of phosphoglycerate dehydrogenase consistently increases during progression of CIN toward cancer. Gene expression changes occurring during CIN progression were investigated using microarray analysis, for the first time, in CIN2 and CIN3 keratinocytes naturally infected with HPV16. Phosphoglycerate dehydrogenase is likely to be associated with tumorigenesis and may be a potential prognostic marker for CIN progression. J. Cell. Physiol. © 2014 Wiley Periodicals, Inc.
      PubDate: 2014-09-09T17:48:46.34043-05:0
      DOI: 10.1002/jcp.24808
  • The Coiled‐Coil Domain Containing 80 (ccdc80) Gene Regulates
           gadd45β2 Expression in the Developing Somites of Zebrafish as a New
           Player of the Hedgehog Pathway
    • Authors: Isabella Della Noce; Silvia Carra, Chiara Brusegan, Rosina Critelli, Andrea Frassine, Carlo De Lorenzo, Antonio Giordano, Gianfranco Bellipanni, Erica Villa, Franco Cotelli, Anna Pistocchi, Filippo Schepis
      Abstract: The Coiled‐Coil Domain Containing 80 (CCDC80) gene has been identified as strongly induced in rat thyroid PC CL3 cells immortalized by the adenoviral E1A gene. In human, CCDC80 is a potential oncosoppressor due to its down‐regulation in several tumor cell lines and tissues and it is expressed in almost all tissues. CCDC80 has homologous in mouse, chicken and zebrafish. We cloned the zebrafish ccdc80 and analyzed its expression and function during embryonic development. The in silico translated zebrafish protein shares high similarity with its mammalian homologous, with nuclear localization signals and a signal peptide. Gene expression analysis demonstrates that zebrafish ccdc80 is maternally and zygotically expressed throughout the development. In particular, ccdc80 is strongly expressed in the notochord and it is under the regulation of the Hedgehog pathway. In this work we investigated the functional effects of ccdc80‐loss‐of‐function during embryonic development and verified its interaction with gadd45β2 in somitogenesis. J. Cell. Physiol. © 2014 Wiley Periodicals, Inc.
      PubDate: 2014-09-09T17:47:10.174514-05:
      DOI: 10.1002/jcp.24810
  • Vitamin D Attenuates Cytokine‐Induced Remodeling in Human Fetal
           Airway Smooth Muscle Cells
    • Authors: Rodney D. Britt; Arij Faksh, Elizabeth R. Vogel, Michael A. Thompson, Vivian Chu, Hitesh C. Pandya, Yassine Amrani, Richard J. Martin, Christina M. Pabelick, Y.S. Prakash
      Abstract: Asthma in the pediatric population remains a significant contributor to morbidity and increasing healthcare costs. Vitamin D3 insufficiency and deficiency have been associated with development of asthma. Recent studies in models of adult airway diseases suggest that the bioactive Vitamin D3 metabolite, calcitriol (1,25‐dihydroxyvitamin D3; 1,25(OH)2D3), modulates responses to inflammation; however this concept has not been explored in developing airways in the context of pediatric asthma. We used human fetal airway smooth muscle (ASM) cells as a model of the early postnatal airway to explore how calcitriol modulates remodeling induced by pro‐inflammatory cytokines. Cells were pre‐treated with calcitriol and then exposed to TNFα or TGFβ for up to 72 h. Matrix metalloproteinase (MMP) activity, production of extracellular matrix (ECM), and cell proliferation were assessed. Calcitriol attenuated TNFα enhancement of MMP‐9 expression and activity. Additionally, calcitriol attenuated TNFα and TGFβ‐induced collagen III expression and deposition, and separately, inhibited proliferation of fetal ASM cells induced by either inflammatory mediator. Analysis of signaling pathways suggested that calcitriol effects in fetal ASM involve ERK signaling, but not other major inflammatory pathways. Overall, our data demonstrate that calcitriol can blunt multiple effects of TNFα and TGFβ in developing airway, and point to a potentially novel approach to alleviating structural changes in inflammatory airway diseases of childhood. J. Cell. Physiol. © 2014 Wiley Periodicals, Inc.
      PubDate: 2014-09-09T17:41:49.922249-05:
      DOI: 10.1002/jcp.24814
  • Isoform‐Specific SCFFbw7 Ubiquitination Mediates Differential
           Regulation of PGC‐1α
    • Authors: Julie S. Trausch‐Azar; Mona Abed, Amir Orian, Alan L. Schwartz
      Abstract: The E3 ubiquitin ligase and tumor suppressor SCFFbw7 exists as three isoforms that govern the degradation of a host of critical cell regulators, including c‐Myc, cyclin E, and PGC‐1α. Peroxisome proliferator activated receptor‐gamma coactivator 1α (PGC‐1α) is a transcriptional coactivator with broad effects on cellular energy metabolism. Cellular PGC‐1α levels are tightly controlled in a dynamic state by the balance of synthesis and rapid degradation via the ubiquitin‐proteasome system. Yet, isoform‐specific functions of SCFFbw7 are yet to be determined. Here, we show that the E3 ubiquitin ligase, SCFFbw7, regulates cellular PGC‐1α levels via two independent, isoform specific, mechanisms. The cytoplasmic isoform (SCFFbw7β) reduces cellular PGC‐1α levels via accelerated ubiquitin‐proteasome degradation. In contrast, the nuclear isoform (SCFFbw7α) increases cellular PGC‐1α levels and protein stability via inhibition of ubiquitin‐proteasomal degradation. When nuclear Fbw7α proteins are redirected to the cytoplasm, cellular PGC‐1α protein levels are reduced through accelerated ubiquitin‐proteasomal degradation. We find that SCFFbw7β catalyzes high molecular weight PGC‐1α‐ubiquitin conjugation, whereas SCFFbw7α produces low molecular weight PGC‐1α‐ubiquitin conjugates that are not effective degradation signals. Thus, selective ubiquitination by specific Fbw7 isoforms represents a novel mechanism that tightly regulates cellular PGC‐1α levels. Fbw7 isoforms mediate degradation of a host of regulatory proteins. The E3 ubiquitin ligase, Fbw7, mediates PGC‐1α levels via selective isoform‐specific ubiquitination. Fbw7β reduces cellular PGC‐1α via ubiquitin‐mediated degradation, whereas Fbw7α increases cellular PGC‐1α via ubiquitin‐mediated stabilization. J. Cell. Physiol. © 2014 Wiley Periodicals, Inc.
      PubDate: 2014-09-09T17:40:19.811917-05:
      DOI: 10.1002/jcp.24812
  • pRb2/p130 is Delocalized in Cytoplasm in Diffuse Gastric Cancer: A
           Preliminary Study
    • Authors: Letizia Cito; Paola Indovina, Iris Maria Forte, Domenico Di Marzo, Pasquale Somma, Daniela Barone, Antonella Penon, Danila Penon, Elisa Ceccherini, Pietro Micheli, Luca Saragoni, Marina Di Domenico, Antonia Feola, Franco Roviello, Eliseo Mattioli, Giovan Giacomo Giordano, Antonio Giordano
      Abstract: pRb2/p130 phosphorylation occurs at G0/G1 transition, when cells exit from quiescence. This phenomenon leads to E2F4 and E2F5 transcription factors releasing, allowing them to activate their target genes. Hence, pRb2/p130 represents one of the most important cell cycle controller because of transcription factors binding/releasing capability. It was previously shown that the loss of expression, or function, of pRb2/p130 is involved in different neoplasms development, due to related loss of control of E2F4 and E2F5 transcription activity. Here we analyzed gastric cancer tissue samples of diffuse histotype, comparing them with the normal counterpart. Yet, we did not find any change of expression levels. Rather, we found a cytoplasmic localization of pRb2/p130 in cancer tissue samples whereas, in normal counterparts, we found the expected nuclear localization. This occurrence lead to hypothesize an almost completely uncontrolled transcriptional activity of E2F4 and E2F5 in this kind of neoplasm, actually contributing to maintenance of transformed status. Although in limited number of cases, our data support the necessity of further investigations to verify the possibility of using pRb2/p130 as diagnostic marker of diffuse gastric cancer. J. Cell. Physiol. © 2014 Wiley Periodicals, Inc.
      PubDate: 2014-09-09T17:40:00.89997-05:0
      DOI: 10.1002/jcp.24805
  • GATA‐1 Deficiency Rescues Trabecular but not Cortical Bone in OPG
           Deficient Mice
    • Authors: Tomas E. Meijome; R. Adam Hooker, Ying‐Hua Cheng, Whitney Walker, Mark C. Horowitz, Robyn K. Fuchs, Melissa A. Kacena
      Abstract: GATA‐1low/low mice have an increase in megakaryocytes (MKs) and trabecular bone. The latter is thought to result from MKs directly stimulating osteoblastic bone formation while simultaneously inhibiting osteoclastogenesis. Osteoprotegerin (OPG) is known to inhibit osteoclastogenesis and OPG‐/‐ mice have reduced trabecular and cortical bone due to increased osteoclastogenesis. Interestingly, GATA‐1low/low mice have increased OPG levels. Here we sought to determine whether GATA‐1 knockdown in OPG‐/‐ mice could rescue the observed osteoporotic bone phenotype. GATA‐1low/low mice were bred with OPG‐/‐ mice and bone phenotype assessed. GATA‐1low/low X OPG‐/‐ mice have increased cortical bone porosity, similar to OPG‐/‐ mice. Both OPG‐/‐ and GATA‐1low/low X OPG‐/‐ mice, were found to have increased osteoclasts localized to cortical bone, possibly producing the observed elevated porosity. Biomechanical assessment indicates that OPG‐/‐ and GATA‐1low/low X OPG‐/‐ femurs are weaker and less stiff than C57BL/6 or GATA‐1low/low femurs. Notably, GATA‐1low/low X OPG‐/‐ mice had trabecular bone parameters that were not different from C57BL/6 values, suggesting that GATA‐1 deficiency can partially rescue the trabecular bone loss observed with OPG deficiency. The fact that GATA‐1 deficiency appears to be able to partially rescue the trabecular, but not the cortical bone phenotype suggests that MKs can locally enhance trabecular bone volume, but that MK secreted factors cannot access cortical bone sufficiently to inhibit osteoclastogenesis or that OPG itself is required to inhibit osteoclastogenesis in cortical bone. J. Cell. Physiol. © 2014 Wiley Periodicals, Inc.
      PubDate: 2014-09-09T17:37:45.37493-05:0
      DOI: 10.1002/jcp.24803
  • Review: Cell Dynamics in Malignant Pleural Effusions
    • Authors: Enrico Giarnieri; Gianfranco Bellipanni, Marcella Macaluso, Rita Mancini, Adam Carl Holstein, Carla Milanese, Maria Rosaria Giovagnoli, Antonio Giordano, Giuseppe Russo
      Abstract: Malignant pleural effusions (MPEs) are a common manifestation found in patients with lung cancer. After cytological and histological confirmation of malignancy, talc pleurodesis still remains the treatment of choice in patients with MPEs resistant to chemotherapy. Despite this, primary challenges include reduced quality of life and life expectancy in general. Therefore, a better understanding of the cell biology of MPEs, along with improvements in treatment is greatly needed. It has recently been demonstrated that malignant pleural effusions (MPEs) may represent an excellent source for identification of molecular mechanisms within the tumor and its environment. The present review summarizes the current understanding of MPEs cells and tumor microenvironment, and particularly focuses on dissecting the cross‐talk between MPEs and epithelial to mesenchymal transition (EMT), inflammation and cancer stem cells. J. Cell. Physiol. © 2014 Wiley Periodicals, Inc.
