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Sample records for epithelial growth factor

  1. Epithelial expression of keratinocytes growth factor in oral precancer lesions

    PubMed Central

    Jimson, Sudha; Murali, S.; Zunt, Susan L.; Goldblatt, Lawrence I.; Srinivasan, Mythily

    2016-01-01

    Background: Keratinocyte growth factor (KGF) is a potent epithelial mitogen that acts by binding the KGF receptors (KGFRs) expressed on epithelial cells and regulates proliferation and differentiation. The objective of this study was to investigate the expression of KGF in the epithelium in oral precancer. Materials and Methods: Archival tissues of oral submucous fibrosis (SMF) and leukoplakia were assessed for epithelial KGF expression by immunohistochemistry and real-time quantitative polymerase chain reaction. Results: KGF was predominantly expressed in the basal and parabasal cells in the epithelium of SMF tissues. KGF transcript in the epithelial cells increased with increasing severity of epithelial dysplasia in oral leukoplakia. Conclusion: Although widely reported as a product secreted by the mesenchymal cells, our data suggest that the KGF is also expressed in oral epithelial cells much like the expression in ovarian epithelial cells. Based on the localization of KGF in cells at the epithelial mesenchymal junction and that of the reported presence of KGFR in oral keratinocytes, a potential mechanism involving paracrine and autocrine interactions of KGF and KGFR in early stages of oral precancer is postulated. PMID:27274338

  2. Bone morphogenetic protein-4 strongly potentiates growth factor-induced proliferation of mammary epithelial cells

    SciTech Connect

    Montesano, Roberto Sarkoezi, Rita; Schramek, Herbert

    2008-09-12

    Bone morphogenetic proteins (BMPs) are multifunctional cytokines that elicit pleiotropic effects on biological processes such as cell proliferation, cell differentiation and tissue morphogenesis. With respect to cell proliferation, BMPs can exert either mitogenic or anti-mitogenic activities, depending on the target cells and their context. Here, we report that in low-density cultures of immortalized mammary epithelial cells, BMP-4 did not stimulate cell proliferation by itself. However, when added in combination with suboptimal concentrations of fibroblast growth factor (FGF)-2, FGF-7, FGF-10, epidermal growth factor (EGF) or hepatocyte growth factor (HGF), BMP-4 potently enhanced growth factor-induced cell proliferation. These results reveal a hitherto unsuspected interplay between BMP-4 and growth factors in the regulation of mammary epithelial cell proliferation. We suggest that the ability of BMP-4 to potentiate the mitogenic activity of multiple growth factors may contribute to mammary gland ductal morphogenesis as well as to breast cancer progression.

  3. Effect of Growth Factors on the Proliferation and Gene Expression of Human Meibomian Gland Epithelial Cells

    PubMed Central

    Liu, Shaohui; Kam, Wendy R.; Ding, Juan; Hatton, Mark P.; Sullivan, David A.

    2013-01-01

    Purpose. We hypothesize that growth factors, including epidermal growth factor (EGF) and bovine pituitary extract (BPE), induce proliferation, but not differentiation (e.g., lipid accumulation), of human meibomian gland epithelial cells. We also hypothesize that these actions involve a significant upregulation of genes linked to cell cycle processes, and a significant downregulation of genes associated with differentiation. Our objective was to test these hypotheses. Methods. Immortalized human meibomian gland and conjunctival epithelial cells were cultured for varying time periods in the presence or absence of EGF, BPE, EGF + BPE, or serum, followed by cell counting, neutral lipid staining, or RNA isolation for molecular biological procedures. Results. Our studies show that growth factors stimulate a significant, time-dependent proliferation of human meibomian gland epithelial cells. These effects are associated with a significant upregulation of genes linked to cell cycle, DNA replication, ribosomes, and translation, and a significant decrease in those related to cell differentiation, tissue development, lipid metabolic processes, and peroxisome proliferator-activated receptor signaling. Serum-induced differentiation, but not growth factor-related proliferation, elicits a pronounced lipid accumulation in human meibomian gland epithelial cells. This lipogenic response is unique, and is not duplicated by human conjunctival epithelial cells. Conclusions. Our results demonstrate that EGF and BPE stimulate human meibomian gland epithelial cells to proliferate. Further, our findings show that action is associated with an upregulation of cell cycle and translation ontologies, and a downregulation of genetic pathways linked to differentiation and lipid biosynthesis. PMID:23493293

  4. Intracellular insulin-like growth factor-1 induces Bcl-2 expression in airway epithelial cells.

    PubMed

    Chand, Hitendra S; Harris, Jennifer Foster; Mebratu, Yohannes; Chen, Yangde; Wright, Paul S; Randell, Scott H; Tesfaigzi, Yohannes

    2012-05-01

    Bcl-2, a prosurvival protein, regulates programmed cell death during development and repair processes, and it can be oncogenic when cell proliferation is deregulated. The present study investigated what factors modulate Bcl-2 expression in airway epithelial cells and identified the pathways involved. Microarray analysis of mRNA from airway epithelial cells captured by laser microdissection showed that increased expression of IL-1β and insulin-like growth factor-1 (IGF-1) coincided with induced Bcl-2 expression compared with controls. Treatment of cultured airway epithelial cells with IL-1β and IGF-1 induced Bcl-2 expression by increasing Bcl-2 mRNA stability with no discernible changes in promoter activity. Silencing the IGF-1 expression using short hairpin RNA showed that intracellular IGF-1 (IC-IGF-1) was increasing Bcl-2 expression. Blocking epidermal growth factor receptor or IGF-1R activation also suppressed IC-IGF-1 and abolished the Bcl-2 induction. Induced expression and colocalization of IC-IGF-1 and Bcl-2 were observed in airway epithelial cells of mice exposed to LPS or cigarette smoke and of patients with cystic fibrosis and chronic bronchitis but not in the respective controls. These studies demonstrate that IC-IGF-1 induces Bcl-2 expression in epithelial cells via IGF-1R and epidermal growth factor receptor pathways, and targeting IC-IGF-1 could be beneficial to treat chronic airway diseases. PMID:22461702

  5. Hepatocyte Growth Factor Inhibits Epithelial to Myofibroblast Transition in Lung Cells via Smad7

    PubMed Central

    Shukla, Manasi N.; Rose, Jane L.; Ray, Rabindranath; Lathrop, Kira L.; Ray, Anuradha; Ray, Prabir

    2009-01-01

    Idiopathic pulmonary fibrosis is a lethal parenchymal lung disease characterized by denudation of the lung epithelium, fibroblast proliferation, and collagen deposition. Cellular changes underlying disease progression involve injury to alveolar epithelial cells, epithelial to mesenchymal transition, proliferation of α-smooth muscle actin (α-SMA)–expressing myofibroblasts and of fibroblasts resulting in enhanced deposition of extracellular matrix proteins. Hepatocyte growth factor (HGF) inhibits progression of bleomycin-induced pulmonary fibrosis in mice. The mechanism underlying the inhibitory effect of HGF was investigated in an in vitro model. We show that HGF markedly antagonizes basal and transforming growth factor (TGF)-β–induced expression of myofibroblast markers such as α-SMA, collagen type 1, and fibronectin in rat alveolar epithelial cells. HGF also inhibited TGF-β–induced α-SMA expression in primary murine alveolar epithelial cells. Since TGF-β is known to regulate α-SMA expression, the effect of HGF on components of TGF-β signaling was investigated. HGF induced expression of Smad7, an inhibitor of TGF-β signaling, in a mitogen-activated protein kinase–dependent manner. HGF also induced the nuclear export of Smad7 and Smad ubiquitin regulatory factor 1 (Smurf1) to the cytoplasm. HGF-dependent decrease in α-SMA was abolished with specific siRNAs targeted to Smad7. Thus, induction of Smad7 by HGF serves to limit acquisition of the myofibroblast phenotype in alveolar epithelial cells. PMID:18988920

  6. Transforming Growth Factor-β2 Induces Bronchial Epithelial Mucin Expression in Asthma

    PubMed Central

    Chu, Hong Wei; Balzar, Silvana; Seedorf, Gregory J.; Westcott, Jay Y.; Trudeau, John B.; Silkoff, Phil; Wenzel, Sally E.

    2004-01-01

    The transforming growth factor (TGF)-β family is important for tissue repair in pathological conditions including asthma. However, little is known about the impact of either TGF-β1 or TGF-β2 on asthmatic airway epithelial mucin expression. We evaluated bronchial epithelial TGF-β1 and TGF-β2 expression and their effects on mucin expression, and the role of TGF-β1 or TGF-β2 in interleukin (IL)-13-induced mucin expression. Epithelial TGF-β1, TGF-β2, and mucin expression were evaluated in endobronchial biopsies from asthmatics and normal subjects. The effects of TGF-β1 or TGF-β2 on mucin MUC5AC protein and mRNA expression, and the impact of IL-13 on epithelial TGF-β1, TGF-β2, and MUC5AC were determined in cultured bronchial epithelial cells from endobronchial brushings of both subject groups. In biopsy tissue, epithelial TGF-β2 expression levels were higher than TGF-β1 in both asthmatics and normals. TGF-β2, but not TGF-β1, was increased in asthmatics compared with normals, and significantly correlated with mucin expression. TGF-β2, but not TGF-β1, increased mucin expression in cultured epithelial cells from both subject groups. IL-13 increased the release of TGF-β2, but not TGF-β1, from epithelial cells. A neutralizing TGF-β2 antibody partially inhibited IL-13-induced mucin expression. These data suggest that TGF-β2 production by asthmatic bronchial epithelial cells may increase airway mucin expression. IL-13-induced mucin expression may occur in part through TGF-β2 up-regulation. PMID:15466377

  7. Transforming growth factor-beta2 induces bronchial epithelial mucin expression in asthma.

    PubMed

    Chu, Hong Wei; Balzar, Silvana; Seedorf, Gregory J; Westcott, Jay Y; Trudeau, John B; Silkoff, Phil; Wenzel, Sally E

    2004-10-01

    The transforming growth factor (TGF)-beta family is important for tissue repair in pathological conditions including asthma. However, little is known about the impact of either TGF-beta1 or TGF-beta2 on asthmatic airway epithelial mucin expression. We evaluated bronchial epithelial TGF-beta1 and TGF-beta2 expression and their effects on mucin expression, and the role of TGF-beta1 or TGF-beta2 in interleukin (IL)-13-induced mucin expression. Epithelial TGF-beta1, TGF-beta2, and mucin expression were evaluated in endobronchial biopsies from asthmatics and normal subjects. The effects of TGF-beta1 or TGF-beta2 on mucin MUC5AC protein and mRNA expression, and the impact of IL-13 on epithelial TGF-beta1, TGF-beta2, and MUC5AC were determined in cultured bronchial epithelial cells from endobronchial brushings of both subject groups. In biopsy tissue, epithelial TGF-beta2 expression levels were higher than TGF-beta1 in both asthmatics and normals. TGF-beta2, but not TGF-beta1, was increased in asthmatics compared with normals, and significantly correlated with mucin expression. TGF-beta2, but not TGF-beta1, increased mucin expression in cultured epithelial cells from both subject groups. IL-13 increased the release of TGF-beta2, but not TGF-beta1, from epithelial cells. A neutralizing TGF-beta2 antibody partially inhibited IL-13-induced mucin expression. These data suggest that TGF-beta2 production by asthmatic bronchial epithelial cells may increase airway mucin expression. IL-13-induced mucin expression may occur in part through TGF-beta2 up-regulation. PMID:15466377

  8. Involvement of Connective Tissue Growth Factor (CTGF) in Insulin-like Growth Factor-I (IGF1) Stimulation of Proliferation of a Bovine Mammary Epithelial Cell Line

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Insulin-like growth factor I (IGF1) plays an important role in mammary gland development and lactation in part by stimulating proliferation of the milk-producing epithelial cells. In this study, we used the bovine mammary epithelial cell line MAC-T cells as a model to understand the mechanism by whi...

  9. Growth factor induced proliferation, migration, and lumen formation of rat endometrial epithelial cells in vitro.

    PubMed

    Islam, Md Rashedul; Yamagami, Kazuki; Yoshii, Yuka; Yamauchi, Nobuhiko

    2016-06-17

    Endometrial modulation is essential for the preservation of normal uterine physiology, and this modulation is driven by a number of growth factors. The present study investigated the mitogenic, motogenic, and morphogenic effects of epidermal growth factor (EGF) and hepatocyte growth factor (HGF) on rat endometrial epithelial (REE) cells. The REE cells were isolated and cultured and then characterized based on their morphology and their expression of epithelial cell markers. The MTT assay revealed that EGF and HGF induce proliferation of REE cells. Consistent with increased proliferation, we found that the cell cycle regulatory factor Cyclin D1 was also upregulated upon EGF and HGF addition. REE cell migration was prompted by EGF, as observed with the Oris Cell Migration Assay. The morphogenic impact of growth factors on REE cells was studied in a three-dimensional BD Matrigel cell culture system, wherein these growth factors also increased the frequency of lumen formation. In summary, we show that EGF and HGF have a stimulatory effect on REE cells, promoting proliferation, cell migration, and lumen formation. Our findings provide important insights that further the understanding of endometrial regeneration and its regulation. PMID:26946922

  10. Growth factor induced proliferation, migration, and lumen formation of rat endometrial epithelial cells in vitro

    PubMed Central

    ISLAM, Md. Rashedul; YAMAGAMI, Kazuki; YOSHII, Yuka; YAMAUCHI, Nobuhiko

    2016-01-01

    Endometrial modulation is essential for the preservation of normal uterine physiology, and this modulation is driven by a number of growth factors. The present study investigated the mitogenic, motogenic, and morphogenic effects of epidermal growth factor (EGF) and hepatocyte growth factor (HGF) on rat endometrial epithelial (REE) cells. The REE cells were isolated and cultured and then characterized based on their morphology and their expression of epithelial cell markers. The MTT assay revealed that EGF and HGF induce proliferation of REE cells. Consistent with increased proliferation, we found that the cell cycle regulatory factor Cyclin D1 was also upregulated upon EGF and HGF addition. REE cell migration was prompted by EGF, as observed with the Oris Cell Migration Assay. The morphogenic impact of growth factors on REE cells was studied in a three-dimensional BD Matrigel cell culture system, wherein these growth factors also increased the frequency of lumen formation. In summary, we show that EGF and HGF have a stimulatory effect on REE cells, promoting proliferation, cell migration, and lumen formation. Our findings provide important insights that further the understanding of endometrial regeneration and its regulation. PMID:26946922

  11. Differential growth factor induction and modulation of human gastric epithelial regeneration

    SciTech Connect

    Tetreault, Marie-Pier; Chailler, Pierre; Rivard, Nathalie; Menard, Daniel . E-mail: Daniel.Menard@USherbrooke.ca

    2005-05-15

    While several autocrine/paracrine growth factors (GFs) can all stimulate epithelial regeneration in experimentally wounded primary gastric cultures, clinical relevance for their non-redundant cooperative actions in human gastric ulcer healing is suggested by the sequential pattern of GF gene induction in vivo. Using new HGE cell lines able to form a coherent monolayer with tight junctions as well as using primary human gastric epithelial cultures, we show that EGF, TGF{alpha}, HGF and IGFs accelerate epithelial restitution upon wounding, independently of the TGF{beta} pathway (as opposed to intestinal cells). However, they differently modulate cell behavior: TGF{alpha} exerts strong effects (even more than EGF) on cytoplasmic spreading and non-oriented protruding activity of bordering cells whereas HGF preferentially coordinates single lamella formation, cell elongation and migration into the wound. IGF-I and IGF-II rather induce the alignment of bordering cells and maintain a compact monolayer front. The number of mitotic cells maximally increases with EGF, followed by TGF{alpha} and IGF-I,-II. The current study demonstrates that GFs differentially regulate the regeneration of human gastric epithelial cells through specific modulation of cell shape adaptation, migration and proliferation, further stressing that a coordination of GF activities would be necessary for the normal progression of post-wounding epithelial repair.

  12. Trafficking of epidermal growth factor receptor ligands in polarized epithelial cells.

    PubMed

    Singh, Bhuminder; Coffey, Robert J

    2014-01-01

    A largely unilamellar epithelial layer lines body cavities and organ ducts such as the digestive tract and kidney tubules. This polarized epithelium is composed of biochemically and functionally separate apical and basolateral surfaces. The epidermal growth factor receptor (EGFR) signaling pathway is a critical regulator of epithelial homeostasis and is perturbed in a number of epithelial disorders. It is underappreciated that in vivo EGFR signaling is most often initiated by cell-surface delivery and processing of one of seven transmembrane ligands, resulting in release of the soluble form that binds EGFR. In polarized epithelial cells, EGFR is restricted largely to the basolateral surface, and apical or basolateral ligand delivery therefore has important biological consequences. In vitro approaches have been used to study the biosynthesis, cell-surface delivery, proteolytic processing, and release of soluble EGFR ligands in polarized epithelial cells. We review these results, discuss their relevance to normal physiology, and demonstrate the pathophysiological consequences of aberrant trafficking. These studies have uncovered a rich diversity of apico-basolateral trafficking mechanisms among the EGFR ligands, provided insights into the pathogenesis of an inherited magnesium-wasting disorder of the kidney (isolated renal hypomagnesemia), and identified a new mode of EGFR ligand signaling via exosomes. PMID:24215440

  13. Trafficking of Epidermal Growth Factor Receptor Ligands in Polarized Epithelial Cells

    PubMed Central

    Singh, Bhuminder; Coffey, Robert J.

    2014-01-01

    A largely unilamellar epithelial layer lines body cavities and organ ducts such as the digestive tract and kidney tubules. This polarized epithelium is composed of biochemically and functionally separate apical and basolateral surfaces. The epidermal growth factor receptor (EGFR) signaling pathway is a critical regulator of epithelial homeostasis and is perturbed in a number of epithelial disorders. It is underappreciated that in vivo EGFR signaling is most often initiated by cell-surface delivery and processing of one of seven transmembrane ligands, resulting in release of the soluble form that binds EGFR. In polarized epithelial cells, EGFR is restricted largely to the basolateral surface, and apical or basolateral ligand delivery therefore has important biological consequences. In vitro approaches have been used to study the biosynthesis, cell-surface delivery, proteolytic processing, and release of soluble EGFR ligands in polarized epithelial cells. We review these results, discuss their relevance to normal physiology, and demonstrate the pathophysiological consequences of aberrant trafficking. These studies have uncovered a rich diversity of apico-basolateral trafficking mechanisms among the EGFR ligands, provided insights into the pathogenesis of an inherited magnesium-wasting disorder of the kidney (isolated renal hypomagnesemia), and identified a new mode of EGFR ligand signaling via exosomes. PMID:24215440

  14. Keratinocyte growth factor 1 inhibits wound edge epithelial cell apoptosis in vitro.

    PubMed

    Firth, James D; Putnins, Edward E

    2004-01-01

    The ability of keratinocyte growth factor 1 to modulate apoptosis in the absence of proliferation was studied in vitro. A HaCaT scrape wound model was developed in which dense monolayers prior to wounding were cultured to quiescence in defined media with hydroxyurea at concentrations that blocked proliferation without loss of cell viability. Scrape wounding was then found to induce apoptosis, originating at the wound edge, but subsequently radiating away over a 24 h period to encompass areas not originally damaged. Keratinocyte growth factor 1 inhibited this radial progression of apoptosis in a concentration-dependent manner up to 20 ng per mL with induced migration present at the wound edge. The extent of this rescue was modulated by the concentration of Ca2+ prior to wounding. In control wound cultures apoptotic bodies were found in cells adjacent to the wound interface but were greatly reduced in keratinocyte-growth-factor-1-treated groups. Keratinocyte growth factor 1 receptor expression was significantly induced within two to three cell widths of the scraped wound edge, at levels far exceeding those found at the leading edge of a nonwounded epithelial sheet. Tumor necrosis factor alpha (1-5 ng per mL) or Escherichia coli lipopolysaccharide (10-50 ng per mL) exacerbated scrape-induced early apoptosis (1-4 h), but was largely ameliorated by coculture with keratinocyte growth factor 1. Keratinocyte growth factor 1 protection was associated with a reduction in both caspase-3 activation and cytokeratin-19 loss. Protected wound edges were also associated with the maintenance of e-cadherin expression and induction of beta1 integrin and actin stress fiber organization. These results suggest that keratinocyte growth factor 1 may play a role in limiting mechanically induced apoptotic processes at the epithelial wound edge in a manner that is distinct from its proliferative function. PMID:14962112

  15. Human papilloma virus DNAs immortalize normal human mammary epithelial cells and reduce their growth factor requirements

    SciTech Connect

    Band, V.; Zajchowski, D.; Kulesa, V.; Sager, R. )

    1990-01-01

    Human papilloma virus (HPV) types 16 and 18 are most commonly associated with cervical carcinoma in patients and induce immortalization of human keratinocytes in culture. HPV has not been associated with breast cancer. This report describes the immortalization of normal human mammary epithelial cells (76N) by plasmid pHPV18 or pHPV16, each containing the linearized viral genome. Transfectants were grown continuously for more than 60 passages, whereas 76N cells senesce after 18-20 passages. The transfectants also differ from 76N cells in cloning in a completely defined medium called D2 and growing a minimally supplemented defined medium (D3) containing epidermal growth factor. All transfectant tested contain integrated HPV DNA, express HPV RNA, and produce HPV E7 protein. HPV transfectants do not form tumors in a nude mouse assay. It is concluded that products of the HPV genome induce immortalization of human breast epithelial cells and reduce their growth factor requirements. This result raises the possibility that HPV might be involved in breast cancer. Furthermore, other tissue-specific primary epithelial cells that are presently difficult to grown and investigate may also be immortalized by HPV.

  16. Vasopressin regulation of epithelial colonic proliferation and permeability is mediated by pericryptal platelet-derived growth factor A.

    PubMed

    Miró, Lluïsa; Pérez-Bosque, Anna; Maijó, Mònica; Naftalin, Richard J; Moretó, Miquel

    2014-10-01

    Arginine vasopressin (AVP) has trophic effects on the rat distal colon, increasing the growth of pericryptal myofibroblasts and reducing the colonic crypt wall permeability. This study aimed to reproduce in vitro the effects of AVP observed in vivo using cultures of human CCD-18Co myofibroblasts and T84 colonic epithelial cells. Proliferation of myofibroblasts was quantified by bromodeoxyuridine incorporation; the expression of platelet-derived growth factor A (PDGFA), platelet-derived growth factor B, epidermal growth factor, transforming growth factor-β and vascular endothelial growth factor was measured by PCR and the expression of epithelial junction proteins by Western blot. Arginine vasopressin stimulated myofibroblast proliferation and the expression of PDGFA without affecting the expression of platelet-derived growth factor B, epidermal growth factor, transforming growth factor-β or vascular endothelial growth factor. These effects were prevented when AVP receptor inhibitors were present in the medium. Pre-incubation of CCD-18Co cells with anti-PDGF antibody or with an inhibitor of the PDGF receptor abolished the effects of AVP. When colonocytes were incubated with medium obtained from myofibroblasts incubated with AVP, both cell proliferation and the expression of epithelial junction proteins increased; however, direct incubation of colonocytes with AVP did not modify these variables. These results demonstrate that AVP stimulates myofibroblast proliferation and induces PDGFA secretion, implying that PDGFA mediates local myofibroblast proliferation by an autocrine feedback loop and regulates epithelial proliferation and permeability by a paracrine mechanism. PMID:25085844

  17. Controlled-release of epidermal growth factor from cationized gelatin hydrogel enhances corneal epithelial wound healing.

    PubMed

    Hori, Kuniko; Sotozono, Chie; Hamuro, Junji; Yamasaki, Kenta; Kimura, Yu; Ozeki, Makoto; Tabata, Yasuhiko; Kinoshita, Shigeru

    2007-04-01

    We designed a new ophthalmic drug-delivery system for epidermal growth factor (EGF) from the biodegradable hydrogel of cationized gelatin. We placed a cationized gelatin hydrogel (CGH) with incorporated (125)I-labelled EGF in the conjunctival sac of mice and measured the residual radioactivity at different times to evaluate the in vivo profile of EGF release. Approximately 60-67% and 10-12% of EGF applied initially remained 1 and 7 days after application, respectively; whereas EGF delivered in topically applied solution or via EGF impregnation of soft contact lenses disappeared within the first day. We also placed CGH films with 5.0 mug of incorporated EGF on round corneal defects in rabbits to evaluate the healing process using image analysis software and to assess epithelial proliferation immunohistochemically by counting the number of Ki67-positive cells. The application of a CGH film with incorporated EGF resulted in a reduction in the epithelial defect in rabbit corneas accompanied by significantly enhanced epithelial proliferation compared with the reduction seen after the topical application of EGF solution or the placement of an EGF-free CGH film. The controlled release of EGF from a CGH placed over a corneal epithelial defect accelerated ocular surface wound healing. PMID:17289206

  18. Stromal inhibition of prostatic epithelial cell proliferation not mediated by transforming growth factor beta.

    PubMed Central

    Kooistra, A.; van den Eijnden-van Raaij, A. J.; Klaij, I. A.; Romijn, J. C.; Schröder, F. H.

    1995-01-01

    The paracrine influence of prostatic stroma on the proliferation of prostatic epithelial cells was investigated. Stromal cells from the human prostate have previously been shown to inhibit anchorage-dependent as well as anchorage-independent growth of the prostatic tumour epithelial cell lines PC-3 and LNCaP. Antiproliferative activity, mediated by a diffusible factor in the stromal cell conditioned medium, was found to be produced specifically by prostatic stromal cells. In the present study the characteristics of this factor were examined. It is demonstrated that prostate stroma-derived inhibiting factor is an acid- and heat-labile, dithiothreitol-sensitive protein. Although some similarities with type beta transforming growth factor (TGF-beta)-like inhibitors are apparent, evidence is presented that the factor is not identical to TGF-beta or to the TGF-beta-like factors activin and inhibin. Absence of TGF-beta activity was shown by the lack of inhibitory response of the TGF-beta-sensitive mink lung cell line CCL-64 to prostate stromal cell conditioned medium and to concentrated, partially purified preparations of the inhibitor. Furthermore, neutralising antibodies against TGF-beta 1 or TGF-beta 2 did not cause a decline in the level of PC-3 growth inhibition caused by partially purified inhibitor. Using Northern blot analyses, we excluded the involvement of inhibin or activin. It is concluded that the prostate stroma-derived factor may be a novel growth inhibitor different from any of the currently described inhibiting factors. Images Figure 5 PMID:7543773

  19. Autocrine ligands of the epithelial growth factor receptor mediate inflammatory responses to diesel exhaust particles

    PubMed Central

    2014-01-01

    Background Diesel exhaust is associated with cardiovascular and respiratory mortality and morbidity. Acute exposure leads to increased IL-8 expression and airway neutrophilia, however the mechanism of this response is unknown. Objectives: As cigarette smoke-induced IL-8 expression by epithelial cells involves transactivation of the epidermal growth factor receptor (EGFR), we studied the effects of diesel exhaust particles (DEP) on IL-8 release and the role of the EGFR. Methods Primary bronchial epithelial cells (PBEC) were exposed to DEPs or carbon black. IL-8 and EGFR ligand expression (transforming growth factor alpha (TGFα), heparin-binding EGF-like growth factor, and amphiregulin (AR)) were assessed by quantitative RT-PCR and ELISA. Results DEP, but not carbon black, caused a dose-dependent increase in mitogen-activated protein kinase (MAPK) activation and IL-8 expression, however above 50 μg/ml there was an increase in cytotoxicity. At 50 μg/ml, DEPs stimulated transcription and release of IL-8 and EGFR ligands. IL-8 release was blocked by EGFR neutralizing antibodies, an EGFR-selective tyrosine kinase inhibitor and by the metalloprotease inhibitor, GM6001, which blocks EGFR ligand shedding. Neutralizing antibodies to AR, TGFα and heparin-binding (HB)-EGF reduced DEP-induced IL-8 by >50%. Conclusion Expression of IL-8 in response to DEPs is dependent on EGFR activation and that autocrine production of EGFR ligands makes a substantial contribution to this response. Capsule Summary: This study identifies a mechanism whereby diesel particles stimulates IL-8 release from bronchial epithelial cells. This mechanism may help to explain the recruitment of neutrophils into the airways of people exposed to particulate air pollution. PMID:24555532

  20. Analysis of Epithelial-Mesenchymal Transition Induced by Transforming Growth Factor β.

    PubMed

    Valcourt, Ulrich; Carthy, Jonathon; Okita, Yukari; Alcaraz, Lindsay; Kato, Mitsuyasu; Thuault, Sylvie; Bartholin, Laurent; Moustakas, Aristidis

    2016-01-01

    In recent years, the importance of the cell biological process of epithelial-mesenchymal transition (EMT) has been established via an exponentially growing number of reports. EMT has been documented during embryonic development, tissue fibrosis, and cancer progression in vitro, in animal models in vivo and in human specimens. EMT relates to many molecular and cellular alterations that occur when epithelial cells undergo a switch in differentiation that generates mesenchymal-like cells with newly acquired migratory and invasive properties. In addition, EMT relates to a nuclear reprogramming similar to the one occurring in the generation of induced pluripotent stem cells. Via such a process, EMT is gradually established to promote the generation and maintenance of adult tissue stem cells which under disease states such as cancer, are known as cancer stem cells. EMT is induced by developmental growth factors, oncogenes, radiation, and hypoxia. A prominent growth factor that causes EMT is transforming growth factor β (TGF-β).A series of molecular and cellular techniques can be applied to define and characterize the state of EMT in diverse biological samples. These methods range from DNA and RNA-based techniques that measure the expression of key EMT regulators and markers of epithelial or mesenchymal differentiation to functional assays of cell mobility, invasiveness and in vitro stemness. This chapter focuses on EMT induced by TGF-β and provides authoritative protocols and relevant reagents and citations of key publications aiming at assisting newcomers that enter this prolific area of biomedical sciences, and offering a useful reference tool to pioneers and aficionados of the field. PMID:26520123

  1. Transforming growth factor-beta during carcinogenesis: the shift from epithelial to mesenchymal signaling.

    PubMed

    Matsuzaki, Koichi; Okazaki, Kazuichi

    2006-04-01

    Transforming growth factor-beta (TGF-beta) activates not only TGF-beta type I receptor (TbetaRI) but also c-Jun N-terminal kinase (JNK), changing unphosphorylated Smad3 to its phosphoisoforms: C-terminally phosphorylated Smad3 (pSmad3C) and linker phosphorylated Smad3 (pSmad3L). While the TbetaRI/pSmad3C pathway inhibits growth of normal epithelial cells, JNK/pSmad3L-mediated signaling is involved in invasion by activated mesenchymal cells. During sporadic human colorectal carcinogenesis, TGF-beta signaling confers a selective advantage on tumor cells by shifting from the TbetaRI/pSmad3C pathway characteristic of mature epithelial cells to the JNK/pSmad3L pathway, which is more characteristic of the state of flux shown by the activated mesenchymal cells. JNK acts as a regulator of TGF-beta signaling by increasing the basal level of pSmad3L available for action in the nuclei of the invasive adenocarcinoma, in the meantime shutting down TGF-beta-dependent nuclear activity of pSmad3C. Loss of epithelial homeostasis and acquisition of a migratory, mesenchymal phenotype are essential for tumor invasion. From the viewpoint of TGF-beta signaling, a key therapeutic aim in cancer would be restoration of the lost tumor suppressor function observed in normal colorectal epithelial cells at the expense of effects promoting aggressive behavior of the adenocarcinoma. Specific inhibitors of the JNK/pSmad3L pathway might prove useful in this respect. In the case of molecularly targeted therapy for human cancer, pSmad3L and pSmad3C could be assessed as biomarkers to evaluate the likely benefit from specific inhibition of the JNK/pSmad3L pathway. PMID:16741607

  2. Keratinocyte growth factor induces proliferation of hepatocytes and epithelial cells throughout the rat gastrointestinal tract.

    PubMed Central

    Housley, R M; Morris, C F; Boyle, W; Ring, B; Biltz, R; Tarpley, J E; Aukerman, S L; Devine, P L; Whitehead, R H; Pierce, G F

    1994-01-01

    Keratinocyte growth factor (KGF), a member of the fibroblast growth factor (FGF) family, was identified as a specific keratinocyte mitogen after isolation from a lung fibroblast line. Recently, recombinant (r)KGF was found to influence proliferation and differentiation patterns of multiple epithelial cell lineages within skin, lung, and the reproductive tract. In the present study, we designed experiments to identify additional target tissues, and focused on the rat gastrointestinal (GI) system, since a putative receptor, K-sam, was originally identified in a gastric carcinoma. Expression of KGF receptor and KGF mRNA was detected within the entire GI tract, suggesting the gut both synthesized and responded to KGF. Therefore, rKGF was administered to adult rats and was found to induce markedly increased proliferation of epithelial cells from the foregut to the colon, and of hepatocytes, one day after systemic treatment. Daily treatment resulted in the marked selective induction of mucin-producing cell lineages throughout the GI tract in a dose-dependent fashion. Other cell lineages were either unaffected (e.g., Paneth cells), or relatively decreased (e.g., parietal cells, enterocytes) in rKGF-treated rats. The direct effect of rKGF was confirmed by demonstrating markedly increased carcinoembryonic antigen production in a human colon carcinoma cell line, LIM1899. Serum levels of albumin were specifically and significantly elevated after daily treatment. These results demonstrate rKGF can induce epithelial cell activation throughout the GI tract and liver. Further, endogenous KGF may be a normal paracrine mediator of growth within the gut. Images PMID:7962522

  3. The Role of Endogenous Epidermal Growth Factor Receptor Ligands in Mediating Corneal Epithelial Homeostasis

    PubMed Central

    Peterson, Joanne L.; Phelps, Eric D.; Doll, Mark A.; Schaal, Shlomit; Ceresa, Brian P.

    2014-01-01

    Purpose. To provide a comprehensive study of the biological role and therapeutic potential of six endogenous epidermal growth factor receptor (EGFR) ligands in corneal epithelial homeostasis. Methods. Kinetic analysis and dose response curves were performed by using in vitro and in vivo wound-healing assays. Biochemical assays were used to determine receptor expression and activity. Human tears were collected and quantitatively analyzed by multianalyte profiling for endogenous EGFR ligands. Results. Epidermal growth factor receptor ligands improved wound closure and activated EGFR, but betacellulin (BTC) was the most efficacious promoter of wound healing in vitro. In contrast, only epidermal growth factor (EGF) promoted wound healing in vivo. Human tears from 25 healthy individuals showed EGFR ligands at these average concentrations: EGF at 2053 ± 312.4 pg/mL, BTC at 207 ± 39.4 pg/mL, heparin-binding EGF at 44 ± 5.8 pg/mL, amphiregulin at 509 ± 28.8 pg/mL, transforming growth factor-α at 84 ± 19 pg/mL, and epiregulin at 52 ± 15 pg/mL. Conclusions. Under unwounded conditions, only EGF was present at concentrations near the ligand's Kd for the receptor, indicating it is the primary mediator of corneal epithelial homeostasis. Other ligands were present but at concentrations 11- to 7500-fold less their Kd, preventing significant ligand binding. Further, the high levels of EGF and its predicted binding preclude receptor occupancy by exogenous ligand and can explain the discrepancy between the in vitro and in vivo data. Therefore, therapeutic use of EGFR ligands may be unpredictable and impractical. PMID:24722692

  4. Transforming growth factor-beta stimulates epithelial-mesenchymal transformation in the proepicardium.

    PubMed

    Olivey, Harold E; Mundell, Nathan A; Austin, Anita F; Barnett, Joey V

    2006-01-01

    The proepicardium (PE) migrates over the heart and forms the epicardium. A subset of these PE-derived cells undergoes epithelial-mesenchymal transformation (EMT) and gives rise to cardiac fibroblasts and components of the coronary vasculature. We report that transforming growth factor-beta (TGFbeta) 1 and TGFbeta2 increase EMT in PE explants as measured by invasion into a collagen gel, loss of cytokeratin expression, and redistribution of ZO1. The type I TGFbeta receptors ALK2 and ALK5 are both expressed in the PE. However, only constitutively active (ca) ALK2 stimulates PE-derived epithelial cell activation, the first step in transformation, whereas caALK5 stimulates neither activation nor transformation in PE explants. Overexpression of Smad6, an inhibitor of ALK2 signaling, inhibits epithelial cell activation, whereas BMP7, a known ligand for ALK2, has no effect. These data demonstrate that TGFbeta stimulates transformation in the PE and suggest that ALK2 partially mediates this effect. PMID:16245329

  5. The interplay of matrix metalloproteinases, morphogens and growth factors is necessary for branching of mammary epithelial cells

    SciTech Connect

    Simian, M.; Harail, Y.; Navre, M.; Werb, Z.; Lochter, A.; Bissell, M.J.