      PubDate: 2014-09-09T17:31:30.748438-05:
      DOI: 10.1002/jcp.24806
  • Role of Acinus in Regulating Retinoic Acid‐responsive Gene
           Pre‐mRNA Splicing
    • Authors: Fang Wang; Kenneth J. Soprano, Dianne Robert Soprano
      Abstract: Acinus‐S' is a co‐repressor for retinoic acid receptor (RAR)‐dependent gene transcription and has been suggested to be involved in RNA processing. In this study the role of Acinus isoforms in regulating pre‐mRNA splicing was explored using in vivo splicing assays. Both Acinus‐L and Acinus‐S', with the activity of Acinus‐L higher than that of Acinus‐S', increase the splicing of a retinoic acid (RA)‐responsive minigene containing a weak 5' splice site but not a RA‐responsive minigene containing a strong 5' splice site. RA treatment further enhances the splicing of the weak 5' splice site by Acinus in a dose‐ and time‐dependent manner, suggesting a RA‐dependent activity in addition to a RA‐independent activity of Acinus. The RA‐independent effect of Acinus occurs to varying degrees using minigene constructs containing several different promoters while the RA‐dependent splicing activity of Acinus is specific for transcripts derived from the minigene driven by a RA response element (RARE)‐containing promoter. This suggests that the ligand‐dependent splicing activity of Acinus is related to the RA‐activated RAR bound to the RARE. The RRM domain is necessary for the RA‐dependent splicing activity of Acinus and the RA‐independent splicing activity of Acinus is repressed by RNPS1. Importantly, measurement of the splicing of endogenous human RARβ and Bcl‐x in vivo demonstrates that Acinus stimulates the use of the weaker alternative 5' splice site of these two genes in a RA‐dependent manner for RARβ and a RA‐independent manner for Bcl‐x. Taken together, these studies demonstrate that Acinus functions in both RAR‐dependent splicing and RAR‐dependent transcription. J. Cell. Physiol. © 2014 Wiley Periodicals, Inc.
      PubDate: 2014-09-09T17:31:18.818058-05:
      DOI: 10.1002/jcp.24804
  • Palmitate‐stimulated monocytes induce adhesion molecule expression
           in endothelial cells via IL‐1 signaling pathway
    • Authors: Yosuke Shikama; Nanako Aki, Akiko Hata, Miho Nishimura, Seiichi Oyadomari, Makoto Funaki
      Abstract: Increased intake of saturated fatty acids (SFAs), such as palmitate (Pal), is linked to a higher risk of type 2 diabetes and cardiovascular disease. Although recent studies have investigated the direct effects of SFAs on inflammatory responses in vascular endothelial cells, it remains unknown whether SFAs also induce these responses mediated by circulating cells. In this study, especially focused on adhesion molecules and monocytes, we investigated the indirect effects of Pal on expression and release of ICAM‐1 and E‐selectin in vascular endothelial cells. Phorbol 12‐myristate 13‐acetate (PMA)‐treated THP‐1 (pTHP‐1) cells and human monocytes were stimulated with various free fatty acids (FFAs). SFAs, but not unsaturated fatty acids (UFAs), increased interleukin (IL)‐1β secretion and decreased IL‐1 receptor antagonist (IL‐1Ra) secretion, resulting in an increase in the IL‐1β/IL‐1Ra secretion ratio. UFAs dose‐dependently inhibited the increase in IL‐1β secretion and decrease in IL‐1Ra secretion induced by Pal. Moreover, in human aortic and vein endothelial cells, expression and release of ICAM‐1 and E‐selectin were induced by treatment with conditioned medium collected from Pal‐stimulated pTHP‐1 cells and human monocytes, but not by Pal itself. The up‐regulated expression and release of adhesion molecules by the conditioned medium were mostly abolished by recombinant human IL‐1Ra supplementation. These results suggest that the Pal‐induced increase in the ratio of IL‐1β/IL‐1Ra secretion in monocytes up‐regulates endothelial adhesion molecules, which could enhance leukocyte adhesion to endothelium. This study provides further evidence that IL‐1β neutralization through receptor antagonism may be useful for preventing the onset and development of cardiovascular disease. J. Cell. Physiol. © 2014 Wiley Periodicals, Inc.
      PubDate: 2014-09-09T05:08:38.88911-05:0
      DOI: 10.1002/jcp.24797
  • microRNA‐151‐3p Regulates Slow Muscle Gene Expression by
           Targeting ATP2a2 in Skeletal Muscle Cells
    • Authors: Huan Wei; Zhongwen Li, Xiaobin Wang, Jie Wang, Weijun Pang, Gongshe Yang, Qingwu W. Shen
      Abstract: MicroRNAs (miRNAs) are a group of small noncoding RNAs that regulate the stability or translation of cognate mRNAs at the posttranscriptional level. Accumulating evidence indicates that miRNAs play important roles in many aspects of muscle function, including muscle growth and development, regeneration, contractility, and muscle fiber type plasticity. In the current study, we examined the function of miR‐151‐3p in myoblast proliferation and differentiation. Results show that overexpression of miR‐151‐3p not only upregulates myoblast proliferation, but also decreases slow muscle gene expression (such as MHC‐β/slow and slow muscle troponin I) in both C2C12 myotubes and in primary cultures. Alternatively, inhibition of miR‐151‐3p by antisense RNA was found to upregulate MHC‐β/slow expression, indicating that miR‐151‐3p plays a role in muscle fiber type determination. Further investigation into the underlying mechanisms revealed for the first time that miR‐151‐3p directly targets ATP2a2, a gene encoding for a slow skeletal and cardiac muscle specific Ca2+ ATPase, SERCA2 thus downregulating slow muscle gene expression. Mechanisms by which the alteration in SERCA2 expression induces changes in other slow muscle gene expression levels needs to be defined in future research. J. Cell. Physiol. © 2014 Wiley Periodicals, Inc.
      PubDate: 2014-09-09T05:07:57.752466-05:
      DOI: 10.1002/jcp.24793
  • miR‐34c regulates the permeability of blood‐tumor barrier via
           MAZ‐mediated expression changes of ZO‐1, occludin and
    • Authors: Lini Zhao; Ping Wang, Yunhui Liu, Jun Ma, Yixue Xue
      Abstract: The purposes of this study were to investigate the potential roles of miR‐34c in regulating blood‐tumor barrier (BTB) functions and its possible molecular mechanisms. The over‐expression of miR‐34c significantly impaired the integrity and increased the permeability of BTB, which were detected in an in vitro BTB model by transendothelial electric resistance and horseradish peroxidase flux assays respectively. Meanwhile, real‐time quantitative PCR (qRT‐PCR), Western blot and immunofluorescence assays successively demonstrated down‐regulation of ZO‐1, occludin and claudin‐5. And miR‐34c silencing uncovered the opposite results. Dual‐luciferase reporter assays results revealed myc‐associated zinc‐finger protein (MAZ) is a target gene of miR‐34c. Besides, mRNA and protein expressions of MAZ were reversely regulated by miR‐34c. The down‐expression of MAZ significantly impaired the integrity and increased the permeability of BTB as well as down‐regulated the expressions of ZO‐1, occludin and claudin‐5. And chromatin immunoprecipitation verified that MAZ interacted with “GGGCGGG”, “CCCTCCC” and “GGGAGGG” DNA sequence of ZO‐1, occludin and claudin‐5 promoter respectively. The over‐expression or silencing of either miR‐34c or MAZ was performed simultaneously to further explore their functional relations, and results elucidated that miR‐34c and MAZ displayed reverse regulatory effects on the integrity and permeability of BTB as well as the expressions of ZO‐1, occludin and claudin‐5. In conclusion, our present study indicated that miR‐34c regulated the permeability of BTB via MAZ‐mediated expression changes of ZO‐1, occludin and claudin‐5. J. Cell. Physiol. © 2014 Wiley Periodicals, Inc.
      PubDate: 2014-09-09T04:51:55.133271-05:
      DOI: 10.1002/jcp.24799
  • Sphingosine‐1‐phosphate mediates COX‐2 expression and
           PGE2/IL‐6 secretion via c‐Src‐dependent AP‐1
    • Authors: Chih‐Kai Hsu; I‐Ta Lee, Chih‐Chung Lin, Li‐Der Hsiao, Chuen‐Mao Yang
      Abstract: Sphingosine‐1‐phosphate (S1P) has been shown to regulate cyclooxygenase‐2 (COX‐2)/prostaglandin E2 (PGE2) expression and IL‐6 secretion in various respiratory diseases. However, the mechanisms underlying S1P‐induced COX‐2 expression and PGE2 production in human tracheal smooth muscle cells (HTSMCs) remain unclear. Here we demonstrated that S1P markedly induced COX‐2 expression. S1P also induced PGE2 and IL‐6 secretion which were reduced by the inhibitors of COX‐2 (NS‐398 and celecoxib). Pretreatment with the inhibitor of S1PR1 (W123), S1PR3 (CAY10444), c‐Src (PP1), PYK2 (PF431396), MEK1/2 (U0126), p38 MAPK (SB202190), JNK1/2 (SP600125), or AP‐1 (Tanshinone IIA) and transfection with siRNA of S1PR1, S1PR3, c‐Src, PYK2, p38, p42, JNK2, c‐Jun, or c‐Fos reduced S1P‐induced COX‐2 expression and PGE2/IL‐6 secretion. Moreover, S1P induced c‐Src, PYK2, p42/p44 MAPK, JNK1/2, p38 MAPK, and c‐Jun phosphorylation. We observed that S1P‐induced p42/p44 MAPK and JNK1/2, but not p38 MAPK activation was mediated via a c‐Src/PYK2‐dependent pathway. S1P also enhanced c‐Fos, but not c‐Jun mRNA and protein expression and the AP‐1 promoter activity. S1P‐induced c‐Fos mRNA and protein expression, c‐Jun phosphorylation, and AP‐1 promoter activity was reduced by W123, CAY10444, PP1, PF431396, U0126, SP600125, or SB202190. These results demonstrated that S1P‐induced COX‐2 expression and PGE2/IL‐6 generation was mediated through S1PR1/3/c‐Src/PYK2/p42/p44 MAPK‐ or JNK1/2‐ and S1PR1/3/c‐Src/p38 MAPK‐dependent AP‐1 activation. J. Cell. Physiol. © 2014 Wiley Periodicals, Inc.