    2002-03-06

    The mammary gland develops its adult form by a process referred to as branching morphogenesis. Many factors have been reported to affect this process. We have used cultured primary mammary epithelial organoids and mammary epithelial cell lines in three-dimensional collagen gels to elucidate which growth factors, matrix metalloproteinases (MMPs) and mammary morphogens interact in branching morphogenesis. Branching stimulated by stromal fibroblasts, epidermal growth factor, fibroblast growth factor 7, fibroblast growth factor 2 and hepatocyte growth factor was strongly reduced by inhibitors of MMPs, indicating the requirement of MMPs for three-dimensional growth involved in morphogenesis. Recombinant stromelysin 1/MMP-3 alone was sufficient to drive branching in the absence of growth factors in the organoids. Plasmin also stimulated branching; however, plasmin-dependent branching was abolished by both inhibitors of plasmin and MMPs, suggesting that plasmin activates MMPs. To differentiate between signals for proliferation and morphogenesis, we used a cloned mammary epithelial cell line that lacks epimorphin, an essential mammary morphogen. Both epimorphin and MMPs were required for morphogenesis, but neither was required for epithelial cell proliferation. These results provide direct evidence for a critical role of MMPs in branching in mammary epithelium and suggest that, in addition to epimorphin, MMP activity is a minimum requirement for branching morphogenesis in the mammary gland.

  6. Involvement of hepatocyte growth factor-induced epithelial-mesenchymal transition in human adenomyosis.

    PubMed

    Khan, Khaleque Newaz; Kitajima, Michio; Hiraki, Koichi; Fujishita, Akira; Nakashima, Masahiro; Masuzaki, Hideaki

    2015-02-01

    Adenomyosis is commonly believed to arise from the basalis endometrium. As an estromedin growth factor, hepatocyte growth factor (HGF) exhibits multiple functions in endometriosis, a disease commonly believed to arise from the functionalis endometrium. Here, we investigated the role of HGF in the occurrence of epithelial-mesenchymal transition (EMT) in adenomyosis. Full-thickness-biopsy specimens from endometrium to myometrium were collected after hysterectomy from women with and without adenomyosis. The relationship between HGF and E-cadherin (epithelial cell marker) and N-cadherin (mesenchymal cell markers) was examined at the gene and protein levels using endometrial epithelial cells (EECs) in culture and tissues by quantitative RT-PCR and immunohistochemistry. The gene and protein expressions of two transcriptional repressors of E-cadherin, SLUG and SNAIL, were examined using Ishikawa cells and in response to HGF and estrogen (E2). HGF down-regulated E-cadherin and up-regulated N-cadherin mRNA expression in EECs, and an inverse relationship in protein expression between HGF and E-cadherin was observed in basalis endometria derived from women with diffuse and focal adenomyosis. HGF induced morphological changes of EECs from a cobblestone-like appearance to spindle-shaped cells and promoted migration of EECs. Ishikawa cells exhibited up-regulation of SLUG/SNAIL gene expression in response to both HGF and E2 with an additive effect between them. HGF- and E2-promoted SLUG/SNAIL gene expression was significantly abrogated after pretreatment of cells with anti-HGF antibody or ICI 182720, an estrogen receptor antagonist. HGF may be involved in gland invagination deep into the myometrium by inducing EMT at the endo-myometrial junction in women with adenomyosis. PMID:25505196

  7. Transforming growth factor-alpha promotes mammary tumorigenesis through selective survival and growth of secretory epithelial cells.

    PubMed Central

    Smith, G. H.; Sharp, R.; Kordon, E. C.; Jhappan, C.; Merlino, G.

    1995-01-01

    Transforming growth factor (TGF)-alpha stimulates the growth and development of mammary epithelial cells and is implicated in the pathogenesis of human breast cancer. In this report we evaluate the consequences of overexpressing TGF-alpha in the mammary gland of transgenic mice and examine associated cellular mechanisms. When operating on a FVB/N genetic background (line MT100), TGF-alpha induced the stochastic development of mammary adenomas and adenocarcinomas f secretory epithelial origin in 64% of multiparous females. In contrast, tumors were exceedingly rare in virgin MT100 females, MT100 males, and multiparous FVB/N females. In MT100 females multiple foci of hyperplastic secretory lesions preceded the development of frank tumors; these initial lesions appeared during the involution period after the first lactation. Serial transplantation of these hyperplasias indicated an absence of proliferative immortality. Nevertheless, they gave rise to tumors at a low frequency and after a prolonged latency in virgin hosts; in multiparous hosts, tumors developed earlier and at a high incidence. The TGF-alpha transgene was highly expressed in hyperplasias and tumors but not in virgin and nonlesion-bearing tissue, suggesting that TGF-alpha overexpression provides a selective growth advantage. TGF-alpha also induced at lactation a 6.4-fold increase in DNA synthesis in MT100 epithelial cells, many of which were binucleated. MT100 mammary tissue experienced an obvious delay in involution, resulting in the postlactational survival of a significant population of unregressed secretory epithelial cells. In contrast, another line of transgenic mice on a CD-1 genetic background (MT42), in which TGF-alpha overexpression induced liver but not mammary tumors, failed to demonstrate postlactational epithelial cell survival. These data show that TGF-alpha promotes mammary tumorigenesis in multiparous MT100 mice by stimulating secretory epithelial cell proliferation during lactation and

  8. Effects of anti-vascular endothelial growth factor monoclonal antibody (bevacizumab) on lens epithelial cells

    PubMed Central

    Jun, Jong Hwa; Sohn, Wern-Joo; Lee, Youngkyun; Kim, Jae-Young

    2016-01-01

    The molecular and cellular effects of anti-vascular endothelial growth factor monoclonal antibody (bevacizumab) on lens epithelial cells (LECs) were examined using both an immortalized human lens epithelial cell line and a porcine capsular bag model. After treatment with various concentrations of bevacizumab, cell viability and proliferation patterns were evaluated using the water-soluble tetrazolium salt assay and 5-bromo-2′-deoxyuridine enzyme-linked immunosorbent assay, respectively. The scratch assay and Western blot analysis were employed to validate the cell migration pattern and altered expression levels of signaling molecules related to the epithelial–mesenchymal transition (EMT). Application of bevacizumab induced a range of altered cellular events in a concentration-dependent manner. A 0.1–2 mg/mL concentration demonstrated dose-dependent increase in proliferation and viability of LECs. However, 4 mg/mL decreased cell proliferation and viability. Cell migrations displayed dose-dependent retardation from 0.1 mg/mL bevacizumab treatment. Transforming growth factor-β2 expression was markedly increased in a dose-dependent manner, and α-smooth muscle actin, matrix metalloproteinase-9, and vimentin expression levels showed dose-dependent changes in a B3 cell line. Microscopic observation of porcine capsular bag revealed changes in cellular morphology and a decline in cell density compared to the control after 2 mg/mL treatment. The central aspect of posterior capsule showed delayed confluence, and the factors related to EMT revealed similar expression patterns to those identified in the cell line. Based on these results, bevacizumab modulates the proliferation and viability of LECs and induces morphological alterations through the modulation of expression patterns of specific factors related to the EMT. PMID:27418802

  9. Recurrent exposure to nicotine differentiates human bronchial epithelial cells via epidermal growth factor receptor activation

    SciTech Connect

    Martinez-Garcia, Eva; Irigoyen, Marta; Anso, Elena; Martinez-Irujo, Juan Jose; Rouzaut, Ana

    2008-05-01

    Cigarette smoking is the major preventable cause of lung cancer in developed countries. Nicotine (3-(1-methyl-2-pyrrolidinyl)-pyridine) is one of the major alkaloids present in tobacco. Besides its addictive properties, its effects have been described in panoply of cell types. In fact, recent studies have shown that nicotine behaves as a tumor promoter in transformed epithelial cells. This research focuses on the effects of acute repetitive nicotine exposure on normal human bronchial epithelial cells (NHBE cells). Here we show that treatment of NHBE cells with recurrent doses of nicotine up to 500 {mu}M triggered cell differentiation towards a neuronal-like phenotype: cells emitted filopodia and expressed neuronal markers such as neuronal cell adhesion molecule, neurofilament-M and the transcription factors neuronal N and Pax-3. We also demonstrate that nicotine treatment induced NF-kB translocation to the nucleus, phosphorylation of the epidermal growth factor receptor (EGFR), and accumulation of heparin binding-EGF in the extracellular medium. Moreover, addition of AG1478, an inhibitor of EGFR tyrosine phosphorylation, or cetuximab, a monoclonal antibody that precludes ligand binding to the same receptor, prevented cell differentiation by nicotine. Lastly, we show that differentiated cells increased their adhesion to the extracellular matrix and their protease activity. Given that several lung pathologies are strongly related to tobacco consumption, these results may help to better understand the damaging consequences of nicotine exposure.

  10. Fibroblast Growth Factor 2 Is Required for Epithelial Recovery, but Not for Pulmonary Fibrosis, in Response to Bleomycin

    PubMed Central

    Guzy, Robert D.; Stoilov, Ivan; Elton, Timothy J.; Mecham, Robert P.

    2015-01-01

    The pathogenesis of pulmonary fibrosis involves lung epithelial injury and aberrant proliferation of fibroblasts, and results in progressive pulmonary scarring and declining lung function. In vitro, fibroblast growth factor (FGF) 2 promotes myofibroblast differentiation and proliferation in cooperation with the profibrotic growth factor, transforming growth factor-β1, but the in vivo requirement for FGF2 in the development of pulmonary fibrosis is not known. The bleomycin model of lung injury and pulmonary fibrosis was applied to Fgf2 knockout (Fgf2−/−) and littermate control mice. Weight loss, mortality, pulmonary fibrosis, and histology were analyzed after a single intranasal dose of bleomycin. Inflammation was evaluated in bronchoalveolar lavage (BAL) fluid, and epithelial barrier integrity was assessed by measuring BAL protein and Evans Blue dye permeability. Fgf2 is expressed in mouse and human lung epithelial and inflammatory cells, and, in response to bleomycin, Fgf2−/− mice have significantly increased mortality and weight loss. Analysis of BAL fluid and histology show that pulmonary fibrosis is unaltered, but Fgf2−/− mice fail to efficiently resolve inflammation, have increased BAL cellularity, and, importantly, deficient recovery of epithelial integrity. Fgf2−/− mice similarly have deficient recovery of club cell secretory protein+ bronchial epithelium in response to naphthalene. We conclude that FGF2 is not required for bleomycin-induced pulmonary fibrosis, but rather is essential for epithelial repair and maintaining epithelial integrity after bleomycin-induced lung injury in mice. These data identify that FGF2 acts as a protective growth factor after lung epithelial injury, and call into question the role of FGF2 as a profibrotic growth factor in vivo. PMID:24988442

  11. Involvement of Toll-like receptor 2 and epidermal growth factor receptor signaling in epithelial expression of airway remodeling factors.

    PubMed

    Homma, Tetsuya; Kato, Atsushi; Sakashita, Masafumi; Norton, James E; Suh, Lydia A; Carter, Roderick G; Schleimer, Robert P

    2015-04-01

    Staphylococcus aureus (SA) colonization and infection is common, and may promote allergic or inflammatory airway diseases, such as asthma, cystic fibrosis, and chronic rhinosinusitis by interacting with airway epithelial cells. Airway epithelial cells not only comprise a physical barrier, but also play key roles in immune, inflammatory, repair, and remodeling responses upon encounters with pathogens. To elucidate the impact of SA on epithelial-mediated remodeling of allergic airways, we tested the hypothesis that SA can enhance the remodeling process. Normal human bronchial epithelial (NHBE) cells were stimulated with heat-killed SA (HKSA) or transforming growth factor (TGF) α. Cell extracts were collected to measure mRNA (real-time RT-PCR) and signaling molecules (Western blot); supernatants were collected to measure protein (ELISA) after 24 hours of stimulation. Epidermal growth factor receptor (EGFR) signaling inhibition experiments were performed using a specific EGFR kinase inhibitor (AG1478) and TGF-α was blocked with an anti-TGF-α antibody. HKSA induced both mRNA and protein for TGF-α and matrix metalloproteinase (MMP) 1 from NHBE cells by a Toll-like receptor 2-dependent mechanism. Recombinant human TGF-α also induced mRNA and protein for MMP-1 from NHBE cells; anti-TGF-α antibody inhibited HKSA-induced MMP-1, suggesting that endogenous TGF-α mediates the MMP-1 induction by HKSA. HKSA-induced MMP-1 expression was suppressed when a specific EGFR kinase inhibitor was added, suggesting that EGFR signaling was mediating the HKSA-induced MMP-1 release. Exposure or colonization by SA in the airway may enhance the remodeling of tissue through a TGF-α-dependent induction of MMP-1 expression, and may thereby promote remodeling in airway diseases in which SA is implicated, such as asthma and chronic rhinosinusitis. PMID:25180535

  12. Involvement of Toll-Like Receptor 2 and Epidermal Growth Factor Receptor Signaling in Epithelial Expression of Airway Remodeling Factors

    PubMed Central

    Kato, Atsushi; Sakashita, Masafumi; Norton, James E.; Suh, Lydia A.; Carter, Roderick G.; Schleimer, Robert P.

    2015-01-01

    Staphylococcus aureus (SA) colonization and infection is common, and may promote allergic or inflammatory airway diseases, such as asthma, cystic fibrosis, and chronic rhinosinusitis by interacting with airway epithelial cells. Airway epithelial cells not only comprise a physical barrier, but also play key roles in immune, inflammatory, repair, and remodeling responses upon encounters with pathogens. To elucidate the impact of SA on epithelial-mediated remodeling of allergic airways, we tested the hypothesis that SA can enhance the remodeling process. Normal human bronchial epithelial (NHBE) cells were stimulated with heat-killed SA (HKSA) or transforming growth factor (TGF) α. Cell extracts were collected to measure mRNA (real-time RT-PCR) and signaling molecules (Western blot); supernatants were collected to measure protein (ELISA) after 24 hours of stimulation. Epidermal growth factor receptor (EGFR) signaling inhibition experiments were performed using a specific EGFR kinase inhibitor (AG1478) and TGF-α was blocked with an anti–TGF-α antibody. HKSA induced both mRNA and protein for TGF-α and matrix metalloproteinase (MMP) 1 from NHBE cells by a Toll-like receptor 2–dependent mechanism. Recombinant human TGF-α also induced mRNA and protein for MMP-1 from NHBE cells; anti–TGF-α antibody inhibited HKSA-induced MMP-1, suggesting that endogenous TGF-α mediates the MMP-1 induction by HKSA. HKSA-induced MMP-1 expression was suppressed when a specific EGFR kinase inhibitor was added, suggesting that EGFR signaling was mediating the HKSA-induced MMP-1 release. Exposure or colonization by SA in the airway may enhance the remodeling of tissue through a TGF-α–dependent induction of MMP-1 expression, and may thereby promote remodeling in airway diseases in which SA is implicated, such as asthma and chronic rhinosinusitis. PMID:25180535

  13. The effect of retinal pigment epithelial cell patch size on growth factor expression

    SciTech Connect

    Vargis, Elizabeth A.; Peterson, Cristen B.; Morrell-Falvey, Jennifer L.; Retterer, Scott T.; Collier, Charles Patrick

    2014-01-30

    The spatial organization of retinal pigment epithelial (RPE) cells grown in culture was controlled using micropatterning techniques in order to examine the effect of patch size on cell health and differentiation. Understanding this effect is a critical step in the development of multiplexed high throughput fluidic assays and provides a model for replicating disease states associated with the deterioration of retinal tissue during age-related macular degeneration (AMD). Microcontact printing of fibronectin on polystyrene and glass substrates was used to promote cell attachment, forming RPE patches of controlled size and shape. These colonies mimic the effect of atrophy and loss-of-function that occurs in the retina during degenerative diseases such as AMD. After 72 hours of cell growth, levels of vascular endothelial growth factor (VEGF), an important biomarker of AMD, were measured. Cells were counted and morphological indicators of cell viability and tight junction formation were assessed via fluorescence microscopy. As a result, up to a twofold increase of VEGF expression per cell was measured as colony size decreased, suggesting that the local microenvironment of, and connections between, RPE cells influences growth factor expression leading to the initiation and progression of diseases such as AMD.

  14. The effect of retinal pigment epithelial cell patch size on growth factor expression

    DOE PAGESBeta

    Vargis, Elizabeth A.; Peterson, Cristen B.; Morrell-Falvey, Jennifer L.; Retterer, Scott T.; Collier, Charles Patrick

    2014-01-30

    The spatial organization of retinal pigment epithelial (RPE) cells grown in culture was controlled using micropatterning techniques in order to examine the effect of patch size on cell health and differentiation. Understanding this effect is a critical step in the development of multiplexed high throughput fluidic assays and provides a model for replicating disease states associated with the deterioration of retinal tissue during age-related macular degeneration (AMD). Microcontact printing of fibronectin on polystyrene and glass substrates was used to promote cell attachment, forming RPE patches of controlled size and shape. These colonies mimic the effect of atrophy and loss-of-function thatmore » occurs in the retina during degenerative diseases such as AMD. After 72 hours of cell growth, levels of vascular endothelial growth factor (VEGF), an important biomarker of AMD, were measured. Cells were counted and morphological indicators of cell viability and tight junction formation were assessed via fluorescence microscopy. As a result, up to a twofold increase of VEGF expression per cell was measured as colony size decreased, suggesting that the local microenvironment of, and connections between, RPE cells influences growth factor expression leading to the initiation and progression of diseases such as AMD.« less

  15. Controlling Retinal Pigment Epithelial Cell Patch Size Influences Growth Factor Expression

    DOE PAGESBeta

    Vargis, Elizabeth A; Peterson, Cristen B; Morrell-Falvey, Jennifer L; Retterer, Scott T; Collier, Pat

    2014-01-01

    The spatial organization of retinal pigment epithelial (RPE) cells grown in culture was controlled using micropatterning techniques in order to examine the effect of patch size on cell health and differentiation. Understanding this effect is a critical step in the development of multiplexed high throughput fluidic assays and provides a model for replicating disease states associated with the deterioration of retinal tissue during age-related macular degeneration (AMD). Microcontact printing of fibronectin on polystyrene and glass substrates was used to promote cell attachment, forming RPE patches of controlled size and shape. These colonies mimic the effect of atrophy and loss-of-function thatmore » occurs in the retina during degenerative diseases such as AMD. After 72 hours of cell growth, levels of vascular endothelial growth factor (VEGF), an important biomarker of AMD, were measured. Cells were counted and morphological indicators of cell viability and tight junction formation were assessed via fluorescence microscopy. Up to a twofold increase of VEGF expression per cell was measured as colony size decreased, suggesting that the local microenvironment of, and connections between, RPE cells influences growth factor expression leading to the initiation and progression of diseases such as AMD.« less

  16. Requirement of HDAC6 for Transforming Growth Factor-β1-induced Epithelial-Mesenchymal Transition*

    PubMed Central

    Shan, Bin; Yao, Tso-pang; Nguyen, Hong T.; Zhuo, Ying; Levy, Dawn R.; Klingsberg, Ross C.; Tao, Hui; Palmer, Michael L.; Holder, Kevin N.; Lasky, Joseph A.

    2008-01-01

    The aberrant expression of transforming growth factor (TGF)-β1 in the tumor microenvironment and fibrotic lesions plays a critical role in tumor progression and tissue fibrosis by inducing epithelial-mesenchymal transition (EMT). EMT promotes tumor cell motility and invasiveness. How EMT affects motility and invasion is not well understood. Here we report that HDAC6 is a novel modulator of TGF-β1-induced EMT. HDAC6 is a microtubule-associated deacetylase that predominantly deacetylates nonhistone proteins, including α-tubulin, and regulates cell motility. We showed that TGF-β1-induced EMT is accompanied by HDAC6-dependent deacetylation of α-tubulin. Importantly, inhibition of HDAC6 by small interfering RNA or the small molecule inhibitor tubacin attenuated the TGF-β1-induced EMT markers, such as the aberrant expression of epithelial and mesenchymal peptides, as well as the formation of stress fibers. Reduced expression of HDAC6 also impaired the activation of SMAD3 in response to TGF-β1. Conversely, inhibition of SMAD3 activation substantially impaired HDAC6-dependent deacetylation of α-tubulin as well as the expression of EMT markers. These findings reveal a novel function of HDAC6 in EMT by intercepting the TGF-β-SMAD3 signaling cascade. Our results identify HDAC6 as a critical regulator of EMT and a potential therapeutic target against pathological EMT, a key event for tumor progression and fibrogenesis. PMID:18499657

  17. Effects of butyrate on the expression of insulin-like growth factor binding proteins in bovine kidney epithelial cells

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Sodium butyrate induces cell cycle arrest and apoptosis in bovine kidney epithelial cells primarily via down-regulating cell cycle-related gene expression and enhancing expression of pro-apoptotic genes. The insulin-like growth factor (IGF) system plays an essential role in these processes as well a...

  18. An IL6-correlated signature in serous epithelial ovarian cancer associates with growth factor response

    PubMed Central

    2013-01-01

    Background Epithelial ovarian cancer (EOC) is one of the most lethal gynecological cancers; the majority of EOC is the serous histotype and diagnosed at advanced stage. IL6 is the cytokine that has been found most frequently associated with carcinogenesis and progression of serous EOCs. IL6 is a growth-promoting and anti-apoptotic factor, and high plasma levels of IL6 in advanced stage EOCs correlate with poor prognosis. The objective of the present study was to identify IL6 co-regulated genes and gene network/s in EOCs. Results We applied bioinformatics tools on 7 publicly available data sets containing the gene expression profiles of 1262 EOC samples. By Pearson's correlation analysis we identified, in EOCs, an IL6-correlated gene signature containing 40 genes mainly associated with proliferation. 33 of 40 genes were also significantly correlated in low malignant potential (LMP) EOCs, while 7 genes, named C5AR1, FPR1, G0S2, IL8, KLF2, MMP19, and THBD were IL6-correlated only in advanced stage EOCs. Among the 40-gene signature EGFR ligand HBEGF, genes of the EGR family members and genes encoding for negative feedback regulators of growth factor signaling were included. The results obtained by Gene Set Enrichment and Ingenuity Pathway Analyses enabled the identification, respectively, of gene sets associated with ‘early growth factor response’ for the 40-gene signature, and a biological network related to ‘thrombosis and cardiovascular disease’ for the 7-gene signature. In agreement with these results, selected genes from the identified signatures were validated in vitro by real time RT-PCR in serous EOC cell lines upon stimulation with EGF. Conclusions Serous EOCs, independently of their aggressiveness, co-regulate IL6 expression together with that of genes associated to growth factor signaling, arguing for the hypothesis that common mechanism/s driven by EGFR ligands characterize both advanced-stage and LMP EOCs. Only advanced-stage EOCs appeared to be

  19. Basic fibroblast growth factor promotes proliferation of rat glomerular visceral epithelial cells in vitro.

    PubMed Central

    Takeuchi, A.; Yoshizawa, N.; Yamamoto, M.; Sawasaki, Y.; Oda, T.; Senoo, A.; Niwa, H.; Fuse, Y.

    1992-01-01

    Glomerular visceral epithelial cells (vGEC) play an important role in the synthesis of the glomerular basement membrane (GBM), and together with glomerular endothelial cells and the GBM, in glomerular ultrafiltration. Therefore clarification of the properties of vGEC is essential to investigations of glomerular morphology and function in both physiologic and pathologic conditions. This article demonstrates that basic fibroblast growth factor (bFGF) is mitogenic to vGEC in vitro. Its effect was found at concentrations as low as 1.25 ng/ml, and was synergistic with epidermal growth factor (EGF). In contrast, EGF by itself had no demonstrable mitogenic effect at concentrations of 1.25-100 ng/ml. In addition, mRNA for bFGF was identified in cultured vGEC by the method of reverse transcriptase polymerase chain reaction and the immunoreactivity of bFGF was found in GEC of the Sprague-Dawley rat kidney. These results suggest that bFGF stimulates the proliferation of vGEC in an autocrine manner in vivo. A unique relationship similar to that observed in endothelial cells may also exist among bFGF, vGEC, and the extracellular matrix (ECM). In a word, bFGF may be produced by vGEC and stored in the ECM, that is the GBM, and may be one factor that stimulates vGEC to proliferate when vGEC are injured and lost in vivo. Images Figure 1 Figure 2 Figure 3 Figure 6 Figure 8 PMID:1632456

  20. Transforming growth factor-β1 and cigarette smoke inhibit the ability of β2-agonists to enhance epithelial permeability.

    PubMed

    Unwalla, Hoshang J; Ivonnet, Pedro; Dennis, John S; Conner, Gregory E; Salathe, Matthias

    2015-01-01

    Chronic bronchitis, caused by cigarette smoke exposure, is characterized by mucus hypersecretion and reduced mucociliary clearance (MCC). Effective MCC depends, in part, on adequate airway surface liquid. Cystic fibrosis transmembrane conductance regulator (CFTR) provides the necessary osmotic gradient for serosal to mucosal fluid transport through its ability to both secrete Cl(-) and regulate paracellular permeability, but CFTR activity is attenuated in chronic bronchitis and in smokers. β2-adrenergic receptor (β2-AR) agonists are widely used for managing chronic obstructive pulmonary disease, and can activate CFTR, stimulate ciliary beat frequency, and increase epithelial permeability, thereby stimulating MCC. Patients with chronic airway diseases and cigarette smokers demonstrate increased transforming growth factor (TGF)-β1 signaling, which suppresses β2-agonist-mediated CFTR activation and epithelial permeability increases. Restoring CFTR function in these diseases can restore the ability of β2-agonists to enhance epithelial permeability. Human bronchial epithelial cells, fully redifferentiated at the air-liquid interface, were used for (14)C mannitol flux measurements, Ussing chamber experiments, and quantitative RT-PCR. β2-agonists enhance epithelial permeability by activating CFTR via the β2-AR/adenylyl cyclase/cAMP/protein kinase A pathway. TGF-β1 inhibits β2-agonist-mediated CFTR activation and epithelial permeability enhancement. Although TGF-β1 down-regulates both β2-AR and CFTR mRNA, functionally it only decreases CFTR activity. Cigarette smoke exposure inhibits β2-agonist-mediated epithelial permeability increases, an effect reversed by blocking TGF-β signaling. β2-agonists enhance epithelial permeability via CFTR activation. TGF-β1 signaling inhibits β2-agonist-mediated CFTR activation and subsequent increased epithelial permeability, potentially limiting the ability of β2-agonists to facilitate paracellular transport in disease

  1. Transforming Growth Factor-β1 and Cigarette Smoke Inhibit the Ability of β2-Agonists to Enhance Epithelial Permeability

    PubMed Central

    Ivonnet, Pedro; Dennis, John S.; Conner, Gregory E.; Salathe, Matthias

    2015-01-01

    Chronic bronchitis, caused by cigarette smoke exposure, is characterized by mucus hypersecretion and reduced mucociliary clearance (MCC). Effective MCC depends, in part, on adequate airway surface liquid. Cystic fibrosis transmembrane conductance regulator (CFTR) provides the necessary osmotic gradient for serosal to mucosal fluid transport through its ability to both secrete Cl− and regulate paracellular permeability, but CFTR activity is attenuated in chronic bronchitis and in smokers. β2-adrenergic receptor (β2-AR) agonists are widely used for managing chronic obstructive pulmonary disease, and can activate CFTR, stimulate ciliary beat frequency, and increase epithelial permeability, thereby stimulating MCC. Patients with chronic airway diseases and cigarette smokers demonstrate increased transforming growth factor (TGF)-β1 signaling, which suppresses β2-agonist–mediated CFTR activation and epithelial permeability increases. Restoring CFTR function in these diseases can restore the ability of β2-agonists to enhance epithelial permeability. Human bronchial epithelial cells, fully redifferentiated at the air–liquid interface, were used for 14C mannitol flux measurements, Ussing chamber experiments, and quantitative RT-PCR. β2-agonists enhance epithelial permeability by activating CFTR via the β2-AR/adenylyl cyclase/cAMP/protein kinase A pathway. TGF-β1 inhibits β2-agonist–mediated CFTR activation and epithelial permeability enhancement. Although TGF-β1 down-regulates both β2-AR and CFTR mRNA, functionally it only decreases CFTR activity. Cigarette smoke exposure inhibits β2-agonist–mediated epithelial permeability increases, an effect reversed by blocking TGF-β signaling. β2-agonists enhance epithelial permeability via CFTR activation. TGF-β1 signaling inhibits β2-agonist–mediated CFTR activation and subsequent increased epithelial permeability, potentially limiting the ability of β2-agonists to facilitate paracellular transport in

  2. Normal Human Lung Epithelial Cells Inhibit Transforming Growth Factor-β Induced Myofibroblast Differentiation via Prostaglandin E2

    PubMed Central

    Epa, Amali P.; Thatcher, Thomas H.; Pollock, Stephen J.; Wahl, Lindsay A.; Lyda, Elizabeth; Kottmann, R. M.; Phipps, Richard P.; Sime, Patricia J.

    2015-01-01

    Introduction Idiopathic pulmonary fibrosis (IPF) is a chronic progressive disease with very few effective treatments. The key effector cells in fibrosis are believed to be fibroblasts, which differentiate to a contractile myofibroblast phenotype with enhanced capacity to proliferate and produce extracellular matrix. The role of the lung epithelium in fibrosis is unclear. While there is evidence that the epithelium is disrupted in IPF, it is not known whether this is a cause or a result of the fibroblast pathology. We hypothesized that healthy epithelial cells are required to maintain normal lung homeostasis and can inhibit the activation and differentiation of lung fibroblasts to the myofibroblast phenotype. To investigate this hypothesis, we employed a novel co-culture model with primary human lung epithelial cells and fibroblasts to investigate whether epithelial cells inhibit myofibroblast differentiation. Measurements and Main Results In the presence of transforming growth factor (TGF)-β, fibroblasts co-cultured with epithelial cells expressed significantly less α-smooth muscle actin and collagen and showed marked reduction in cell migration, collagen gel contraction, and cell proliferation compared to fibroblasts grown without epithelial cells. Epithelial cells from non-matching tissue origins were capable of inhibiting TGF-β induced myofibroblast differentiation in lung, keloid and Graves’ orbital fibroblasts. TGF-β promoted production of prostaglandin (PG) E2 in lung epithelial cells, and a PGE2 neutralizing antibody blocked the protective effect of epithelial cell co-culture. Conclusions We provide the first direct experimental evidence that lung epithelial cells inhibit TGF-β induced myofibroblast differentiation and pro-fibrotic phenotypes in fibroblasts. This effect is not restricted by tissue origin, and is mediated, at least in part, by PGE2. Our data support the hypothesis that the epithelium plays a crucial role in maintaining lung homeostasis

  3. Vascular endothelial growth factor antagonism restores epithelial barrier dysfunction via affecting zonula occludens proteins

    PubMed Central

    YUKSEL, HASAN; YILMAZ, OZGE; KARAMAN, MERAL; FIRINCI, FATIH; TURKELI, AHMET; KANIK, ESRA TOPRAK; INAN, SEVINC

    2015-01-01

    Epithelial barrier dysfunction is important in the pathogenesis of asthma and allergic responses, and is therefore a therapeutic target. The aim of the present study was to investigate the effects of dexamethasone, a classic therapeutic agent, an anti-tumor necrosis factor agent (etanercept), which is used to treat difficult cases of asthma, and an anti-vascular endothelial growth factor (VEGF) agent (bevacizumab), which is an angiogenesis inhibitor, on zonula occludens (ZO) proteins in an experimental asthma model. The experimental model of asthma was developed using intraperitoneal (IP) and inhaled administration of ovalbumin in 38 BALB/c mice, which were divided into four groups. The control group (n=6) did not receive any treatment, while the four remaining groups (n=8 per group) received an IP injection of saline, etanercept, bevacizumab or dexamethasone, respectively. Occludin, claudin and junctional adhesion molecule (JAM) were immunohistochemically stained in the left middle lobe samples using an indirect avidin-peroxidase method, after which the staining was semiquantified with H-scores. Statistically significant differences were observed in the occludin, claudin and JAM H-scores among the four groups (P<0.001). In the untreated asthma, etanercept, bevacizumab and dexamethasone groups, the median H-scores for occludin were 93, 177, 280 and 198, respectively, while the H-scores for claudin were 82, 193.5, 274 and 202.5, respectively, and the median H-scores for JAM were 130, 210, 288 and 210, respectively. Pairwise comparisons revealed that all three ZO protein H-scores were significantly lower in the saline group when compared with each treatment group. However, the H-scores of the ZO proteins were not significantly different between the etanercept and dexamethasone groups. Furthermore, the bevacizumab group exhibited higher H-scores for all the proteins compared with the dexamethasone group. Therefore, antagonism of VEGF with bevacizumab restores the

  4. Respiratory epithelial cell expression of human transforming growth factor-alpha induces lung fibrosis in transgenic mice.

    PubMed Central

    Korfhagen, T R; Swantz, R J; Wert, S E; McCarty, J M; Kerlakian, C B; Glasser, S W; Whitsett, J A

    1994-01-01

    Increased production of EGF or TGF-alpha by the respiratory epithelial cells has been associated with the pathogenesis of various forms of lung injury. Growth factors and cytokines are thought to act locally, via paracrine and autocrine mechanisms, to stimulate cell proliferation and matrix deposition by interstitial lung cells resulting in pulmonary fibrosis. To test whether TGF-alpha mediates pulmonary fibrotic responses, we have generated transgenic mice expressing human TGF-alpha under control of regulatory regions of the human surfactant protein C (SP-C) gene. Human TGF-alpha mRNA was expressed in pulmonary epithelial cells in the lungs of the transgenic mice. Adult mice bearing the SP-C-TGF-alpha transgene developed severe pulmonary fibrosis. Fibrotic lesions were observed in peribronchial, peribronchiolar, and perivascular regions, as well as subjacent to pleural surfaces. Lesions consisted of fibrous tissue that included groups of epithelial cells expressing endogenous SP-C mRNA, consistent with their identification as distal respiratory epithelial cells. Peripheral fibrotic regions consisted of thickened pleura associated with extensive collagen deposition. Alveolar architecture was disrupted in the transgenic mice with loss of alveoli in the lung parenchyma. Pulmonary epithelial cell expression of TGF-alpha in transgenic mice disrupts alveolar morphogenesis and produces fibrotic lesions mediated by paracrine signaling between respiratory epithelial and interstitial cells of the lung. Images PMID:8163670

  5. Transforming growth factor-alpha in vivo stimulates epithelial cell proliferation in digestive tissues of suckling rats.

    PubMed Central

    Hormi, K; Lehy, T

    1996-01-01

    BACKGROUND: The role that exogenous transforming growth factor-alpha (TGF-alpha) may exert on cell proliferation in vivo is poorly understood. AIM: To investigate the effect of rat TGF-alpha on epithelial cell proliferation in all suckling rat digestive tissues and to compare it with that of rat epidermal growth factor (EGF). ANIMAL AND METHODS: TGF-alpha and EGF were given three times daily either subcutaneously (10 or 20 micrograms/kg) or intraperitoneally (100 micrograms/kg) to rats from the ninth postnatal day. Cell proliferation was assessed through 5-bromo- 2-deoxyuridine incorporation and estimation of labelling indices. RESULTS: For both growth factors, the highest dose given for only two days significantly increased stomach and intestinal weights compared with controls (p < 0.05 to p < 0.001). The proliferative responded depended on the dose given, colonic mucosa being the most sensitive whereas oxyntic mucosa remained unresponsive. TGF-alpha was as potent as EGF in stimulating epithelial cell proliferation in antral, duodenal, and colonic mucosae. However, EGF was more active on oesophageal and jejunal cell proliferation whereas TGF-alpha was more active on pancreatic exocrine cell proliferation and the differences between the two growth factor treated groups were significant. CONCLUSIONS: These results prove for the first time the stimulating effect in vivo of exogenous rat TGF-alpha on epithelial cell proliferation in rat digestive tissues during the developmental period and support a functional role for TGF-alpha at that time. PMID:8944561

  6. Induction of epithelial-mesenchymal transition in alveolar epithelial cells by transforming growth factor-beta1: potential role in idiopathic pulmonary fibrosis.

    PubMed

    Willis, Brigham C; Liebler, Janice M; Luby-Phelps, Katherine; Nicholson, Andrew G; Crandall, Edward D; du Bois, Roland M; Borok, Zea

    2005-05-01

    The hallmark of idiopathic pulmonary fibrosis (IPF) is the myofibroblast, the cellular origin of which in the lung is unknown. We hypothesized that alveolar epithelial cells (AECs) may serve as a source of myofibroblasts through epithelial-mesenchymal transition (EMT). Effects of chronic exposure to transforming growth factor (TGF)-beta1 on the phenotype of isolated rat AECs in primary culture and a rat type II cell line (RLE-6TN) were evaluated. Additionally, tissue samples from patients with IPF were evaluated for cells co-expressing epithelial (thyroid transcription factor (TTF)-1 and pro-surfactant protein-B (pro-SP-B), and mesenchymal (alpha-smooth muscle actin (alpha-SMA)) markers. RLE-6TN cells exposed to TGF-beta1 for 6 days demonstrated increased expression of mesenchymal cell markers and a fibroblast-like morphology, an effect augmented by tumor necrosis factor-alpha (TNF-alpha). Exposure of rat AECs to TGF-beta1 (100 pmol/L) resulted in increased expression of alpha-SMA, type I collagen, vimentin, and desmin, with concurrent transition to a fibroblast-like morphology and decreased expression of TTF-1, aquaporin-5 (AQP5), zonula occludens-1 (ZO-1), and cytokeratins. Cells co-expressing epithelial markers and alpha-SMA were abundant in lung tissue from IPF patients. These results suggest that AECs undergo EMT when chronically exposed to TGF-beta1, raising the possibility that epithelial cells may serve as a novel source of myofibroblasts in IPF. PMID:15855634

  7. Nemo-like kinase regulates the expression of vascular endothelial growth factor (VEGF) in alveolar epithelial cells

    PubMed Central

    Ke, Hengning; Masoumi, Katarzyna Chmielarska; Ahlqvist, Kristofer; Seckl, Michael J.; Rydell-Törmänen, Kristina; Massoumi, Ramin

    2016-01-01

    The canonical Wnt signaling can be silenced either through β-catenin-mediated ubiquitination and degradation or through phosphorylation of Tcf and Lef by nemo-like kinase (NLK). In the present study, we generated NLK deficient animals and found that these mice become cyanotic shortly before death because of lung maturation defects. NLK−/− lungs exhibited smaller and compressed alveoli and the mesenchyme remained thick and hyperplastic. This phenotype was caused by epithelial activation of vascular endothelial growth factor (VEGF) via recruitment of Lef1 to the promoter of VEGF. Elevated expression of VEGF and activation of the VEGF receptor through phosphorylation promoted an increase in the proliferation rate of epithelial and endothelial cells. In summary, our study identifies NLK as a novel signaling molecule for proper lung development through the interconnection between epithelial and endothelial cells during lung morphogenesis. PMID:27035511

  8. Nemo-like kinase regulates the expression of vascular endothelial growth factor (VEGF) in alveolar epithelial cells.

    PubMed

    Ke, Hengning; Masoumi, Katarzyna Chmielarska; Ahlqvist, Kristofer; Seckl, Michael J; Rydell-Törmänen, Kristina; Massoumi, Ramin

    2016-01-01

    The canonical Wnt signaling can be silenced either through β-catenin-mediated ubiquitination and degradation or through phosphorylation of Tcf and Lef by nemo-like kinase (NLK). In the present study, we generated NLK deficient animals and found that these mice become cyanotic shortly before death because of lung maturation defects. NLK-/- lungs exhibited smaller and compressed alveoli and the mesenchyme remained thick and hyperplastic. This phenotype was caused by epithelial activation of vascular endothelial growth factor (VEGF) via recruitment of Lef1 to the promoter of VEGF. Elevated expression of VEGF and activation of the VEGF receptor through phosphorylation promoted an increase in the proliferation rate of epithelial and endothelial cells. In summary, our study identifies NLK as a novel signaling molecule for proper lung development through the interconnection between epithelial and endothelial cells during lung morphogenesis. PMID:27035511

  9. Disruption of E-Cadherin by Matrix Metalloproteinase Directly Mediates Epithelial-Mesenchymal Transition Downstream of Transforming Growth Factor-β1 in Renal Tubular Epithelial Cells

    PubMed Central

    Zheng, Guoping; Lyons, James Guy; Tan, Thian Kui; Wang, Yiping; Hsu, Tzu-Ting; Min, Danqing; Succar, Lena; Rangan, Gopala K.; Hu, Min; Henderson, Beric R.; Alexander, Stephen I.; Harris, David C.H.

    2009-01-01

    Epithelial-mesenchymal transition (EMT) plays an important role in organ fibrosis, including that of the kidney. Loss of E-cadherin expression is a hallmark of EMT; however, whether the loss of E-cadherin is a consequence or a cause of EMT remains unknown, especially in the renal system. In this study, we show that transforming growth factor (TGF)-β1-induced EMT in renal tubular epithelial cells is dependent on proteolysis. Matrix metalloproteinase-mediated E-cadherin disruption led directly to tubular epithelial cell EMT via Slug. TGF-β1 induced the proteolytic shedding of E-cadherin, which caused the nuclear translocation of β-catenin, the transcriptional induction of Slug, and the repression of E-cadherin transcription in tubular epithelial cells. These findings reveal a direct role for E-cadherin and for matrix metalloproteinases in causing EMT downstream of TGF-β1 in fibrotic disease. Specific inhibition rather than activation of matrix metalloproteinases may offer a novel approach for treatment of fibrotic disease. PMID:19590041

  10. Niclosamide inhibits epithelial-mesenchymal transition and tumor growth in lapatinib-resistant human epidermal growth factor receptor 2-positive breast cancer.

    PubMed

    Liu, Junjun; Chen, Xiaosong; Ward, Toby; Mao, Yan; Bockhorn, Jessica; Liu, Xiaofei; Wang, Gen; Pegram, Mark; Shen, Kunwei

    2016-02-01

    Acquired resistance to lapatinib, a human epidermal growth factor receptor 2 kinase inhibitor, remains a clinical problem for women with human epidermal growth factor receptor 2-positive advanced breast cancer, as metastasis is commonly observed in these patients. Niclosamide, an anti-helminthic agent, has recently been shown to exhibit cytotoxicity to tumor cells with stem-like characteristics. This study was designed to identify the mechanisms underlying lapatinib resistance and to determine whether niclosamide inhibits lapatinib resistance by reversing epithelial-mesenchymal transition. Here, two human epidermal growth factor receptor 2-positive breast cancer cell lines, SKBR3 and BT474, were exposed to increasing concentrations of lapatinib to establish lapatinib-resistant cultures. Lapatinib-resistant SKBR3 and BT474 cells exhibited up-regulation of the phenotypic epithelial-mesenchymal transition markers Snail, vimentin and α-smooth muscle actin, accompanied by activation of nuclear factor-кB and Src and a concomitant increase in stem cell marker expression (CD44(high)/CD24(low)), compared to naive lapatinib-sensitive SKBR3 and BT474 cells, respectively. Interestingly, niclosamide reversed epithelial-mesenchymal transition, induced apoptosis and inhibited cell growth by perturbing aberrant signaling pathway activation in lapatinib-resistant human epidermal growth factor receptor 2-positive cells. The ability of niclosamide to alleviate stem-like phenotype development and invasion was confirmed. Collectively, our results demonstrate that lapatinib resistance correlates with epithelial-mesenchymal transition and that niclosamide inhibits lapatinib-resistant cell viability and epithelial-mesenchymal transition. These findings suggest a role of niclosamide or derivatives optimized for more favorable bioavailability not only in reversing lapatinib resistance but also in reducing metastatic potential during the treatment of human epidermal growth factor receptor

  11. Transforming growth factor-beta induces loss of epithelial character and smooth muscle cell differentiation in epicardial cells.

    PubMed

    Compton, Leigh A; Potash, Dru A; Mundell, Nathan A; Barnett, Joey V

    2006-01-01

    During embryogenesis, epicardial cells undergo epithelial-mesenchymal transformation (EMT), invade the myocardium, and differentiate into components of the coronary vasculature, including smooth muscle cells. We tested the hypothesis that transforming growth factor-beta (TGFbeta) stimulates EMT and smooth muscle differentiation of epicardial cells. In epicardial explants, TGFbeta1 and TGFbeta2 induce loss of epithelial morphology, cytokeratin, and membrane-associated Zonula Occludens-1 and increase the smooth muscle markers calponin and caldesmon. Inhibition of activin receptor-like kinase (ALK) 5 blocks these effects, whereas constitutively active (ca) ALK5 increases cell invasion by 42%. Overexpression of Smad 3 did not mimic the effects of caALK5. Inhibition of p160 rho kinase or p38 MAP kinase prevented the loss of epithelial morphology in response to TGFbeta, whereas only inhibition of p160 rho kinase blocked TGFbeta-stimulated caldesmon expression. These data demonstrate that TGFbeta stimulates loss of epithelial character and smooth muscle differentiation in epicardial cells by means of a mechanism that requires ALK5 and p160 rho kinase. PMID:16258965

  12. Vitamin D inhibition of pro-fibrotic effects of transforming growth factor β1 in lung fibroblasts and epithelial cells

    PubMed Central

    Ramirez, Allan M.; Wongtrakool, Cherry; Welch, Teresa; Steinmeyer, Andreas; Zügel, Ulrich; Roman, Jesse

    2010-01-01

    The mechanisms that control fibroproliferation and matrix deposition in lung fibrosis remain unclear. We speculate that vitamin D deficiency may contribute to pulmonary fibrosis since vitamin D deficiency has been implicated in several diseases. First, we confirmed the presence of vitamin D receptors (VDR) in cultured NIH/3T3 and lung fibroblasts. Fibroblasts transfected with a vitamin D response element – reporter construct and exposed to the active vitamin D metabolite, 1,25(OH)2D3, showed increased promoter activity indicating VDR functionality in these cells. Testing the effects of 1,25(OH)2D3 on fibroblasts treated with transforming growth factor β1 (TGFβ1), considered a driver of many fibrotic disorders, we found that 1,25(OH)2D3 inhibited TGFβ1-induced fibroblast proliferation in a dose-dependent fashion. 1,25(OH)2D3 also inhibited TGFβ1 stimulation of α-smooth muscle actin expression and polymerization and prevented the upregulation of fibronectin and collagen in TGFβ1-treated fibroblasts. Finally, we examined how 1,25(OH)2D3 affects epithelial-mesenchymal transformation of lung epithelial cells upon exposure to TGFβ1. We showed that the TGFβ1-induced upregulation of mesenchymal cell markers and abnormal expression of epithelial cell markers were blunted by 1,25(OH)2D3. These observations suggest that under TGFβ1 stimulation, 1,25(OH)2D3 inhibits the profibrotic phenotype of lung fibroblasts and epithelial cells. PMID:19931390

  13. Lung fibroblasts accelerate wound closure in human alveolar epithelial cells through hepatocyte growth factor/c-Met signaling

    PubMed Central

    Correll, Kelly; Schiel, John A.; Finigan, Jay H.; Prekeris, Rytis; Mason, Robert J.

    2014-01-01

    There are 190,600 cases of acute lung injury/acute respiratory distress syndrome (ALI/ARDS) each year in the United States, and the incidence and mortality of ALI/ARDS increase dramatically with age. Patients with ALI/ARDS have alveolar epithelial injury, which may be worsened by high-pressure mechanical ventilation. Alveolar type II (ATII) cells are the progenitor cells for the alveolar epithelium and are required to reestablish the alveolar epithelium during the recovery process from ALI/ARDS. Lung fibroblasts (FBs) migrate and proliferate early after lung injury and likely are an important source of growth factors for epithelial repair. However, how lung FBs affect epithelial wound healing in the human adult lung has not been investigated in detail. Hepatocyte growth factor (HGF) is known to be released mainly from FBs and to stimulate both migration and proliferation of primary rat ATII cells. HGF is also increased in lung tissue, bronchoalveolar lavage fluid, and serum in patients with ALI/ARDS. Therefore, we hypothesized that HGF secreted by FBs would enhance wound closure in alveolar epithelial cells (AECs). Wound closure was measured using a scratch wound-healing assay in primary human AEC monolayers and in a coculture system with FBs. We found that wound closure was accelerated by FBs mainly through HGF/c-Met signaling. HGF also restored impaired wound healing in AECs from the elderly subjects and after exposure to cyclic stretch. We conclude that HGF is the critical factor released from FBs to close wounds in human AEC monolayers and suggest that HGF is a potential strategy for hastening alveolar repair in patients with ALI/ARDS. PMID:24748602

  14. E-cadherin Surface Levels in Epithelial Growth Factor-stimulated Cells Depend on Adherens Junction Protein Shrew-1

    PubMed Central

    Gross, Julia Christina; Schreiner, Alexander; Engels, Knut

    2009-01-01

    Gain- and loss-of-function studies indicate that the adherens junction protein shrew-1 acts as a novel modulator of E-cadherin internalization induced by epithelial growth factor (EGF) or E-cadherin function-blocking antibody during epithelial cell dynamics. Knocking down shrew-1 in MCF-7 carcinoma cells preserves E-cadherin surface levels upon EGF stimulation. Overexpression of shrew-1 leads to preformation of an E-cadherin/EGF receptor (EGFR) HER2/src-kinase/shrew-1 signaling complex and accelerated E-cadherin internalization. Shrew-1 is not sufficient to stimulate E-cadherin internalization, but facilitates the actions of EGFR and thus may promote malignant progression in breast cancer cells with constitutive EGFR stimulation by reducing surface E-cadherin expression. PMID:19515834

  15. Inhibition of Transforming Growth Factor-{beta} Signaling in Normal Lung Epithelial Cells Confers Resistance to Ionizing Radiation

    SciTech Connect

    Reeves, Anna; Zagurovskaya, Marianna; Gupta, Seema; Shareef, Mohammed M.; Mohiuddin, Mohammed; Ahmed, Mansoor M. . E-mail: mmahmed@geisinger.edu

    2007-05-01

    Purpose: To address the functional role of radiation-induced transforming growth factor-{beta} (TGF-{beta}) signaling in a normal epithelial background, we selected a spontaneously immortalized lung epithelial cell line derived from the normal lung tissue of a dominant-negative mutant of the TGF-{beta} RII ({delta}RII) transgenic mouse that conditionally expressed {delta}RII under the control of the metallothionein promoter (MT-1), and assessed this cell line's response to radiation. Methods and Materials: A spontaneously immortalized lung epithelial cell culture (SILECC) was established and all analyses were performed within 50 passages. Colony-forming and terminal transferase dUPT nick end labeling (TUNEL) assays were used to assess clonogenic inhibition and apoptosis, respectively. Western-blot analysis was performed to assess the kinetics of p21, bax, and RII proteins. Transforming growth factor-{beta}-responsive promoter activity was measured using dual-luciferase reporter assay. Results: Exposure to ZnSO{sub 4} inhibited TGF-{beta} signaling induced either by recombinant TGF-{beta}1 or ionizing radiation. The SILECC, treated with either ZnSO{sub 4} or neutralizing antibody against TGF-{beta}, showed a significant increase in radio-resistance compared to untreated cells. Furthermore, the expression of {delta}RII inhibited the radiation-induced up-regulation of the TGF-{beta} effector gene p21{sup waf1/cip1}. Conclusions: Our findings imply that inhibition of radiation-induced TGF-{beta} signaling via abrogation of the RII function enhances the radio-resistance of normal lung epithelial cells, and this can be directly attributed to the loss of TGF-{beta} signaling function.

  16. Induction by transforming growth factor-β1 of epithelial to mesenchymal transition is a rare event in vitro

    PubMed Central

    Brown, Kimberly A; Aakre, Mary E; Gorska, Agnieska E; Price, James O; Eltom, Sakina E; Pietenpol, Jennifer A; Moses, Harold L

    2004-01-01

    Introduction Transforming growth factor (TGF)-β1 is proposed to inhibit the growth of epithelial cells in early tumorigenesis, and to promote tumor cell motility and invasion in the later stages of carcinogenesis through the induction of an epithelial to mesenchymal transition (EMT). EMT is a multistep process that is characterized by changes in cell morphology and dissociation of cell–cell contacts. Although there is growing interest in TGF-β1-mediated EMT, the phenotype is limited to only a few murine cell lines and mouse models. Methods To identify alternative cell systems in which to study TGF-β1-induced EMT, 18 human and mouse established cell lines and cultures of two human primary epithelial cell types were screened for TGF-β1-induced EMT by analysis of cell morphology, and localization of zonula occludens-1, E-cadherin, and F-actin. Sensitivity to TGF-β1 was also determined by [3H]thymidine incorporation, flow cytometry, phosphorylation of Smad2, and total levels of Smad2 and Smad3 in these cell lines and in six additional cancer cell lines. Results TGF-β1 inhibited the growth of most nontransformed cells screened, but many of the cancer cell lines were insensitive to the growth inhibitory effects of TGF-β1. In contrast, TGF-β1 induced Smad2 phosphorylation in the majority of cell lines, including cell lines resistant to TGF-β1-mediated cell cycle arrest. Of the cell lines screened only two underwent TGF-β1-induced EMT. Conclusion The results presented herein show that, although many cancer cell lines have lost sensitivity to the growth inhibitory effect of TGF-β1, most show evidence of TGF-β1 signal transduction, but only a few cell lines undergo TGF-β1-mediated EMT. PMID:15084245

  17. Transforming growth factor-beta mediates intestinal healing and susceptibility to injury in vitro and in vivo through epithelial cells.

    PubMed

    Beck, Paul L; Rosenberg, Ian M; Xavier, Ramnik J; Koh, Theodore; Wong, Josée F; Podolsky, Daniel K

    2003-02-01

    In vitro studies suggest that transforming growth factor (TGF)-beta has potent effects on gastrointestinal mucosal integrity, wound repair, and neoplasia. However, the multiplicity of actions of this peptide on many different cell types confounds efforts to define the role of TGF-beta within the intestinal epithelium in vivo. To delineate these effects selective blockade of intestinal epithelial TGF-beta activity was undertaken through targeted expression of a dominant-negative (DN) TGF-beta RII to intestinal epithelial cells in vitro and in vivo. Stable intestinal epithelial cell (IEC)-6 lines overexpressing TGF-beta RII-DN (nucleotides -7 to 573) were established. Transgenic mice overexpressing TGF-beta RII-DN under the regulation of a modified liver fatty acid-binding promoter (LFABP-PTS4) were constructed. In vitro healing was assessed by wounding of confluent monolayers. Colitis was induced by the addition of dextran sodium sulfate (2.5 to 7.5% w/v) to their drinking water. Overexpression of TGF-beta RII-DN in intestinal epithelial cell-6 cells resulted in a marked reduction in cell migration and TGF-beta-stimulated wound healing in vitro. TGF-beta RII-DN transgenic mice did not exhibit baseline intestinal inflammation or changes in survival, body weight, epithelial cell proliferation, aberrant crypt foci, or tumor formation. TGF-beta RII-DN mice were markedly more susceptible to dextran sodium sulfate-induced colitis and exhibited impaired recovery after colonic injury. TGF-beta is required for intestinal mucosal healing and TGF-beta modulation of the intestinal epithelium plays a central role in determining susceptibility to injury. PMID:12547717

  18. Potentiation of growth factor signaling by insulin-like growth factor-binding protein-3 in breast epithelial cells requires sphingosine kinase activity.

    PubMed

    Martin, Janet L; Lin, Mike Z; McGowan, Eileen M; Baxter, Robert C

    2009-09-18

    We have investigated the mechanism underlying potentiation of epidermal growth factor receptor (EGFR) and type 1 insulin-like growth factor receptor (IGFR1) signaling by IGF-binding protein-3 (IGFBP-3) in MCF-10A breast epithelial cells, focusing on a possible involvement of the sphingosine kinase (SphK) system. IGFBP-3 potentiated EGF-stimulated EGF receptor activation and DNA synthesis, and this was blocked by inhibitors of SphK activity or small interference RNA-mediated silencing of SphK1, but not SphK2, expression. Similarly, IGFR1 phosphorylation and DNA synthesis stimulated by LR3-IGF-I (an IGF-I analog not bound by IGFBP-3), were enhanced by IGFBP-3, and this was blocked by SphK1 silencing. SphK1 expression and activity were stimulated by IGFBP-3 approximately 2-fold over 24 h. Silencing of sphingosine 1-phosphate receptor 1 (S1P1) or S1P3, but not S1P2, abolished the effect of IGFBP-3 on EGF-stimulated EGFR activation. The effects of IGFBP-3 could be reproduced with exogenous S1P or medium conditioned by cells treated with IGFBP-3, and this was also blocked by inhibition of S1P1 and S1P3. These data indicate that potentiation of growth factor signaling by IGFBP-3 in MCF-10A cells requires SphK1 activity and S1P1/S1P3, suggesting that S1P, the product of SphK activity and ligand for S1P1 and S1P3, is the "missing link" mediating IGF and EGFR transactivation and cell growth stimulation by IGFBP-3. PMID:19633297

  19. Transforming Growth Factor β-1 Stimulates Profibrotic Epithelial Signaling to Activate Pericyte-Myofibroblast Transition in Obstructive Kidney Fibrosis

    PubMed Central

    Wu, Ching-Fang; Chiang, Wen-Chih; Lai, Chun-Fu; Chang, Fan-Chi; Chen, Yi-Ting; Chou, Yu-Hsiang; Wu, Ting-Hui; Linn, Geoffrey R.; Ling, Hong; Wu, Kwan-Dun; Tsai, Tun-Jun; Chen, Yung-Ming; Duffield, Jeremy S.; Lin, Shuei-Liong

    2014-01-01

    Pericytes have been identified as the major source of precursors of scar-producing myofibroblasts during kidney fibrosis. The underlying mechanisms triggering pericyte-myofibroblast transition are poorly understood. Transforming growth factor β-1 (TGF-β1) is well recognized as a pluripotent cytokine that drives organ fibrosis. We investigated the role of TGF-β1 in inducing profibrotic signaling from epithelial cells to activate pericyte-myofibroblast transition. Increased expression of TGF-β1 was detected predominantly in injured epithelium after unilateral ureteral obstruction, whereas downstream signaling from the TGF-β1 receptor increased in both injured epithelium and pericytes. In mice with ureteral obstruction that were treated with the pan anti–TGF-β antibody (1D11) or TGF-β receptor type I inhibitor (SB431542), kidney pericyte-myofibroblast transition was blunted. The consequence was marked attenuation of fibrosis. In addition, epithelial cell cycle G2/M arrest and production of profibrotic cytokines were both attenuated. Although TGF-β1 alone did not trigger pericyte proliferation in vitro, it robustly induced α smooth muscle actin (α-SMA). In cultured kidney epithelial cells, TGF-β1 stimulated G2/M arrest and production of profibrotic cytokines that had the capacity to stimulate proliferation and transition of pericytes to myofibroblasts. In conclusion, this study identified a novel link between injured epithelium and pericyte-myofibroblast transition through TGF-β1 during kidney fibrosis. PMID:23142380

  20. Transforming Growth Factor β/activin signalling induces epithelial cell flattening during Drosophila oogenesis

    PubMed Central

    Brigaud, Isabelle; Duteyrat, Jean-Luc; Chlasta, Julien; Le Bail, Sandrine; Couderc, Jean-Louis; Grammont, Muriel

    2015-01-01

    ABSTRACT Although the regulation of epithelial morphogenesis is essential for the formation of tissues and organs in multicellular organisms, little is known about how signalling pathways control cell shape changes in space and time. In the Drosophila ovarian epithelium, the transition from a cuboidal to a squamous shape is accompanied by a wave of cell flattening and by the ordered remodelling of E-cadherin-based adherens junctions. We show that activation of the TGFβ pathway is crucial to determine the timing, the degree and the dynamic of cell flattening. Within these cells, TGFβ signalling controls cell-autonomously the formation of Actin filament and the localisation of activated Myosin II, indicating that internal forces are generated and used to remodel AJ and to promote cytoskeleton rearrangement. Our results also reveal that TGFβ signalling controls Notch activity and that its functions are partly executed through Notch. Thus, we demonstrate that the cells that undergo the cuboidal-to-squamous transition produce active cell-shaping mechanisms, rather than passively flattening in response to a global force generated by the growth of the underlying cells. Thus, our work on TGFβ signalling provides new insights into the mechanisms through which signal transduction cascades orchestrate cell shape changes to generate proper organ structure. PMID:25681395

  1. FGF10: A multifunctional mesenchymal-epithelial signaling growth factor in development, health, and disease.

    PubMed

    Itoh, Nobuyuki

    2016-04-01

    The FGF family comprises 22 members with diverse functions in development and health. FGF10 specifically activates FGFR2b in a paracrine manner with heparan sulfate as a co-factor. FGF10and FGFR2b are preferentially expressed in the mesenchyme and epithelium, respectively. FGF10 is a mesenchymal signaling molecule in the epithelium. FGF10 knockout mice die shortly after birth due to the complete absence of lungs as well as fore- and hindlimbs. FGF10 is also essential for the development of multiple organs. The phenotypes of Fgf10 knockout mice are very similar to those of FGFR2b knockout mice, indicating that FGF10 acts as a ligand that is specific to FGFR2b in mouse multi-organ development. FGF10 also plays roles in epithelial-mesenchymal transition, the repair of tissue injury, and embryonic stem cell differentiation. In humans, FGF10 loss-of-function mutations result in inherited diseases including aplasia of lacrimal and salivary gland, lacrimo-auriculo-dento-digital syndrome, and chronic obstructive pulmonary disease. FGF10 is also involved in the oncogenicity of pancreatic and breast cancers. Single nucleotide polymorphisms in FGF10 are also potential risk factors for limb deficiencies, cleft lip and palate, and extreme myopia. These findings indicate that FGF10 is a crucial paracrine signal from the mesenchyme to epithelium for development, health, and disease. PMID:26559461

  2. Investigating impact of Vascular Endothelial Growth Factor Polymorphisms in Epithelial Ovarian Cancers: A Study in the Indian Population

    PubMed Central

    Bhagat, Rahul; Chennagiri Srinivasamurthy, Premalata; Venketeshiah Reddihalli, Pallavi; Gawari, Ramesh

    2015-01-01

    Epithelial ovarian cancer is one of the increasingly incident malignancies that is notorious because of its evasiveness for early diagnosis and high mortality rates. Epithelial ovarian cancers are highly dependent on pathologic vasculature and Vascular Endothelial Growth Factor is known to be one of the most efficient angiogenic factors. Polymorphisms of the VEGF gene, in this study, were assessed for association with the malignancy and other clinico-pathological factors. 300 case samples and 320 age and mensus status matched controls were inculcated into the study. rs699947, rs833061, rs1570360, rs2010963, rs1413711 and rs3025039 were the six single nucleotide polymorphisms that were scrutinized. Genotyping was carried out by polymerase chain reaction and restriction fragment length polymorphism. rs 3025039 showed immense promise as a marker for disease aggression and recurrence and a factor for poor prognosis. rs699947 showed least association with the disease and clinico-pathologic factors studied. rs833061, rs 1570360 showed significant association with some clinico-pathological factors such as bilateral affliction of ovaries and post operative CA-125 levels. rs2010963 associated with presence of ascites in higher volumes. The SNPs under consideration showed no formidable linkage in our study samples. A haplotype analysis (excluding rs699947 and rs1413711) revealed 5 frontrunners being present in >85% of the population with TGGC and CGCC associating significantly as protective and risk factors respectively. These haplotypes showed a dose dependent additive effect of their seeming functionality. This study is unique and a first of its kind carried out in the Indian population of South-east Asia. PMID:26158818

  3. Investigating impact of Vascular Endothelial Growth Factor Polymorphisms in Epithelial Ovarian Cancers: A Study in the Indian Population.

    PubMed

    Janardhan, Bhaskari; Vaderhobli, Shilpa; Bhagat, Rahul; Chennagiri Srinivasamurthy, Premalata; Venketeshiah Reddihalli, Pallavi; Gawari, Ramesh; Krishnamoorthy, Lakshmi

    2015-01-01

    Epithelial ovarian cancer is one of the increasingly incident malignancies that is notorious because of its evasiveness for early diagnosis and high mortality rates. Epithelial ovarian cancers are highly dependent on pathologic vasculature and Vascular Endothelial Growth Factor is known to be one of the most efficient angiogenic factors. Polymorphisms of the VEGF gene, in this study, were assessed for association with the malignancy and other clinico-pathological factors. 300 case samples and 320 age and mensus status matched controls were inculcated into the study. rs699947, rs833061, rs1570360, rs2010963, rs1413711 and rs3025039 were the six single nucleotide polymorphisms that were scrutinized. Genotyping was carried out by polymerase chain reaction and restriction fragment length polymorphism. rs 3025039 showed immense promise as a marker for disease aggression and recurrence and a factor for poor prognosis. rs699947 showed least association with the disease and clinico-pathologic factors studied. rs833061, rs 1570360 showed significant association with some clinico-pathological factors such as bilateral affliction of ovaries and post operative CA-125 levels. rs2010963 associated with presence of ascites in higher volumes. The SNPs under consideration showed no formidable linkage in our study samples. A haplotype analysis (excluding rs699947 and rs1413711) revealed 5 frontrunners being present in >85% of the population with TGGC and CGCC associating significantly as protective and risk factors respectively. These haplotypes showed a dose dependent additive effect of their seeming functionality. This study is unique and a first of its kind carried out in the Indian population of South-east Asia. PMID:26158818

  4. P21 Activated Kinase-1 Mediates Transforming Growth Factor β1-Induced Prostate Cancer Cell Epithelial to Mesenchymal Transition

    PubMed Central

    Al-Azayzih, Ahmad; Gao, Fei; Somanath, Payaningal R.

    2015-01-01

    Transforming growth factor beta (TGFβ) is believed to play a dual role in prostate cancer. Molecular mechanism by which TGFβ1 suppresses early prostate tumor growth and induces epithelial-to-mesenchymal transition (EMT) in advanced stages is not known. We determined if P21-activated kinase1 (Pak1), which mediates cytoskeletal remodeling is necessary for the TGFβ1 induced prostate cancer EMT. Effects of TGFβ1 on control prostate cancer PC3 and DU145 cells and those with IPA 3 and siRNA mediated Pak1 inhibition were tested for prostate tumor xenograft in vivo and EMT in vitro. TGFβ1 inhibited PC3 tumor xenograft growth via activation of P38-MAPK and caspase-3, 9. Long-term stimulation with TGFβ1 induced PC3 and DU145 cell scattering and increased expression of EMT markers such as Snail and N-cadherin through tumor necrosis factor receptor-associated factor-6 (TRAF6)-mediated activation of Rac1/Pak1 pathway. Selective inhibition of Pak1 using IPA 3 or knockdown using siRNA both significantly inhibited TGFβ1-induced prostate cancer cell EMT and expression of mesenchymal markers. Our study demonstrated that TGFβ1 induces apoptosis and EMT in prostate cancer cells via activation of P38-MAPK and Rac1/Pak1 respectively. Our results reveal the potential therapeutic benefits of targeting TGFβ1-Pak1 pathway for advanced-stage prostate cancer. PMID:25746720

  5. ECM and FGF-dependent assay of embryonic submandibular gland epithelial morphogenesis: investigating growth factor/matrix regulation of gene expression during development

    PubMed Central

    Rebustini, Ivan T.; Hoffman, Matthew P.

    2012-01-01

    Epithelial-mesenchyme interactions during organogenesis are regulated by dynamic and reciprocal interactions between growth factors and extracellular matrix (ECM) components. Mouse embryonic submandibular gland (SMG) epithelium, isolated from its endogenous mesenchyme, undergoes branching morphogenesis when cultured ex vivo in a basement membrane extract in serum-free medium with growth factor stimulation. The resulting three-dimensional epithelial morphogenesis in the defined culture system makes this a useful model to analyze cell-cell and cell-matrix interactions, growth factor-mediated signaling and gene expression, proliferation, apoptosis, migration, lumen formation, and epithelial morphogenesis in a primary organ culture system. SMG epithelial culture is robust, reproducible, uses small amounts of reagents, and changes in gene expression are measured by real-time PCR using a limited amount of embryonic tissue. In this chapter, we describe a detailed protocol for isolating primary embryonic SMG epithelium and setting up an ECM and growth factor-dependent, serum-free assay of epithelial morphogenesis, with subsequent analysis of gene expression by real-time PCR. PMID:19247608

  6. Insulin-like growth factor binding protein-2 mediates the inhibition of DNA synthesis by transforming growth factor-beta in mink lung epithelial cells.