      PubDate: 2014-09-09T04:50:38.124258-05:
      DOI: 10.1002/jcp.24795
  • CRM197 in combination with shRNA interference of VCAM‐1 displays
           enhanced inhibitory effects on human glioblastoma cells
    • Authors: Yi Hu; Xing Lin, Ping Wang, Yi‐Xue Xue, Zhen Li, Li‐Bo Liu, Bo Yu, Tian‐Da Feng, Yun‐Hui Liu
      Abstract: CRM197 is a naturally nontoxic diphtheria toxin mutant that binds and inhibits heparin‐binding epidermal growth factor‐like growth factor. CRM197 serves as carrier protein for vaccine and other therapeutic agents. CRM197 also inhibits the growth, migration, invasion and induces apoptosis in various tumors. Vascular cell adhesion molecule‐1 (VCAM‐1) is an important cell surface adhesion molecule associated with malignancy of gliomas. In this work, we aimed to investigate the role and mechanism of CRM197 combined with shRNA interference of VCAM‐1 (shRNA‐VCAM‐1) on the migration, invasion and apoptosis of glioblastoma cells. U87 and U251 human glioblastoma cells were treated with CRM197 (10 μg/ml) and shRNA interfering technology was employed to silence VCAM‐1 expression. Cell viability, migration, invasiveness and apoptosis were assessed with CCK8, Transwell and Annexin V‐PE/7‐AAD staining. Activation of cleaved caspase‐3, 8 and 9, activity of matrix metalloproteinase‐2/9 (MMP‐2/9) and expression of phosphorylated Akt (p‐Akt) were also checked. Results showed that CRM197 and shRNA‐VCAM‐1 not only significantly inhibited the cell proliferation, migration, invasion, but also promoted the apoptosis of U87 and U251 cells. Combined treatment of both displayed enhanced inhibitory effects on the malignant biological behavior of glioma cells. The activation of cleaved caspase‐3, 8, 9 was promoted, activity of MMP‐2 and MMP‐9 and expression of p‐Akt were inhibited significantly by the treatment of CRM197 and shRNA‐VCAM‐1 alone or in combination, indicating that the combination of CRM197 with shRNA‐VCAM‐1 additively inhibited the malignant behavior of human glioblastoma cells via activating caspase‐3, 8, 9 as well as inhibiting MMP‐2, MMP‐9 and Akt pathway. J. Cell. Physiol. © 2014 Wiley Periodicals, Inc.
      PubDate: 2014-09-09T04:47:53.443367-05:
      DOI: 10.1002/jcp.24798
  • MSCs modified with ACE2 restore endothelial function following LPS
           challenge by inhibiting the activation of RAS
    • Authors: Hong‐li He; Ling Liu, Qi‐hong Chen, Shi‐xia Cai, Ji‐bin Han, Shu‐ling Hu, Pan Chun, Yi Yang, Feng‐mei Guo, Ying‐zi Huang, Hai‐bo Qiu
      Abstract: Angiotensin (Ang) II plays an important role in the process of endothelial dysfunction in acute lung injury(ALI) and is degraded by angiotensin‐converting enzyme2 (ACE2). However, treatmentsthat target ACE2 to injured endothelium and promote endothelial repair of ALI are lacking. Mesenchymal stem cells (MSCs) are capable of homing to the injured site and delivering a protective gene. Our study aimed to evaluate the effects of genetically modified MSCs, which overexpress the ACE2 protein in a sustained manner via a lentiviral vector, on Ang II production in endothelium and in vitro repair of LPS‐induced endothelial injury.We found that the efficiency of lentiviral vector transduction of MSCs was as high as 97.8% and was well maintained over 30 passages. MSCs modified with ACE2 showed a sustained high expression of ACE2 mRNA and protein. The modified MSCs secreted soluble ACE2 protein into the culture medium, which reduced the concentration of Ang II and increased the production of Ang 1–7. MSCs modified with ACE2 were more effective at restoring endothelial function than were unmodified MSCs, as shown by the enhanced survival of endothelial cells; the downregulated production of inflammatory mediators, including ICAM‐1, VCAM‐1, TNF‐α, and IL‐6; reduced paracellular permeability; and increased expression of VE‐cadherin. These data demonstrate that MSCs modified to overexpress the ACE2 gene can produce biologically active ACE2 protein over a sustained period of time and have an enhanced ability to promote endothelial repair after LPS challenge.These results encourage further testing of the beneficial effects of ACE2‐modified MSCs in an ALI animal model. J. Cell. Physiol. © 2014 Wiley Periodicals, Inc.
      PubDate: 2014-09-09T04:40:42.77059-05:0
      DOI: 10.1002/jcp.24794
  • Interleukin‐17 induces CC chemokine receptor 6 expression and cell
           migration in colorectal cancer cells
    • Authors: Chih‐Chien Chin; Cheng‐Nan Chen, Hsing‐Chun Kuo, Chung‐Sheng Shi, Meng Chiao Hsieh, Yi‐Hung Kuo, Shui‐Yi Tung, Kam‐Fai Lee, Wen‐Shih Huang
      Abstract: The CC chemokine receptor 6 (CCR6) and its ligand CCL20 are involved in human colorectal cancer (CRC) carcinogenesis and can promote the progression of CRC. In addition, interleukin‐17 (IL‐17), produced by a T cell subset named “Th17,” has been identified as an important player in inflammatory responses, and has emerged as a mediator in inflammation‐associated cancer. However, the relevance of IL‐17 in the development and progression of CRC still remains to be explored. This study aimed to investigate the effect of IL‐17 on the cell migration of CRC cells. Human CRC HCT‐116 cells were used to study the effect of IL‐17 on CCR6 expression and cell migration in CRC cells. IL‐17 treatment induced migration of HCT‐116 cells across the Boyden chamber membrane and increased the expression level of the CCR6. Inhibition of CCR6 by small interfering RNA (siRNA) and neutralizing antibody inhibited IL‐17‐induced cell migration. By using specific inhibitors and short hairpin RNA (shRNA), we demonstrated that the activation of ERK and p38 pathways are critical for IL‐17‐induced CCR6 expression and cell migration. Promoter activity and transcription factor ELISA assays showed that IL‐17 increased NF‐κB‐DNA binding activity in HCT‐116 cells. Inhibition of NF‐κB activation by specific inhibitors and siRNA blocked the IL‐17‐induced CCR6 expression. Our findings support the hypothesis that CCR6 up‐regulation stimulated by IL‐17 may play an active role in CRC cell migration. J. Cell. Physiol. © 2014 Wiley Periodicals, Inc.
      PubDate: 2014-09-09T04:40:26.168474-05:
      DOI: 10.1002/jcp.24796
  • Acetylcholine Inhibits Tumor Necrosis Factor α Activated Endoplasmic
           Reticulum Apoptotic Pathway via EGFR‐PI3K Signaling in
    • Authors: Yi Miao; Xue‐Yuan Bi, Mei Zhao, Hong‐Ke Jiang, Jin‐Jun Liu, Dong‐Ling Li, Xiao‐Jiang Yu, Yong‐Hua Yang, Ning Huang, Wei‐Jin Zang
      Abstract: Previous findings have shown that acetycholine (ACh) decreased hypoxia‐induced tumor necrosis factor alpha (TNF α) production, thus protected against cardiomyocyte injury. However, whether and how ACh affects TNF α‐induced endoplasmic reticulum (ER) stress and cell apoptosis remain poorly defined. This study was aimed at determining the effect of ACh in H9c2 cells after TNF α stimulation. Presence of ER stress was verified using the ER stress protein markers glucose regulatory protein 78 (GRP78) and C/EBP homologous protein (CHOP). Cell apoptosis was shown by caspase‐3 activation and terminal deoxynucleotidyl transferase mediated dUTP‐biotin nick end labeling. Exogenously administered ACh significantly decreased these TNF α‐induced changes. Moreover, when the cells were exposed to nonspecific muscarinic receptor (M AChR) inhibitor atropine, methoctramine (M2 AChR inhibitor) or the epidermal growth factor receptor (EGFR) inhibitor AG1478, the cardioprotection elicited by ACh was diminished. Furthermore, the above effects were also blocked by M2 AChR or EGFR siRNA, indicating that EGFR transactivation by M2 AChR may be the major pathway responsible for the benefits of ACh. In addition, LY294002, a phosphatidylinositol‐3‐kinase (PI3K) inhibitor, displayed the similar trends as AG1478, suggesting that PI3K/Akt signaling may be the downstream of EGFR in ACh‐elicited anti‐apoptotic property. Together, these data indicate that EGFR‐PI3K/Akt signaling is involved in M2 AChR‐mediated ER apoptotic pathway suppression and the subsequent survival of H9c2 cardiomyocytes. We have identified a novel pathway underlying the cardioprotection afforded by ACh. J. Cell. Physiol. © 2014 Wiley Periodicals, Inc.
      PubDate: 2014-09-09T04:35:37.86881-05:0
      DOI: 10.1002/jcp.24800
  • Roles of SATB2 in Site‐Specific Stemness, Autophagy and Senescence
           of Bone Marrow Mesenchymal Stem Cells
      Abstract: Craniofacial bone marrow mesenchymal stem cells (BMSCs) display some site‐specific properties that differ from those of BMSCs derived from the trunk and appendicular skeleton, but the characteristics of craniofacial BMSCs and the mechanisms that underlie their properties are not completely understood. Previous studies indicated that special AT‐rich binding protein 2 (SATB2) may be a potential regulator of craniofacial skeletal patterning and site‐specific osteogenic capacity. Here, we investigated the stemness, autophagy, and anti‐aging capacity of mandible‐derived BMSCs (M‐BMSCs) and tibia‐derived BMSCs (T‐BMSCs) and explored the role of SATB2 in regulating these properties. M‐BMSCs not only possessed stronger expression of SATB2 and stemness markers (pluripotency genes, such as Nanog, OCT‐4, Sox2, and Nestin) but also exhibited stronger autophagy and anti‐aging capacities under normal or hypoxia/serum deprivation conditions compared to T‐BMSCs. Exogenous expression of SATB2 in T‐BMSCs significantly enhanced the expression of pluripotency genes as well as autophagy and anti‐aging capacity. Moreover, SATB2 markedly enhanced osteogenic differentiation of BMSCs in vitro, and promoted bone defect regeneration and the survival of BMSCs that were transplanted into mandibles with critical size defects. Mechanistically, SATB2 upregulates pluripotency genes and autophagy‐related genes, which in turn activate the mechanistic target of rapamycin signaling pathway. Collectively, our results provide novel evidence that site‐specific BMSCs have distinct biological properties and suggest that SATB2 plays a potential role in regulating the stemness, autophagy, and anti‐aging properties of craniofacial BMSCs. The application of SATB2 to manipulate stem cells for the reconstruction of bone defects might represent a new approach. J. Cell. Physiol. © 2014 Wiley Periodicals, Inc.
      PubDate: 2014-09-09T04:34:09.725711-05:
      DOI: 10.1002/jcp.24792
  • Gene therapies for cancer: strategies, challenges and successes
    • Authors: Swadesh K Das; Mitchell E. Menezes, Shilpa Bhatia, Xiang‐Yang Wang, Luni Emdad, Devanand Sarkar, Paul B. Fisher
      Abstract: Gene therapy, which involves replacement of a defective gene with a functional, healthy copy of that gene, is a potentially beneficial cancer treatment approach particularly over chemotherapy, which often lacks selectivity and can cause non‐specific toxicity. Despite significant progress pre‐clinically with respect to both enhanced targeting and expression in a tumor‐selective manner several hurdles still prevent success in the clinic, including non‐specific expression, low‐efficiency delivery and biosafety. Various innovative genetic approaches are under development to reconstruct vectors/transgenes to make them safer and more effective. Utilizing cutting‐edge delivery technologies, gene expression can now be targeted in a tissue‐ and organ‐specific manner. With these advances, gene therapy is poised to become amenable for routine cancer therapy with potential to elevate this methodology as a first line therapy for neoplastic diseases. This review discusses recent advances in gene therapy and their impact on a pre‐clinical and clinical level. J. Cell. Physiol. © 2014 Wiley Periodicals, Inc.
      PubDate: 2014-09-06T03:27:34.725787-05:
      DOI: 10.1002/jcp.24791
  • The Cooperation of CREB and NFAT is Required for PTHrP‐Induced RANKL
           Expression in Mouse Osteoblastic Cells
    • Authors: Hyun‐Jung Park; Kyunghwa Baek, Jeong‐Hwa Baek, Hyung‐Ryong Kim
      Pages: n/a - n/a
      Abstract: Parathyroid hormone‐related protein (PTHrP) is known to induce the expression of receptor activator of NF‐κB ligand (RANKL) in stromal cells/osteoblasts. However, the signaling pathways involved remain controversial. In the present study, we investigated the role of cAMP/protein kinase A (PKA) and calcineurin/NFAT pathways in PTHrP‐induced RANKL expression in C2C12 and primary cultured mouse calvarial cells. PTHrP‐mediated induction of RANKL expression was significantly inhibited by H89 and FK506, an inhibitor of PKA and calcineurin, respectively. PTHrP upregulated CREB phosphorylation and the transcriptional activity of NFAT. Knockdown of CREB or NFATc1 blocked PTHrP‐induced RANKL expression. PTHrP increased the activity of the RANKL promoter reporter that contains approximately 2 kb mouse RANKL promoter DNA sequences. Insertions of mutations in CRE‐like element or in NFAT‐binding element abrogated PTHrP‐induced RANKL promoter activity. Chromatin immunoprecipitation assays showed that PTHrP increased the binding of CREB and NFATc1/NFATc3 to their cognate binding elements in the RANKL promoter. Inhibition of cAMP/PKA and its downstream ERK activity suppressed PTHrP‐induced expression and transcriptional activity of NFATc1. CREB knockdown prevented PTHrP induction of NFATc1 expression. Furthermore, NFATc1 and CREB were co‐immunoprecipitated. Mutations in CRE‐like element completely blocked NFATc1‐induced transactivation of the RANKL promoter reporter; however, mutations in NFAT‐binding element partially suppressed CREB‐induced RANKL promoter activity. Overexpression of CREB increased NFATc1 binding to the RANKL promoter and vice versa. These results suggest that PTHrP‐induced RANKL expression depends on the activation of both cAMP/PKA and calcineurin/NFAT pathways, and subsequently, CREB and NFAT cooperate to transactivate the mouse RANKL gene. J. Cell. Physiol. © 2014 Wiley Periodicals, Inc.
      PubDate: 2014-09-04T05:04:05.489794-05:
      DOI: 10.1002/jcp.24790
  • Influence of Nuclear Blebs and Micronuclei Status on the Growth Kinetics
           of Human Mesenchymal Stem Cells
    • Authors: Shikha Sharma; Ramesh Bhonde
      Pages: n/a - n/a
      Abstract: Therapeutic potential of mesenchymal stem cells (MSCs) demands assurance of the quality, safety, and genetic stability. Nuclear blebs (NBs) and Micronuclei (MNs) are considered as biomarkers for cancer and an increase in their numbers is associated with malignancy and other pathologic disorders. However, the status of NBs and MNs in MSCs is not known. Hence we examined the frequency of NBs and MNs in MSCs from umbilical cord (UC‐MSCs) and placenta (PD‐MSCs) and found a difference in the number of NBs and MNs depending on the source of the MSCs. The number of NBs and MNs was always found to be less in UC‐MSCs as compared to PD‐MSCs obtained from the same patient. Furthermore, we observed that the number of NBs was inversely proportional to the proliferation rate of the cells. The cryopreservation of these MSCs over 6 months also led to increase in the number of NBs and MNs thus slowing down their rate of proliferation on revival. MSCs from both the sources exhibiting high NBs and MNs showed longer S phase and G2‐M arrest and increase in senescent cells without altering their CD‐marker profile and differentiation potential. This feature was consistent with the upregulation of cell cycle checkpoint genes (p53, p21, Gadd45, ATM, ATR, chek2, p27, p16, and p10). In conclusion, our data demonstrates for the first time the importance of checking the occurrence of NBs and MNs in MSCs before using them for cellular therapy as a quality control measure to check their genetic stability. J. Cell. Physiol. © 2014 Wiley Periodicals, Inc.
      PubDate: 2014-09-04T05:03:32.484741-05:
      DOI: 10.1002/jcp.24789
  • Pin1 Plays a Critical Role as a Molecular Switch in Canonical BMP
    • Authors: Won‐Joon Yoon; Rabia Islam, Young‐Dan Cho, Kyung‐Min Ryu, Hye‐Rim Shin, Kyung‐Mi Woo, Jeong‐Hwa Baek, Hyun‐Mo Ryoo
      Pages: n/a - n/a
      Abstract: Pin1 is a peptidyl prolyl cis‐trans isomerase that specifically binds to the phosphoserine–proline or phosphothreonine–proline motifs of numerous proteins. Previously, we reported that Pin1 deficiency resulted in defects in osteoblast differentiation during early bone development. In this study, we found that adult Pin1‐deficient mice developed osteoporotic phenotypes compared to age‐matched controls. Since BMP2 stored in the bone matrix plays a critical role in adult bone maintenance, we suspected that BMP R‐Smads (Smad1 and Smad5) could be critical targets for Pin1 action. Pin1 specifically binds to the phosphorylated linker region of Smad1, which leads to structural modification and stabilization of the Smad1 protein. In this process, Pin1‐mediated conformational modification of Smad1 directly suppresses the Smurf1 interaction with Smad1, thereby promoting sustained activation of the Smad1 molecule. Our data demonstrate that post‐phosphorylational prolyl isomerization of Smad1 is a converging signal to stabilize the Smad1 molecule against the ubiquitination process mediated by Smurf1. Therefore, Pin1 is a critical molecular switch in the determination of Smad1 fate, opposing the death signal transmitted to the Smad1 linker region by phosphorylation cascades after its nuclear localization and transcriptional activation. Thus, Pin1 could be developed as a major therapeutic target in many skeletal diseases. J. Cell. Physiol. © 2014 Wiley Periodicals, Inc.
      PubDate: 2014-09-04T04:55:34.928623-05:
      DOI: 10.1002/jcp.24787
  • Methotrexate‐Induced Bone Marrow Adiposity is Mitigated by Folinic
           Acid Supplementation Through the Regulation of Wnt/β‐Catenin
    • Authors: Kristen R. Georgiou; Rethi Raghu Nadhanan, Chia‐Ming Fan, Cory J. Xian
      Pages: n/a - n/a
      Abstract: Antimetabolite Methotrexate (MTX) is commonly used in childhood oncology. As a dihydrofolate reductase inhibitor it exerts its action through the reduction of cellular folate, thus its intensive use is associated with damage to soft tissues, bone marrow and bone. In the clinic, MTX is administered with folinic acid (FA) supplementation to alleviate some of this soft tissue damage. However, whether and how FA alleviates damage to the bone and bone marrow requires further investigation. As the Wnt/β‐catenin signalling pathway is critical for commitment and differentiation of mesenchymal stem cells down the osteogenic or adipogenic lineage, its deregulation has been found associated with increased marrow adiposity following MTX treatment. In order to elucidate whether FA supplementation prevents MTX‐induced bone marrow adiposity by regulating Wnt/β‐catenin signalling, young rats were given saline or 0.75mg/kg MTX once daily for 5 days, receiving saline or 0.75mg/kg FA 6h after MTX. FA rescue alleviated the MTX‐induced bone marrow adiposity, as well as well as inducing up‐regulation of Wnt10b mRNA and β‐catenin protein expression in the bone. Furthermore, FA blocked up‐regulation of the secreted Wnt antagonist sFRP‐1 mRNA expression. Moreover, secreted sFRP‐1 protein in the bone marrow and its expression by osteoblasts and adipocytes was found increased following MTX treatment. This potentially indicates that sFRP‐1 is a major regulator of defective Wnt/β‐catenin signalling following MTX treatment. This study provides evidence that folate depletion caused by MTX chemotherapy results in increased bone marrow adiposity, and that FA rescue alleviates these defects by up‐regulating Wnt/β‐catenin signalling in the bone. J. Cell. Physiol. © 2014 Wiley Periodicals, Inc.
      PubDate: 2014-09-04T04:44:20.12122-05:0
      DOI: 10.1002/jcp.24788
  • Palmitate induces ER stress and autophagy in H9c2 cells: implications for
           apoptosis and adiponectin resistance
    • Authors: Min Park; Anna Sabetski, Yee Kwan Chan, Subat Turdi, Gary Sweeney
      Abstract: The association between obesity and heart failure is well documented and recent studies have indicated that understanding the physiological role of autophagy will be of great significance. Cardiomyocyte apoptosis is one component of cardiac remodeling which leads to heart failure and in this study we used palmitate‐treated H9c2 cells as an in vitro model of lipotoxicity to investigate the role of autophagy in cell death. Temporal analysis revealed that palmitate (100μM) treatment induced a gradual increase of intracellular lipid accumulation as well as apoptotic cell death. Palmitate induced autophagic flux, determined via increased LC3‐II formation and p62 degradation as well as by detecting reduced colocalization of GFP with RFP in cells overexpressing tandem fluorescent GFP/RFP‐LC3. The increased level of autophagy indicated by these measures were confirmed using transmission electron microscopy (TEM). Upon inhibiting autophagy using bafilomycin we observed an increased level of cell death assessed by Annexin V/PI staining, detection of active caspase‐3 and MTT cell viability assay. Interestingly, using TEM and p‐PERK or p‐eIF2α detection we observed increased endoplasmic reticulum (ER) stress in response to palmitate. Autophagy was induced as an adaptive response against ER stress since it was sensitive to ER stress inhibition. Palmitate‐induced ER stress also induced adiponectin resistance, assessed via AMPK phosphorylation, via reducing APPL1 expression. This effect was independent of palmitate‐induced autophagy. In summary, our data indicate that palmitate induces autophagy subsequent to ER stress and that this confers a prosurvival effect against lipotoxicity‐induced cell death. Palmitate‐induced ER stress also led to adiponecin resistance. J. Cell. Physiol. © 2014 Wiley Periodicals, Inc.