    PubMed

    Dong, Feng; Wu, Hai-Bin; Hong, Jiang; Rechler, Matthew M

    2002-01-01

    Insulin-like growth factor binding protein-3 (IGFBP-3) has been proposed to mediate the growth inhibitory effects of transforming growth factor (TGF)-beta in breast and prostate cancer cells. Both TGF-beta and exogenous IGFBP-3 inhibit DNA synthesis in Mv1 mink lung epithelial cells (CCL64). The present study asks whether IGFBPs synthesized by CCL64 cells mediate growth inhibition by TGF-beta. CCL64 cells synthesize and secrete a single 34-kDa IGFBP that was identified as IGFBP-2 by immunoprecipitation and immunodepletion. Recombinant bovine IGFBP-2 inhibited CCL64 DNA synthesis in serum-free media in an IGF-independent manner. Coincubation with Leu(60)-IGF-I, an IGF-I analog that binds to IGFBPs with higher affinity than to IGF-I receptors, decreased the inhibition by bIGFBP-2. Leu(60)-IGF-I also decreased the inhibition of CCL64 DNA synthesis by TGF-beta by up to 70%, whereas Long-R3-IGF-I, an IGF-I analog with higher affinity for IGF-I receptors than for IGFBPs, did not decrease inhibition, suggesting that the effect of Leu(60)-IGF-I resulted from its forming complexes with endogenous IGFBPs. Leu(60)-IGF-I did not decrease TGF-beta stimulation of a Smad3-dependent reporter gene. Following incubation of intact CCL64 cells with bIGFBP-2 at 0 degrees C, bIGFBP-2 was recovered in membrane fractions; membrane association was abolished by coincubation with Leu(60)-IGF-I. If exogenous and secreted IGFBP-2 must bind to CCL64 cells to inhibit DNA synthesis, Leu(60)-IGF-I might reduce the inhibition of DNA synthesis by bIGFBP-2 or TGF-beta by inhibiting the association of IGFBP-2 in the media with CCL64 cells. Since TGF-beta does not increase IGFBP-2 abundance, we propose that TGF-beta sensitizes CCL64 cells to the latent growth inhibitory activity of endogenous IGFBP-2 by potentiating an intracellular IGFBP-2 signaling pathway or by promoting the association of secreted IGFBP-2 with the plasma membrane. PMID:11807812

  7. Complexity in Interpretation of Embryonic Epithelial-Mesenchymal Transition in Response to Transforming Growth Factor-β Signaling

    PubMed Central

    Ahmed, Shaheen

    2007-01-01

    Epithelial-mesenchymal transition (EMT) is a highly conserved and fundamental process that governs morphogenesis in development and may also contribute to cancer metastasis. Transforming growth factor (TGF-β) is a potent inducer of EMT in various developmental and tumor systems. The analysis of TGF-β signal transduction pathways is now considered a critically important area of biology, since many defects occur in these pathways in embryonic development. The complexity of TGF-β signal transduction networks is overwhelming due to the large numbers of interacting constituents, complicated feedforward, feedback and crosstalk circuitry mechanisms that they involve in addition to the cellular kinetics and enzymatics that contribute to cell signaling. As a result of this complexity, apparently simple but highly important questions remain unanswered, that is, how do epithelial cells respond to such TGF-β signals? System biology and cellular kinetics play a crucial role in cellular function; omissions of such a critical contributor may lead to inaccurate understanding of embryonic EMT. In this review, we identify and explain why certain conditions need to be considered for a true representation of TGF-β signaling in vivo to better understand the controlled, yet delicate mechanism of embryonic EMT. PMID:17587819

  8. Ionizing Radiation Promotes Migration and Invasion of Cancer Cells Through Transforming Growth Factor-Beta-Mediated Epithelial-Mesenchymal Transition

    SciTech Connect

    Zhou Yongchun; Liu Junye; Li Jing; Zhang Jie; Xu Yuqiao; Zhang Huawei; Qiu Lianbo; Ding Guirong; Su Xiaoming; Mei Shi; Guo Guozhen

    2011-12-01

    Purpose: To examine whether ionizing radiation enhances the migratory and invasive abilities of cancer cells through transforming growth factor (TGF-{beta})-mediated epithelial-mesenchymal transition (EMT). Methods and Materials: Six cancer cell lines originating from different human organs were irradiated by {sup 60}Co {gamma}-ray at a total dose of 2 Gy, and the changes associated with EMT, including morphology, EMT markers, migration and invasion, were observed by microscope, Western blot, immunofluorescence, scratch assay, and transwell chamber assay, respectively. Then the protein levels of TGF-{beta} in these cancer cells were detected by enzyme-linked immunosorbent assay, and the role of TGF-{beta} signaling pathway in the effect of ionizing radiation on EMT was investigate by using the specific inhibitor SB431542. Results: After irradiation with {gamma}-ray at a total dose of 2 Gy, cancer cells presented the mesenchymal phenotype, and compared with the sham-irradiation group the expression of epithelial markers was decreased and of mesenchymal markers was increased, the migratory and invasive capabilities were strengthened, and the protein levels of TGF-{beta} were enhanced. Furthermore, events associated with EMT induced by IR in A549 could be reversed through inhibition of TGF-{beta} signaling. Conclusions: These results suggest that EMT mediated by TGF-{beta} plays a critical role in IR-induced enhancing of migratory and invasive capabilities in cancer cells.

  9. Osmotic Induction of Angiogenic Growth Factor Expression in Human Retinal Pigment Epithelial Cells

    PubMed Central

    Reichenbach, Andreas; Wiedemann, Peter; Kohen, Leon; Bringmann, Andreas

    2016-01-01

    Background Although systemic hypertension is a risk factor of age-related macular degeneration, antihypertensive medications do not affect the risk of the disease. One condition that induces hypertension is high intake of dietary salt resulting in increased blood osmolarity. In order to prove the assumption that, in addition to hypertension, high osmolarity may aggravate neovascular retinal diseases, we determined the effect of extracellular hyperosmolarity on the expression of angiogenic cytokines in cultured human retinal pigment epithelial (RPE) cells. Methodology/Principal Findings Hyperosmolarity was induced by the addition of 100 mM NaCl or sucrose to the culture medium. Hypoxia and oxidative stress were induced by the addition of the hypoxia mimetic CoCl2 and H2O2, respectively. Alterations in gene expression were determined with real-time RT-PCR. Secretion of bFGF was evaluated by ELISA. Cell viability was determined by trypan blue exclusion. Nuclear factor of activated T cell 5 (NFAT5) expression was knocked down with siRNA. Hyperosmolarity induced transcriptional activation of bFGF, HB-EGF, and VEGF genes, while the expression of other cytokines such as EGF, PDGF-A, TGF-β1, HGF, and PEDF was not or moderately altered. Hypoxia induced increased expression of the HB-EGF, EGF, PDGF-A, TGF-β1, and VEGF genes, but not of the bFGF gene. Oxidative stress induced gene expression of HB-EGF, but not of bFGF. The hyperosmotic expression of the bFGF gene was dependent on the activation of p38α/β MAPK, JNK, PI3K, and the transcriptional activity of NFAT5. The hyperosmotic expression of the HB-EGF gene was dependent on the activation of p38α/β MAPK, ERK1/2, and JNK. The hyperosmotic expression of bFGF, HB-EGF, and VEGF genes was reduced by inhibitors of TGF-β1 superfamily activin receptor-like kinase receptors and the FGF receptor kinase, respectively. Hyperosmolarity induced secretion of bFGF that was reduced by inhibition of autocrine/paracrine TGF-β1

  10. Autocrine epidermal growth factor signaling stimulates directionally persistent mammary epithelial cell migration

    SciTech Connect

    Maheshwari, Gargi; Wiley, H Steven ); Lauffenburger, Douglas A.

    2001-12-24

    Autocrine receptor/ligand signaling loops were first identified in tumor cells, where it was found that transformation of cells resulted in overexpression of certain growth factors leading to unregulated proliferation of the tumor cells (Sporn and Todaro, 1980). However, in the ensuing decades autocrine signaling has been found to operate in numerous physiological situations (Sporn and Roberts, 1992), including wound healing (Tokumaru et al., 2000), angiogenesis (Seghezzi et al., 1998), and tissue organization during development (Wasserman and Freeman, 1998) and reproductive cycles (Xie et al., 1997). Although it is becoming evident that autocrine loops play crucial roles in regulation of cell function within tissue contexts, it is unclear whether their effects on cell responses are different from the effects of the same ligand presented in exogenous or paracrine manner.

  11. Transforming Growth Factor-β1 Induced Epithelial Mesenchymal Transition is blocked by a chemical antagonist of translation factor eIF4E

    PubMed Central

    Smith, K. A.; Zhou, B.; Avdulov, S.; Benyumov, A.; Peterson, M.; Liu, Y.; Okon, A.; Hergert, P.; Braziunas, J.; Wagner, C. R.; Borok, Z.; Bitterman, P. B.

    2015-01-01

    The epithelial to mesenchymal transition (EMT) imparts disease-defining properties to epithelial cells in cancer and organ fibrosis. Prior studies identify EMT control points at the level of transcription and translation, and indicate that activation of translation initiation factor 4E (eIF4E) is involved in the mechanisms coordinating these two levels of control. Here we show that 4Ei-1, a specific chemical antagonist of the eIF4E-mRNA cap interaction, potently inhibits transforming growth factor beta 1 (TGF-β1) mediated EMT in lung epithelial cells. Upon treatment with TGF-β1, we observed a rapid recruitment of Snail1 mRNA into the actively translated polysome pool accompanied by accumulation of the EMT transcription factor Snail1 in the nucleus. 4Ei-1 blocks ribosome recruitment to the Snail1 transcript thereby preventing accumulation of the Snail1 protein in the nucleus. Our findings establish an obligatory role for upstream translational control of downstream Snail1-mediated transcriptional events in TGF-β1 induced EMT, and provide proof of concept for efforts to pharmacologically modulate the eIF4E-cap interaction as a means to inhibit pathological EMT in the setting of cancer and organ fibrosis. PMID:26678431

  12. Notch induces myofibroblast differentiation of alveolar epithelial cells via transforming growth factor-{beta}-Smad3 pathway.

    PubMed

    Aoyagi-Ikeda, Kana; Maeno, Toshitaka; Matsui, Hiroki; Ueno, Manabu; Hara, Kenichiro; Aoki, Yasuhiro; Aoki, Fumiaki; Shimizu, Takehisa; Doi, Hiroshi; Kawai-Kowase, Keiko; Iso, Tatsuya; Suga, Tatsuo; Arai, Masashi; Kurabayashi, Masahiko

    2011-07-01

    Notch is an ancient cell-signaling system that regulates the specification of cell fate. This study examined the role of Notch in the epithelial-mesenchymal transition (EMT) and myofibroblast differentiation of cultured RLE-6TN cells (i.e., rat alveolar epithelial cells). The activation of Notch, either by ectopic expression of the Notch intracellular domain or by the co-culture of RLE-6TN cells with L-Jagged1 cells, induces the expression of smooth muscle α-actin (SMA) and other mesenchymal marker genes (collagen I and vimentin), and reduces the expression of epithelial marker genes (E-cadherin, occludin, and zonula occludens-1). The pharmacologic inhibition of the endogenous Notch signal significantly inhibited the transforming growth factor-β (TGF-β)-induced expression of SMA. Cell migratory capacity was increased by Notch. Luciferase assays revealed that the CC(A/T)(6)GG (CArG) box and the TGF-β control element (TCE) are required for Notch-induced SMA gene transcription. DNA microarray analysis revealed that members of the TGF-β family as well as Jagged1 were induced in RLE-6TN cells by Notch. Western blot analysis showed that Notch induced the phosphorylation of Smad3, and the TGF-β receptor type I/activin receptor-like kinase 5 (ALK5) kinase inhibitor SB431542 markedly reduced the Notch-induced expression of SMA. Enzyme-linked immunosorbent assays confirmed the production of TGF-β1 from RLE-6TN cells by Notch. Immunohistochemistry of a bleomycin-induced model of pulmonary fibrosis and lung specimens from patients with idiopathic interstitial pneumonias showed that Notch was strongly expressed in myofibroblasts, identified as SMA-positive cells. These data indicate that Notch induces myofibroblast differentiation through a TGF-β-Smad3 pathway that activates SMA gene transcription in a CArG-dependent and TCE-dependent manner in alveolar epithelial cells. Our data also imply that Notch induces the EMT phenotype, with increased migratory behavior in

  13. Epigallocatechin-3-gallate attenuates transforming growth factor-β1 induced epithelial-mesenchymal transition via Nrf2 regulation in renal tubular epithelial cells.

    PubMed

    Wang, Yanqiu; Liu, Na; Su, Xuesong; Zhou, Guangyu; Sun, Guangping; Du, Feng; Bian, Xiaohui; Wang, Bowen

    2015-03-01

    Transforming growth factor-β1 (TGF-β1) induced epithelial-mesenchymal transition (EMT) plays an important role in renal fibrotic process regulation. Epigallocatechin-3-gallate (EGCG) exerts a protective effect against acute renal damage through its anti-oxidative effect by activating the Nrf2 signaling pathway. This study aims to investigate whether EGCG prevents TGF-β1 induced EMT and whether this effect acts via the Nrf2-mediated suppression of TGF-β1 signaling. MTT was used for cytotoxicity of EGCG examination and Western blotting and immunofluorescence were used for protein expression analysis. Results showed that EGCG prevented TGF-β1 mediated EMT and Smad 2 and Smad 3 phosphorylation in a dose dependent manner in NRK-52E cells. In addition, EGCG increased Nrf2 nuclear accumulation. Overexpression of Nrf2 blocked the phosphorylation of Smad 2 and Smad 3 mediated by TGF-β1 and decreased protein expression of plasminogen activator inhibitor 1 (PAI-1) and α-smooth muscle actin (α-SMA). Furthermore, siRNA-mediated knockdown of Nrf2 gene completely blocked the effects of EGCG, indicated by the reduced expressions of type I collagen (Col-I) and α-SMA were restored. In summary, EGCG inhibits TGF-β1 induced EMT and fibrotic proteins expression by Nrf2 activation. This study reveals a possible underlying mechanism of the renal protective effects of EGCG, and may provide a potential candidate to renal fibrosis therapy. PMID:25776510

  14. A novel inhibitor of cyclin-Cdk activity detected in transforming growth factor beta-arrested epithelial cells.

    PubMed Central

    Slingerland, J M; Hengst, L; Pan, C H; Alexander, D; Stampfer, M R; Reed, S I

    1994-01-01

    Transforming growth factor beta (TGF-beta) is a potent inhibitor of epithelial cell growth. Cyclins E and A in association with Cdk2 have been shown to play a role in the G1-to-S phase transition in mammalian cells. We have studied the effects of TGF-beta-mediated growth arrest on G1/S cyclins E and A. Inhibition of cyclin A-associated kinase by TGF-beta is primarily due to a decrease in cyclin A mRNA and protein. By contrast, while TGF-beta inhibits accumulation of cyclin E mRNA, the reduction in cyclin E protein is minimal. Instead, we find that the activation of cyclin E-associated kinase that normally accompanies the G1-to-S phase transition is inhibited. A novel inhibitor of cyclin-Cdk complexes was detected in TGF-beta-treated cell lysates. Inhibition is mediated by a heat-stable protein that targets both Cdk2 and Cdc2 kinases. In G0-arrested cells, a similar inhibitor of Cdk2 kinase was detected. These data suggest the existence of an inhibitor of cyclin-dependent kinases induced under different conditions to mediate antiproliferative responses. Images PMID:8196612

  15. Characterization of mechanical behavior of an epithelial monolayer in response to epidermal growth factor stimulation

    SciTech Connect

    Yang, Ruiguo; Chen, Jennifer Y.; Xi, Ning; Lai, King Wai Chiu; Qu, Chengeng; Fung, Carmen Kar Man; Penn, Lynn S.; Xi, Jun

    2012-03-10

    Cell signaling often causes changes in cellular mechanical properties. Knowledge of such changes can ultimately lead to insight into the complex network of cell signaling. In the current study, we employed a combination of atomic force microscopy (AFM) and quartz crystal microbalance with dissipation monitoring (QCM-D) to characterize the mechanical behavior of A431 cells in response to epidermal growth factor receptor (EGFR) signaling. From AFM, which probes the upper portion of an individual cell in a monolayer of cells, we observed increases in energy dissipation, Young's modulus, and hysteresivity. Increases in hysteresivity imply a shift toward a more fluid-like mechanical ordering state in the bodies of the cells. From QCM-D, which probes the basal area of the monolayer of cells collectively, we observed decreases in energy dissipation factor. This result suggests a shift toward a more solid-like state in the basal areas of the cells. The comparative analysis of these results indicates a regionally specific mechanical behavior of the cell in response to EGFR signaling and suggests a correlation between the time-dependent mechanical responses and the dynamic process of EGFR signaling. This study also demonstrates that a combination of AFM and QCM-D is able to provide a more complete and refined mechanical profile of the cells during cell signaling. -- Highlights: Black-Right-Pointing-Pointer The EGF-induced cellular mechanical response is regionally specific. Black-Right-Pointing-Pointer The EGF-induced cellular mechanical response is time and dose dependent. Black-Right-Pointing-Pointer A combination of AFM and QCM-D provides a more complete mechanical profile of cells.

  16. MATRIX METALLOPROTEINS (MMP)-MEDIATED PHOSPHORYLATION OF THE EPIDERMAL GROWTH FACTOR RECEPTOR (EGFR) IN HUMAN AIRWAY EPITHELIAL CELLS (HAEC) EXPOSED TO ZINC (ZN)

    EPA Science Inventory

    Matrix Metalloproteinase (MMP)-Mediated Phosphorylation of The Epidermal Growth Factor Receptor (EGFR) in Human Airway Epithelial Cells (HAEC) Exposed to Zinc (Zn)
    Weidong Wu, James M. Samet, Robert Silbajoris, Lisa A. Dailey, Lee M. Graves, and Philip A. Bromberg
    Center fo...

  17. Evaluation of transforming growth factor-β1 suppress Pokemon/epithelial-mesenchymal transition expression in human bladder cancer cells.

    PubMed

    Li, Wei; Kidiyoor, Amritha; Hu, Yangyang; Guo, Changcheng; Liu, Min; Yao, Xudong; Zhang, Yuanyuan; Peng, Bo; Zheng, Junhua

    2015-02-01

    Transforming growth factor-β1 (TGF-β1) plays a dual role in apoptosis and in proapoptotic responses in the support of survival in a variety of cells. The aim of this study was to determine the function of TGF-β1 in bladder cancer cells and the relationship with POK erythroid myeloid ontogenic factor (Pokemon). TGF-β1 and its receptors mediate several tumorigenic cascades that regulate cell proliferation, migration, and survival of bladder cancer cells. Bladder cancer cells T24 were treated with different levels of TGF-β1. Levels of Pokemon, E-cadherin, Snail, MMP2, MMP9, Twist, VEGF, and β-catenin messenger RNA (mRNA) and protein were examined by real-time quantitative fluorescent PCR and Western blot analysis, respectively. The effects of TGF-β1 on epithelial-mesenchymal transition of T24 cells were evaluated with wound-healing assay, proliferation of T24 was evaluated with reference to growth curves with MTT assay, and cell invasive ability was investigated by Transwell assay. Data show that Pokemon was inhibited by TGF-β1 treatment; the gene and protein of E-cadherin and β-catenin expression level showed decreased markedly after TGF-β1 treatment (P < 0.05). While the bladder cancer cell after TGF-β1 treatment showed a significantly reduced wound-closing efficiency at 6, 12, and 24 h, mechanistic analyses demonstrated that different levels of TGF-β1 promotes tumor cell growth, migration, and invasion in bladder cancer cells (P < 0.01, P < 0.05, respectively). In summary, our findings suggest that TGF-β1 may inhibit the expression of Pokemon, β-catenin, and E-cadherin. The high expression of TGF-β1 leads to an increase in the phenotype and apical-base polarity of epithelial cells. These changes of cells may result in the recurrence and progression of bladder cancer at last. Related mechanism is worthy of further investigation. PMID:25722217

  18. Osmotic induction of placental growth factor in retinal pigment epithelial cells in vitro: contribution of NFAT5 activity.

    PubMed

    Hollborn, Margrit; Reichmuth, Konrad; Prager, Philipp; Wiedemann, Peter; Bringmann, Andreas; Kohen, Leon

    2016-08-01

    One risk factor of neovascular age-related macular degeneration is systemic hypertension; hypertension is mainly caused by extracellular hyperosmolarity after consumption of dietary salt. In retinal pigment epithelial (RPE) cells, high extracellular osmolarity induces vascular endothelial growth factor (VEGF)-A (Hollborn et al. in Mol Vis 21:360-377, 2015). The aim of the present study was to determine whether extracellular hyperosmolarity and chemical hypoxia trigger the expression of further VEGF family members including placental growth factor (PlGF) in human RPE cells. Hyperosmotic media were made up by addition of 100 mM NaCl or sucrose. Chemical hypoxia was induced by CoCl2. Gene expression was quantified by real-time RT-PCR, and secretion of PlGF-2 was investigated with ELISA. Nuclear factor of activated T cell 5 (NFAT5) was depleted using siRNA. Extracellular hyperosmolarity triggered expression of VEGF-A, VEGF-D, and PlGF genes, and secretion of PlGF-2. Hypoosmolarity decreased PlGF gene expression. Hypoxia induced expression of VEGF-A, VEGF-B, VEGF-D, and PlGF genes. Extracellular hyperosmolarity and hypoxia produced additive PlGF gene expression. Both hyperosmolarity and hypoxia induced expression of KDR and FLT-4 receptor genes, while hyperosmolarity caused neuropilin-2 and hypoxia neuropilin-1 gene expression. The hyperosmotic, but not the hypoxic, PlGF gene expression was in part mediated by NFAT5. The expression of PlGF in RPE cells depends on the extracellular osmolarity. The data suggest that high consumption of dietary salt may exacerbate the angiogenic response of RPE cells in the hypoxic retina via transcriptional activation of various VEGF family member genes. PMID:27230578

  19. Transactivation of the epidermal growth factor receptor mediates muscarinic stimulation of focal adhesion kinase in intestinal epithelial cells.

    PubMed

    Calandrella, Sean O; Barrett, Kim E; Keely, Stephen J

    2005-04-01

    We have previously shown that the Gq protein coupled receptor (GqPCR) agonist, carbachol (CCh), transactivates and recruits epidermal growth factor receptor (EGFr)-dependent signaling mechanisms in intestinal epithelial cells. Increasing evidence suggests that GqPCR agonists can also recruit focal adhesion-dependent signaling pathways in some cell types. Therefore, the aim of the present study was to investigate if CCh stimulates activation of the focal adhesion-associated protein, focal adhesion kinase (FAK), in intestinal epithelia and, if so, to examine the signaling mechanisms involved. Experiments were carried out on monolayers of T84 cells grown on permeable supports. CCh rapidly induced tyrosine phosphorylation of FAK in T84 cells. This effect was accompanied by phosphorylation of another focal adhesion-associated protein, paxillin, and association of FAK with paxillin. CCh-stimulated FAK phosphorylation was inhibited by a chelator of intracellular Ca2+, BAPTA/AM (20 microM), and was mimicked by thapsigargin (2 microM), which mobilizes intracellular Ca2+ in a receptor-independent fashion. CCh also induced association of FAK with the EGFr and FAK phosphorylation was attenuated by an EGFr inhibitor, tyrphostin AG1478, and an inhibitor of Src family kinases, PP2. The actin cytoskeleton disruptor, cytochalasin D (20 microM), abolished FAK phosphorylation in response to CCh but did not alter CCh-induced EGFr or ERK MAPK activation. In summary, these data demonstrate that agonists of GqPCRs have the ability to induce FAK activation in intestinal epithelial cells. GqPCR-induced FAK activation is mediated by via a pathway involving transactivation of the EGFr and alterations in the actin cytoskeleton. PMID:15389641

  20. Environmental Particulate (PM2.5) Augments Stiffness-Induced Alveolar Epithelial Cell Mechanoactivation of Transforming Growth Factor Beta

    PubMed Central

    Dysart, Marilyn M.; Galvis, Boris R.; Russell, Armistead G.; Barker, Thomas H.

    2014-01-01

    Dysfunctional pulmonary homeostasis and repair, including diseases such as pulmonary fibrosis (PF), chronic obstructive pulmonary disease (COPD), and tumorigenesis have been increasing over the past decade, a fact that heavily implicates environmental influences. Several investigations have suggested that in response to increased transforming growth factor - beta (TGFβ) signaling, the alveolar type II (ATII) epithelial cell undergoes phenotypic changes that may contribute to the complex pathobiology of PF. We have previously demonstrated that increased tissue stiffness associated with PF is a potent extracellular matrix (ECM) signal for epithelial cell activation of TGFβ. The work reported here explores the relationship between tissue stiffness and exposure to environmental stimuli in the activation of TGFβ. We hypothesized that exposure of ATII cells to fine particulate matter (PM2.5) will result in enhanced cell contractility, TGFβ activation, and subsequent changes to ATII cell phenotype. ATII cells were cultured on increasingly stiff substrates with or without addition of PM2.5. Exposure to PM2.5 resulted in increased activation of TGFβ, increased cell contractility, and elongation of ATII cells. Most notably, on 8 kPa substrates, a stiffness greater than normal but less than established fibrotic lung, addition of PM2.5 resulted in increased cortical cell stiffness, enhanced actin staining and cell elongation; a result not seen in the absence of PM2.5. Our work suggests that PM2.5 exposure additionally enhances the existing interaction between ECM stiffness and TGFβ that has been previously reported. Furthermore, we show that this additional enhancement is likely a consequence of intracellular reactive oxygen species (ROS) leading to increased TGFβ signaling events. These results highlight the importance of both the micromechanical and biochemical environment in lung disease initiation and suggest that individuals in early stages of lung remodeling

  1. Dynamic Sialylation in Transforming Growth Factor-β (TGF-β)-induced Epithelial to Mesenchymal Transition*

    PubMed Central

    Du, Jun; Hong, Senlian; Dong, Lu; Cheng, Bo; Lin, Liang; Zhao, Bing; Chen, Ye-Guang; Chen, Xing

    2015-01-01

    Epithelial-mesenchymal transition (EMT) is a fundamental process in embryonic development and organ formation. Aberrant regulation of EMT often leads to tumor progression. Changes in cell surface sialylation have recently been implicated in mediating EMT. Herein we report the visualization of dynamic changes of sialylation and glycoproteomic analysis of newly synthesized sialylated proteins in EMT by metabolic labeling of sialylated glycans with azides, followed by click labeling with fluorophores or affinity tags. We discovered that sialylation was down-regulated during EMT but then reverted and up-regulated in the mesenchymal state after EMT, accompanied by mRNA expression level changes of genes involved in the sialic acid biosynthesis. Quantitative proteomic analysis identified a list of sialylated proteins whose biosynthesis was dynamically regulated during EMT. Sialylation of cell surface adherent receptor integrin β4 was found to be down-regulated, which may regulate integrin functions during EMT. Furthermore, a global sialylation inhibitor was used to probe the functional role of sialylation during EMT. We found that inhibition of sialylation promoted EMT. Taken together, our findings suggest the important role of sialylation in regulating EMT and imply its possible function in related pathophysiological events, such as cancer metastasis. PMID:25809486

  2. Lipopolysaccharide (LPS)-Induced Biliary Epithelial Cell NRas Activation Requires Epidermal Growth Factor Receptor (EGFR)

    PubMed Central

    Trussoni, Christy E.; Tabibian, James H.; Splinter, Patrick L.; O’Hara, Steven P.

    2015-01-01

    Cholangiocytes (biliary epithelial cells) actively participate in microbe-induced proinflammatory responses in the liver and contribute to inflammatory and infectious cholangiopathies. We previously demonstrated that cholangiocyte TLR-dependent NRas activation contributes to proinflammatory/ proliferative responses. We test the hypothesis that LPS-induced activation of NRas requires the EGFR. SV40-transformed human cholangiocytes (H69 cells), or low passage normal human cholangiocytes (NHC), were treated with LPS in the presence or absence of EGFR or ADAM metallopeptidase domain 17 (TACE) inhibitors. Ras activation assays, quantitative RT-PCR, and proliferation assays were performed in cells cultured with or without inhibitors or an siRNA to Grb2. Immunofluorescence for phospho-EGFR was performed on LPS-treated mouse samples and specimens from patients with primary sclerosing cholangitis, primary biliary cirrhosis, hepatitis C, and normal livers. LPS-treatment induced an association between the TLR/MyD88 and EGFR/Grb2 signaling apparatus, NRas activation, and EGFR phosphorylation. NRas activation was sensitive to EGFR and TACE inhibitors and correlated with EGFR phosphorylation. The TACE inhibitor and Grb2 depletion prevented LPS-induced IL6 expression (p<0.05) and proliferation (p<0.01). Additionally, cholangiocytes from LPS-treated mouse livers and human primary sclerosing cholangitis (PSC) livers exhibited increased phospho-EGFR (p<0.01). Moreover, LPS-induced mouse cholangiocyte proliferation was inhibited by concurrent treatment with the EGFR inhibitor, Erlotinib. Our results suggest that EGFR is essential for LPS-induced, TLR4/MyD88-mediated NRas activation and induction of a robust proinflammatory cholangiocyte response. These findings have implications not only for revealing the signaling potential of TLRs, but also implicate EGFR as an integral component of cholangiocyte TLR-induced proinflammatory processes. PMID:25915403

  3. Lipopolysaccharide (LPS)-Induced Biliary Epithelial Cell NRas Activation Requires Epidermal Growth Factor Receptor (EGFR).

    PubMed

    Trussoni, Christy E; Tabibian, James H; Splinter, Patrick L; O'Hara, Steven P

    2015-01-01

    Cholangiocytes (biliary epithelial cells) actively participate in microbe-induced proinflammatory responses in the liver and contribute to inflammatory and infectious cholangiopathies. We previously demonstrated that cholangiocyte TLR-dependent NRas activation contributes to proinflammatory/ proliferative responses. We test the hypothesis that LPS-induced activation of NRas requires the EGFR. SV40-transformed human cholangiocytes (H69 cells), or low passage normal human cholangiocytes (NHC), were treated with LPS in the presence or absence of EGFR or ADAM metallopeptidase domain 17 (TACE) inhibitors. Ras activation assays, quantitative RT-PCR, and proliferation assays were performed in cells cultured with or without inhibitors or an siRNA to Grb2. Immunofluorescence for phospho-EGFR was performed on LPS-treated mouse samples and specimens from patients with primary sclerosing cholangitis, primary biliary cirrhosis, hepatitis C, and normal livers. LPS-treatment induced an association between the TLR/MyD88 and EGFR/Grb2 signaling apparatus, NRas activation, and EGFR phosphorylation. NRas activation was sensitive to EGFR and TACE inhibitors and correlated with EGFR phosphorylation. The TACE inhibitor and Grb2 depletion prevented LPS-induced IL6 expression (p<0.05) and proliferation (p<0.01). Additionally, cholangiocytes from LPS-treated mouse livers and human primary sclerosing cholangitis (PSC) livers exhibited increased phospho-EGFR (p<0.01). Moreover, LPS-induced mouse cholangiocyte proliferation was inhibited by concurrent treatment with the EGFR inhibitor, Erlotinib. Our results suggest that EGFR is essential for LPS-induced, TLR4/MyD88-mediated NRas activation and induction of a robust proinflammatory cholangiocyte response. These findings have implications not only for revealing the signaling potential of TLRs, but also implicate EGFR as an integral component of cholangiocyte TLR-induced proinflammatory processes. PMID:25915403

  4. Epithelial-to-mesenchymal transition in breast cancer: a role for insulin-like growth factor I and insulin-like growth factor–binding protein 3?

    PubMed Central

    Zielinska, Hanna A; Bahl, Amit; Holly, Jeff MP; Perks, Claire M

    2015-01-01

    Evidence indicates that for most human cancers the problem is not that gene mutations occur but is more dependent upon how the body deals with damaged cells. It has been estimated that only about 1% of human cancers can be accounted for by unmistakable hereditary cancer syndromes, only up to 5% can be accounted for due to high-penetrance, single-gene mutations, and in total only 5%–15% of all cancers may have a major genetic component. The predominant contribution to the causation of most sporadic cancers is considered to be environmental factors contributing between 58% and 82% toward different cancers. A nutritionally poor lifestyle is associated with increased risk of many cancers, including those of the breast. As nutrition, energy balance, macronutrient composition of the diet, and physical activity levels are major determinants of insulin-like growth factor (IGF-I) bioactivity, it has been proposed that, at least in part, these increases in cancer risk and progression may be mediated by alterations in the IGF axis, related to nutritional lifestyle. Localized breast cancer is a manageable disease, and death from breast cancer predominantly occurs due to the development of metastatic disease as treatment becomes more complicated with poorer outcomes. In recent years, epithelial-to-mesenchymal transition has emerged as an important contributor to breast cancer progression and malignant transformation resulting in tumor cells with increased potential for migration and invasion. Furthermore, accumulating evidence suggests a strong link between components of the IGF pathway, epithelial-to-mesenchymal transition, and breast cancer mortality. Here, we highlight some recent studies highlighting the relationship between IGFs, IGF-binding protein 3, and epithelial-to-mesenchymal transition. PMID:25632238

  5. Transforming Growth Factor-β-Induced RBFOX3 Inhibition Promotes Epithelial-Mesenchymal Transition of Lung Cancer Cells

    PubMed Central

    Kim, Yong-Eun; Kim, Jong Ok; Park, Ki-Sun; Won, Minho; Kim, Kyoon Eon; Kim, Kee K.

    2016-01-01

    The RNA-binding protein Rbfox3 is a well-known splicing regulator that is used as a marker for post-mitotic neurons in various vertebrate species. Although recent studies indicate a variable expression of Rbfox3 in non-neuronal tissues, including lung tissue, its cellular function in lung cancer remains largely unknown. Here, we report that the number of RBFOX3-positive cells in tumorous lung tissue is lower than that in normal lung tissue. As the transforming growth factor-β (TGF-β) signaling pathway is important in cancer progression, we investigated its role in RBFOX3 expression in A549 lung adenocarcinoma cells. TGF-β1 treatment inhibited RBFOX3 expression at the transcriptional level. Further, RBFOX3 depletion led to a change in the expression levels of a subset of proteins related to epithelial-mesenchymal transition (EMT), such as E-cadherin and Claudin-1, during TGF-β1-induced EMT. In immunofluorescence microscopic analysis, mesenchymal morphology was more prominent in RBFOX3-depleted cells than in control cells. These findings show that TGF-β-induced RBFOX3 inhibition plays an important role in EMT and propose a novel role for RBFOX3 in cancer progression. PMID:27432190

  6. Role of mesenchymal-epithelial transition amplification in resistance to anti-epidermal growth factor receptor agents

    PubMed Central

    Morgillo, Floriana; De Mello, Ramon Andrade; Mountzios, Giannis

    2015-01-01

    All patients with epidermal growth factor receptor (EGFR) mutant advanced non-small cell lung cancer (NSCLC) treated with an EGFR-tyrosine kinase inhibitor (EGFR-TKI) such as gefitinib, erlotinib or afatinib will progress after a median of 9-12 months. So far, development of a secondary T790M mutation represents the most common (approximately 60%) mechanism of resistance to these drugs. The relative rarity of mesenchymal-epithelial transition (MET) amplification in NSCLC suggests that this event plays a limited role in primary resistance to EGFR-TKI. In contrast, MET gene amplification has been detected as a secondary event representing one of the most relevant mechanisms involved in the acquired resistance to EGFR-TKIs both in preclinical and clinical studies. The aim of this review is to discuss the role of MET amplification as a mechanism of resistance to anti-EGFR therapies and to review strategies which aim at overcoming this mechanism of resistance, including studies assessing drug combinations targeting both EGFR and MET pathways. PMID:25992380

  7. Aptamer-conjugated gold nanorod for photothermal ablation of epidermal growth factor receptor-overexpressed epithelial cancer

    NASA Astrophysics Data System (ADS)

    Choi, Jihye; Park, Yeonji; Choi, Eun Bi; Kim, Hyun-Ouk; Kim, Dong Joo; Hong, Yoochan; Ryu, Sung-Ho; Lee, Jung Hwan; Suh, Jin-Suck; Yang, Jaemoon; Huh, Yong-Min; Haam, Seungjoo

    2014-05-01

    Biomarker-specific photothermal nanoparticles that can efficiently sense markers that are overexpressed in distinguished adenocarcinomas have attracted much interest in an aspect of efficacy increase of cancer treatment. We demonstrated a promising prospect of a smart photothermal therapy agent employing anti-epidermal growth factor receptor aptamer (AptEGFR)-conjugated polyethylene glycol (PEG) layted gold nanorods (AptEGFR-PGNRs). The cetyltrimethylammonium bromide bilayer on GNRs was replaced with heterobifunctional PEG (COOH-PEG-SH) not only to serve as a biocompatible stabilizer and but also to conjugate Apt. Subsequently, to direct photothermal therapy agent toward epithelial cancer cells, the carboxylated PEGylated GNRs (PGNRs) were further functionalized with Apt using carbodiimide chemistry. Then, to assess the potential as biomarker-specific photothermal therapy agent of synthesized Apt-PGNRs, the optical properties, biocompatibility, colloidal stability, binding affinity, and epicellial cancer cell killing efficacy in vitro/in vivo under near-infrared laser irradiation were investigated. As a result, Apt-PGNRs exhibit excellent tumor targeting ability and feasibility of effective photothermal ablation cancer therapy.

  8. Role of 3'-5'-cyclic adenosine monophosphate on the epidermal growth factor dependent survival in mammary epithelial cells.