      PubDate: 2014-08-28T04:18:15.283126-05:
      DOI: 10.1002/jcp.24781
  • Thymoquinone restores radiation induced TGF‐β expression and
           abrogate s EMT in chemoradiotherapy of breast cancer cells
    • Authors: Shashi Rajput; B. N. Prashanth Kumar, Payel Banik, Sheetal Parida, Mahitosh Mandal
      Abstract: Radiotherapy remains a prime approach to adjuvant therapies in patients with early and advanced breast cancer. In spite of therapeutic success, metastatic progression in patients undergoing therapy, limits its application. However, effective therapeutic strategies to understand the cellular and molecular machinery in inhibiting radiation induced metastatic progression, which is poorly understood so far, need to be strengthened. Ionizing radiation was known to prompt cancer cell's metastatic ability by eliciting Transforming Growth Factor‐beta (TGF‐ beta), a key regulator in epithelial‐mesenchymal transdifferentiation and radio‐resistance. In this viewpoint, we employed thymoquinone as a radiosensitizer to investigate its migration and invas ion reversal abilities in irradiated breast cancer cell lines by assessing their respective attributes. The role of metastasis regulatory molecules like TGF‐β, E‐cadherin and integrin aV and its downstream molecules were determined using RT‐PCR, western blotting, immunofluorescence and extracellular TGF‐β levels affirmed through ELISA assays. These studies affirmed the TGF‐β restoring ability of thymoquinone in radiation driven migration and invasion. Also, results demonstrated that the epithelial markers E‐cadherin and cytokeratin 19 were down‐regulated whereas mesenchymal markers like integrin aV, MMP9 and MMP2 were upregulated by irradiation treatment, however thymoquinone pre‐sensitization has reverted the expression of these proteins backs to control proteins expression. Here, paclitaxel was chosen as an apoptosis inducer in TGF‐β restored cells and confirmed its cytotoxic effects in radiation alone and thymoquinone sensitized irradiated cells. We conclude that this therapeutic modality is effective in preventing radiation induced epithelial‐mesenchymal transdifferentiation and concomitant induction of apoptosis in breast cancer. J. Cell. Physiol. © 2014 Wiley Periodicals, Inc.
      PubDate: 2014-08-28T04:16:32.026714-05:
      DOI: 10.1002/jcp.24780
  • PLCâ1a and PLCâ1b selective regulation and cyclin D3 modulation
           reduced by Kinamycin F during K562 cell differentiation
    • Authors: Alberto Bavelloni; Gary I. Dmitrienko, Valerie J. Goodfellow, Ahmad Ghavami, Manuela Piazzi, William Blalock, Francesca Chiarini, Lucio Cocco, Irene Faenza
      Abstract: Here we report that both PLCâ1a and PLCâ1b are relevant regulators of erythropoiesis in that kinamycin F, a potent inducer of γ‐globin production in K562 cells, caused a selectively reduction of both PLCâ1 isozymes even though the results point out that the effect of the drug is mainly directed toward the expression of the PLCâ1a isoform. We have identified a different role for the two isozymes as regulators of K562 differentiation process induced by kinamycin F. The overexpression of PLCâ1b induced an increase in γ‐globin expression even in the absence of kinamycin F. Moreover during K562 differentiation, cyclin D3 level is regulated by PLCâ1 signaling pathway. Namely the amplification of the expression of the PLCâ1a, but not of PLCâ1b, is able to maintain high levels of expression of cyclin D3 even after treatment with kinamycin F. This could be due to their different distribution in the cell compartments since the amount of PLCâ1b is mainly present in the nucleus in respect to PLCâ1a. Our data indicate that the amplification of PLCâ1a expression, following treatment with kinamycin F, confers a real advantage to K562 cells viability and protects cells themselves from apoptosis. J. Cell. Physiol. © 2014 Wiley Periodicals, Inc.
      PubDate: 2014-08-27T06:10:17.697556-05:
      DOI: 10.1002/jcp.24776
  • Initial Characterization of Osteoblast Differentiation and Loss of RUNX2
           Stability in the Newly Established SK11 Human Embryonic Stem
           Cell‐Derived Cell Line
    • Authors: Jiali Yu; Helty Adisetiyo, Gillian H. Little, C.Thomas Vangsness, Jianjie Jiang, Hal Sternberg, Michael D. West, Baruch Frenkel
      Abstract: We describe a novel model for investigation of genetically normal human osteoblasts in culture. SK11 is a clonal progenitor cell line derived from human embryonic stem cells. Initially selected based on the expression of chondrogenic markers when differentiated in micromass culture, SK11 cultures display typical mRNA expression patterns of bone phenotypic genes under osteogenic conditions. These include Osterix, α1(I) collagen, Alkaline phosphatase, Osteonectin, Osteopontin and Osteocalcin. Similar to well‐characterized murine osteoblast cultures, the osteoblast master regulator RUNX2 was present during the first few days after plating, but the protein disappeared during the first week of culture. Loss of RUNX2 expression is considered an important regulatory feature for osteoblast maturation. Indeed, following ∼2 weeks of differentiation, SK11 cultures exhibited robust calcium deposition, evidenced by alizarin red staining. We also introduced a lentiviral vector encoding doxycycline (dox)‐inducible FLAG‐tagged RUNX2 into SK11 cells. Dox‐mediated enhancement of RUNX2 expression resulted in accelerated mineralization, which was further increased by co‐treatment with BMP‐2. Like the endogenous RUNX2, expression of the virally coded FLAG‐RUNX2 was lost during the first week of culture despite persistent dox treatment. By following RUNX2 decay after dox withdrawal from day‐5 versus day‐3 cultures, we demonstrated a developmentally regulated decrease in RUNX2 stability. Availability of culture models for molecular investigation of genetically normal human osteoblasts is important because differences between murine and human osteoblasts, demonstrated here for the regulation of Matrix Gla Protein, may have significant biomedical implications. J. Cell. Physiol. © 2014 Wiley Periodicals, Inc.
      PubDate: 2014-08-27T06:05:41.538286-05:
      DOI: 10.1002/jcp.24773
  • MicroRNA‐Based Biotechnology for Plant Improvement
    • Authors: Baohong Zhang; Qinglian Wang
      First page: 1
      Abstract: MicroRNAs (miRNAs) are an extensive class of newly discovered endogenous small RNAs, which negatively regulate gene expression at the post‐transcription levels. As the application of next‐generation deep sequencing and advanced bioinformatics, the miRNA‐related study has been expended to non‐model plant species and the number of identified miRNAs has dramatically increased in the past years. miRNAs play a critical role in almost all biological and metabolic processes, and provide a unique strategy for plant improvement. Here, we first briefly review the discovery, history, and biogenesis of miRNAs, then focus more on the application of miRNAs on plant breeding and the future directions. Increased plant biomass through controlling plant development and phase change has been one achievement for miRNA‐based biotechnology; plant tolerance to abiotic and biotic stress was also significantly enhanced by regulating the expression of an individual miRNA. Both endogenous and artificial miRNAs may serve as important tools for plant improvement. J. Cell. Physiol. 230: 1–15, 2014. © 2014 Wiley Periodicals, Inc.
      PubDate: 2014-09-29T19:20:39.347243-05:
      DOI: 10.1002/jcp.24685
  • 3D Cell Culture Systems: Advantages and Applications
    • Authors: Maddaly Ravi; V. Paramesh, S.R. Kaviya, E. Anuradha, F.D. Paul Solomon
      First page: 16
      Abstract: Cell cultures are important material of study for the variety of advantages that they offer. Both established continuous cell lines and primary cell cultures continue to be invaluable for basic research and for direct applications. Technological advancements are necessary to address emerging complex challenges and the way cells are cultured in vitro is an area of intense activity. One important advancement in cell culture techniques has been the introduction of three dimensional culture systems. This area is one of the fastest growing experimental approaches in life sciences. Augmented with advancements in cell imaging and analytical systems, as well as the applications of new scaffolds and matrices, cells have been increasingly grown as three dimensional models. Such cultures have proven to be closer to in vivo natural systems, thus proving to be useful material for many applications. Here, we review the three dimensional way of culturing cells, their advantages, the scaffolds and matrices currently available, and the applications of such cultures in major areas of life sciences. J. Cell. Physiol. 230: 16–26, 2014. © 2014 Wiley Periodicals, Inc.
      PubDate: 2014-09-29T19:20:40.479944-05:
      DOI: 10.1002/jcp.24683
  • CXCR4/CXCR7 Molecular Involvement in Neuronal and Neural Progenitor
           Migration: Focus in CNS Repair
    • Authors: José Joaquín Merino; Victor Bellver‐Landete, María Jesús Oset‐Gasque, Beatriz Cubelos
      First page: 27
      Abstract: In the adult brain, neural progenitor cells (NPCs) reside in the subventricular zone (SVZ) of the lateral ventricles, the dentate gyrus and the olfactory bulb. Following CNS insult, NPCs from the SVZ can migrate along the rostral migratory stream (RMS), a migration of NPCs that is directed by proinflammatory cytokines. Cells expressing CXCR4 follow a homing signal that ultimately leads to neuronal integration and CNS repair, although such molecules can also promote NPC quiescence. The ligand, SDF1 alpha (or CXCL12) is one of the chemokines secreted at sites of injury that it is known to attract NSC‐derived neuroblasts, cells that express CXCR4. In function of its concentration, CXCL12 can induce different responses, promoting NPC migration at low concentrations while favoring cell adhesion via EGF and the alpha 6 integrin at high CXCL12 concentrations. However, the preclinical effectiveness of chemokines and their relationship with NPC mobilization requires further study, particularly with respect to CNS repair. NPC migration may also be affected by the release of cytokines or chemokines induced by local inflammation, through autocrine or paracrine mechanisms, as well as through erythropoietin (EPO) or nitric oxide (NO) release. CXCL12 activity requires G‐coupled proteins and the availability of its ligand may be modulated by its binding to CXCR7, for which it shows a stronger affinity than for CXCR4. J. Cell. Physiol. 230: 27–42, 2014. © 2014 Wiley Periodicals, Inc.
      PubDate: 2014-09-29T19:20:36.122763-05:
      DOI: 10.1002/jcp.24695
  • Tumor Vasculature Is Regulated by FGF/FGFR Signaling‐Mediated
           Angiogenesis and Bone Marrow‐Derived Cell Recruitment: This
           Mechanism Is Inhibited by SSR128129E, the First Allosteric Antagonist of
    • Authors: Pierre Fons; Geneviève Gueguen‐Dorbes, Jean‐Pascal Herault, Fabien Geronimi, Joël Tuyaret, Dol Frédérique, Paul Schaeffer, Cécile Volle‐Challier, Jean‐Marc Herbert, Françoise Bono
      First page: 43
      Abstract: Tumor angiogenesis is accompanied by vasculogenesis, which is involved in the differentiation and mobilization of human bone marrow cells. In order to further characterize the role of vasculogenesis in the tumor growth process, the effects of FGF2 on the differentiation of human bone marrow AC133+ cells (BM‐AC133+) into vascular precursors were studied in vitro. FGF2, like VEGFA, induced progenitor cell differentiation into cell types with endothelial cell characteristics. SSR128129E, a newly discovered specific FGFR antagonist acting by allosteric interaction with FGFR, abrogated FGF2‐induced endothelial cell differentiation, showing that FGFR signaling is essential during this process. To assess the involvement of the FGF/FRGR signaling in vivo, the pre‐clinical model of Lewis lung carcinoma (LL2) in mice was used. Subcutaneous injection of LL2 cells into mice induced an increase of circulating EPCs from peripheral blood associated with tumor growth and an increase of intra‐tumoral vascular index. Treatment with the FGFR antagonist SSR128129E strongly decreased LL2 tumor growth as well as the intra‐tumoral vascular index (41% and 50% decrease vs. vehicle‐treated mice respectively, P 
      PubDate: 2014-09-29T19:20:37.657767-05:
      DOI: 10.1002/jcp.24656
  • Histone Deacetylase Inhibition Destabilizes the Multi‐Potent State
           of Uncommitted Adipose‐Derived Mesenchymal Stromal Cells
    • Authors: Amel Dudakovic; Emily T. Camilleri, Eric A. Lewallen, Meghan E. McGee‐Lawrence, Scott M. Riester, Sanjeev Kakar, Martin Montecino, Gary S. Stein, Hyun‐Mo Ryoo, Allan B. Dietz, Jennifer J. Westendorf, Andre J. van Wijnen
      First page: 52
      Abstract: Human adipose‐derived mesenchymal stromal cells (AMSCs) grown in platelet lysate are promising agents for therapeutic tissue regeneration. Here, we investigated whether manipulation of epigenetic events by the clinically relevant histone deacetylase inhibitor suberoylanilide hydroxamic acid (SAHA) alters differentiation of AMSCs. The multipotency of AMSCs was validated by their ability to differentiate into osteogenic, chondrogenic, and adipogenic lineages. High‐throughput RNA sequencing and RT‐qPCR established that human histone deacetylases (HDAC1 to HDAC11, and SIRT1 to SIRT7) are differentially expressed in AMSCs. SAHA induces hyper‐acetylation of histone H3 and H4, stimulates protein expression of the HDAC‐responsive gene SLC9A3R1/NHERF1 and modulates the AKT/FOXO1 pathway. Biologically, SAHA interferes with osteogenic, chondrogenic and adipogenic lineage commitment of multipotent AMSCs. Mechanistically, SAHA‐induced loss of differentiation potential of uncommitted AMSCs correlates with multiple changes in the expression of principal transcription factors that control mesenchymal or pluripotent states. We propose that SAHA destabilizes the multi‐potent epigenetic state of uncommitted human AMSCs by hyper‐acetylation and perturbation of key transcription factor pathways. Furthermore, AMSCs grown in platelet lysate may provide a useful biological model for screening of new HDAC inhibitors that control the biological fate of human mesenchymal stromal cells. J. Cell. Physiol. 230: 52–62, 2014. © 2014 Wiley Periodicals, Inc.