    PubMed

    Grinman, Diego Y; Romorini, Leonardo; Presman, Diego M; Rocha-Viegas, Luciana; Coso, Omar A; Davio, Carlos; Pecci, Adali

    2016-01-01

    Epidermal growth factor (EGF) has been suggested to play a key role in the maintenance of epithelial cell survival during lactation. Previously, we demonstrated that EGF dependent activation of PI3K pathway prevents apoptosis in confluent murine HC11 cells cultured under low nutrient conditions. The EGF protective effect is associated with increased levels of the antiapoptotic protein Bcl-XL. Here, we identify the EGF-dependent mechanism involved in cell survival that converges in the regulation of bcl-X expression by activated CREB. EGF induces Bcl-XL expression through activation of a unique bcl-X promoter, the P1; being not only the PI3K/AKT signaling pathway but also the increase in cAMP levels and the concomitant PKA/CREB activation necessary for both bcl-XL upregulation and apoptosis avoidance. Results presented in this work suggest the existence of a novel connection between the EGF receptor and the adenylate cyclase that would have an impact in preventing apoptosis under low nutrient conditions. PMID:26522133

  9. Insulin-like growth factor II blocks apoptosis of N-myc2-expressing woodchuck liver epithelial cells.

    PubMed Central

    Yang, D; Faris, R; Hixson, D; Affigne, S; Rogler, C E

    1996-01-01

    N-myc2 and insulin-like growth factor II (IGF-II) are coordinately overexpressed in the great majority of altered hepatic foci, which are the earliest precancerous lesions observed in the liver of woodchuck hepatitis virus carrier woodchucks, and these genes continue to be overexpressed in hepatocellular carcinomas (HCCs). We have investigated the function of these genes in woodchuck hepatocarcinogenesis by using a woodchuck liver epithelial cell line (WC-3). WC-3 cells react positively with a monoclonal antibody (12.8.5) against woodchuck oval cells, suggesting a lineage relationship with oval cells. Overexpression of N-myc2 in three WC-3 cell lines caused their morphological transformation and increased their growth rate and saturation density in medium containing 10% serum. Removal of serum from the medium increased cell death of the N-myc2-expressing lines, whereas cell death in control lines was minimal. The death of N-myc2-expressing WC-3 cells was accompanied by nucleosomal fragmentation of cellular DNA, and DAPI (4',6-diamidino-2-phenylindole) staining revealed condensation and fragmentation of the nuclei, suggesting that N-myc2-expressing WC-3 cells undergo apoptosis in the absence of serum. In colony regression assays, conducted in the absence of serum, control colonies were stable, while N-myc2-expressing colonies regressed to various degrees. Addition of recombinant human IGF-II to the serum-free medium blocked both cell death and colony regression in all the N-myc2-expressing lines. Therefore, coordinate overexpression of N-myc2 and IGF-II in woodchuck altered hepatic foci may allow cells which otherwise might die to survive and progress to hepatocellular carcinoma. PMID:8709253

  10. A Lactobacillus rhamnosus GG-derived Soluble Protein, p40, Stimulates Ligand Release from Intestinal Epithelial Cells to Transactivate Epidermal Growth Factor Receptor*

    PubMed Central

    Yan, Fang; Liu, Liping; Dempsey, Peter J.; Tsai, Yu-Hwai; Raines, Elaine W.; Wilson, Carole L.; Cao, Hailong; Cao, Zheng; Liu, LinShu; Polk, D. Brent

    2013-01-01

    p40, a Lactobacillus rhamnosus GG (LGG)-derived soluble protein, ameliorates intestinal injury and colitis, reduces apoptosis, and preserves barrier function by transactivation of the EGF receptor (EGFR) in intestinal epithelial cells. The aim of this study is to determine the mechanisms by which p40 transactivates the EGFR in intestinal epithelial cells. Here we show that p40-conditioned medium activates EGFR in young adult mouse colon epithelial cells and human colonic epithelial cell line, T84 cells. p40 up-regulates a disintegrin and metalloproteinase domain-containing protein 17 (ADAM17) catalytic activity, and broad spectrum metalloproteinase inhibitors block EGFR transactivation by p40 in these two cell lines. In ADAM17-deficient mouse colonic epithelial (ADAM17−/− MCE) cells, p40 transactivation of EGFR is blocked, but can be rescued by re-expression with WT ADAM17. Furthermore, p40 stimulates release of heparin binding (HB)-EGF, but not transforming growth factor (TGF)α or amphiregulin, in young adult mouse colon cells and ADAM17−/− MCE cells overexpressing WT ADAM17. Knockdown of HB-EGF expression by siRNA suppresses p40 effects on transactivating EGFR and Akt, preventing apoptosis, and preserving tight junction function. The effects of p40 on HB-EGF release and ADAM17 activation in vivo are examined after administration of p40-containing pectin/zein hydrogel beads to mice. p40 stimulates ADAM17 activity and EGFR activation in colonic epithelial cells and increases HB-EGF levels in blood from WT mice, but not from mice with intestinal epithelial cell-specific ADAM17 deletion. Thus, these data define a mechanism of a probiotic-derived soluble protein in modulating intestinal epithelial cell homeostasis through ADAM17-mediated HB-EGF release, leading to transactivation of EGFR. PMID:24043629

  11. Visceral glomerular epithelial cells can proliferate in vivo and synthesize platelet-derived growth factor B-chain.

    PubMed Central

    Floege, J.; Johnson, R. J.; Alpers, C. E.; Fatemi-Nainie, S.; Richardson, C. A.; Gordon, K.; Couser, W. G.

    1993-01-01

    In glomerular diseases associated with antibody- and complement-mediated injury to endothelial and mesangial cells, cell proliferation is an important early response that precedes matrix accumulation and glomerulosclerosis. In contrast, in diseases in which the visceral glomerular epithelial cell (vGEC) is the principal target of injury, cell proliferation is not a recognized consequence, although vGECs respond in vitro to a variety of growth factors and inflammatory mediators. To explore the possibility that low levels of vGEC proliferation may occur and participate in such diseases, serial studies were done in the passive Heymann nephritis model of membranous nephropathy, in which the vGEC is the primary target of antibody- and C5b-9-mediated injury. The results showed mitotic figures and positive staining for the proliferating cell nuclear antigen in cells whose location defined them as vGECs. The proliferating cell nuclear antigen-positive cells at the edge of the capillary wall were confirmed to be vGECs by double-immunostaining with antibodies to SPARC/osteonectin, which preferentially label vGECs in passive Heymann nephritis. Proliferation of vGECs in vivo was preceded by increased glomerular expression of platelet-derived growth factor (PDGF) B-chain protein and messenger RNA, both of which localized to vGECs. PDGF B-chain protein and messenger RNA were also detected in cultured vGECs. PDGF receptor beta-subunit protein or messenger RNA could not be demonstrated in vGECs in vivo or in vitro, and no growth response of cultured vGECs to PDGF was noted. These results suggest that proliferation of vGECs does occur in a model of glomerular injury induced by antibody and C5b-9 on vGECs. VGEC proliferation and production of PDGF may be involved in the restoration of the capillary wall but could also contribute to local capillary wall injury and proliferation of other cells in Bowman's capsule, interstitium, and tubules. Images Figure 1 Figure 2 Figure 4 Figure 5

  12. Differential regulation of epidermal growth factor receptor by hydrogen peroxide and flagellin in cultured lung alveolar epithelial cells.

    PubMed

    Nishi, Hiroyuki; Maeda, Noriko; Izumi, Shunsuke; Higa-Nakamine, Sayomi; Toku, Seikichi; Kakinohana, Manabu; Sugahara, Kazuhiro; Yamamoto, Hideyuki

    2015-02-01

    In previous studies, we found that stimulation of Toll-like receptor 5 (TLR5) by flagellin induced the activation of mitogen-activated protein kinase (MAPK)-activated protein kinase-2 (MAPKAPK-2) through activation of the p38 MAPK pathway in cultured alveolar epithelial A549 cells. Our studies strongly suggested that MAPKAPK-2 phosphorylated epidermal growth factor receptor (EGFR) at Ser1047. It has been reported that phosphorylation of Ser1047 after treatment with tumor necrosis factor α (TNFα) induced the internalization of EGFR. In the present study, we first found that treatment of A549 cells with hydrogen peroxide induced the activation of MAPKAPK-2 and phosphorylation of EGFR at Ser1047 within 30 min. This was different from flagellin treatment because hydrogen peroxide treatment induced the phosphorylation of EGFR at Tyr1173 as well as Ser1047, indicating the activation of EGFR. We also found that KN93, an inhibitor of CaM kinase II, inhibited the hydrogen peroxide-induced phosphorylation of EGFR at Ser1047 through inhibition of the activation of the p38 MAPK pathway. Furthermore, we examined the internalization of EGFR by three different methods. Flow cytometry with an antibody against the extracellular domain of EGFR and biotinylation of cell surface proteins revealed that flagellin, but not hydrogen peroxide, decreased the amount of cell-surface EGFR. In addition, activation of extracellular signal-regulated kinase by EGF treatment was reduced by flagellin pre-treatment. These results strongly suggested that hydrogen peroxide activated the p38 MAPK pathway via activation of CaM kinase II and that flagellin and hydrogen peroxide regulate the functions of EGFR by different mechanisms. PMID:25542757

  13. Lack of Radiation Dose or Quality Dependence of Epithelial-to-Mesenchymal Transition (EMT) Mediated by Transforming Growth Factor {beta}

    SciTech Connect

    Andarawewa, Kumari L.; Costes, Sylvain V.; Fernandez-Garcia, Ignacio; Chou, William S.; Ravani, Shraddha A.; Park, Howard; Barcellos-Hoff, Mary Helen

    2011-04-01

    Purpose: Epithelial-to-mesenchymal transition (EMT) is a phenotype that alters cell morphology, disrupts morphogenesis, and increases motility. Our prior studies have shown that the progeny of human mammary epithelial cells (HMECs) irradiated with 2 Gy undergoes transforming growth factor {beta} (TGF-{beta})-mediated EMT. In this study we determined whether radiation dose or quality affected TGF-{beta}-mediated EMT. Methods and Materials: HMECs were cultured on tissue culture plastic or in Matrigel (BD Biosciences, San Jose, CA) and exposed to low or high linear energy transfer (LET) and TGF-{beta} (400 pg/mL). Image analysis was used to measure membrane-associated E-cadherin, a marker of functional epithelia, or fibronectin, a product of mesenchymal cells, as a function of radiation dose and quality. Results: E-cadherin was reduced in TGF-{beta}-treated cells irradiated with low-LET radiation doses between 0.03 and 2 Gy compared with untreated, unirradiated cells or TGF-{beta} treatment alone. The radiation quality dependence of TGF-{beta}-mediated EMT was determined by use of 1 GeV/amu (gigaelectron volt / atomic mass unit) {sup 56}Fe ion particles at the National Aeronautics and Space Administration's Space Radiation Laboratory. On the basis of the relative biological effectiveness of 2 for {sup 56}Fe ion particles' clonogenic survival, TGF-{beta}-treated HMECs were irradiated with equitoxic 1-Gy {sup 56}Fe ion or 2-Gy {sup 137}Cs radiation in monolayer. Furthermore, TGF-{beta}-treated HMECs irradiated with either high- or low-LET radiation exhibited similar loss of E-cadherin and gain of fibronectin and resulted in similar large, poorly organized colonies when embedded in Matrigel. Moreover, the progeny of HMECs exposed to different fluences of {sup 56}Fe ion underwent TGF-{beta}-mediated EMT even when only one-third of the cells were directly traversed by the particle. Conclusions: Thus TGF-{beta}-mediated EMT, like other non-targeted radiation effects, is

  14. Snail and Slug Promote Epithelial-Mesenchymal Transition through β-Catenin–T-Cell Factor-4-dependent Expression of Transforming Growth Factor-β3

    PubMed Central

    Medici, Damian; Hay, Elizabeth D.

    2008-01-01

    Members of the Snail family of transcription factors have been shown to induce epithelial-mesenchymal transition (EMT), a fundamental mechanism of embryogenesis and progressive disease. Here, we show that Snail and Slug promote formation of β-catenin–T-cell factor (TCF)-4 transcription complexes that bind to the promoter of the TGF-β3 gene to increase its transcription. Subsequent transforming growth factor (TGF)-β3 signaling increases LEF-1 gene expression causing formation of β-catenin–lymphoid enhancer factor (LEF)-1 complexes that initiate EMT. TGF-β1 or TGF-β2 stimulates this signaling mechanism by up-regulating synthesis of Snail and Slug. TGF-β1- and TGF-β2-induced EMT were found to be TGF-β3 dependent, establishing essential roles for multiple TGF-β isoforms. Finally, we determined that β-catenin–LEF-1 complexes can promote EMT without upstream signaling pathways. These findings provide evidence for a unified signaling mechanism driven by convergence of multiple TGF-β and TCF signaling molecules that confers loss of cell–cell adhesion and acquisition of the mesenchymal phenotype. PMID:18799618

  15. Characterization of insulin-like growth factor-binding proteins secreted by isolated sheep thyroid epithelial cells.

    PubMed

    Wang, J F; Becks, G P; Buckingham, K D; Hill, D J

    1990-06-01

    We have characterized the insulin-like growth factor-binding proteins (IGF-BPs) released by isolated sheep thyroid epithelial cells. Thyroid follicles were isolated with collagenase and cultured in Coon's modified F-12 M (0H medium) supplemented with insulin, cortisol, transferrin, glycyl-histidyl-lysine and somatostatin (5H medium) and TSH (6H medium). Conditioned 0H medium specifically bound both 125I-labelled IGF-I and -II, although binding capacity was reduced following acid-gel filtration to separate endogenous IGF-BP complexes, suggesting some destruction of BPs. The binding of 125I-labelled IGF-I or -II to conditioned (0H) medium was progressively displaced by increasing amounts of unlabelled homologous peptides, while fractionation on concanavalin A-Sepharose showed that the IGF-BPs consisted of both glycoprotein and non-glycoprotein components. The molecular sizes of the IGF-BPs were resolved by separation of 0H medium on SDS-PAGE and ligand blot analysis with 125I-labelled IGF-I or -II. Conditioned medium contained four specific binding species for IGF-II of 19, 30, 38 and 46 kDa; all but the smallest also binding radiolabelled IGF-I. Prior fractionation on concanavalin A-Sepharose showed that the 46 kDa binding species was a glycoprotein. Competition studies with increasing concentrations of unlabelled IGF-I or -II during ligand blotting suggested that the 46 and 30 kDa binding species had a greater affinity for IGF-II than IGF-I, while the 38 kDa had a greater relative affinity for IGF-I. Incubation of cells in 5H medium reduced the abundance of the 46 kDa binding protein, while incubation in 6H medium decreased the release of all binding protein species. Results show that isolated thyroid follicles released several forms of IGF-BP with differing relative affinities for IGF-I and -II. Gross changes seen in the presence of BPs between 0H, 5H and 6H media suggest acute hormonal control of release. PMID:1695663

  16. Novel epidermal growth factor receptor pathway mediates release of human β-defensin 3 from Helicobacter pylori-infected gastric epithelial cells.

    PubMed

    Muhammad, Jibran S; Zaidi, Syed F; Zhou, Yue; Sakurai, Hiroaki; Sugiyama, Toshiro

    2016-04-01

    Persistent Helicobacter pylori (H. pylori) infection in hostile gastric mucosa can result in gastric diseases. Helicobacter pylori induces to express antimicrobial peptides from gastric epithelial cells, especially human β-defensin 3 (hBD3), as an innate immune response, and this expression of hBD3 is mediated by epidermal growth factor receptor (EGFR) activation. In this study, we found that phosphorylation of a serine residue of EGFR via transforming growth factor β-activated kinase-1 (TAK1), and subsequent p38α activation is essential for H. pylori-induced hBD3 release from gastric epithelial cells. We showed that this pathway was dependent on H. pylori type IV secretion system and was independent of H. pylori-derived CagA or peptidoglycan. H. pylori infection induced phosphorylation of serine residue of EGFR, and this phosphorylation was followed by internalization of EGFR; consequently, hBD3 was released at an early phase of the infection. In the presence of TAK1 or p38α inhibitors, synthesis of hBD3 was completely inhibited. Similar results were observed in EGFR-, TAK1- or p38α-knockdown cells. However, NOD1 knockdown in gastric epithelial cells did not inhibit hBD3 induction. Our study has firstly demonstrated that this novel EGFR activating pathway functioned to induce hBD3 at an early phase of H. pylori infection. PMID:26733497

  17. Inhibition of cellular proliferation and modulation of insulin-like growth factor binding proteins by retinoids in a bovine mammary epithelial cell line.

    PubMed

    Woodward, T L; Turner, J D; Hung, H T; Zhao, X

    1996-06-01

    Retinoids are potent inhibitors of growth and tumor progression in many mammary carcinoma cell lines, though regulation of growth in nontumorigenic mammary epithelial cells by retinoids is less clear. Here, we have characterized the inhibition of MAC-T (a nontransformed bovine mammary epithelial cell line) cellular proliferation by retinoids and their role in regulating insulin-like growth factor binding proteins (IGFBPs). Retinoic acid (RA) (100 nM) was a potent inhibitor of MAC-T cell proliferation. Retinol was 10-100 times less effective. Neither retinoid could completely arrest growth at noncytotoxic concentrations. Retinoic acid inhibited cellular proliferation by 1 h (P < .05), but inhibition was fivefold greater by 24 h (P < .01). This second stage of growth inhibition (after 12 h) was dependent upon protein synthesis. However, RA-induced inhibition of cellular proliferation did not persist, with thymidine incorporation increasing toward control levels by 4 days in culture. Retinoic acid was less effective in inhibiting thymidine incorporation when cells were stimulated with insulin, des(1-3) IGF-I, or Long(R3) IGF-I when compared to cells stimulated with native IGF-I or serum. Inhibition of proliferation by RA was associated with increased levels of IGFBP-2 in conditioned media and in plasma membrane preparations. Treatment with insulin or des(1-3) IGF-I resulted in the appearance of IGFBP-3 in conditioned media and on the cell surface. However, RA significantly reduced IGFBP-3 levels in conditioned media and eliminated IGFBP-3 associated with the plasma membrane. Thus, RA is a potent but transient inhibitor of bovine mammary epithelial cell proliferation, and this growth inhibition is correlated with increased IGFBP-2 accumulation and inhibition of IGF-I stimulated IGFBP-3 protein secretion. PMID:8655603

  18. Epidermal Growth Factor Removal or Tyrphostin AG1478 Treatment Reduces Goblet Cells & Mucus Secretion of Epithelial Cells from Asthmatic Children Using the Air-Liquid Interface Model

    PubMed Central

    Parker, Jeremy C.; Douglas, Isobel; Bell, Jennifer; Comer, David; Bailie, Keith; Skibinski, Grzegorz; Heaney, Liam G.; Shields, Michael D.

    2015-01-01

    Rationale Epithelial remodelling in asthma is characterised by goblet cell hyperplasia and mucus hypersecretion for which no therapies exist. Differentiated bronchial air-liquid interface cultures from asthmatic children display high goblet cell numbers. Epidermal growth factor and its receptor have been implicated in goblet cell hyperplasia. Objectives We hypothesised that EGF removal or tyrphostin AG1478 treatment of differentiating air-liquid interface cultures from asthmatic children would result in a reduction of epithelial goblet cells and mucus secretion. Methods In Aim 1 primary bronchial epithelial cells from non-asthmatic (n = 5) and asthmatic (n = 5) children were differentiated under EGF-positive (10ng/ml EGF) and EGF-negative culture conditions for 28 days. In Aim 2, cultures from a further group of asthmatic children (n = 5) were grown under tyrphostin AG1478, a tyrosine kinase inhibitor, conditions. All cultures were analysed for epithelial resistance, markers of differentiation using immunocytochemistry, ELISA for MUC5AC mucin secretion and qPCR for MUC5AC mRNA. Results In cultures from asthmatic children the goblet cell number was reduced in the EGF negative group (p = 0.01). Tyrphostin AG1478 treatment of cultures from asthmatic children had significant reductions in goblet cells at 0.2μg/ml (p = 0.03) and 2μg/ml (p = 0.003) as well as mucus secretion at 2μg/ml (p = 0.04). Conclusions We have shown in this preliminary study that through EGF removal and tyrphostin AG1478 treatment the goblet cell number and mucus hypersecretion in differentiating air-liquid interface cultures from asthmatic children is significantly reduced. This further highlights the epidermal growth factor receptor as a potential therapeutic target to inhibit goblet cell hyperplasia and mucus hypersecretion in asthma. PMID:26057128

  19. Alveolar Epithelial Cells Are Critical in Protection of the Respiratory Tract by Secretion of Factors Able To Modulate the Activity of Pulmonary Macrophages and Directly Control Bacterial Growth

    PubMed Central

    Petursdottir, Dagbjort H.; Periolo, Natalia; Fernández, Carmen

    2013-01-01

    The respiratory epithelium is a physical and functional barrier actively involved in the clearance of environmental agents. The alveolar compartment is lined with membranous pneumocytes, known as type I alveolar epithelial cells (AEC I), and granular pneumocytes, type II alveolar epithelial cells (AEC II). AEC II are responsible for epithelial reparation upon injury and ion transport and are very active immunologically, contributing to lung defense by secreting antimicrobial factors. AEC II also secrete a broad variety of factors, such as cytokines and chemokines, involved in activation and differentiation of immune cells and are able to present antigen to specific T cells. Another cell type important in lung defense is the pulmonary macrophage (PuM). Considering the architecture of the alveoli, a good communication between the external and the internal compartments is crucial to mount effective responses. Our hypothesis is that being in the interface, AEC may play an important role in transmitting signals from the external to the internal compartment and in modulating the activity of PuM. For this, we collected supernatants from AEC unstimulated or stimulated in vitro with lipopolysaccharide (LPS). These AEC-conditioned media were used in various setups to test for the effects on a number of macrophage functions: (i) migration, (ii) phagocytosis and intracellular control of bacterial growth, and (iii) phenotypic changes and morphology. Finally, we tested the direct effect of AEC-conditioned media on bacterial growth. We found that AEC-secreted factors had a dual effect, on one hand controlling bacterial growth and on the other hand increasing macrophage activity. PMID:23147039

  20. β-Galactoside α2,6-Sialyltranferase 1 Promotes Transforming Growth Factor-β-mediated Epithelial-Mesenchymal Transition*

    PubMed Central

    Lu, Jishun; Isaji, Tomoya; Im, Sanghun; Fukuda, Tomohiko; Hashii, Noritaka; Takakura, Daisuke; Kawasaki, Nana; Gu, Jianguo

    2014-01-01

    β-Galactoside α2,6-sialyltranferase 1 (ST6GAL1) catalyzes the addition of terminal α2,6-sialylation to N-glycans. Increased expression of ST6GAL1 has been reported in diverse carcinomas and highly correlates with tumor progression. Here, we report that St6gal1 transcription and α2,6-sialylated N-glycans are up-regulated during TGF-β-induced epithelial-mesenchymal transition (EMT) in GE11 cells, requiring the Sp1 element within the St6gal1 promoter. Knockdown of St6gal1 strongly suppressed TGF-β-induced EMT with a concomitant increase in E-cadherin expression, a major determinant of epithelial cell adherens junctions. Conversely, overexpression of ST6GAL1 increased the turnover of cell surface E-cadherin and promoted TGF-β-induced EMT. Overexpressing β-galactoside α2,3-sialyltranferase 4 had little influence on EMT, indicating specificity for α2,6-sialylation. The basal mesenchymal phenotype of MDA-MB-231 human breast cancer cells was partially reversed by ST6GAL1 silencing. Moreover, ST6GAL1 knockdown inhibited the phosphorylation of Akt, but not Smad2, suggesting that ST6GAL1 contributes to EMT through a non-Smad signaling pathway. Taken together, our data indicate that ST6GAL1 promotes TGF-β-dependent EMT as well as maintenance of the mesenchymal state by growth signaling, providing a plausible mechanism whereby up-regulated ST6GAL1 may promote malignant progression. PMID:25344606

  1. Mechanisms of hepatocyte growth factor-mediated signaling in differentiation of pancreatic ductal epithelial cells into insulin-producing cells

    SciTech Connect

    Li, Xin-Yu; Zhan, Xiao-Rong; Lu, Chong; Liu, Xiao-Min; Wang, Xiao-Chen

    2010-07-30

    Research highlights: {yields} A hypothesis that the differentiation of PDEC is through MAPKs or PI3K/AKT pathways. {yields} Determine if kinases (ERK1/2, p38, JNK, and AKT) are activated in these pathways. {yields} Determine signal pathway(s) that may effect on HGF-induced differentiation of PDEC. {yields} PI3K-AKT pathway is involved in the differentiation of PDECs induced by HGF. {yields} MEK-ERK pathway effect on the proliferation of PDECs but not the differentiation. -- Abstract: Pancreatic ductal epithelial cells (PDECs) were induced to differentiate into insulin-producing cells by hepatocyte growth factor (HGF) in our previous study, but the mechanism through which this induction occurs is still unknown. HGF is a ligand that activates a tyrosine kinase encoded by the c-Met proto-oncogene. This activation is followed by indirect activation of multiple downstream signal transduction pathways (including MAPKs and the PI3K/AKT signaling pathways) that initiate various biological effects. Therefore, we speculated that the differentiation of PDECs is through either the MAPK signaling pathway or the PI3K/AKT signaling pathway. To test this hypothesis, isolated PDECs from adult rats were stimulated by adding HGF to their medium for 28 days. Then, the expression levels of several protein kinases, including MAPKs (ERK1/2, p38, and JNK) and AKT, were determined by Western blotting to determine if specific protein kinases are activated in these pathways. Subsequently, re-isolated from adult rats and cultured PDECs were pre-treated with specific inhibitors of proteins shown to be activated in these signaling pathways; these cells were then induced to differentiate by the addition of HGF. The expression levels of protein kinases were determined by Western blotting, and the differentiation rate of insulin-positive cells was determined by flow cytometry. The change of PDEC differentiation rates were compared between the groups in which cells with or without inhibitors

  2. Targeting matriptase in breast cancer abrogates tumor progression via impairment of stromal-epithelial growth factor signaling

    PubMed Central

    Zoratti, Gina L.; Tanabe, Lauren M.; Varela, Fausto A.; Murray, Andrew S.; Bergum, Christopher; Colombo, Eloic; Lang, Julie; Molinolo, Alfredo A.; Leduc, Richard; Marsault, Eric; Boerner, Julie; List, Karin

    2015-01-01

    Matriptase is an epithelia-specific membrane-anchored serine protease that has received considerable attention in recent years due to its consistent dysregulation in human epithelial tumors, including breast cancer. Mice with reduced levels of matriptase display a significant delay in oncogene-induced mammary tumor formation and blunted tumor growth. The abated tumor growth is associated with a decrease in cancer cell proliferation. Here we demonstrate by genetic deletion and silencing that the proliferation impairment in matriptase deficient breast cancer cells is caused by their inability to initiate activation of the c-Met signaling pathway in response to fibroblast-secreted pro-HGF. Similarly, inhibition of matriptase catalytic activity using a selective small-molecule inhibitor abrogates the activation of c-Met, Gab1 and AKT, in response to pro-HGF, which functionally leads to attenuated proliferation in breast carcinoma cells. We conclude that matriptase is critically involved in breast cancer progression and represents a potential therapeutic target in breast cancer. PMID:25873032

  3. Lactobacillus GG-fermented milk prevents DSS-induced colitis and regulates intestinal epithelial homeostasis through activation of epidermal growth factor receptor

    PubMed Central

    Yoda, Kazutoyo; Miyazawa, Kenji; Hosoda, Masataka; Hiramatsu, Masaru; Yan, Fang; He, Fang

    2014-01-01

    Background Fermented milk is considered one of the best sources for efficient consumption of probiotic strains by hosts to promote good health. The purpose of this study was to investigate the effects of orally administering LGG-fermented milk (LGG milk) on intestinal inflammation and injury and to study the mechanisms of LGG milk's action. Methods LGG milk and non-LGG-fermented milk (non-LGG milk) were administered through gavage to mice before and during dextran sodium sulfate (DSS)-induced intestinal injury and colitis. Inflammatory/injury score and colon length were assessed. Intestinal epithelial cells were treated with the soluble fraction of LGG milk to detect its effects on the epidermal growth factor receptor (EGFR) and its down stream target, Akt activation, cytokine-induced apoptosis, and hydrogen peroxide (H2O2)-induced disruption of tight junctions. Results LGG milk treatment significantly reduced DSS-induced colonic inflammation and injury, and colon shortening in mice, compared to that in non-LGG milk-treated and untreated mice. The soluble fraction of LGG milk, but not non-LGG milk, stimulated activation of EGFR and Akt in a concentration-dependent manner, suppressed cytokine-induced apoptosis, and attenuated H2O2-induced disruption of tight junction complex in the intestinal epithelial cells. These effects of LGG milk were blocked by the EGFR kinase inhibitor. LGG milk, but not non-LGG milk, contained two soluble proteins, p40 and p75, which have been reported to promote survival and growth of intestinal epithelial cells through activation of EGFR. Depletion of p40 and p75 from LGG milk abolished the effects of LGG milk on prevention of cytokine-induced apoptosis and H2O2-induced disruption of tight junctions. Conclusions These results suggest that LGG milk may regulate intestinal epithelial homeostasis and potentially prevent intestinal inflammatory diseases through activation of EGFR by LGG-derived proteins. PMID:23468308

  4. Arctigenin suppresses transforming growth factor-β1-induced expression of monocyte chemoattractant protein-1 and the subsequent epithelial-mesenchymal transition through reactive oxygen species-dependent ERK/NF-κB signaling pathway in renal tubular epithelial cells.

    PubMed

    Li, A; Wang, J; Zhu, D; Zhang, X; Pan, R; Wang, R

    2015-01-01

    Transforming growth factor-β1 (TGF-β1) induces expression of the proinflammatory and profibrotic cytokine monocyte chemoattractant protein-1 (MCP-1) in tubular epithelial cells (TECs) and thereby contributes to the tubular epithelial-mesenchymal transition (EMT), which in turn leads to the progression of tubulointerstitial inflammation into tubulointerstitial fibrosis. Exactly how TGF-β1 causes MCP-1 overexpression and subsequent EMT is not well understood. Using human tubular epithelial cultures, we found that TGF-β1 upregulated the expression of reduced nicotinamide adenine dinucleotide phosphate oxidases 2 and 4 and their regulatory subunits, inducing the production of reactive oxygen species. These reactive species activated a signaling pathway mediated by extracellular signal-regulated kinase (ERK1/2) and nuclear factor-κB (NF-κB), which upregulated expression of MCP-1. Incubating cultures with TGF-β1 was sufficient to induce hallmarks of EMT, such as downregulation of epithelial marker proteins (E-cadherin and zonula occludens-1), induction of mesenchymal marker proteins (α-smooth muscle actin, fibronectin, and vimentin), and elevated cell migration and invasion in an EMT-like manner. Overexpressing MCP-1 in cells exposed to TGF-β1 exacerbated these EMT-like changes. Pretreating cells with the antioxidant and anti-inflammatory compound arctigenin (ATG) protected them against these TGF-β1-induced EMT-like changes; the compound worked by inhibiting the ROS/ERK1/2/NF-κB pathway to decrease MCP-1 upregulation. These findings suggest ATG as a new therapeutic candidate to inhibit or even reverse tubular EMT-like changes during progression to tubulointerstitial fibrosis, and they provide the first clues to how ATG may work. PMID:25968940

  5. Effect of freezing on lens epithelial cell growth.

    PubMed

    Fukaya, Y; Hara, T; Hara, T; Iwata, S

    1988-05-01

    The effect of freezing on the growth of rat lens epithelial cells was studied in vitro. We found that 80% of the lens epithelial cells died after freezing at -45 degrees C for two hours and that the surviving cells could grow with the addition of growth factors or when placed on a sheet of type 4 collagen, but not when placed on a plain plastic culture dish. These results suggest that the surviving cells are at the Go phase of the cell cycle and that type 4 collagen or growth factors can initiate cell division. PMID:3294380

  6. Enhanced wound healing by recombinant Escherichia coli Nissle 1917 via human epidermal growth factor receptor in human intestinal epithelial cells: therapeutic implication using recombinant probiotics.

    PubMed

    Choi, Hye Jin; Ahn, Jung Hoon; Park, Seong-Hwan; Do, Kee Hun; Kim, Juil; Moon, Yuseok

    2012-03-01

    The gastrointestinal mucosa has a remarkable ability to repair damage with the support of epidermal growth factor (EGF), which stimulates epithelial migration and proliferative reepithelialization. For the treatment of mucosal injuries, it is important to develop efficient methods for the localized delivery of mucoactive biotherapeutics. The basic idea in the present study came from the assumption that an intestinal probiotic vehicle can carry and deliver key recombinant medicinal proteins to the injured epithelial target in patients with intestinal ulcerative diseases, including inflammatory bowel disease. The study was focused on the use of the safe probiotic E. coli Nissle 1917, which was constructed to secrete human EGF in conjunction with the lipase ABC transporter recognition domain (LARD). Using the in vitro physically wounded monolayer model, ABC transporter-mediated EGF secretion by probiotic E. coli Nissle 1917 was demonstrated to enhance the wound-healing migration of human enterocytes. Moreover, the epithelial wound closure was dependent on EGF receptor-linked activation, which exclusively involved the subsequent signaling pathway of the mitogen-activated protein kinase kinase (MEK) extracellular-related kinases 1 and 2 (ERK1/2). In particular, the migrating frontier of the wounded edge displayed the strongest EGF receptor-linked signaling activation in the presence of the recombinant probiotic. The present study provides a basis for the clinical application of human recombinant biotherapeutics via an efficient, safe probiotic vehicle. PMID:22184415

  7. Transforming Growth Factor Beta 1 Induces Tight Junction Disruptions and Loss of Transepithelial Resistance Across Porcine Vas Deferens Epithelial Cells1

    PubMed Central

    Pierucci-Alves, Fernando; Yi, Sheng; Schultz, Bruce D.

    2011-01-01

    ABSTRACT Epithelial cells lining the male excurrent duct contribute to male fertility by employing a number of physiological mechanisms that generate a luminal microenvironment conducive to spermatozoa maturation and storage. Among these mechanisms, male duct epithelia establish intercellular tight junctions that constitute a barrier to paracellular diffusion of water, solutes, large molecules, and cells. Mechanisms regulating the male duct epithelial barrier remain unidentified. Transforming growth factor beta (TGFB) is a regulatory cytokine present in high concentrations in human semen. This study examined whether TGFB has any effects on epithelial function exhibited by primary cultures of porcine vas deferens epithelia. TGFB1 exposure caused a 70%–99% decrease in basal transepithelial electrical resistance (RTE, a sensitive indicator of barrier integrity), while a significant decrease in anion secretory response to forskolin was detected at the highest levels of TGFB1 exposure employed. SB431542, a selective TGFB receptor I (TGFBR1) inhibitor, prevented decreases in barrier function. Results also demonstrated that TGFB1 exposure modifies the distribution pattern of tight junction proteins occludin and claudin 7. TGFBR1 is localized at the apical border of the native porcine vas deferens epithelium. Pharmacological inhibition of mitogen-activated protein kinase (MAPK) 11 (also known as p38-MAPK) did not alter the effect of TGFB1 on RTE significantly. These data suggest that epithelia lining the vas deferens are subject to disruptions in the physical barrier if active TGFB becomes bioavailable in the luminal fluid, which might be expected to compromise fertility. PMID:21957188

  8. INHIBITION OF PROTEIN TYROSINE PHOSPHATASE ACTIVITY MEDIATES EPIDERMAL GROWTH FACTOR RECEPTOR SIGNALING IN HUMAN AIRWAY EPITHELIAL CELLS

    EPA Science Inventory

    Epidemiological studies have implicated zinc in the toxicity of ambient particulate matter (PM) inhalation. We previously showed that exposure to metal-laden PM inhibits protein tyrosine phosphatase (PTP) activity in human primary bronchial epithelial cells (HAEC) and leads t...