      PubDate: 2014-09-29T19:20:38.524201-05:
      DOI: 10.1002/jcp.24680
  • microRNA‐Mediated Survivin Control of Pluripotency
    • Authors: Kristina Kapinas; Heesun Kim, Matthew Mandeville, Lori A. Martin‐Buley, Carlo M. Croce, Jane B. Lian, Andre J. van Wijnen, Janet L. Stein, Dario C. Altieri, Gary S. Stein
      First page: 63
      Abstract: Understanding the mechanisms that sustain pluripotency in human embryonic stem cells (hESCs) is an active area of research that may prove useful in regenerative medicine and will provide fundamental information relevant to development and cancer. hESCs and cancer cells share the unique ability to proliferate indefinitely and rapidly. Because the protein survivin is uniquely overexpressed in virtually all human cancers and in hESCs, we sought to investigate its role in supporting the distinctive capabilities of these cell types. Results presented here suggest that survivin contributes to the maintenance of pluripotency and that post‐transcriptional control of survivin isoform expression is selectively regulated by microRNAs. miR‐203 has been extensively studied in human tumors, but has not been characterized in hESCs. We show that miR‐203 expression and activity is consistent with the expression and subcellular localization of survivin isoforms that in turn modulate expression of the Oct4 and Nanog transcription factors to sustain pluripotency. This study contributes to understanding of the complex regulatory mechanisms that govern whether hESCs proliferate or commit to lineages. J. Cell. Physiol. 230: 63–70, 2014. © 2014 Wiley Periodicals, Inc.
      PubDate: 2014-09-29T19:20:44.220553-05:
      DOI: 10.1002/jcp.24681
  • Brain Natriuretic Peptide and C‐Type Natriuretic Peptide Maintain
           Porcine Oocyte Meiotic Arrest
    • Authors: Wenqiang Zhang; Ye Yang, Wei Liu, Qian Chen, Huarong Wang, Xiao Wang, Yanhao Zhang, Meijia Zhang, Guoliang Xia
      First page: 71
      Abstract: Recent studies have shown that C‐type natriuretic peptide (CNP) serves as a key control system during mouse oocyte maturation. We used pig models (in vitro and in vivo) to explore the role played by the natriuretic peptide family in porcine oocyte maturation. We reported the expression and location of natriuretic peptide system in different stages of porcine antral follicles. Atrial natriuretic peptide (ANP) and CNP were expressed primarily in granulosa cells, whereas brain natriuretic peptide (BNP) and natriuretic peptide receptor‐B (NPRB) receptor were expressed in granulosa cells (both cumulus and mural granulosa cells) and thecal internal cells, and the natriuretic peptide receptor‐A (NPRA) receptor predominantly in thecal cells. Upon in vitro culture, BNP and CNP maintained meiotic arrest of oocytes associated with cumulus cells. The expression levels of BNP, CNP, and the NPRB receptor increased upon treatment of prepubertal gilts with pregnant mare's serum gonadotropin and decreased upon subsequent human chorionic gonadotropin injection. Such dynamic changes in the expression of natriuretic peptides and their receptor paralleled the proportions of oocytes exhibiting nuclear maturation in vivo. These data indicated that BNP and CNP co‐contributed to maintaining porcine meiotic arrest under physiological condition and lutenizing hormone (LH) relieved this inhibitory effect by decreasing the expression levels of BNP and CNP in vivo. Our present work, combined with previous data, improved the understanding of the oocyte meiotic arrest mechanisms and further revealed that natriuretic peptides serve as oocyte maturation inhibitor (OMI) to inhibit oocyte maturation in mammals. J. Cell. Physiol. 230: 71–81, 2014. © 2014 Wiley Periodicals, Inc.
      PubDate: 2014-09-29T19:20:40.089506-05:
      DOI: 10.1002/jcp.24682
  • EGFL7 Is Expressed in Bone Microenvironment and Promotes Angiogenesis via
           ERK, STAT3, and Integrin Signaling Cascades
    • Authors: Shek Man Chim; Vincent Kuek, Siu To Chow, Bay Sie Lim, Jennifer Tickner, Jinmin Zhao, Rosa Chung, Yu‐Wen Su, Ge Zhang, Wendy Erber, Cory J. Xian, Vicki Rosen, Jiake Xu
      First page: 82
      Abstract: Angiogenesis plays a pivotal role in bone formation, remodeling, and fracture healing. The regulation of angiogenesis in the bone microenvironment is highly complex and orchestrated by intercellular communication between bone cells and endothelial cells. Here, we report that EGF‐like domain 7 (EGFL7), a member of the epidermal growth factor (EGF) repeat protein superfamily is expressed in both the osteoclast and osteoblast lineages, and promotes endothelial cell activities. Addition of exogenous recombinant EGFL7 potentiates SVEC (simian virus 40‐transformed mouse microvascular endothelial cell line) cell migration and tube‐like structure formation in vitro. Moreover, recombinant EGFL7 promotes angiogenesis featuring web‐like structures in ex vivo fetal mouse metatarsal angiogenesis assay. We show that recombinant EGFL7 induces phosphorylation of extracellular signal‐regulated kinase 1/2 (ERK1/2), signal transducer and activator of transcription 3 (STAT3), and focal adhesion kinase (FAK) in SVEC cells. Inhibition of ERK1/2 and STAT3 signaling impairs EGFL7‐induced endothelial cell migration, and angiogenesis in fetal mouse metatarsal explants. Bioinformatic analyses indicate that EGFL7 contains a conserved RGD/QGD motif and EGFL7‐induced endothelial cell migration is significantly reduced in the presence of RGD peptides. Moreover, EGFL7 gene expression is significantly upregulated during growth plate injury repair. Together, these results demonstrate that EGFL7 expressed by bone cells regulates endothelial cell activities through integrin‐mediated signaling. This study highlights the important role that EGFL7, like EGFL6, expressed in bone microenvironment plays in the regulation of angiogenesis in bone. J. Cell. Physiol. 230: 82–94, 2014. © 2014 Wiley Periodicals, Inc.
      PubDate: 2014-09-29T19:20:45.034279-05:
      DOI: 10.1002/jcp.24684
  • A Functional Transient Receptor Potential Vanilloid 4 (TRPV4) Channel Is
           Expressed in Human Endothelial Progenitor Cells
    • Authors: Silvia Dragoni; Germano Guerra, Alessandra Fiorio Pla, Giuseppe Bertoni, Alessandra Rappa, Valentina Poletto, Cinzia Bottino, Adele Aronica, Francesco Lodola, Maria Pia Cinelli, Umberto Laforenza, Vittorio Rosti, Franco Tanzi, Luca Munaron, Francesco Moccia
      First page: 95
      Abstract: Endothelial progenitor cells (EPCs) are mobilized into circulation to replace damaged endothelial cells and recapitulate the vascular network of injured tissues. Intracellular Ca2+ signals are key to EPC activation, but it is yet to be elucidated whether they are endowed with the same blend of Ca2+‐permeable channels expressed by mature endothelial cells. For instance, endothelial colony forming cells (ECFCs), the only EPC subset truly committed to acquire a mature endothelial phenotype, lack canonical transient receptor potential channels 3, 5 and 6 (TRPC3, 5 and 6), which are widely distributed in vascular endothelium; on the other hand, they express a functional store‐operated Ca2+ entry (SOCE). The present study was undertaken to assess whether human circulating EPCs possess TRP vanilloid channel 4 (TRPV4), which plays a master signalling role in mature endothelium, by controlling both vascular remodelling and arterial pressure. We found that EPCs express both TRPV4 mRNA and protein. Moreover, both GSK1016790A (GSK) and phorbol myristate acetate and, two widely employed TRPV4 agonists, induced intracellular Ca2+ signals uniquely in presence of extracellular Ca2+. GSK‐ and PMA‐induced Ca2+ elevations were inhibited by RN‐1734 and ruthenium red, which selectively target TRPV4 in mature endothelium. However, TRPV4 stimulation with GSK did not cause EPC proliferation, while the pharmacological blockade of TRPV4 only modestly affected EPC growth in the presence of a growth factor‐enriched culture medium. Conversely, SOCE inhibition with BTP‐2, La3+ and Gd3+ dramatically decreased cell proliferation. These data indicate that human circulating EPCs possess a functional TRPV4 protein before their engraftment into nascent vessels. J. Cell. Physiol. 230: 95–104, 2014. © 2014 Wiley Periodicals, Inc.
      PubDate: 2014-09-29T19:20:35.314563-05:
      DOI: 10.1002/jcp.24686
  • EGF Regulates Claudin‐2 and ‐4 Expression Through Src and
           STAT3 in MDCK Cells
    • Authors: Vicky García‐Hernández; Catalina Flores‐Maldonado, Ruth Rincon‐Heredia, Odette Verdejo‐Torres, José Bonilla‐Delgado, Ivan Meneses‐Morales, Patricio Gariglio, Rubén G. Contreras
      First page: 105
      Abstract: Epidermal Growth Factor (EGF) is a key regulator of epithelial paracellular permeability, a property that depends on tight junctions (TJ) and can be evaluated through the measurement of the transepithelial electrical resistance (TER). EGF increases the TER of MDCK monolayers by inducing ERK1/2‐dependent downregulation of claudin‐2 (CLDN‐2) and upregulation of claudin‐4 (CLDN‐4). Because either increments or decrements in TER often involve Src activation and epithelial cell differentiation occasionally depends on STAT3, here we investigated whether EGF might control CLDN‐2 downregulation and CLDN‐4 upregulation through those proteins. We found that EGF induces Src activation necessary for the reduction of CLDN‐2 at the TJ, the degradation of this CLDN, the reduction of the cellular levels of its mRNA and the resulting increase of TER. EGF‐induced changes on CLDN‐2 protein and mRNA also depend on STAT3 activity. This growth factor increases the levels of STAT3 phosphorylated at Y705 in the nucleus, a process that depends on Src activation. Interestingly, Src and STAT3 activation do not exclusively mediate the EGF‐induced downregulation of CLDN‐2, but they are also implicated in the EGF‐induced CLDN‐4 transcription, translation, and exocytic fusion into TJ. Our results indicate that EGF controls the levels of CLDN‐2 and ‐4 proteins and mRNAs through Src and STAT3 activity. J. Cell. Physiol. 230: 105–115, 2014. © 2014 Wiley Periodicals, Inc.