  9. Ca2+ channels in retinal pigment epithelial cells regulate vascular endothelial growth factor secretion rates in health and disease

    PubMed Central

    Rosenthal, Rita; Heimann, Heinrich; Agostini, Hansjürgen; Martin, Gottfried; Hansen, Lutz Lothar

    2007-01-01

    Purpose Choroidal neovascularization (CNV) is the most severe complication in age-related macular degeneration. The major angiogenic factor involved is vascular endothelial growth factor (VEGF) secreted by the retinal pigment epithelium (RPE). Since RPE cells express neuroendocrine L-type Ca2+ channels we investigated their involvement in VEGF secretion in normal RPE cells and RPE cells from patients with CNV. Methods Freshly isolated and cultured RPE cells were studied using the patch-clamp technique and ELISA-based secretion assays. Results Both freshly isolated and cultured cells showed whole-cell Ba2+ currents with properties of L-type Ca2+ currents: high activation threshold, sensitivity to dihydropyridines (10 μM nifedipine) and slow inactivation. VEGF-A secretion was elevated by BayK8644 (10 μM) or basic fibroblast growth factor (bFGF, 10 ng/ml), both of which are able to activate L-type channels. Cells from CNV tissue also showed nifedipine-sensitive Ba2+ currents, which displayed a voltage-dependent activation at more negative potentials, faster inactivation and changed regulation by tyrosine kinase pp60c-src. The CNV RPE cells showed higher VEGF secretion rates which were reduced by nifedipine. Conclusions Thus, L-type Ca2+ channels in normal RPE cells regulate the secretion of VEGF. RPE cells from eyes with CNV maintain a VEGF secretion regulated by nifedipine-sensitve Ca2+ channels which might be of importance for the development of CNV. PMID:17417605

  10. Cooperation of the ErbB2 receptor and transforming growth factor β in induction of migration and invasion in mammary epithelial cells

    PubMed Central

    Seton-Rogers, Sarah E.; Lu, Yu; Hines, Lisa M.; Koundinya, Malvika; LaBaer, Joshua; Muthuswamy, Senthil K.; Brugge, Joan S.

    2004-01-01

    MCF10A mammary epithelial cells form growth-arrested structures when cultured in three-dimensional basement membrane gels. Activation of the receptor tyrosine kinase ErbB2 induces formation of proliferative structures that share properties with noninvasive early stage lesions. We conducted a genetic screen to identify cDNAs that can cooperate with ErbB2 to induce migration in these cells, with the hypothesis that they would represent candidate “second hits” in the development of invasive breast carcinomas. We found that expression of transforming growth factor (TGF)β1 and TGFβ3 in cells expressing activated ErbB2 induces migration in transwell chambers and invasive behavior in both basement membrane cultures and invasion chambers. The ability of ErbB2 to cooperate with TGFβ correlated with sustained, elevated activation of extracellular signal-regulated kinase (Erk)-mitogen-activated protein kinase. Pharmacological reduction of Erk activity inhibited the cooperative effect of TGFβ and ErbB2 on migration and expression of activated Erk kinase was sufficient to cooperate with TGFβ to induce migration and invasion, suggesting that sustained Erk activation is critical for ErbB2/TGFβ cooperation. In addition, we show that costimulation of ErbB2 and TGFβ induces autocrine secretion of factors that are sufficient to induce migration, but not invasion, by means of both epidermal growth factor receptor-dependent and -independent processes. These results support the role of TGFβ as a pro-invasion factor in the progression of breast cancers with activated ErbB2 and suggest that activation of the Erk and epidermal growth factor receptor pathways are key in mediating these events. PMID:14739340

  11. Inhibition of Plasminogen Activator Inhibitor-1 Attenuates Transforming Growth Factor-β-Dependent Epithelial Mesenchymal Transition and Differentiation of Fibroblasts to Myofibroblasts

    PubMed Central

    Omori, Keitaro; Hattori, Noboru; Senoo, Tadashi; Takayama, Yusuke; Masuda, Takeshi; Nakashima, Taku; Iwamoto, Hiroshi; Fujitaka, Kazunori; Hamada, Hironobu; Kohno, Nobuoki

    2016-01-01

    Transforming growth factor-β (TGF-β) is central during the pathogenesis of pulmonary fibrosis, in which the plasminogen activator inhibitor-1 (PAI-1) also has an established role. TGF-β is also known to be the strongest inducer of PAI-1. To investigate the link between PAI-1 and TGF-β in fibrotic processes, we evaluated the effect of SK-216, a PAI-1-specific inhibitor, in TGF-β-dependent epithelial-mesenchymal transition (EMT) and fibroblast to myofibroblast differentiation. In human alveolar epithelial A549 cells, treatment with TGF-β induced EMT, whereas co-treatment with SK-216 attenuated the occurrence of EMT. The inhibition of TGF-β-induced EMT by SK-216 was also confirmed in the experiment using murine epithelial LA-4 cells. Blocking EMT by SK-216 inhibited TGF-β-induced endogenous production of PAI-1 and TGF-β in A549 cells as well. These effects of SK-216 were not likely mediated by suppressing either Smad or ERK pathways. Using human lung fibroblast MRC-5 cells, we demonstrated that SK-216 inhibited TGF-β-dependent differentiation of fibroblasts to myofibroblasts. We also observed this inhibition by SK-216 in human primary lung fibroblasts. Following these in vitro results, we tested oral administration of SK-216 into mice injected intratracheally with bleomycin. We found that SK-216 reduced the degree of bleomycin-induced pulmonary fibrosis in mice. Although the precise mechanisms underlying the link between TGF-β and PAI-1 regarding fibrotic process were not determined, PAI-1 seems to act as a potent downstream effector on the pro-fibrotic property of TGF-β. In addition, inhibition of PAI-1 activity by a PAI-1 inhibitor exerts an antifibrotic effect even in vivo. These data suggest that targeting PAI-1 as a downstream effector of TGF-β could be a promising therapeutic strategy for pulmonary fibrosis. PMID:26859294

  12. Post Treatment With an FGF Chimeric Growth Factor Enhances Epithelial Cell Proliferation to Improve Recovery From Radiation-Induced Intestinal Damage

    SciTech Connect

    Nakayama, Fumiaki; Hagiwara, Akiko; Umeda, Sachiko; Asada, Masahiro; Goto, Megumi; Oki, Junko; Suzuki, Masashi; Imamura, Toru; Akashi, Makoto

    2010-11-01

    Purpose: A fibroblast growth factor (FGF) 1-FGF2 chimera (FGFC) was created previously and showed greater structural stability than FGF1. This chimera was capable of stimulating epithelial cell proliferation much more strongly than FGF1 or FGF2 even without heparin. Therefore FGFC was expected to have greater biologic activity in vivo. This study evaluated and compared the protective activity of FGFC and FGF1 against radiation-induced intestinal injuries. Methods and Materials: We administered FGFC and FGF1 intraperitoneally to BALB/c mice 24 h before or after total-body irradiation (TBI). The numbers of surviving crypts were determined 3.5 days after TBI with gamma rays at doses ranging from 8 to 12 Gy. Results: The effect of FGFC was equal to or slightly superior to FGF1 with heparin. However, FGFC was significantly more effective in promoting crypt survival than FGF1 (p < 0.01) when 10 {mu}g of each FGF was administered without heparin before irradiation. In addition, FGFC was significantly more effective at promoting crypt survival (p < 0.05) than FGF1 even when administered without heparin at 24 h after TBI at 10, 11, or 12 Gy. We found that FGFC post treatment significantly promoted 5-bromo-2'-deoxyuridine incorporation into crypts and increased crypt depth, resulting in more epithelial differentiation. However, the number of apoptotic cells in FGFC-treated mice decreased to almost the same level as that in FGF1-treated mice. Conclusions: These findings suggest that FGFC strongly enhanced radioprotection with the induction of epithelial proliferation without exogenous heparin after irradiation and is useful in clinical applications for both the prevention and post treatment of radiation injuries.

  13. Assessment of gene expression of intracellular calcium channels, pumps and exchangers with epidermal growth factor-induced epithelial-mesenchymal transition in a breast cancer cell line

    PubMed Central

    2013-01-01

    Background Epithelial-mesenchymal transition (EMT) is a process implicated in cancer metastasis that involves the conversion of epithelial cells to a more mesenchymal and invasive cell phenotype. In breast cancer cells EMT is associated with altered store-operated calcium influx and changes in calcium signalling mediated by activation of cell surface purinergic receptors. In this study, we investigated whether MDA-MB-468 breast cancer cells induced to undergo EMT exhibit changes in mRNA levels of calcium channels, pumps and exchangers located on intracellular calcium storing organelles, including the Golgi, mitochondria and endoplasmic reticulum (ER). Methods Epidermal growth factor (EGF) was used to induce EMT in MDA-MB-468 breast cancer cells. Serum-deprived cells were treated with EGF (50 ng/mL) for 12 h and gene expression was assessed using quantitative RT-PCR. Results and conclusions These data reveal no significant alterations in mRNA levels of the Golgi calcium pump secretory pathway calcium ATPases (SPCA1 and SPCA2), or the mitochondrial calcium uniporter (MCU) or Na+/Ca2+ exchanger (NCLX). However, EGF-induced EMT was associated with significant alterations in mRNA levels of specific ER calcium channels and pumps, including (sarco)-endoplasmic reticulum calcium ATPases (SERCAs), and inositol 1,4,5-trisphosphate receptor (IP3R) and ryanodine receptor (RYR) calcium channel isoforms. The most prominent change in gene expression between the epithelial and mesenchymal-like states was RYR2, which was enriched 45-fold in EGF-treated MDA-MB-468 cells. These findings indicate that EGF-induced EMT in breast cancer cells may be associated with major alterations in ER calcium homeostasis. PMID:23890218

  14. High-Mobility Group Box 1 Mediates Epithelial-to-Mesenchymal Transition in Pulmonary Fibrosis Involving Transforming Growth Factor-β1/Smad2/3 Signaling.

    PubMed

    Li, Liu-Cheng; Li, De-Lin; Xu, Liang; Mo, Xiao-Ting; Cui, Wen-Hui; Zhao, Ping; Zhou, Wen-Cheng; Gao, Jian; Li, Jun

    2015-09-01

    Epithelial-to-mesenchymal transition (EMT) is a crucial event in the cellular origin of myofibroblasts that secrete extracellular matrix in the progression of pulmonary fibrosis (PF). High-mobility group box 1 (HMGB1) is a novel mediator of EMT. However, whether this process involves the recognized transforming growth factor-β1 (TGF-β1)/Smad signaling that also contributes to EMT in PF has not yet been elucidated. Here, we developed a model of PF induced by bleomycin (BLM) in rats and conducted several simulation experiments in A549 (human) and RLE-6TN (rat) alveolar epithelial cell (AEC) lines to unravel the role of TGF-β1/Smad2/3 signaling in HMGB1-mediated EMT. We found that the levels of serum HMGB1 and lung hydroxyproline were severely elevated after BLM administration. Moreover, the protein expression of HMGB1, TGF-β1, phosphorylated Smad2/3 (p-Smad2/3), and mesenchymal markers including α-smooth muscle actin, vimentin, and type I collagen were significantly increased with the reduced protein expression of an epithelial marker (E-cadherin) in the rat model by Western blot or immunohistochemical analysis. In addition, the uptake of both exogenous TGF-β1 and HMGB1 by AECs could induce EMT; meanwhile, HMGB1 dramatically enhanced TGF-β1 expression and triggered Smad2/3 phosphorylation. In contrast, TGF-β1 deficiency evidently ameliorated HMGB1-mediated EMT with reduced p-Smad2/3 in A549 cells. It provides new insights that HMGB1 release from injured lungs promotes AEC damage through induction of the EMT process, in which TGF-β1/Smad2/3 signaling is activated and contributes to PF. These results suggest that HMGB1 may constitute a therapeutic target for developing antifibrotic agents for abnormal lung remodeling. PMID:26126535

  15. Insulin-like growth factor I and risk of epithelial invasive ovarian cancer by tumour characteristics: results from the EPIC cohort

    PubMed Central

    Ose, J; Fortner, R T; Schock, H; Peeters, P H; Onland-Moret, N C; Bueno-de-Mesquita, H B; Weiderpass, E; Gram, I T; Overvad, K; Tjonneland, A; Dossus, L; Fournier, A; Baglietto, L; Trichopoulou, A; Benetou, V; Trichopoulos, D; Boeing, H; Masala, G; Krogh, V; Matiello, A; Tumino, R; Popovic, M; Obón-Santacana, M; Larrañaga, N; Ardanaz, E; Sánchez, M-J; Menéndez, V; Chirlaque, M-D; Travis, R C; Khaw, K-T; Brändstedt, J; Idahl, A; Lundin, E; Rinaldi, S; Kuhn, E; Romieu, I; Gunter, M J; Merritt, M A; Riboli, E; Kaaks, R

    2015-01-01

    Background: Prospective studies on insulin-like growth factor I (IGF-I) and epithelial ovarian cancer (EOC) risk are inconclusive. Data suggest risk associations vary by tumour characteristics. Methods: We conducted a nested case–control study in the European Prospective Investigation into Cancer and Nutrition (EPIC) to evaluate IGF-I concentrations and EOC risk by tumour characteristics (n=565 cases). Multivariable conditional logistic regression models were used to estimate associations. Results: We observed no association between IGF-I and EOC overall or by tumour characteristics. Conclusions: In the largest prospective study to date was no association between IGF-I and EOC risk. Pre-diagnostic serum IGF-I concentrations may not influence EOC risk. PMID:25349976

  16. Hepatocyte growth factor-induced up-regulation of Twist drives epithelial-mesenchymal transition in a canine mammary tumour cell line.

    PubMed

    Yoshida, Kota; Choisunirachon, Nan; Saito, Tomochika; Matsumoto, Kaori; Saeki, Kohei; Mochizuki, Manabu; Nishimura, Ryohei; Sasaki, Nobuo; Nakagawa, Takayuki

    2014-12-01

    Epithelial-mesenchymal transition (EMT) is a crucial step in tumour progression. However, the molecular mechanisms underlying EMT in canine tumours remain to be elucidated. In this study, the similarity or difference in the molecular mechanism of EMT in canine cells was evaluated and compared with that reported in human and mouse cells. We used eight cell lines derived from canine mammary cancers. Stimulation with hepatocyte growth factor (HGF) increased cell motility and changed EMT-related markers towards mesenchyme in CHMm cell line. These changes were accompanied by an increase in Twist expression and did not occur in CHMm transfected with Twist siRNA, indicating that Twist plays a key role in this phenomenon in CHMm. However, the down-regulation of E-cadherin was not observed by HGF stimulation. Further studies are required to elucidate the difference between human and canine Twist. PMID:25278141

  17. Transforming growth factor-β1 regulated phosphorylated AKT and interferon gamma expressions are associated with epithelial cell survival in rhesus macaque colon explants.

    PubMed

    Pahar, Bapi; Pan, Diganta; Lala, Wendy; Kenway-Lynch, Carys S; Das, Arpita

    2015-05-01

    Transforming growth factor-β1 (TGF-β1) is an important immunoregulatory cytokine that plays an obligate role in regulating T-cell functions. Here, we demonstrated the role of TGF-β1 in regulating the survival of intestinal epithelial cells (ECs) in rhesus colon explant cultures using either anti-TGF-β1 antibody or recombinant TGF-β1 proteins. Neutralization of endogenous TGF-β1 using anti-TGF-β1 antibodies induced apoptosis of both intestinal ECs and lamina propria (LP) cells. Additionally, endogenous TGF-β1 blocking significantly increased expression of IFNγ, TNFα, CD107a and Perforin in LP cells compared to media and isotype controls. A significant decrease in pAKT expression was detected in anti-TGF-β1 MAbs treated explants compared to isotype and rTGF-β1 protein treated explants. Our results demonstrated TGF-β1 regulated pAKT and IFNγ expressions were associated with epithelial cell survival in rhesus macaque colon explants and suggest a potential role of mucosal TGF-β1 in regulating intestinal homeostasis and EC integrity. PMID:25769244

  18. Transforming growth factor-β1 regulated phosphorylated AKT and interferon gamma expressions are associated with epithelial cell survival in rhesus macaque colon explants

    PubMed Central

    Pahar, Bapi; Pan, Diganta; Lala, Wendy; Kenway-Lynch, Carys S.; Das, Arpita

    2015-01-01

    Transforming growth factor-β1 (TGF-β1) is an important immunoregulatory cytokine that plays an obligate role in regulating T-cell functions. Here, we demonstrated the role of TGF-β1 in regulating the survival of intestinal epithelial cells (ECs) in rhesus colon explant cultures using either anti-TGF-β1 antibody or recombinant TGF-β1 proteins. Neutralization of endogenous TGF-β1 using anti-TGF-β1 antibodies induced apoptosis of both intestinal ECs and lamina propria (LP) cells. Additionally, endogenous TGF-β1 blocking significantly increased expression of IFNγ, TNFα, CD107a and Perforin in LP cells compared to media and isotype controls. A significant decrease in pAKT expression was detected in anti-TGF-β1 MAbs treated explants compared to isotype and rTGF-β1 protein treated explants. Our results demonstrated TGF-β1 regulated pAKT and IFNγ expressions were associated with epithelial cell survival in rhesus macaque colon explants and suggest a potential role of mucosal TGF-β1 in regulating intestinal homeostasis and EC integrity. PMID:25769244

  19. Hepatocyte Growth Factor and MET Support Mouse Enteric Nervous System Development, the Peristaltic Response, and Intestinal Epithelial Proliferation in Response to Injury

    PubMed Central

    Avetisyan, Marina; Wang, Hongtao; Schill, Ellen Merrick; Bery, Saya; Grider, John R.; Hassell, John A.; Stappenbeck, Thaddeus

    2015-01-01

    Factors providing trophic support to diverse enteric neuron subtypes remain poorly understood. We tested the hypothesis that hepatocyte growth factor (HGF) and the HGF receptor MET might support some types of enteric neurons. HGF and MET are expressed in fetal and adult enteric nervous system. In vitro, HGF increased enteric neuron differentiation and neurite length, but only if vanishingly small amounts (1 pg/ml) of glial cell line-derived neurotrophic factor were included in culture media. HGF effects were blocked by phosphatidylinositol-3 kinase inhibitor and by MET-blocking antibody. Both of these inhibitors and MEK inhibition reduced neurite length. In adult mice, MET was restricted to a subset of calcitonin gene-related peptide-immunoreactive (IR) myenteric plexus neurons thought to be intrinsic primary afferent neurons (IPANs). Conditional MET kinase domain inactivation (Metfl/fl; Wnt1Cre+) caused a dramatic loss of myenteric plexus MET-IR neurites and 1–1′-dioctodecyl-3,3,3′,3′-tetramethylindocarbocyamine perchlorate (DiI) labeling suggested reduced MET-IR neurite length. In vitro, Metfl/fl; Wnt1Cre+ mouse bowel had markedly reduced peristalsis in response to mucosal deformation, but normal response to radial muscle stretch. However, whole-bowel transit, small-bowel transit, and colonic-bead expulsion were normal in Metfl/fl; Wnt1Cre+ mice. Finally, Metfl/fl; Wnt1Cre+ mice had more bowel injury and reduced epithelial cell proliferation compared with WT animals after dextran sodium sulfate treatment. These results suggest that HGF/MET signaling is important for development and function of a subset IPANs and that these cells regulate intestinal motility and epithelial cell proliferation in response to bowel injury. SIGNIFICANCE STATEMENT The enteric nervous system has many neuronal subtypes that coordinate and control intestinal activity. Trophic factors that support these neuron types and enhance neurite growth after fetal development are not well

  20. Interleukin 24 is induced by the RET/PTC3 oncoprotein and is an autocrine growth factor for epithelial cells.

    PubMed

    Shinohara, Shogo; Rothstein, Jay L

    2004-09-30

    Thyroid cancers, like hematological malignancies, are commonly associated with chromosomal translocations leading to the formation of fusion proteins. Through altered signaling by fusion proteins, cell death and survival pathways are disrupted and the physiological balance of cell-cell communication may be lost. A consequence of this disruption is the release of factors by stressed cells that alert the host. One type of host response is leukocytic infiltration that may develop into chronic inflammation or autoimmune disease. Although inflammation can be associated with neoplastic tissue, the mechanism driving this process is largely unknown. Therefore, to address the mechanism of cancer inflammation we investigated the effects of an oncogene in a murine model system. A comprehensive genetic analysis revealed several soluble factors that were induced by RET/papillary thyroid carcinoma (PTC)3 gene expression including several proinflammatory cytokines, chemokines and immunologically relevant costimulatory molecules. Following a large genetic screen using RP3-expressing thyroid cells, we identified a highly abundant transcript and later identified it as interleukin 24 (Il24), a cytokine with diverse tumor suppressor and inflammatory activities. We show that RET/PTC3 induces Il24 expression in rat thyrocytes and that this expression is dependent on the signaling properties of its tyrosine kinase. Likewise, RET/PTC3 induces large amounts of Il24 following expression in murine thyrocytes, but its expression is dramatically reduced in poorly differentiated carcinomas, a finding that parallels the loss of RET/PTC3 expression. Consistent with its behavior as a tumor suppressor, the loss of Il24 coincided with the loss of RET/PTC3 in poorly differentiated mouse tumors. A functional role of Il24 in the autocrine growth/survival of RET/PTC3-expressing thyroid cells was identified helping to support its role in cellular transformation. These data suggest that the induction of

  1. Vascular endothelial growth factor polymorphisms and a synchronized examination of plasma and tissue expression in epithelial ovarian cancers.

    PubMed

    Bhaskari, J; Premalata, C S; Shilpa, V; Rahul, B; Pallavi, V R; Ramesh, G; Krishnamoorthy, Lakshmi

    2016-01-01

    In this study, we have analyzed six genetic polymorphisms of the VEGF-A gene and correlated the genetic data with plasma and tissue expression of VEGF-A in epithelial ovarian carcinomas. A total of 130 cases including 95 malignant carcinomas, 17 low malignant potential and 18 benign tumours were studied. rs699947, rs833061, rs1570360, rs2010963, rs1413711 and rs3025039 were studied by polymerase chain reaction and restriction fragment length polymorphism (PCR-RFLP). Plasma levels of VEGF-A were estimated by enzyme-linked immunosorbent assay (ELISA) and tissue expression of VEGF-A by immunohistochemistry (IHC). Four polymorphisms of the above excluding rs699947 and rs3025039 showed significant association with malignancy, and we observed the presence of positive correlation between haplotype CCGGCC and increased expression of VEGF-A in both plasma and tissues which also correlated with poor prognosis and recurrence suggesting a probable increase in resistance to treatment in such carriers. Highly upregulated tissue expression of VEGF-A was seen in all epithelial ovarian carcinomas with intensity of expression increasing from benign to malignant cases. ELISA data from our study showed an increase in circulating levels of VEGF-A in malignancies. VEGF-A plasma levels can be employed as a biomarker for high-grade malignancy in epithelial ovarian cancers alongside tissue expression and CA-125 levels. This study is unique due to the fact that a simultaneous analysis of plasma and tissue expression has been demonstrated and is a first such study in epithelial ovarian cancers and representing the Indian population (South-east Asian) synchronized with genetic polymorphism data as well. PMID:26264619

  2. Epidermal growth factor: Porcine uterine luminal epithelial cell migratory signal during the peri-implantation period of pregnancy.

    PubMed

    Jeong, Wooyoung; Jung, Seoungo; Bazer, Fuller W; Song, Gwonhwa; Kim, Jinyoung

    2016-01-15

    The majority of early conceptus mortality in pregnancy occurs during the peri-implantation period, suggesting that this period is important for conceptus viability and the establishment of pregnancy. Successful establishment of pregnancy in all mammalian species depends on the orchestrated molecular events that transpire at the conceptus-uterine interface during the peri-implantation period of pregnancy. This maternal-conceptus interaction is especially crucial in pigs because they have a non-invasive epitheliochorial placentation during a protracted peri-implantation period. During the pre-implantation period of pregnancy, conceptus survival and the establishment of pregnancy depend on the developing conceptus receiving an adequate supply of histotroph which contains a wide range of nutrients and growth factors. Evidence links epidermal growth factor (EGF) to embryogenesis or implantation in various mammalian species. EGF exhibits potential growth-promoting activities on the conceptus and endometrium; however, in the case of pigs, little is known its functions, especially their regulatory mechanisms at the maternal-conceptus interface. EGF receptor (EGFR) mRNA and protein are abundant in endometrial luminal (LE) and glandular (GE) epithelia and conceptus trophectoderm on Days 13-14 of pregnancy, suggesting that EGF provides an autocrine signal to uterine LE and GE just prior to implantation. Therefore, the objectives of this study were to determine: 1) the potential intracellular signaling pathways responsible for the activities of EGF in porcine uterine LE (pLE) cells; and 2) the changes in cellular activities induced by EGF. EGF treatment of pLE cells increased the abundance of phosphorylated (p)-ERK1/2, p-P70RSK and p-RPS6 compared to that for control cells. Furthermore, EGF-stimulated phosphorylation of ERK1/2 MAPK was inhibited in pLE cells transfected with an EGFR siRNA compared with control siRNA-transfected pLE cells. Moreover, EGF stimulated migration of

  3. Upregulated epidermal growth factor receptor expression following near-infrared irradiation simulating solar radiation in a three-dimensional reconstructed human corneal epithelial tissue culture model

    PubMed Central

    Tanaka, Yohei; Nakayama, Jun

    2016-01-01

    Background and objective Humans are increasingly exposed to near-infrared (NIR) radiation from both natural (eg, solar) and artificial (eg, electrical appliances) sources. Although the biological effects of sun and ultraviolet (UV) exposure have been extensively investigated, the biological effect of NIR radiation is still unclear. We previously reported that NIR as well as UV induces photoaging and standard UV-blocking materials, such as sunglasses, do not sufficiently block NIR. The objective of this study was to investigate changes in gene expression in three-dimensional reconstructed corneal epithelial tissue culture exposed to broad-spectrum NIR irradiation to simulate solar NIR radiation that reaches human tissues. Materials and methods DNA microarray and quantitative real-time polymerase chain reaction analysis were used to assess gene expression levels in a three-dimensional reconstructed corneal epithelial model composed of normal human corneal epithelial cells exposed to water-filtered broad-spectrum NIR irradiation with a contact cooling (20°C). The water-filter allowed 1,000–1,800 nm wavelengths and excluded 1,400–1,500 nm wavelengths. Results A DNA microarray with >62,000 different probes showed 25 and 150 genes that were up- or downregulated by at least fourfold and twofold, respectively, after NIR irradiation. In particular, epidermal growth factor receptor (EGFR) was upregulated by 19.4-fold relative to control cells. Quantitative real-time polymerase chain reaction analysis revealed that two variants of EGFR in human corneal epithelial tissue were also significantly upregulated after five rounds of 10 J/cm2 irradiation (P<0.05). Conclusion We found that NIR irradiation induced the upregulated expression of EGFR in human corneal cells. Since over half of the solar energy reaching the Earth is in the NIR region, which cannot be adequately blocked by eyewear and thus can induce eye damage with intensive or long-term exposure, protection from both

  4. Secreted or nonsecreted forms of acidic fibroblast growth factor produced by transfected epithelial cells influence cell morphology, motility, and invasive potential.

    PubMed Central

    Jouanneau, J; Gavrilovic, J; Caruelle, D; Jaye, M; Moens, G; Caruelle, J P; Thiery, J P

    1991-01-01

    Addition of exogenous acidic fibroblast growth factor (aFGF) to NBT-II epithelial carcinoma cells results in fibroblastic transformation and cell motility. We have generated aFGF-producing NBT-II cells by transfection with recombinant expression vectors containing human aFGF cDNA, or the human aFGF cDNA coupled to a signal peptide (SP) sequence. The effects of the nonsecreted and the secreted 16-kDa growth factor on the morphology, motility, and cell invasive potential (gelatinase activity) were compared. aFGF coupled to a SP was actively secreted out of the producing cells. The secretion of aFGF was not necessary for induction of gelatinase activity, as this was observed in NBT-II cells producing aFGF with or without SP. Production of aFGF, whether secreted or not secreted, resulted in increased in vitro motility of most isolated clones; however, there was no correlation between aFGF level and motility rate. The data suggest that expression of aFGF in NBT-II cells induces metastatic potential through an autocrine or intracrine mechanism. Images PMID:1707175

  5. Sofalcone, a gastric mucosa protective agent, increases vascular endothelial growth factor via the Nrf2-heme-oxygenase-1 dependent pathway in gastric epithelial cells

    SciTech Connect

    Shibuya, Akiko; Onda, Kenji; Kawahara, Hirofumi; Uchiyama, Yuka; Nakayama, Hiroko; Omi, Takamasa; Nagaoka, Masayoshi; Matsui, Hirofumi; Hirano, Toshihiko

    2010-07-30

    Research highlights: {yields} Sofalcone increases HO-1 in gastric epithelial cells. {yields} The induction of HO-1 by sofalcone treatment follows the activation of Nrf2. {yields} The production of VEGF by sofalcone treatment is mediated by HO-1 induction. -- Abstract: Sofalcone, 2'-carboxymethoxy-4,4-bis(3-methyl-2-butenyloxy)chalcone, is an anti-ulcer agent that is classified as a gastric mucosa protective agent. Recent studies indicate heat shock proteins such as HSP32, also known as heme-oxygenase-1(HO-1), play important roles in protecting gastrointestinal tissues from several stresses. We have previously reported that sofalcone increases the expression of HO-1 in adipocytes and pre-adipocytes, although the effect of sofalcone on HO-1 induction in gastrointestinal tissues is not clear. In the current study, we investigated the effects of sofalcone on the expression of HO-1 and its functional role in rat gastric epithelial (RGM-1) cells. We found that sofalcone increased HO-1 expression in RGM-1 cells in both time- and concentration-dependent manners. The HO-1 induction was associated with the nuclear translocation of nuclear factor (erythroid-derived 2)-like 2 (Nrf2) in RGM-1 cells. We also observed that sofalcone increased vascular endothelial growth factor (VEGF) production in the culture medium. Treatment of RGM-1 cells with an HO-1 inhibitor (tin-protoporphyrin), or HO-1 siRNA inhibited sofalcone-induced VEGF production, suggesting that the effect of sofalcone on VEGF expression is mediated by the HO-1 pathway. These results suggest that the gastroprotective effects of sofalcone are partly exerted via Nrf2-HO-1 activation followed by VEGF production.

  6. Prognostic impact of KI67, p53, human epithelial growth factor receptor 2, topoisomerase IIalpha, epidermal growth factor receptor, and nm23 expression of ovarian carcinomas and disseminated tumor cells in the bone marrow.

    PubMed

    Schindlbeck, C; Hantschmann, P; Zerzer, M; Jahns, B; Rjosk, D; Janni, W; Rack, B; Sommer, H; Friese, K

    2007-01-01

    Examination of tumor biological factors for prognostic and predictive indicators is not part of routine testing in ovarian cancer. As in other tumors, the detection of hematogenous tumor spread could help to estimate the risk of metastatic disease. We examined the expression of p53, KI67, topoisomerase IIalpha (Top IIa), epidermal growth factor receptor (EGFR), human epithelial growth factor receptor 2 (HER2) and nm23 in tumor tissues from 90 patients with ovarian cancer. All underwent bone marrow (BM) aspiration and screening for disseminated tumor cells in the bone marrow (DTC-BM) at primary diagnosis. BM aspiration, cytospin preparation, and immunocytochemical staining with the anticytokeratin antibody (A45-B/B3) were done following a standardized protocol. The expression of p53, KI67, Top IIa, EGFR, HER2, and nm23 was evaluated by immunohistochemistry on paraffin-embedded tissue samples and classified by percentage of stained cells or immunoreactive score (IRS). The prognostic impact of the individual factors together with standard histologic parameters was calculated by univariate and multivariate analyses. Expression rates for HER2 (2+/3+: 34.5%), KI67 (median 30%), p53 (median IRS 5), and Top IIa (median IRS 4) were relatively high, whereas nm23 (median IRS 2) and EGFR (IRS 0: 61%) showed weak staining. In 21/90 patients (23.3%), DTC-BM (>/=1/2 x 10(6) cells) could be detected. The presence of DTC-BM was inversely related to nodal status (P = .015) but not to the other factors examined. Tumor stage (P = .02), lymph node involvement (P = .003), grade (P = .046), postoperative tumor residue (P < .001), peritoneal seeding (P = .02), and KI67 (P = .046) significantly correlated with overall survival (OS) after a median observation time of 28 months (2-105). The finding of ascites was borderline significant (P = .050). The presence of DTC-BM (P = .04) and KI67 positivity (P = .02) predicted reduced distant disease-free survival. By multivariate analysis

  7. Myeloid-Epithelial-Reproductive Receptor Tyrosine Kinase and Milk Fat Globule Epidermal Growth Factor 8 Coordinately Improve Remodeling After Myocardial Infarction via Local Delivery of Vascular Endothelial Growth Factor

    PubMed Central

    Howangyin, Kiave-Yune; Zlatanova, Ivana; Pinto, Cristina; Ngkelo, Anta; Cochain, Clément; Rouanet, Marie; Vilar, José; Lemitre, Mathilde; Stockmann, Christian; Fleischmann, Bernd K.; Mallat, Ziad

    2016-01-01

    Background— In infarcted heart, improper clearance of dying cells by activated neighboring phagocytes may precipitate the transition to heart failure. We analyzed the coordinated role of 2 major mediators of efferocytosis, the myeloid-epithelial-reproductive protein tyrosine kinase (Mertk) and the milk fat globule epidermal growth factor (Mfge8), in directing cardiac remodeling by skewing the inflammatory response after myocardial infarction. Methods and Results— We generated double-deficient mice for Mertk and Mfge8 (Mertk−/−/Mfge8−/−) and challenged them with acute coronary ligature. Compared with wild-type, Mertk-deficient (Mertk−/−), or Mfge8-deficient (Mfge8−/−) animals, Mertk−/−/Mfge8−/− mice displayed greater alteration in cardiac function and remodeling. Mertk and Mfge8 were expressed mainly by cardiac Ly6CHigh and Low monocytes and macrophages. In parallel, Mertk−/−/Mfge8−/− bone marrow chimeras manifested increased accumulation of apoptotic cells, enhanced fibrotic area, and larger infarct size, as well as reduced angiogenesis. We found that the abrogation of efferocytosis affected neither the ability of circulating monocytes to infiltrate cardiac tissue nor the number of resident Ly6CHigh and Ly6CHow monocytes/macrophages populating the infarcted milieu. In contrast, combined Mertk and Mfge8 deficiency in Ly6CHigh/Ly6CLow monocytes/macrophages either obtained from in vitro differentiation of bone marrow cells or isolated from infarcted hearts altered their capacity of efferocytosis and subsequently blunted vascular endothelial growth factor A (VEGFA) release. Using LysMCre+/VEGFAfl/fl mice, we further identified an important role for myeloid-derived VEGFA in improving cardiac function and angiogenesis. Conclusions— After myocardial infarction, Mertk- and Mfge8-expressing monocyte/macrophages synergistically engage the clearance of injured cardiomyocytes, favoring the secretion of VEGFA to locally repair the

  8. Involvement of epithelial-to-mesenchymal transition and associated transforming growth factor-β/Smad signaling in paraquat-induced pulmonary fibrosis.