      PubDate: 2014-09-29T19:20:38.082263-05:
      DOI: 10.1002/jcp.24687
  • Human Olfactory Bulb Neural Stem Cells expressing hNGF Restore Cognitive
           Deficit in Alzheimer's Disease Rat Model
    • Authors: Hany E. S. Marei; Amany Farag, Asma Althani, Nahla Afifi, Ahmed Abd‐Elmaksoud, Samah Lashen, Shaymaa Rezk, Roberto Pallini, Patrizia Casalbore, Carlo Cenciarelli
      First page: 116
      Abstract: In this study, we aim to demonstrate the fate of allogenic adult human olfactory bulb neural stem/progenitor cells (OBNSC/NPCs) transplanted into the rat hippocampus treated with ibotenic acid (IBO), a neurotoxicant specific to hippocampal cholinergic neurons that are lost in Alzheimer's disease. We assessed their possible ability to survive, integrate, proliferate, and differentiate into different neuronal and glial elements: we also evaluate their possible therapeutic potential, and the mechanism(s) relevant to neuroprotection following their engraftment into the CNS milieu. OBNSC/NPCs were isolated from adult human olfactory bulb patients, genetically engineered to express GFP and human nerve growth factor (hNGF) by lentivirus‐mediated infection, and stereotaxically transplanted into the hippocampus of IBO‐treated animals and controls. Stereological analysis of engrafted OBNSCs eight weeks post transplantation revealed a 1.89 fold increase with respect to the initial cell population, indicating a marked ability for survival and proliferation. In addition, 54.71 ± 11.38%, 30.18 ± 6.00%, and 15.09 ± 5.38% of engrafted OBNSCs were identified by morphological criteria suggestive of mature neurons, oligodendrocytes and astrocytes respectively. Taken together, this work demonstrated that human OBNSCs expressing NGF ameliorate the cognitive deficiencies associated with IBO‐induced lesions in AD model rats, and the improvement can probably be attributed primarily to neuronal and glial cell replacement as well as the trophic influence exerted by the secreted NGF. J. Cell. Physiol. 230: 116–130, 2014. © 2014 Wiley Periodicals, Inc.
      PubDate: 2014-09-29T19:20:43.584524-05:
      DOI: 10.1002/jcp.24688
  • Sorafenib/Regorafenib and Lapatinib Interact to kill CNS Tumor Cells
    • Authors: Hossein A. Hamed; Seyedmehrad Tavallai, Steven Grant, Andrew Poklepovic, Paul Dent
      First page: 131
      Abstract: The present studies were to determine whether the multi‐kinase inhibitor sorafenib or its derivative regorafenib interacted with the ERBB1/ERBB2 inhibitor lapatinib to kill CNS tumor cells. In multiple CNS tumor cell types sorafenib and lapatinib interacted in a greater than additive fashion to cause tumor cell death. Tumor cells lacking PTEN, and anoikis or lapatinib resistant cells were as sensitive to the drug combination as cells expressing PTEN or parental cells, respectively. Similar data were obtained using regorafenib. Treatment of brain cancer cells with [sorafenib + lapatinib] enhanced radiation toxicity. The drug combination increased the numbers of LC3‐GFP vesicles; this correlated with a reduction in endogenous LC3II, and p62 and LAMP2 degradation. Knock down of Beclin1 or ATG5 significantly suppressed drug combination lethality. Expression of c‐FLIP‐s, BCL‐XL, or dominant negative caspase 9 reduced drug combination toxicity; knock down of FADD or CD95 was protective. Expression of both activated AKT and activated MEK1 or activated mTOR was required to strongly suppress drug combination lethality. As both lapatinib and sorafenib are FDA approved agents, our data argue for further determination as to whether lapatinib and sorafenib is a useful glioblastoma therapy. J. Cell. Physiol. 230: 131–139, 2014. © 2014 Wiley Periodicals, Inc.
      PubDate: 2014-09-29T19:20:44.60158-05:0
      DOI: 10.1002/jcp.24689
  • CCN1 Secreted by Tonsil‐Derived Mesenchymal Stem Cells Promotes
           Endothelial Cell Angiogenesis via Integrin αvβ3 and AMPK
    • Authors: Yoon Shin Park; Soojin Hwang, Yoon Mi Jin, Yeonsil Yu, Sung‐Ae Jung, Sung‐Chul Jung, Kyung‐Ha Ryu, Han Su Kim, Inho Jo
      First page: 140
      Abstract: CCN1 is highly expressed in cancer cells and has been identified in the secretome of bone marrow‐derived mesenchymal stem cells (BM‐MSC). Although secreted CCN1 is known to promote angiogenesis, its underlying mechanism remains unclear. Here, we examined whether our recently‐established tonsil‐derived MSC (T‐MSC) secrete CCN1 and, if any, how CCN1 promotes the angiogenesis of human umbilical vein endothelial cells (HUVEC). Compared with untreated control T‐MSC, a higher level of CCN1 was secreted by T‐MSC treated with activin A and sonic hedgehog, drugs known to induce endodermal differentiation. Expectedly, conditioned medium collected from differentiated T‐MSC (DCM) significantly increased HUVEC migration and tube formation compared with that from control T‐MSC (CCM), and these stimulatory effects were reversed by neutralization with anti‐CCN1 antibody. Treatment with recombinant human CCN1 (rh‐CCN1) alone also mimicked the stimulatory effects of DCM. Furthermore, treatment with either DCM or rh‐CCN1 increased the phosphorylation of AMP kinase (AMPK), and ectopic expression of siRNA of the AMPK gene inhibited all observed effects of both DCM and rh‐CCN1. However, no alteration of intracellular ATP levels or phosphorylation of LKB1, a well‐known upstream factor of AMPK activation, was observed under our conditions. Finally, the neutralization of integrin αvβ3 with anti‐integrin αvβ3 antibody almost completely reversed the effects of CCN1 on AMPK phosphorylation, and EC migration and tube formation. Taken together, we demonstrated that T‐MSC increase the secretion of CCN1 in response to endodermal differentiation and that integrin αvβ3 and AMPK mediate CCN1‐induced EC migration and tube formation independent of intracellular ATP levels alteration. J. Cell. Physiol. 230: 140–149, 2014. © 2014 Wiley Periodicals, Inc.
      PubDate: 2014-09-29T19:20:42.220175-05:
      DOI: 10.1002/jcp.24690
  • Effect of CTGF/CCN2 on Osteo/Cementoblastic and Fibroblastic
    • Authors: Asuka Yuda; Hidefumi Maeda, Shinsuke Fujii, Satoshi Monnouchi, Naohide Yamamoto, Naohisa Wada, Katsuaki Koori, Atsushi Tomokiyo, Sayuri Hamano, Daigaku Hasegawa, Akifumi Akamine
      First page: 150
      Abstract: Appropriate mechanical loading during occlusion and mastication play an important role in maintaining the homeostasis of periodontal ligament (PDL) tissue. Connective tissue growth factor (CTGF/CCN2), a matricellular protein, is known to upregulate extracellular matrix production, including collagen in PDL tissue. However, the underlying mechanisms of CTGF/CCN2 in regulation of PDL tissue integrity remain unclear. In this study, we investigated the effect of CTGF/CCN2 on osteo/cementoblastic and fibroblastic differentiation of human PDL stem cells using the cell line 1–11. CTGF/CCN2 expression in rat PDL tissue and human PDL cells (HPDLCs) was confirmed immunohisto/cytochemically. Mechanical loading was found to increase gene expression and secretion of CTGF/CCN2 in HPDLCs. CTGF/CCN2 upregulated the proliferation and migration of 1–11 cells. Furthermore, increased bone/cementum‐related gene expression in this cell line led to mineralization. In addition, combined treatment of 1–11 cells with CTGF/CCN2 and transforming growth factor‐β1 (TGF‐β1) significantly promoted type I collagen and fibronectin expression compared with that of TGF‐β1 treatment alone. Thus, these data suggest the underlying biphasic effects of CTGF/CCN2 in 1–11 cells, inducible osteo/cementoblastic, and fibroblastic differentiation dependent on the environmental condition. CTGF/CCN2 may contribute to preservation of the structural integrity of PDL tissue, implying its potential use as a therapeutic agent for PDL regeneration. J. Cell. Physiol. 230: 150–159, 2014. © 2014 Wiley Periodicals, Inc.
      PubDate: 2014-09-29T19:20:41.455063-05:
      DOI: 10.1002/jcp.24693
  • Intracellular MMP‐2 Activity in Skeletal Muscle Is Associated With
           Type II Fibers
    • Authors: Elin Hadler‐Olsen; Ann Iren Solli, Anne Hafstad, Jan‐Olof Winberg, Lars Uhlin‐Hansen
      First page: 160
      Abstract: Matrix metalloproteinase 2 (MMP‐2) is a proteolytic enzyme implicated in motility, differentiation, and regeneration of skeletal muscle fibers through processing of extracellular substrates. Although MMP‐2 has been found to be localized intracellularly in cardiomyocytes where the enzyme is thought to contribute to post‐ischemic loss of contractility, little is known about intracellular MMP‐2 activity in skeletal muscle fibers. In the present study we demonstrate intracellular MMP‐2 in normal skeletal muscle by immunohistochemical staining. Immunogold electron microscopic analyses indicated that the enzyme was concentrated in Z‐lines of the sarcomers, in the nuclear membrane, and in mitochondria. By use of in situ zymography, we found that gelatinolytic activity in muscle fibers was co‐localized with immunofluorecent staining for MMP‐2. Staining for MMP‐9, the other member of the gelatinase group of the MMPs, was negative. The broad‐spectrum metalloprotease inhibitor EDTA and the selective gelatinase inhibitor CTT2, but not the cysteine inhibitor E64, strongly reduced the gelatinolytic activity. The intracellular gelatinolytic activity was much more prominent in fast twitch type II fibers than in slow twitch type I fibers, and there was a decrease in intracellular gelatinolytic activity and MMP‐2 expression in muscles from mice exposed to high intensity interval training. Together our results indicate that MMP‐2 is part of the intracellular proteolytic network in normal skeletal muscle, especially in fast twitch type II fibers. Further, the results suggest that intracellular MMP‐2 in skeletal muscle fibers is active during normal homeostasis, and affected by the level of physical activity. J. Cell. Physiol. 230: 160–169, 2014. © 2014 Wiley Periodicals, Inc.