    PubMed

    Han, Ying-Ying; Shen, Peng; Chang, Wen-Xiu

    2015-12-01

    Paraquat (PQ) is a highly toxic herbicide which is able to induce pulmonary fibrosis in humans and animals. The epithelial‑to‑mesenchymal transition (EMT) was demonstrated to be an important factor in pulmonary fibrosis. However, it has remained elusive whether PQ induces pulmonary fibrosis via EMT, which was therefore investigated in the present study. In addition, the underlying mechanisms of PQ‑induced EMT were examined in vitro. Hematoxylin and eosin staining of rat lung tissues demonstrated that PQ induced pulmonary fibrosis in vivo. Western blot analysis then revealed that the expression of epithelial cell marker E‑cadherin was significantly decreased, while the expression of mesenchymal markers α‑smooth‑muscle actin and vimentin was significantly increased in rat lung tissues and A549 cells following PQ treatment. Transforming growth factor (TGF)‑β/Smad signaling was also induced by PQ as evidenced by increased expression of TGF‑β1 and Smad2. However, PQ‑induced EMT in A549 cells was abolished by transfection with TGF‑β1‑specific small hairpin RNA. In conclusion, the present study demonstrated that PQ induced EMT in vivo and in vitro, which may be an important process in the development of PQ‑induced pulmonary fibrosis. In addition, TGF-β/Smad signaling was involved in PQ-induced EMT. PMID:26499763

  9. Microvesicles Derived from Human Umbilical Cord Mesenchymal Stem Cells Facilitate Tubular Epithelial Cell Dedifferentiation and Growth via Hepatocyte Growth Factor Induction

    PubMed Central

    Wu, Shuai; Zou, Xiang-yu; Zhang, Guang-yuan; Gu, Di; Miao, Shuai; Zhu, Ying-jian; Sun, Jie; Du, Tao

    2015-01-01

    During acute kidney injury (AKI), tubular cell dedifferentiation initiates cell regeneration; hepatocyte growth factor (HGF) is involved in modulating cell dedifferentiation. Mesenchymal stem cell (MSC)-derived microvesicles (MVs) deliver RNA into injured tubular cells and alter their gene expression, thus regenerating these cells. We boldly speculated that MVs might induce HGF synthesis via RNA transfer, thereby facilitating tubular cell dedifferentiation and regeneration. In a rat model of unilateral AKI, the administration of MVs promoted kidney recovery. One of the mechanisms of action is the acceleration of tubular cell dedifferentiation and growth. Both in vivo and in vitro, rat HGF expression in damaged rat tubular cells was greatly enhanced by MV treatment. In addition, human HGF mRNA present in MVs was delivered into rat tubular cells and translated into the HGF protein as another mechanism of HGF induction. RNase treatment abrogated all MV effects. In the in vitro experimental setting, the conditioned medium of MV-treated injured tubular cells, which contains a higher concentration of HGF, strongly stimulated cell dedifferentiation and growth, as well as Erk1/2 signaling activation. Intriguingly, these effects were completely abrogated by either c-Met inhibitor or MEK inhibitor, suggesting that HGF induction is a crucial contributor to the acceleration of cell dedifferentiation and growth. All these findings indicate that MV-induced HGF synthesis in damaged tubular cells via RNA transfer facilitates cell dedifferentiation and growth, which are important regenerative mechanisms. PMID:25793303

  10. The Marine-Derived Oligosaccharide Sulfate MS80, a Novel Transforming Growth Factor β1 Inhibitor, Reverses Epithelial Mesenchymal Transition Induced by Transforming Growth Factor-β1 and Suppresses Tumor Metastasis.

    PubMed

    Zhou, Ji; You, Wenjie; Sun, Guangqiang; Li, Yixuan; Chen, Bi; Ai, Jing; Jiang, Handong

    2016-10-01

    Metastasis accounts for the majority of cancer-related deaths. Transforming growth factor β (TGF-β) is believed to promote late-stage cancer progression and metastasis by inducing epithelial-mesenchymal transition (EMT). We previously reported that MS80, a novel oligosaccharide sulfate, inhibits TGF-β1-induced pulmonary fibrosis by binding TGF-β1. In our study MS80 effectively inhibited TGF-β/Smad signaling in lung cancer cells, breast cancer cells, and model cell lines. In addition, MS80 inhibited TGF-β1-induced EMT, motility, and invasion in vitro. Moreover, MS80 significantly inhibited lung metastasis in orthotopic 4T1 xenografts. Notably, the MS80 treatment significantly increased the infiltration of CD8(+) T cells and decreased the infiltration of regulatory T cells in primary tumors and spleens in mice bearing 4T1 xenografts. Therefore, MS80 is a novel and promising candidate for treating metastatic malignancies by targeting TGF-β1-induced EMT and mediating immunosuppression. PMID:27432893

  11. D1398G Variant of MET Is Associated with Impaired Signaling of Hepatocyte Growth Factor in Alveolar Epithelial Cells and Lung Fibroblasts.

    PubMed

    Atanelishvili, Ilia; Shirai, Yuichiro; Akter, Tanjina; Noguchi, Atsushi; Ash, Kurt T; Misra, Suniti; Ghatak, Sibnath; Silver, Richard M; Bogatkevich, Galina S

    2016-01-01

    Pulmonary fibrosis represents the terminal stage of a diverse group of lung diseases including scleroderma associated interstitial lung disease. The molecular mechanisms underlying the pathogenesis of lung fibrosis are not well understood and there is a great need for more effective treatment for this lethal disease. We recently discovered a small fragment of hepatocyte growth factor (HGF) receptor MET as a peptide designated "M10," with strong antifibrotic properties. Furthermore, we showed that aspartic acid at position 1398 of MET is essential for M10 generation. The current study was undertaken to investigate the D1398G variant of MET in which aspartic acid at position 1398 was mutated to glycine resulting in loss of M10. We demonstrate that lung fibroblasts, A549, and primary alveolar epithelial cells (AEC) expressing D1398G MET exhibit reduced auto-phosphorylation on tyrosine residues and reduced activation of Ras and MAPK. HGF treatment of scleroderma lung fibroblasts as well as HGF treatment of TGFβ-treated normal lung fibroblasts transfected with wild type MET is associated with decreased collagen, connective tissue growth factor (CTGF, CCN2) and smooth muscle α-actin (SMA). However, HGF has no such effects in cells transfected with MET D1398G. Cisplatin- and FasL-induced apoptosis is significantly reduced in AEC transfected with MET wild type, but not in AEC transfected with MET D1398G. We conclude that the D1398G variant of MET is associated with compromised phosphorylation and impaired HGF signaling in lung fibroblasts and AEC, two cell types implicated in the pathogenesis of pulmonary fibrosis associated with scleroderma. Ongoing studies will explore the frequency of this variant and its relationship to pulmonary outcomes in scleroderma patients. PMID:27584154

  12. Preparation of redispersible liposomal dry powder using an ultrasonic spray freeze-drying technique for transdermal delivery of human epithelial growth factor

    PubMed Central

    Yin, Fei; Guo, Shiyan; Gan, Yong; Zhang, Xinxin

    2014-01-01

    In this work, an ultrasonic spray freeze-drying (USFD) technique was used to prepare a stable liposomal dry powder for transdermal delivery of recombinant human epithelial growth factor (rhEGF). Morphology, particle size, entrapment efficiency, in vitro release, and skin permeability were systematically compared between rhEGF liposomal dry powder prepared using USFD and that prepared using a conventional lyophilization process. Porous and spherical particles with high specific area were produced under USFD conditions. USFD effectively avoided formation of ice crystals, disruption of the bilayer structure, and drug leakage during the liposome drying process, and maintained the stability of the rhEGF liposomal formulation during storage. The reconstituted rhEGF liposomes prepared from USFD powder did not show significant changes in morphology, particle size, entrapment efficiency, or in vitro release characteristics compared with those of rhEGF liposomes before drying. Moreover, the rhEGF liposomal powder prepared with USFD exhibited excellent enhanced penetration in ex vivo mouse skin compared with that for powder prepared via conventional lyophilization. The results suggest that ultrasonic USFD is a promising technique for the production of stable protein-loaded liposomal dry powder for application to the skin. PMID:24729702

  13. Loss of BRCA1 leads to an increase in epidermal growth factor receptor expression in mammary epithelial cells, and epidermal growth factor receptor inhibition prevents estrogen receptor-negative cancers in BRCA1-mutant mice

    PubMed Central

    2011-01-01

    Introduction Women who carry a BRCA1 mutation typically develop "triple-negative" breast cancers (TNBC), defined by the absence of estrogen receptor (ER), progesterone receptor and Her2/neu. In contrast to ER-positive tumors, TNBCs frequently express high levels of epidermal growth factor receptor (EGFR). Previously, we found a disproportionate fraction of progenitor cells in BRCA1 mutation carriers with EGFR overexpression. Here we examine the role of EGFR in mammary epithelial cells (MECs) in the emergence of BRCA1-related tumors and as a potential target for the prevention of TNBC. Methods Cultures of MECs were used to examine EGFR protein levels and promoter activity in response to BRCA1 suppression with inhibitory RNA. EGFR was assessed by immunoblot and immunofluorescence analysis, real-time reverse transcriptase-polymerase chain reaction assay (RT-PCR) and flow cytometry. Binding of epidermal growth factor (EGF) to subpopulations of MECs was examined by Scatchard analysis. The responsiveness of MECs to the EGFR inhibitor erlotinib was assessed in vitro in three-dimensional cultures and in vivo. Mouse mammary tumor virus-Cre recombinase (MMTV-Cre) BRCA1flox/flox p53+/- mice were treated daily with erlotinib or vehicle control, and breast cancer-free survival was analyzed using the Kaplan-Meier method. Results Inhibition of BRCA1 in MECs led to upregulation of EGFR with an inverse correlation of BRCA1 with cellular EGFR protein levels (r2 = 0.87) and to an increase in cell surface-expressed EGFR. EGFR upregulation in response to BRCA1 suppression was mediated by transcriptional and posttranslational mechanisms. Aldehyde dehydrogenase 1 (ALDH1)-positive MECs expressed higher levels of EGFR than ALDH1-negative MECs and were expanded two- to threefold in the BRCA1-inhibited MEC population. All MECs were exquisitely sensitive to EGFR inhibition with erlotinib in vitro. EGFR inhibition in MMTV-Cre BRCA1flox/flox p53+/- female mice starting at age 3 months increased

  14. Change of retinal pigment epithelial atrophy after anti-vascular endothelial growth factor treatment in exudative age-related macular degeneration

    PubMed Central

    Kim, Moosang; Kim, Eung Suk; Seo, Kyung Hoon; Yu, Seung-Young; Kwak, Hyung-Woo

    2016-01-01

    Purpose: The study aimed to investigate the quantitative changes of retinal pigment epithelial (RPE) atrophy during a 24-month follow-up period of anti-vascular endothelial growth factor (VEGF) for exudative age-related macular degeneration (AMD). Materials and Methods: This is a retrospective study. Sixty-five eyes of 62 consecutive patients with naïve exudative AMD who had received treatment with anti-VEGF therapy and followed for more 24 months were enrolled. All patients received three initial monthly injections of anti-VEGF (ranibizumab or bevacizumab), followed by pro re nata or treat-and-extend protocol. Color fundus image, optical coherence tomography, and fundus autofluorescence were evaluated for RPE atrophy. Multiple regression analysis was performed to investigate the predictive factors found during univariate analysis to identify an association with increased RPE atrophic areas. Results: The mean number of anti-VEGF treatments was 9.18. RPE atrophic area was 1.293 ± 1.298 mm2 at baseline and enlarged to 2.394 ± 1.940 mm2 after 24 months, which differed significantly (P = 0.001). Multiple regression analysis revealed that larger areas of RPE atrophy at month 4 and larger numbers of anti-VEGF treatments were associated with increased RPE atrophic areas. Conclusions: RPE atrophy progresses in eyes with exudative AMD during anti-VEGF treatment. Larger areas of RPE atrophy at month 4 and larger numbers of anti-VEGF injections were associated with an increased risk of progression of RPE atrophy the following treatment. These findings may be useful to clinicians using intravitreal anti-VEGF for the treatment of exudative AMD, both for selecting an appropriate treatment plan and for predicting the progression of RPE atrophy. PMID:27488150

  15. Transforming growth factor-β-sphingosine kinase 1/S1P signaling upregulates microRNA-21 to promote fibrosis in renal tubular epithelial cells.

    PubMed

    Liu, Xiujuan; Hong, Quan; Wang, Zhen; Yu, Yanyan; Zou, Xin; Xu, Lihong

    2016-02-01

    Renal fibrosis is a progressive pathological change characterized by tubular cell apoptosis, tubulointerstitial fibroblast proliferation, and excessive deposition of extracellular matrix (ECM). miR-21 has been implicated in transforming growth factor-β (TGF-β)-stimulated tissue fibrosis. Recent studies showed that sphingosine kinase/sphingosine-1-phosphate (SphK/S1P) are also critical for TGF-β-stimulated tissue fibrosis; however, it is not clear whether SphK/S1P interacts with miR-21 or not. In this study, we hypothesized that SphK/S1P signaling is linked to upregulation of miR-21 by TGF-β. To verify this hypothesis, we first determined that miR-21 was highly expressed in renal tubular epithelial cells (TECs) stimulated with TGF-β by using qRT-PCR and Northern blotting. Simultaneously, inhibition of miR-21, mediated by the corresponding antimir, markedly decreased the expression and deposition of type I collagen, fibronectin (Fn), cysteine-rich protein 61 (CCN1), α-smooth muscle actin, and fibroblast-specific protein1 in TGF-β-treated TECs. ELISA and qRT-PCR were used to measure the S1P and SphK1 levels in TECs. S1P production was induced by TGF-β through activation of SphK1. Furthermore, it was observed that TGF-β-stimulated upregulation of miR-21 was abolished by SphK1 siRNA and was restored by the addition of exogenous S1P. Blocking S1PR2 also inhibited upregulation of miR-21. Additionally, miR-21 overexpression attenuated the repression of TGF-β-stimulated ECM deposition and epithelial-mesenchymal transition by SphK1 and S1PR2 siRNA. In summary, our study demonstrates a link between SphK1/S1P and TGF-β-induced miR-21 in renal TECs and may represent a novel therapeutic target in renal fibrosis. PMID:26376826

  16. Epithelial to mesenchymal transition in arsenic-transformed cells promotes angiogenesis through activating β-catenin–vascular endothelial growth factor pathway

    SciTech Connect

    Wang, Zhishan; Humphries, Brock; Xiao, Hua; Jiang, Yiguo; Yang, Chengfeng

    2013-08-15

    Arsenic exposure represents a major health concern increasing cancer risks, yet the mechanism of arsenic carcinogenesis has not been elucidated. We and others recently reported that cell malignant transformation by arsenic is accompanied by epithelial to mesenchymal transition (EMT). However, the role of EMT in arsenic carcinogenesis is not well understood. Although previous studies showed that short term exposure of endothelial cells to arsenic stimulated angiogenesis, it remains to be determined whether cells that were malignantly transformed by long term arsenic exposure have a pro-angiogenic effect. The objective of this study was to investigate the effect of arsenic-transformed human bronchial epithelial cells that underwent EMT on angiogenesis and the underlying mechanism. It was found that the conditioned medium from arsenic-transformed cells strongly stimulated tube formation by human umbilical vein endothelial cells (HUVECs). Moreover, enhanced angiogenesis was detected in mouse xenograft tumor tissues resulting from inoculation of arsenic-transformed cells. Mechanistic studies revealed that β-catenin was activated in arsenic-transformed cells up-regulating its target gene expression including angiogenic-stimulating vascular endothelial growth factor (VEGF). Stably expressing microRNA-200b in arsenic-transformed cells that reversed EMT inhibited β-catenin activation, decreased VEGF expression and reduced tube formation by HUVECs. SiRNA knockdown β-catenin decreased VEGF expression. Adding a VEGF neutralizing antibody into the conditioned medium from arsenic-transformed cells impaired tube formation by HUVECs. Reverse transcriptase-PCR analysis revealed that the mRNA levels of canonical Wnt ligands were not increased in arsenic-transformed cells. These findings suggest that EMT in arsenic-transformed cells promotes angiogenesis through activating β-catenin–VEGF pathway. - Highlights: • Arsenic-transformed cells that underwent EMT displayed a pro

  17. HPV16 Down-Regulates the Insulin-Like Growth Factor Binding Protein 2 to Promote Epithelial Invasion in Organotypic Cultures

    PubMed Central

    Pickard, Adam; McDade, Simon S.; McFarland, Marie; McCluggage, W. Glenn; Wheeler, Cosette M.; McCance, Dennis J.

    2015-01-01

    Cervical cancer is a multi-stage disease caused by human papillomaviruses (HPV) infection of cervical epithelial cells, but the mechanisms regulating disease progression are not clearly defined. Using 3-dimensional organotypic cultures, we demonstrate that HPV16 E6 and E7 proteins alter the secretome of primary human keratinocytes resulting in local epithelial invasion. Mechanistically, absence of the IGF-binding protein 2 (IGFBP2) caused increases in IGFI/II signalling and through crosstalk with KGF/FGFR2b/AKT, cell invasion. Repression of IGFBP2 is mediated by histone deacetylation at the IGFBP2 promoter and was reversed by treatment with histone deacetylase (HDAC) inhibitors. Our in vitro findings were confirmed in 50 invasive cancers and 79 cervical intra-epithelial neoplastic lesions caused by HPV16 infection, where IGFBP2 levels were reduced with increasing disease severity. In summary, the loss of IGFBP2 is associated with progression of premalignant disease, and sensitises cells to pro-invasive IGF signalling, and together with stromal derived factors promotes epithelial invasion. PMID:26107517

  18. Epidermal Growth Factor Receptor Kinase Inhibitors Synergize with TCDD to Induce CYP1A1/1A2 in Human Breast Epithelial MCF10A Cells.

    PubMed

    Joiakim, Aby; Mathieu, Patricia A; Shelp, Catherine; Boerner, Julie; Reiners, John J

    2016-05-01

    CYP1A1andCYP1A2are transcriptionally activated in the human normal breast epithelial cell line MCF10A following exposure to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). Shifting MCF10A cultures to medium deficient in serum and epidermal growth factor (EGF) caused rapid reductions in the activated (i.e., phosphorylated) forms of extracellular regulated kinases (ERKs) and the epidermal growth factor receptor (EGFR). Shifting to serum/EGF-deficient medium also enhanced TCDD-mediated induction of cytochrome P450 (CYP)1A1 Treatment of cells cultured in complete medium with the EGFR inhibitors gefitinib (Iressa), AG1478, and CI-1033 resulted in concentration-dependent reductions of active EGFR and ERKs, and increased CYP1A1 mRNA content ∼3- to 18-fold above basal level. EGFR inhibitors synergized with TCDD and resulted in transient CYP1A1 and CYP1A2 mRNA accumulations ∼8-fold greater (maximum at 5 hours) than that achieved with only TCDD. AG1478, gefitinib, and TCDD individually induced small increases (∼1.2- to 2.5-fold) in CYP1A1 protein content but did not cause additive or synergistic accumulations of CYP1A1 protein when used in combination. The mitogen-activated protein kinase kinase inhibitor PD184352 inhibited ERK and EGFR activation in a concentration-dependent fashion without causing CYP1A1 mRNA accumulation. However, cotreatment with PD184352 potentiated TCDD-mediatedCYP1A1induction. TCDD-mediated induction ofCYP1A1in MCF7-TETon-EGFR cells, a MCF7 variant in which EGFR expression can be controlled, was not affected by the activity status of EGFR or ERKs. Hence, EGFR signaling mutes both basal and ligand-induced expression of two aryl hydrocarbon receptor-responsive P450s in MCF10A cultures. However, these effects are cell context-dependent. Furthermore, CYP1A1 mRNA and protein abundance are not closely coupled in MCF10A cultures. PMID:26953171

  19. Identification of Keratinocyte Growth Factor as a Target of microRNA-155 in Lung Fibroblasts: Implication in Epithelial-Mesenchymal Interactions

    PubMed Central

    Chevalier, Benoit; Puisségur, Marie-Pierre; Lebrigand, Kevin; Robbe-Sermesant, Karine; Bertero, Thomas; Lino Cardenas, Christian L.; Courcot, Elisabeth; Rios, Géraldine; Fourre, Sandra; Lo-Guidice, Jean-Marc; Marcet, Brice; Cardinaud, Bruno; Barbry, Pascal; Mari, Bernard

    2009-01-01

    Background Epithelial-mesenchymal interactions are critical in regulating many aspects of vertebrate embryo development, and for the maintenance of homeostatic equilibrium in adult tissues. The interactions between epithelium and mesenchyme are believed to be mediated by paracrine signals such as cytokines and extracellular matrix components secreted from fibroblasts that affect adjacent epithelia. In this study, we sought to identify the repertoire of microRNAs (miRNAs) in normal lung human fibroblasts and their potential regulation by the cytokines TNF-α, IL-1β and TGF-β. Methodology/Principal Findings MiR-155 was significantly induced by inflammatory cytokines TNF-α and IL-1β while it was down-regulated by TGF-β. Ectopic expression of miR-155 in human fibroblasts induced modulation of a large set of genes related to “cell to cell signalling”, “cell morphology” and “cellular movement”. This was consistent with an induction of caspase-3 activity and with an increase in cell migration in fibroblasts tranfected with miR-155. Using different miRNA bioinformatic target prediction tools, we found a specific enrichment for miR-155 predicted targets among the population of down-regulated transcripts. Among fibroblast-selective targets, one interesting hit was keratinocyte growth factor (KGF, FGF-7), a member of the fibroblast growth factor (FGF) family, which owns two potential binding sites for miR-155 in its 3′-UTR. Luciferase assays experimentally validated that miR-155 can efficiently target KGF 3′-UTR. Site-directed mutagenesis revealed that only one out of the 2 potential sites was truly functional. Functional in vitro assays experimentally validated that miR-155 can efficiently target KGF 3′-UTR. Furthermore, in vivo experiments using a mouse model of lung fibrosis showed that miR-155 expression level was correlated with the degree of lung fibrosis. Conclusions/Significance Our results strongly suggest a physiological function of miR-155 in

  20. Enhancement of Recombinant Adeno-Associated Virus Type 2-Mediated Transgene Expression in a Lung Epithelial Cell Line by Inhibition of the Epidermal Growth Factor Receptor

    PubMed Central

    Smith, Andrew D.; Collaco, Roy F.; Trempe, James P.

    2003-01-01

    Recombinant adeno-associated viruses (rAAVs) have attracted considerable interest as gene delivery systems because they show long-term expression in vivo and transduce numerous cell types. Limitations to successful gene transduction from rAAVs have prompted investigations of a variety of treatments to enhance transgene expression from rAAV vectors. Tyrphostin-1, an epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor, dramatically enhances rAAV transgene expression. Elegant studies have demonstrated that a single-strand D-sequence-binding protein (ssDBP) is phosphorylated by EGFR and binds to the D sequence element in the AAV terminal repeat (TR). Binding of the Tyr-phosphorylated ssDBP prevents conversion of single-stranded vector DNA to a double-strand conformation. We observed dramatic increases in transgene expression in lung epithelial cells (IB3) with tyrphostin treatment. Gel shift analysis of ssDBP revealed that its DNA binding characteristics were unchanged after tyrphostin treatment or adenovirus infection. Tyrphostin stimulated rAAV transgene expression to a greater extent than adenovirus coinfection. Southern hybridizations revealed that the vector DNA remained in the single-strand conformation in tyrphostin-treated cells but double-stranded replicative form monomer DNA was most abundant in adenovirus-infected cells. Northern analyses revealed that tyrphostin treatment enhanced mRNA accumulation more than in adenovirus-infected cultures even though replicative form DNA was undetectable. Analysis of the JNK, ERK, and p38K mitogen-activated protein kinase pathways revealed that tyrphostin treatment stimulated the activity of JNK and p38K. Our data suggest that tyrphostin-induced alteration of stress response pathways results in dramatic enhancement of transcription on linear vector DNA templates in the IB3 cell line. These results expand the downstream targets of the EGFR in regulating rAAV transduction. PMID:12743297

  1. Cross-Talk between Human Mast Cells and Bronchial Epithelial Cells in Plasminogen Activator Inhibitor-1 Production via Transforming Growth Factor-β1

    PubMed Central

    Lee, Sun H.; Kato, Atsushi; Takabayashi, Tetsuji; Kulka, Marianna; Shin, Soon C.; Schleimer, Robert P.

    2015-01-01

    Previous reports suggest that plasminogen activator inhibitor-1 (PAI-1) promotes airway remodeling and that human and mouse mast cells (MCs) are an important source of PAI-1. In the present study we investigated MC–epithelial cell (EC) interactions in the production of PAI-1. We stimulated the human MC line LAD2 with IgE-receptor cross-linking and collected the supernatants. We incubated the human bronchial EC line BEAS-2B with the LAD2 supernatants and measured the level of PAI-1. When the supernatants from IgE-stimulated LAD2 were added to BEAS-2B, there was a significant enhancement of PAI-1 production by BEAS-2B. When we treated the MC supernatants with a transforming growth factor (TGF)-β1 neutralizing antibody, the MC-derived induction of PAI-1 from BEAS-2B was completely abrogated. Although TGF-β1 mRNA was constitutively expressed in resting LAD2, it was not highly induced by IgE-mediated stimulation. Nonetheless, active TGF-β1 protein was significantly increased in LAD2 after IgE-mediated stimulation. Active TGF-β1 produced by primary cultured human MCs was significantly reduced in the presence of a chymase inhibitor, suggesting a role of MC chymase as an activator of latent TGF-β1. This study indicates that stimulation of human MCs by IgE receptor cross-linking triggers activation of TGF-β1, at least in part via chymase, which in turn induces the production of PAI-1 by bronchial ECs. Our data suggest that human MCs may play an important role in airway remodeling in asthma as a direct source of PAI-1 and by activating bronchial ECs to produce further PAI-1 via a TGF-β1–mediated activation pathway. PMID:24987792

  2. Cross-talk between human mast cells and bronchial epithelial cells in plasminogen activator inhibitor-1 production via transforming growth factor-β1.

    PubMed

    Cho, Seong H; Lee, Sun H; Kato, Atsushi; Takabayashi, Tetsuji; Kulka, Marianna; Shin, Soon C; Schleimer, Robert P

    2015-01-01

    Previous reports suggest that plasminogen activator inhibitor-1 (PAI-1) promotes airway remodeling and that human and mouse mast cells (MCs) are an important source of PAI-1. In the present study we investigated MC-epithelial cell (EC) interactions in the production of PAI-1. We stimulated the human MC line LAD2 with IgE-receptor cross-linking and collected the supernatants. We incubated the human bronchial EC line BEAS-2B with the LAD2 supernatants and measured the level of PAI-1. When the supernatants from IgE-stimulated LAD2 were added to BEAS-2B, there was a significant enhancement of PAI-1 production by BEAS-2B. When we treated the MC supernatants with a transforming growth factor (TGF)-β1 neutralizing antibody, the MC-derived induction of PAI-1 from BEAS-2B was completely abrogated. Although TGF-β1 mRNA was constitutively expressed in resting LAD2, it was not highly induced by IgE-mediated stimulation. Nonetheless, active TGF-β1 protein was significantly increased in LAD2 after IgE-mediated stimulation. Active TGF-β1 produced by primary cultured human MCs was significantly reduced in the presence of a chymase inhibitor, suggesting a role of MC chymase as an activator of latent TGF-β1. This study indicates that stimulation of human MCs by IgE receptor cross-linking triggers activation of TGF-β1, at least in part via chymase, which in turn induces the production of PAI-1 by bronchial ECs. Our data suggest that human MCs may play an important role in airway remodeling in asthma as a direct source of PAI-1 and by activating bronchial ECs to produce further PAI-1 via a TGF-β1-mediated activation pathway. PMID:24987792

  3. Protective effects of transforming growth factor β2 in intestinal epithelial cells by regulation of proteins associated with stress and endotoxin responses.

    PubMed

    Nguyen, Duc Ninh; Jiang, Pingping; Jacobsen, Susanne; Sangild, Per T; Bendixen, Emøke; Chatterton, Dereck E W

    2015-01-01

    Transforming growth factor (TGF)-β2 is an important anti-inflammatory protein in milk and colostrum. TGF-β2 supplementation appears to reduce gut inflammatory diseases in early life, such as necrotizing enterocolitis (NEC) in young mice. However, the molecular mechanisms by which TGF-β2 protects immature intestinal epithelial cells (IECs) remain to be more clearly elucidated before interventions in infants can be considered. Porcine IECs PsIc1 were treated with TGF-β2 and/or lipopolysaccharide (LPS), and changes in the cellular proteome were subsequently analyzed using two-dimensional gel electrophoresis-MS and LC-MS-based proteomics. TGF-β2 alone induced the differential expression of 13 proteins and the majority of the identified proteins were associated with stress responses, TGF-β and Toll-like receptor 4 signaling cascades. In particular, a series of heat shock proteins had similar differential trends as previously shown in the intestine of NEC-resistant preterm pigs and young mice. Furthermore, LC-MS-based proteomics and Western blot analyses revealed 20 differentially expressed proteins following treatment with TGF-β2 in LPS-challenged IECs. Thirteen of these proteins were associated with stress response pathways, among which five proteins were altered by LPS and restored by TGF-β2, whereas six were differentially expressed only by TGF-β2 in LPS-challenged IECs. Based on previously reported biological functions, these patterns indicate the anti-stress and anti-inflammatory effects of TGF-β2 in IECs. We conclude that TGF-β2 of dietary or endogenous origin may regulate the IEC responses against LPS stimuli, thereby supporting cellular homeostasis and innate immunity in response to bacterial colonization, and the first enteral feeding in early life. PMID:25668313

  4. RACK1 binds to Smad3 to modulate transforming growth factor-beta1-stimulated alpha2(I) collagen transcription in renal tubular epithelial cells.

    PubMed

    Okano, Kazuhiro; Schnaper, H William; Bomsztyk, Karol; Hayashida, Tomoko

    2006-09-01

    Although it is clear that transforming growth factor-beta1 (TGF-beta1) is critical for renal fibrogenesis, the complexity of the involved mechanisms is increasingly apparent. TGF-beta1 stimulates phosphorylation of Smad2/3 and activates other signaling molecules as well. The molecular link between these other kinases and Smads is not known. We sought new binding partners for Smad3 in renal cells and identified receptor for activated protein kinase C 1 (RACK1) as a novel binding partner of Smad3. The linker region of Smad3 and the tryptophan-aspartic acid repeat 6 and 7 of RACK1 are sufficient for the association. RACK1 also interacts with Smad3 in the human kidney epithelial cell line, HKC. Silencing RACK1 increases transcriptional activity of TGF-beta1-responsive promoter sequences of the Smad binding element (SBE), p3TP-Lux, and alpha2(I) collagen. Conversely, overexpressed RACK1 negatively modulates alpha2(I) collagen transcriptional activity in TGF-beta1-stimulated cells. RACK1 did not affect phosphorylation of Smad3 at the C terminus or in the linker region. However, RACK1 reduced direct binding of Smad3 to the SBE motif. Mutating a RACK1 tyrosine at residue 246, but not at 228, decreased the inhibitory effect of RACK1 on both alpha2(I) collagen promoter activity and Smad binding to SBE induced by TGF-beta1. These results suggest that RACK1 modulates transcription of alpha2(I) collagen by TGF-beta1 through interference with Smad3 binding to the gene promoter. PMID:16849317

  5. Spleen tyrosine kinase mediates high glucose-induced transforming growth factor-{beta}1 up-regulation in proximal tubular epithelial cells

    SciTech Connect

    Yang, Won Seok; Chang, Jai Won; Han, Nam Jeong; Lee, Sang Koo; Park, Su-Kil

    2012-09-10

    The role of spleen tyrosine kinase (Syk) in high glucose-induced intracellular signal transduction has yet to be elucidated. We investigated whether Syk is implicated in high glucose-induced transforming growth factor-{beta}1 (TGF-{beta}1) up-regulation in cultured human proximal tubular epithelial cells (HK-2 cell). High glucose increased TGF-{beta}1 gene expression through Syk, extracellular signal-regulated kinase (ERK), AP-1 and NF-{kappa}B. High glucose-induced AP-1 DNA binding activity was decreased by Syk inhibitors and U0126 (an ERK inhibitor). Syk inhibitors suppressed high glucose-induced ERK activation, whereas U0126 had no effect on Syk activation. High glucose-induced NF-{kappa}B DNA binding activity was also decreased by Syk inhibitors. High glucose increased nuclear translocation of p65 without serine phosphorylation of I{kappa}B{alpha} and without degradation of I{kappa}B{alpha}, but with an increase in tyrosine phosphorylation of I{kappa}B{alpha} that may account for the activation of NF-{kappa}B. Both Syk inhibitors and Syk-siRNA attenuated high glucose-induced I{kappa}B{alpha} tyrosine phosphorylation and p65 nuclear translocation. Depletion of p21-activated kinase 2 (Pak2) by transfection of Pak2-siRNA abolished high glucose-induced Syk activation. In summary, high glucose-induced TGF-{beta}1 gene transcription occurred through Pak2, Syk and subsequent ERK/AP-1 and NF-{kappa}B pathways. This suggests that Syk might be implicated in the diabetic kidney disease.

  6. Lung Fibrosis-associated Surfactant Protein A1 and C Variants Induce Latent Transforming Growth Factor β1 Secretion in Lung Epithelial Cells*

    PubMed Central

    Maitra, Meenakshi; Dey, Moushumi; Yuan, Wen-Cheng; Nathanielsz, Peter W.; Garcia, Christine Kim

    2013-01-01

    Missense mutations of surfactant proteins are recognized as important causes of inherited lung fibrosis. Here, we study rare and common surfactant protein (SP)-A1 and SP-C variants, either discovered in our familial pulmonary fibrosis cohort or described by others. We show that expression of two SP-A1 (R219W and R242*) and three SP-C (I73T, M71V, and L188Q) variant proteins lead to the secretion of the profibrotic latent transforming growth factor (TGF)-β1 in lung epithelial cell lines. The secreted TGF-β1 is capable of autocrine and paracrine signaling and is dependent upon expression of the latent TGF-β1 binding proteins. The dependence upon unfolded protein response (UPR) mediators for TGF-β1 induction differs for each variant. TGF-β1 secretion induced by the expression of the common SP-A1 R219W variant is nearly completely blocked by silencing the UPR transducers IRE-1α and ATF6. In contrast, the secretion of TGF-β1 induced by two rare SP-C mutant proteins (I73T and M71V), is largely unaffected by UPR silencing or by the addition of the small molecular chaperone 4-phenylbutyric acid, implicating a UPR-independent mechanism for these variants. Blocking TGF-β1 secretion reverses cell death of RLE-6TN cells expressing these SP-A1 and SP-C variants suggesting that anti-TGF-β therapeutics may be beneficial to this molecularly defined subgroup of pulmonary fibrosis patients. PMID:23926107

  7. Lung fibrosis-associated surfactant protein A1 and C variants induce latent transforming growth factor β1 secretion in lung epithelial cells.