      PubDate: 2014-09-29T19:20:41.844391-05:
      DOI: 10.1002/jcp.24694
  • Nestin is a Marker of Lung Remodeling Secondary to Myocardial Infarction
           and Type I Diabetes in the Rat
    • Authors: Andréanne Chabot; Marc‐Andre Meus, Patrice Naud, Vanessa Hertig, Jocelyn Dupuis, Louis Villeneuve, Nabel El Khoury, Celine Fiset, Stanley Nattel, Jean‐Francois Jasmin, Angelino Calderone
      First page: 170
      Abstract: Upregulation of the intermediate filament protein nestin was identified in a subpopulation of fibroblasts during reactive and reparative fibrosis and directly contributed to the enhanced proliferative phenotype. The present study tested the hypothesis that nestin was expressed in lung fibroblasts and the pattern of expression represented a distinct marker of pulmonary remodeling secondary to myocardial infarction and type I diabetes. Nestin(+) fibroblasts were detected in rat lungs and a subpopulation exhibited a myofibroblast phenotype delineated by the co‐expression of smooth muscle α‐actin. In the lungs of myocardial infarcted rats, interstitial collagen content and nestin mRNA/protein levels were significantly increased despite the absence of secondary pulmonary hypertension, whereas smooth muscle α‐actin protein expression was unchanged. Exposure of rat pulmonary fibroblasts to pro‐fibrotic stimuli angiotensin II and transforming growth factor‐β significantly increased nestin protein levels. In the lungs of type I diabetic rats, the absence of a reactive fibrotic response was associated with a significant downregulation of nestin mRNA/protein expression. Nestin was reported a target of miR‐125b, albeit miR‐125b levels were unchanged in pulmonary fibroblasts treated with pro‐fibrotic stimuli. Nestin(+) cells lacking smooth muscle α‐actin/collagen staining were also identified in rodent lungs and a transgenic approach revealed that expression of the intermediate filament protein was driven by intron 2 of the nestin gene. The disparate regulation of nestin characterized a distinct pattern of pulmonary remodeling secondary to myocardial infarction and type I diabetes and upregulation of the intermediate filament protein in lung fibroblasts may have facilitated in part the reactive fibrotic response. J. Cell. Physiol. 230: 170–179, 2014. © 2014 Wiley Periodicals, Inc.
      PubDate: 2014-09-29T19:20:37.216127-05:
      DOI: 10.1002/jcp.24696
  • Enhancement of Runx2 Expression Is Potentially Linked to
           β‐Catenin Accumulation in Canine Intervertebral Disc
    • Authors: Munetaka Iwata; Takeshi Aikawa, Takaharu Hakozaki, Kiyotaka Arai, Hiroki Ochi, Hirotaka Haro, Masahiro Tagawa, Yoshinori Asou, Yasushi Hara
      First page: 180
      Abstract: Intervertebral disc degeneration (IVDD) greatly affects the quality of life. The nucleus pulposus (NP) of chondrodystrophic dog breeds (CDBs) is similar to the human NP because the cells disappear with age and are replaced by fibrochondrocyte‐like cells. Because IVDD develops as early as within the first year of life, we used canines as a model to investigate the in vitro mechanisms underlying IVDD. The mechanism underlying age‐related IVDD, however, is poorly understood. Several research groups have suggested that Wnt/β‐catenin signaling plays an important role in IVDD. However, the role of Wnt/β‐catenin signals in IVD cells is not yet well understood. Here, we demonstrate that Wnt/β‐catenin signaling could enhance Runx2 expression in IVDD and lead to IVD calcification. Nucleus pulposus (NP) tissue was obtained from Beagle dogs after evaluation of the degeneration based on magnetic resonance imaging (MRI). Histological analysis showed that lack of Safranin‐O staining, calcified area, and matrix metalloproteinase (MMP) 13‐positive cells increased with progression of the degeneration. Furthermore, the levels of β‐catenin‐ and Runx2‐positive cells also increased. Real‐time reverse‐transcription polymerase chain reaction analysis showed that the MRI signal intensity and mRNA expression levels of β‐catenin and Runx2 are correlated in NP tissues. Moreover, supplementation of LiCl induced β‐catenin accumulation and Runx2 expression. In contrast, FH535 inhibited LiCl‐induced upregulation. These results suggest that Runx2 transcript and protein expression, potentially in combination with β‐catenin accumulation, are enhanced in degenerated and calcified intervertebral discs of CDBs. J. Cell. Physiol. 230: 180–190, 2014. © 2014 Wiley Periodicals, Inc.
      PubDate: 2014-09-29T19:20:38.960554-05:
      DOI: 10.1002/jcp.24697
  • Loss of GLUT4 Induces Metabolic Reprogramming and Impairs Viability of
           Breast Cancer Cells
    • Authors: Pablo Garrido; Fernando G. Osorio, Javier Morán, Estefanía Cabello, Ana Alonso, José M.P. Freije, Celestino González
      First page: 191
      Abstract: Metabolic reprogramming strategies focus on the normalization of metabolism of cancer cells and constitute promising targets for cancer treatment. Here, we demonstrate that the glucose transporter 4 (GLUT4) has a prominent role in basal glucose uptake in MCF7 and MDA‐MB‐231 breast cancer cells. We show that shRNA‐mediated down‐regulation of GLUT4 diminishes glucose uptake and induces metabolic reprogramming by reallocating metabolic flux to oxidative phosphorylation. This reallocation is reflected on an increased activity of the mitochondrial oxidation of pyruvate and lower lactate release. Altogether, GLUT4 inhibition compromises cell proliferation and critically affects cell viability under hypoxic conditions, providing proof‐of‐principle for the feasibility of using pharmacological approaches to inhibit GLUT4 in order to induce metabolic reprogramming in vivo in breast cancer models. J. Cell. Physiol. 230: 191–198, 2014. © 2014 Wiley Periodicals, Inc.
      PubDate: 2014-09-29T19:20:35.7543-05:00
      DOI: 10.1002/jcp.24698
  • Ghrelin Augments the Expressions and Secretions of Proinflammatory
           Adipokines, VEGF120 and MCP‐1, in Differentiated 3T3‐L1
    • Authors: Atsuko Kitahara; Kazuto Takahashi, Rie Moriya, Hirohisa Onuma, Keiko Handa, Yoshikazu Sumitani, Toshiaki Tanaka, Hidenori Katsuta, Susumu Nishida, Takuya Sakurai, Kouichi Inukai, Hideki Ohno, Hitoshi Ishida
      First page: 199
      Abstract: Ghrelin is a physiological‐active peptide with growth hormone‐releasing activity, orexigenic activity, etc. In addition, the recent study has also suggested that ghrelin possesses the pathophysiological abilities related with type 2 diabetes. However, the ghrelin‐direct‐effects implicated in type 2 diabetes on peripheral tissues have been still unclear, whereas its actions on the central nervous system (CNS) appear to induce the development of diabetes. Thus, to assess its peripheral effects correlated with diabetes, we investigated the regulatory mechanisms about adipokines, which play a central role in inducing peripheral insulin resistance, secreted from mature 3T3‐L1 adipocytes stimulated with ghrelin in vitro . The stimulation with 50 nmol/L ghrelin for 24 h resulted in the significant 1.9‐fold increase on vascular endothelial growth factor‐120 (VEGF120) releases (p 
      PubDate: 2014-09-29T19:20:41.023424-05:
      DOI: 10.1002/jcp.24699
  • M‐CSF Priming of Osteoclast Precursors Can Cause
           Osteoclastogenesis‐Insensitivity, Which Can be Prevented and
           Overcome on Bone
    • Authors: Teun J. de Vries; Ton Schoenmaker, David Aerts, Lilyanne C. Grevers, Pedro P.C. Souza, Kamran Nazmi, Mark A. van de Wiel, Bauke Ylstra, Peter L. van Lent, Pieter J.M. Leenen, Vincent Everts
      First page: 210
      Abstract: Osteoclasts and macrophages share progenitors that must receive decisive lineage signals driving them into their respective differentiation routes. Macrophage colony stimulation factor M‐CSF is a common factor; bone is likely the stimulus for osteoclast differentiation. To elucidate the effects of both, shared mouse bone marrow precursor myeloid blast was precultured with M‐CSF on plastic and on bone. M‐CSF priming prior to stimulation with M‐CSF and osteoclast differentiation factor RANKL resulted in a complete loss of osteoclastogenic potential without bone. Such M‐CSF primed cells expressed the receptor RANK, but lacked the crucial osteoclastogenic transcription factor NFATc1. This coincided with a steeply decreased expression of osteoclast genes TRACP and DC‐STAMP, but an increased expression of the macrophage markers F4/80 and CD11b. Compellingly, M‐CSF priming on bone accelerated the osteoclastogenic potential: M‐CSF primed cells that had received only one day M‐CSF and RANKL and were grown on bone already expressed an array of genes that are associated with osteoclast differentiation and these cells differentiated into osteoclasts within 2 days. Osteoclastogenesis‐insensitive precursors grown in the absence of bone regained their osteoclastogenic potential when transferred to bone. This implies that adhesion to bone dictates the fate of osteoclast precursors. Common macrophage‐osteoclast precursors may become insensitive to differentiate into osteoclasts and regain osteoclastogenesis when bound to bone or when in the vicinity of bone. J. Cell. Physiol. 230: 210–225, 2014. © 2014 Wiley Periodicals, Inc.
      PubDate: 2014-09-29T19:20:42.618515-05:
      DOI: 10.1002/jcp.24702
  • Function of Latent TGFβ Binding Protein 4 and Fibulin 5 in
           Elastogenesis and Lung Development
    • Authors: Branka Dabovic; Ian B. Robertson, Lior Zilberberg, Melinda Vassallo, Elaine C. Davis, Daniel B. Rifkin
      First page: 226
      Abstract: Mice deficient in Latent TGFβ Binding Protein 4 (Ltbp4) display a defect in lung septation and elastogenesis. The lung septation defect is normalized by genetically decreasing TGFβ2 levels. However, the elastic fiber assembly is not improved in Tgfb2‐/‐;Ltbp4S‐/‐ compared to Ltbp4S‐/‐ lungs. We found that decreased levels of TGFβ1 or TGFβ3 did not improve lung septation indicating that the TGFβ isoform elevated in Ltbp4S‐/‐ lungs is TGFβ2. Expression of a form of Ltbp4 that could not bind latent TGFβ did not affect lung phenotype indicating that normal lung development does not require the formation of LTBP4‐latent TGFβ complexes. Therefore, the change in TGFβ level in the lungs is not directly related to Ltbp4 deficiency but probably is a consequence of changes in the extracellular matrix. Interestingly, combination of the Ltbp4S‐/‐ mutation with a fibulin‐5 null mutant in Fbln5‐/‐;Ltbp4S‐/‐ mice improves the lung septation compared to Ltbp4S‐/‐ lungs. Large globular elastin aggregates characteristic for Ltbp4S‐/‐ lungs do not form in Fbln5‐/‐;Ltbp4S‐/‐ lungs and EM studies showed that elastic fibers in Fbln5‐/‐;Ltbp4S‐/‐ lungs resemble those found in Fbln5‐/‐ mice. These results are consistent with a role for TGFβ2 in lung septation and for Ltbp4 in regulating fibulin‐5 dependent elastic fiber assembly. J. Cell. Physiol. 230: 226–236, 2014. © 2014 Wiley Periodicals, Inc.
      PubDate: 2014-09-29T19:20:43.198254-05:
      DOI: 10.1002/jcp.24704
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