    PubMed

    Maitra, Meenakshi; Dey, Moushumi; Yuan, Wen-Cheng; Nathanielsz, Peter W; Garcia, Christine Kim

    2013-09-20

    Missense mutations of surfactant proteins are recognized as important causes of inherited lung fibrosis. Here, we study rare and common surfactant protein (SP)-A1 and SP-C variants, either discovered in our familial pulmonary fibrosis cohort or described by others. We show that expression of two SP-A1 (R219W and R242*) and three SP-C (I73T, M71V, and L188Q) variant proteins lead to the secretion of the profibrotic latent transforming growth factor (TGF)-β1 in lung epithelial cell lines. The secreted TGF-β1 is capable of autocrine and paracrine signaling and is dependent upon expression of the latent TGF-β1 binding proteins. The dependence upon unfolded protein response (UPR) mediators for TGF-β1 induction differs for each variant. TGF-β1 secretion induced by the expression of the common SP-A1 R219W variant is nearly completely blocked by silencing the UPR transducers IRE-1α and ATF6. In contrast, the secretion of TGF-β1 induced by two rare SP-C mutant proteins (I73T and M71V), is largely unaffected by UPR silencing or by the addition of the small molecular chaperone 4-phenylbutyric acid, implicating a UPR-independent mechanism for these variants. Blocking TGF-β1 secretion reverses cell death of RLE-6TN cells expressing these SP-A1 and SP-C variants suggesting that anti-TGF-β therapeutics may be beneficial to this molecularly defined subgroup of pulmonary fibrosis patients. PMID:23926107

  8. Transforming growth factor-β1 impairs CFTR-mediated anion secretion across cultured porcine vas deferens epithelial monolayer via the p38 MAPK pathway

    PubMed Central

    Yi, Sheng; Pierucci-Alves, Fernando

    2013-01-01

    The goal of this study was to determine whether transforming growth factor-β1 (TGF-β1) affects epithelial cells lining the vas deferens, an organ that is universally affected in cystic fibrosis male patients. In PVD9902 cells, which are derived from porcine vas deferens epithelium, TGF-β1 exposure significantly reduced short-circuit current (Isc) stimulated by forskolin or a cell membrane-permeant cAMP analog, 8-pCPT-cAMP, suggesting that TGF-β1 affects targets of the cAMP signaling pathway. Electrophysiological results indicated that TGF-β1 reduces the magnitude of current inhibited by cystic fibrosis transmembrane conductance regulator (CFTR) channel blockers. Real-time RT-PCR revealed that TGF-β1 downregulates the abundance of mRNA coding for CFTR, while biotinylation and Western blot showed that TGF-β1 reduces both total CFTR and apical cell surface CFTR abundance. These results suggest that TGF-β1 causes a reduction in CFTR expression, which limits CFTR-mediated anion secretion. TGF-β1-associated attenuation of anion secretion was abrogated by SB431542, a TGF-β1 receptor I inhibitor. Signaling pathway studies showed that the effect of TGF-β1 on Isc was reduced by SB203580, an inhibitor of p38 mitogen-activated protein kinase (MAPK). TGF-β1 exposure also increased the amount of phospho-p38 MAPK substantially. In addition, anisomycin, a p38 MAPK activator, mimicked the effect of TGF-β1, which further suggests that TGF-β1 affects PVD9902 cells through a p38 MAPK pathway. These observations suggest that TGF-β1, via TGF-β1 receptor I and p38 MAPK signaling, reduces CFTR expression to impair CFTR-mediated anion secretion, which would likely compound the effects associated with mild CFTR mutations and ultimately would compromise male fertility. PMID:23903699

  9. Autophagy induction by SIRT6 through attenuation of insulin-like growth factor signaling is involved in the regulation of human bronchial epithelial cell senescence.

    PubMed

    Takasaka, Naoki; Araya, Jun; Hara, Hiromichi; Ito, Saburo; Kobayashi, Kenji; Kurita, Yusuke; Wakui, Hiroshi; Yoshii, Yutaka; Yumino, Yoko; Fujii, Satoko; Minagawa, Shunsuke; Tsurushige, Chikako; Kojima, Jun; Numata, Takanori; Shimizu, Kenichiro; Kawaishi, Makoto; Kaneko, Yumi; Kamiya, Noriki; Hirano, Jun; Odaka, Makoto; Morikawa, Toshiaki; Nishimura, Stephen L; Nakayama, Katsutoshi; Kuwano, Kazuyoshi

    2014-02-01

    Cigarette smoke (CS)-induced cellular senescence has been implicated in the pathogenesis of chronic obstructive pulmonary disease, and SIRT6, a histone deacetylase, antagonizes this senescence, presumably through the attenuation of insulin-like growth factor (IGF)-Akt signaling. Autophagy controls cellular senescence by eliminating damaged cellular components and is negatively regulated by IGF-Akt signaling through the mammalian target of rapamycin (mTOR). SIRT1, a representative sirtuin family, has been demonstrated to activate autophagy, but a role for SIRT6 in autophagy activation has not been shown. Therefore, we sought to investigate the regulatory role for SIRT6 in autophagy activation during CS-induced cellular senescence. SIRT6 expression levels were modulated by cDNA and small interfering RNA transfection in human bronchial epithelial cells (HBECs). Senescence-associated β-galactosidase staining and Western blotting of p21 were performed to evaluate senescence. We demonstrated that SIRT6 expression levels were decreased in lung homogenates from chronic obstructive pulmonary disease patients, and SIRT6 expression levels correlated significantly with the percentage of forced expiratory volume in 1 s/forced vital capacity. CS extract (CSE) suppressed SIRT6 expression in HBECs. CSE-induced HBEC senescence was inhibited by SIRT6 overexpression, whereas SIRT6 knockdown and mutant SIRT6 (H133Y) without histone deacetylase activity enhanced HBEC senescence. SIRT6 overexpression induced autophagy via attenuation of IGF-Akt-mTOR signaling. Conversely, SIRT6 knockdown and overexpression of a mutant SIRT6 (H133Y) inhibited autophagy. Autophagy inhibition by knockdown of ATG5 and LC3B attenuated the antisenescent effect of SIRT6 overexpression. These results suggest that SIRT6 is involved in CSE-induced HBEC senescence via autophagy regulation, which can be attributed to attenuation of IGF-Akt-mTOR signaling. PMID:24367027

  10. The role of epithelial to mesenchymal transition in resistance to epidermal growth factor receptor tyrosine kinase inhibitors in non-small cell lung cancer

    PubMed Central

    Jakobsen, Kristine Raaby; Demuth, Christina; Sorensen, Boe Sandahl

    2016-01-01

    Inhibition of the epidermal growth factor receptor (EGFR) is an important strategy when treating non-small cell lung cancer (NSCLC) patients. However, intrinsic resistance or development of resistance during the course of treatment constitutes a major challenge. The knowledge on EGFR-directed tyrosine kinase inhibitors (TKIs) and their biological effect keeps increasing. Within the group of patients with EGFR mutations some benefit to a much higher degree than others, and for patients lacking EGFR mutations a subset experience an effect. Up to 70% of patients with EGFR mutations and 10–20% of patients without EGFR mutations initially respond to the EGFR-TKI erlotinib, but there is a severe absence of good prognostic markers. Despite initial effect, all patients acquire resistance to EGFR-TKIs. Multiple mechanisms have implications in resistance development, but much is still to be explored. Epithelial to mesenchymal transition (EMT) is a transcriptionally regulated phenotypic shift rendering cells more invasive and migratory. Within the EMT process lays a need for external or internal stimuli to give rise to changes in central signaling pathways. Expression of mesenchymal markers correlates to a bad prognosis and an inferior response to EGFR-TKIs in NSCLC due to the contribution to a resistant phenotype. A deeper understanding of the role of EMT in NSCLC and especially in EGFR-TKI resistance-development constitute one opportunity to improve the benefit of TKI treatment for the individual patient. Many scientific studies have linked the EMT process to EGFR-TKI resistance in NSCLC and our aim is to review the role of EMT in both intrinsic and acquired resistance to EGFR-TKIs. PMID:27186512

  11. Effect of NF-κB p65 antisense oligodeoxynucleotide on transdifferentiation of normal human lens epithelial cells induced by transforming growth factor-β2

    PubMed Central

    Liu, Chao; Wu, Xiao-Li; Wu, Xin-Yi; Zhang, Zhen-Hua; Liu, Xiao-Hua

    2016-01-01

    AIM To study the inhibition of nuclear factor kappa-B p65 (NF-κB p65) antisense oligodeoxynucleotide (ASODN) on transdifferentiation of normal human lens epithelial cells induced by transforming growth factor-β2 (TGF-β2) in vitro. METHODS NF-κB p65 ASODN and NF-κB p65 missense oligodeoxynucleotide (MSODN) were designed and synthesized. Human lens epithelial cell line (HLE B-3) cells were prepared for study and divided into 7 groups. Control group was HLE B-3 cells cultured in vitro in dulbecco's modified eagle medium (DMEM). T1, T2, and T3 group were HLE B-3 cells cultured in vitro in DMEM with 10 ng/mL TGF-β2 for 6h, 12h, 24h respectively. A+T group was HLE B-3 cells cultured with 10 ng/mL TGF-β2 for 24h after transfected by NF-κB p65 ASODN for 24h. M+T group was HLE B-3 cells cultured with 10 ng/mL TGF-β2 for 24h after transfected by NF-κB p65 MSODN for 24h. The negative control group was HLE B-3 cells cultured with 10 ng/mL TGF-β2 for 24h after cultured with transfer agent (HiPerFect) for 24h. Cell morphology was observed at different time points using an inverted microscope. The expression of NF-κB p65 mRNA was detected with reverse transcription-polymerase chain reaction (RT-PCR), and the expression of α-smooth muscle actin (α-SMA) protein was assayed with ELISA. RESULTS With the TGF-β2 stimulation prolongation, the expression of NF-κB p65 mRNA and α-SMA protein increased in T1, T2, T3 groups compared with the control group, and the difference was statistically significant (P<0.05). NF-κB p65 ASODN lowered the expression of NF-κB p65 mRNA and α-SMA protein induced by TGF-β2. NF-κB p65 MSODN and HiPerFect did not lower the expression of NF-κB p65 mRNA and α-SMA protein induced by TGF-β2. The difference between control group and A+T group was not statistically significant (P>0.05), but the difference among A+T group and other groups was statistically significant (P<0.05). CONCLUSION NF-κB p65 ASODN could lower the expression of NF-κB p

  12. Downregulation of gangliotetraosylceramide and β1,3-galactosyltransferase-4 gene expression by Smads during transforming growth factor β-induced epithelial-mesenchymal transition.

    PubMed

    Guo, Jia; Song, Bo; Li, Xiang; Hε, Chenhui; Yang, Ganglong; Yang, Xiaomin; Guan, Feng

    2015-03-01

    The process of epithelial‑mesenchymal transition (EMT) is essential for the proliferation and metastasis of tumor cells. Our previous study demonstrated that the expression of gangliotetraosylceramide (Gg4) and the transcription of UDP‑Gal: β1,3-galactosyltransferase‑4 (β3GalT4), a gene which controls the expression of Gg4, are reduced during transforming growth factor (TGF)‑β‑induced EMT in normal murine mammary gland (NMuMG) cells. The present study revealed that the transcription level of β3GalT4 in patients with breast cancer was decreased compared with healthy control subjects. However, the molecular basis underlying these effects remains to be elucidated. Analysis of the β3GalT4 promoter sequence revealed a putative Smad‑binding element (SBE) of Smad4, which is a transcriptional factor in TGFβ responses and forms a complex with Smad3. To clarify the association between the Smad3/4 complex and the β3GalT4 gene during EMT in NMuMG cells, an electrophoretic mobility shift assay and a chromatin immunoprecipitation assay were performed. The Smad3/4 complex was found to bind directly to the SBE of the β3GalT4 promoter. Overexpression of Smad3 and Smad4 through stable transfection had no notable effect on cell phenotype, but did alter the expression of various EMT protein markers. Treatment with TGFβ reduced the expression of Gg4 and the mRNA levels of the β3GalT4 gene in Smad3‑ and Smad4‑overexpressing cells compared with vector‑transfected cells. Expression of the epithelial markers E‑cadherin and β‑catenin decreased in parallel with the reduction in Gg4. These findings suggested that the activated Smad3/4 complex downregulated the expression of Gg4 and the β3GalT4 gene through translocation into the nucleus and binding to the β3GalT4 promoter. PMID:25384497

  13. Distinct epithelial growth factor receptor mutation profile in non-small-cell lung cancer patients from the Xuanwei area of China

    PubMed Central

    CHEN, YUN; YE, LIJUAN; STANFORD, REBECCA RONGRONG; ZHANG, DONGFANG; ZHANG, XINGSONG; WEI, WANLI

    2016-01-01

    The Xuanwei county in China has a high incidence of lung cancer and related mortality. Previous studies have suggested that these cases may be associated with a distinctive pattern of mutations in the epithelial growth factor receptor (EGFR) gene. In this retrospective study, we investigated the mutation profile of EGFR in non-small-cell lung cancer (NSCLC) tissues from patients in Xuanwei, and the associated clinicopathological characteristics. Specimens from 258 consecutive patients with lung cancer (90 from Xuanwei and 168 from other areas of Yunnan province) were subjected to amplification refractory mutation system-polymerase chain reaction (ARMS-PCR) to detect EGFR mutations. In 67 specimens from Xuanwei, the results were confirmed by direct DNA sequencing for EGFR mutations. Immunohistochemistry (IHC) for the echinoderm microtubule-associated protein-like 4-anaplastic lymphoma kinase (EML4-ALK) fusion protein was performed on all specimens from Xuanwei. We observed that Xuanwei patients presented with distinctive clinicopathological characteristics, including female gender predominance, younger age, higher rate of lymph node metastasis, higher rate of adenocarcinoma histological classification and lower disease stage, and a low rate of the ‘classical’ mutations on EGFR exons 19 and 21 compared with non-Xuanwei patients (7.8 and 21.6% vs. 49.3 and 39.7%, respectively; P<0.05 for combined data). However, a significantly higher percentage of Xuanwei patients harbored co-mutation of EGFR exons 18 and 20 compared with non-Xuanwei patients (45.1 vs. 4.1%, respectively; P<0.0001). Specimens from 2 Xuanwei patients (2.2%) were positive for the EML4-ALK fusion protein; by IHC, neither harbored EGFR mutations. There was no obvious association between EGFR mutations and disease stage or lymph node involvement. Thus, NSCLC patients in Xuanwei presented with a unique EGFR profile of high rates of co-mutation of exons 18 and 20, and low rates of exon 19 or 21

  14. Enhanced Expression of Integrin αvβ3 Induced by TGF-β Is Required for the Enhancing Effect of Fibroblast Growth Factor 1 (FGF1) in TGF-β-Induced Epithelial-Mesenchymal Transition (EMT) in Mammary Epithelial Cells

    PubMed Central

    Ito, Ayano; Okazaki, Mika; Kawaguchi, Naomasa; Hamada, Yoshinosuke; Takada, Yoshikazu; Matsuura, Nariaki

    2015-01-01

    Epithelial-to-mesenchymal transition (EMT) plays a critical role in cancer metastasis, and is regulated by growth factors such as transforming growth factor β (TGF-β) and fibroblast growth factors (FGF) secreted from the stromal and tumor cells. However, the role of growth factors in EMT has not been fully established. Several integrins are upregulated by TGF-β1 during EMT. Integrins are involved in growth factor signaling through integrin-growth factor receptor crosstalk. We previously reported that FGF1 directly binds to integrin αvβ3 and the interaction was required for FGF1 functions such as cell proliferation and migration. We studied the role of αvβ3 induced by TGF-β on TGF-β-induced EMT. Here, we describe that FGF1 augmented EMT induced by TGF-β1 in MCF10A and MCF12A mammary epithelial cells. TGF-β1 markedly amplified integrin αvβ3 and FGFR1 (but not FGFR2). We studied if the enhancing effect of FGF1 on TGF-β1-induced EMT requires enhanced levels of both integrin αvβ3 expression and FGFR1. Knockdown of β3 suppressed the enhancement by FGF1 of TGF-β1-induced EMT in MCF10A cells. Antagonists to FGFR suppressed the enhancing effect of FGF1 on EMT. Integrin-binding defective FGF1 mutant did not augment TGF-β1-induced EMT in MCF10A cells. These findings suggest that enhanced integrin αvβ3 expression in addition to enhanced FGFR1 expression is critical for FGF1 to augment TGF-β1-induced EMT in mammary epithelial cells. PMID:26334633

  15. Aspirin blocks growth of breast tumor cells and tumor-initiating cells and induces reprogramming factors of mesenchymal to epithelial transition.

    PubMed

    Maity, Gargi; De, Archana; Das, Amlan; Banerjee, Snigdha; Sarkar, Sandipto; Banerjee, Sushanta K

    2015-07-01

    Acetylsalicylic acid (ASA), also known as aspirin, a classic, nonsteroidal, anti-inflammatory drug (NSAID), is widely used to relieve minor aches and pains and to reduce fever. Epidemiological studies and other experimental studies suggest that ASA use reduces the risk of different cancers including breast cancer (BC) and may be used as a chemopreventive agent against BC and other cancers. These studies have raised the tempting possibility that ASA could serve as a preventive medicine for BC. However, lack of in-depth knowledge of the mechanism of action of ASA reshapes the debate of risk and benefit of using ASA in prevention of BC. Our studies, using in vitro and in vivo tumor xenograft models, show a strong beneficial effect of ASA in the prevention of breast carcinogenesis. We find that ASA not only prevents breast tumor cell growth in vitro and tumor growth in nude mice xenograft model through the induction of apoptosis, but also significantly reduces the self-renewal capacity and growth of breast tumor-initiating cells (BTICs)/breast cancer stem cells (BCSCs) and delays the formation of a palpable tumor. Moreover, ASA regulates other pathophysiological events in breast carcinogenesis, such as reprogramming the mesenchymal to epithelial transition (MET) and delaying in vitro migration in BC cells. The tumor growth-inhibitory and reprogramming roles of ASA could be mediated through inhibition of TGF-β/SMAD4 signaling pathway that is associated with growth, motility, invasion, and metastasis in advanced BCs. Collectively, ASA has a therapeutic or preventive potential by attacking possible target such as TGF-β in breast carcinogenesis. PMID:25867761

  16. cDNA cloning of a mouse mammary epithelial cell surface protein reveals the existence of epidermal growth factor-like domains linked to factor VIII-like sequences

    SciTech Connect

    Stubbs, J.D.; Bui, A. San Francisco State Univ., CA ); Lekutis, C.; Singer, K.L.; Srinivasan, U.; Parry, G. ); Yuzuki, D. )

    1990-11-01

    A 2.1-kilobase cDNA coding for a surface protein of mammary epithelial cells has been isolated from a mouse mammary gland {lambda}gt11 cDNA library. Sequence analysis of this cDNA reveals an open reading frame of 1,389 base pairs that defines a protein with a molecular mass of 51.5 dKa. Structural analysis of the predicted sequence identifies two putative functional domains of the protein: (i) an N-terminal cysteine-rich region that is similar to epidermal growth factor-like domains of Drosophila Notch-1 protein and (ii) a large segment of the sequence that exhibited 54.5% identify with C-terminal domains of human coagulation factors VIII and V. These similarities in structure are used to predict the possible functions of the protein and its means of interaction with the cell surface. mRNA expression was detectable in mammary tissue from nonpregnant animals but was maximal in the lactating gland. In cultured cells, mRNA levels also correlated with the degree of cellular differentiation.

  17. Transforming growth factor-β1 induces epithelial-to-mesenchymal transition in human lung cancer cells via PI3K/Akt and MEK/Erk1/2 signaling pathways.

    PubMed

    Chen, Xiao-Feng; Zhang, Hui-Jun; Wang, Hai-Bing; Zhu, Jun; Zhou, Wen-Yong; Zhang, Hui; Zhao, Ming-Chuan; Su, Jin-Mei; Gao, Wen; Zhang, Lei; Fei, Ke; Zhang, Hong-Tao; Wang, He-Yong

    2012-04-01

    Metastasis of tumor cells is associated with epithelial-to-mesenchymal transition (EMT), which is a process whereby epithelial cells lose their polarity and acquire new features of mesenchyme. EMT has been reported to be induced by transforming growth factor-β1 (TGF-β1), but its mechanism remains elusive. In this study, we performed a study to investigate whether PI3K/Akt and MAPK/Erk1/2 signaling pathways involved in EMT in the human lung cancer A549 cells. The results showed that after treated with TGF-β1 for 48 h, A549 cells displayed more fibroblast-like shape, lost epithelial marker E-cadherin and increased mesenchymal markers Vimentin and Fibronectin. Moreover, TGF-β1-induced EMT after 48 h was accompanied by increased of cell migration and change of Akt and Erk1/2 phosphorylation. In addition, EMT was reversed by PI3K inhibitor LY294002 and MEK1/2 inhibitor U0126, which suggested that A549 cells under stimulation of TGF-β1 undergo a switch into mesenchymal cells and PI3K/Akt and MAPK/Erk1/2 signaling pathways serve to regulate TGF-β1-induced EMT of A549 cells. PMID:21713404

  18. Vascular endothelial growth factor expression correlates with serum CA125 and represents a useful tool in prediction of refractoriness to platinum-based chemotherapy and ascites formation in epithelial ovarian cancer

    PubMed Central

    Wei, Ai-Qun; Robertson, Gregory; Morris, David L.

    2015-01-01

    There is an increasing need for the identification of novel biological markers and potential therapeutic targets in epithelial ovarian cancer (EOC). Given the critical role of growth factors in the biology of EOC, we aimed in the present study to evaluate the intratumoral expressions of vascular endothelial growth factor (VEGF) and fibroblast growth factor (FGF) proteins and their clinical relevance in a cohort of 100 patients with EOC. All patients received platinum-based chemotherapy after surgery. A comparative immunohistochemical study of normal ovarian and EOC tissues showed that both growth factors were expressed at higher levels in tumor samples. In our statistical analysis, while no association existed between the FGF expression status and the clinicopathological characteristics of patients, intratumoral VEGF was identified as a potential biomarker for the prediction of ascites formation. In addition, the expression status of VEGF appeared to independently predict overall survival and response to chemotherapy. Furthermore, a direct association was demonstrated between the pre-treatment VEGF expression and serum CA125 after three cycles of chemotherapy. In sum, we report for the first time to our knowledge the correlation between intratumoral VEGF and serum CA125 in EOC. Our data also shows the prognostic value of VEGF expression in EOC. These results suggest the potential value of intratumoral VEGF in patient stratification. Dual inhibition of VEGF and CA125 might bring about a better outcome for patients with EOC. PMID:26143638

  19. Control of Francisella tularensis Intracellular Growth by Pulmonary Epithelial Cells

    PubMed Central

    Maggio, Savannah; Takeda, Kazuyo; Stark, Felicity; Meierovics, Anda I.; Yabe, Idalia; Cowley, Siobhan C.

    2015-01-01

    The virulence of F. tularensis is often associated with its ability to grow in macrophages, although recent studies show that Francisella proliferates in multiple host cell types, including pulmonary epithelial cells. Thus far little is known about the requirements for killing of F. tularensis in the non-macrophage host cell types that support replication of this organism. Here we sought to address this question through the use of a murine lung epithelial cell line (TC-1 cells). Our data show that combinations of the cytokines IFN-γ, TNF, and IL-17A activated murine pulmonary epithelial cells to inhibit the intracellular growth of the F. tularensis Live Vaccine Strain (LVS) and the highly virulent F. tularensis Schu S4 strain. Although paired combinations of IFN-γ, TNF, and IL-17A all significantly controlled LVS growth, simultaneous treatment with all three cytokines had the greatest effect on LVS growth inhibition. In contrast, Schu S4 was more resistant to cytokine-induced growth effects, exhibiting significant growth inhibition only in response to all three cytokines. Since one of the main antimicrobial mechanisms of activated macrophages is the release of reactive nitrogen intermediates (RNI) via the activity of iNOS, we investigated the role of RNI and iNOS in Francisella growth control by pulmonary epithelial cells. NOS2 gene expression was significantly up-regulated in infected, cytokine-treated pulmonary epithelial cells in a manner that correlated with LVS and Schu S4 growth control. Treatment of LVS-infected cells with an iNOS inhibitor significantly reversed LVS killing in cytokine-treated cultures. Further, we found that mouse pulmonary epithelial cells produced iNOS during in vivo respiratory LVS infection. Overall, these data demonstrate that lung epithelial cells produce iNOS both in vitro and in vivo, and can inhibit Francisella intracellular growth via reactive nitrogen intermediates. PMID:26379269

  20. c-Jun N-terminal kinase 1 promotes transforming growth factor-β1-induced epithelial-to-mesenchymal transition via control of linker phosphorylation and transcriptional activity of Smad3.

    PubMed

    Velden, Jos L J van der; Alcorn, John F; Guala, Amy S; Badura, Elsbeth C H L; Janssen-Heininger, Yvonne M W

    2011-04-01

    Transforming growth factor (TGF)-β1 is a key mediator of lung remodeling and fibrosis. Epithelial cells are both a source of and can respond to TGF-β1 with epithelial-to-mesenchymal transition (EMT). We recently determined that TGF-β1-induced EMT in lung epithelial cells requires the presence of c-Jun N-terminal kinase (JNK) 1. Because TGF-β1 signals via Smad complexes, the goal of the present study was to determine the impact of JNK1 on phosphorylation of Smad3 and Smad3-dependent transcriptional responses in lung epithelial cells. Evaluation of JNK1-deficient lung epithelial cells demonstrated that TGF-β1-induced terminal phosphorylation of Smad3 was similar, whereas phosphorylation of mitogen-activated protein kinase sites in the linker regions of Smad3 was diminished, in JNK1-deficient cells compared with wild-type cells. In comparison to wild-type Smad3, expression of a mutant Smad3 in which linker mitogen-activated protein kinase sites were ablated caused a marked attenuation in JNK1 or TGF-β1-induced Smad-binding element transcriptional activity, and expression of plasminogen activator inhibitor-1, fibronectin-1, high-mobility group A2, CArG box-binding factor-A, and fibroblast-specific protein-1, genes critical in the process of EMT. JNK1 enhanced the interaction between Smad3 and Smad4, which depended on linker phosphorylation of Smad3. Conversely, Smad3 with phosphomimetic mutations in the linker domain further enhanced EMT-related genes and proteins, even in the absence of JNK1. Finally, we demonstrated a TGF-β1-induced interaction between Smad3 and JNK1. Collectively, these results demonstrate that Smad3 phosphorylation in the linker region and Smad transcriptional activity are directly or indirectly controlled by JNK1, and provide a putative mechanism whereby JNK1 promotes TGF-β1-induced EMT. PMID:20581097

  1. Photodynamic treatment of epithelial tissue derived from patients with endometrial cancer: a contribution to the role of laminin and epidermal growth factor receptor in photodynamic therapy

    NASA Astrophysics Data System (ADS)

    Ziolkowski, Piotr P.; Symonowicz, Krzysztof; Osiecka, Beata J.; Rabczynski, Jerzy; Gerber, Jerzy

    1999-07-01

    Photodynamic therapy (PDT) was used to treat endometrial G1 cancer tissue derived from patients who had undergone a total hysterectomy and bilateral salpingo-oophorectomy. After surgical treatment the cancerous tissue was kept in a medium containing Dulbecco solution, fetal calf serum, and antibiotics. The tissue was then exposed to hematoporphyrin derivative (0.1 mg/l) and 24 h later exposed to light (total light dose--18 J/sq cm). Necrosis depth was evaluated 24 h later using a light microscope. In order to assess the possible role of the basal membrane component laminin, as well as epidermal growth factor receptor susceptibility to PDT, immunohistochemical studies were carried out. Additionally, nucleolar organizer regions evaluation was performed. Our experiment confirmed that PDT results in the necrosis in the treated endometrial cancer, while not affecting the laminin in the cancerous tissue. In contrast, PDT strongly affects the epidermal growth factor receptor and nucleolar organizer regions in cancer cells. We suggest that laminin may contribute to the prevention of cancer dissemination in the cases where PDT has to be repeated, and that after PDT the cells become less susceptible to a mitogen, like, e.g., epidermal growth factor.

  2. DUSP10 regulates intestinal epithelial cell growth and colorectal tumorigenesis.

    PubMed

    Png, C W; Weerasooriya, M; Guo, J; James, S J; Poh, H M; Osato, M; Flavell, R A; Dong, C; Yang, H; Zhang, Y

    2016-01-14

    Dual specificity phosphatase 10 (DUSP10), also known as MAP kinase phosphatase 5 (MKP5), negatively regulates the activation of MAP kinases. Genetic polymorphisms and aberrant expression of this gene are associated with colorectal cancer (CRC) in humans. However, the role of DUSP10 in intestinal epithelial tumorigenesis is not clear. Here, we showed that DUSP10 knockout (KO) mice had increased intestinal epithelial cell (IEC) proliferation and migration and developed less severe colitis than wild-type (WT) mice in response to dextran sodium sulphate (DSS) treatment, which is associated with increased ERK1/2 activation and Krüppel-like factor 5 (KLF5) expression in IEC. In line with increased IEC proliferation, DUSP10 KO mice developed more colon tumours with increased severity compared with WT mice in response to administration of DSS and azoxymethane (AOM). Furthermore, survival analysis of CRC patients demonstrated that high DUSP10 expression in tumours was associated with significant improvement in survival probability. Overexpression of DUSP10 in Caco-2 and RCM-1 cells inhibited cell proliferation. Our study showed that DUSP10 negatively regulates IEC growth and acts as a suppressor for CRC. Therefore, it could be targeted for the development of therapies for colitis and CRC. PMID:25772234

  3. Heterogeneity and stochastic growth regulation of biliary epithelial cells dictate dynamic epithelial tissue remodeling.

    PubMed

    Kamimoto, Kenji; Kaneko, Kota; Kok, Cindy Yuet-Yin; Okada, Hajime; Miyajima, Atsushi; Itoh, Tohru

    2016-01-01

    Dynamic remodeling of the intrahepatic biliary epithelial tissue plays key roles in liver regeneration, yet the cellular basis for this process remains unclear. We took an unbiased approach based on in vivo clonal labeling and tracking of biliary epithelial cells in the three-dimensional landscape, in combination with mathematical simulation, to understand their mode of proliferation in a mouse liver injury model where the nascent biliary structure formed in a tissue-intrinsic manner. An apparent heterogeneity among biliary epithelial cells was observed: whereas most of the responders that entered the cell cycle upon injury exhibited a limited and tapering growth potential, a select population continued to proliferate, making a major contribution in sustaining the biliary expansion. Our study has highlighted a unique mode of epithelial tissue dynamics, which depends not on a hierarchical system driven by fixated stem cells, but rather, on a stochastically maintained progenitor population with persistent proliferative activity. PMID:27431614

  4. ACCRETA COMPLICATING COMPLETE PLACENTA PREVIA IS CHARACTERIZED BY REDUCED SYSTEMIC LEVELS OF VASCULAR ENDOTHELIAL GROWTH FACTOR AND EPITHELIAL-TO-MESENCHYMAL TRANSITION OF THE INVASIVE TROPHOBLAST

    PubMed Central

    Wehrum, Mark J.; Buhimschi, Irina A.; Salafia, Carolyn; Thung, Stephen; Bahtiyar, Mert O.; Werner, Erica F.; Campbell, Katherine H.; Laky, Christine; Sfakianaki, Anna K.; Zhao, Guomao; Funai, Edmund F.; Buhimschi, Catalin S.

    2011-01-01

    OBJECTIVE To characterize serum angiogenic factor profile of women with complete placenta previa and determine if invasive trophoblast differentiation characteristic of accreta, increta or percreta shares features of epitehelial-mesenchymal-transition (EMT). STUDY DESIGN We analyzed gestational age matched serum samples from 90 pregnant women with either complete placenta previa (n=45) or uncomplicated pregnancies (n=45). Vascular-endothelial-growth-factor (VEGF), placental-growth-factor (PlGF) and soluble fms-like-tyrosine-kinase-1 (sFlt-1) were immunoassayed. VEGF and phosphotyrosine (P-Tyr) immunoreactivity was surveyed in histological specimens relative to expression of vimentin and cytokeratin-7. RESULTS Women with previa and invasive placentation [accreta (n=5); increta (n=6); percreta (n=2)] had lower systemic VEGF (invasive previa: median [IQR]: 0.8[0.02–3.4] vs. control: 6.5[2.7–10.5] pg/mL, P=0.02). VEGF and P-Tyr immunostaining predominated in the invasive extravillous trophoblasts (EVT) which co-expressed vimentin and cytokeratin-7, a EMT feature and tumor-like cell phenotype. CONCLUSIONS Lower systemic free VEGF and a switch of the interstitial EVT to a metastable cell phenotype characterize placenta previa with excessive myometrial invasion. PMID:21316642

  5. Reciprocal interactions between Beta1-integrin and epidermal growth factor in three-dimensional basement membrane breast cultures: A different perspective in epithelial biology

    SciTech Connect

    Wang, F.; Weaver, V.M.; Petersen, O.W.; Larabell, C.A.; Dedhar, S.; Briand, P.; Lupu, R.; Bissell, M.J.

    1998-09-30

    Anchorage and growth factor independence are cardinal features of the transformed phenotype. Although it is logical that the two pathways must be coregulated in normal tissues to maintain homeostasis, this has not been demonstrated directly. We showed previously that down-modulation of {beta}1-integrin signaling reverted the malignant behavior of a human breast tumor cell line (T4-2) derived from phenotypically normal cells (HMT-3522) and led to growth arrest in a threedimensional (3D) basement membrane assay in which the cells formed tissue-like acini (14). Here, we show that there is a bidirectional cross-modulation of {beta}1-integrin and epidermal growth factor receptor (EGFR) signaling via the mitogenactivated protein kinase (MAPK) pathway. The reciprocal modulation does not occur in monolayer (2D) cultures. Antibodymediated inhibition of either of these receptors in the tumor cells, or inhibition of MAPK kinase, induced a concomitant downregulation of both receptors, followed by growth-arrest and restoration of normal breast tissue morphogenesis. Crossmodulation and tissue morphogenesis were associated with attenuation of EGF-induced transient MAPK activation. To specifically test EGFR and {beta}1-integrin interdependency, EGFR was overexpressed in nonmalignant cells, leading to disruption of morphogenesis and a compensatory up-regulation of {beta}1-integrin expression, again only in 3D. Our results indicate that when breast cells are spatially organized as a result of contact with basement membrane, the signaling pathways become coupled and bidirectional. They further explain why breast cells fail to differentiate in monolayer cultures in which these events are mostly uncoupled. Moreover, in a subset of tumor cells in which these pathways are misregulated but functional, the cells could be 'normalized' by manipulating either pathway.

  6. A lactobacillus rhamnosus GG-derived soluble protein, p40, stimulates ligand release from intestinal epithelial cells to transactivate epidermal growth factor receptor

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Protein p40, a Lactobacillus rhamnosus GG (LGG)-derived soluble protein, ameliorates intestinal injury and colitis, reduces apoptosis and preserves barrier function by activation of EGF receptor (EGFR) in intestinal epithelial cells. The aim of this study was to determine the mechanisms by which p40...

  7. Induction of epithelial to mesenchymal transition (EMT) and inhibition on adipogenesis: Two different sides of the same coin? Feasible roles and mechanisms of transforming growth factor β1 (TGF-β1) in age-related thymic involution.

    PubMed

    Tan, Jianxin; Wang, Yajun; Zhang, Nannan; Zhu, Xike

    2016-08-01

    Age-related thymic involution is characterized by a loss of thymic epithelial cells (TECs) and a concomitant increase in adipocytes, but the mechanisms involved in thymic adipogenesis are still not clear. Transforming growth factor β1 (TGF-β1) is a pleiotropic cytokine that has been reported to be up-regulated with age in thymic stromal cells in both human and mouse. However, the exact role of TGF-β1 in age-related thymic involution remains to be further elucidated. On the basis of previous findings, we propose a novel hypothesis that TGF-β1 functions a dual role in age-related thymic involution. On one hand, up-regulation of TGF-β1 promotes epithelial to mesenchymal transition (EMT) process in TECs via activating forkhead box protein C2 (FoxC2). On the other hand, TGF-β1 inhibits the transdifferentiation of EMT-derived mesenchymal cells to adipocytes in the thymus. If confirmed, our hypothesis will not only provide further evidence supporting that the transdifferentiation of TECs into pre-adipocytes represents a source of thymic adiposity during age-related thymic involution, but also uncover a unique role of TGF-β1 in the transdifferentiation of TECs into pre-adipocytes. Collectively, the inhibition of TGF-β1 may serve as a strategy to hinder age-related thymic involution or even to restore thymic function in the elderly. PMID:27189906

  8. Phosphatidylinositol 3-kinase, protein kinase B and ribosomal S6 kinases in the stimulation of thyroid epithelial cell proliferation by cAMP and growth factors in the presence of insulin.

    PubMed

    Coulonval, K; Vandeput, F; Stein, R C; Kozma, S C; Lamy, F; Dumont, J E

    2000-06-01

    The proliferation of most normal cells depends on the co-operation of several growth factors and hormones, each with a specific role, but the key events involved in the action of each necessary stimulant remain largely uncharacterized. In the present study, the pathways involved in the mechanism(s) of co-operation have been investigated in primary cultures of dog thyroid epithelial cells. In this physiologically relevant system, thyroid stimulating hormone (TSH) acting through cAMP, epidermal growth factor (EGF) and phorbol esters (such as PMA) induce DNA synthesis. Their effect requires stimulation of the insulin-like growth factor-1 (IGF-1) receptor by either IGF-1 or insulin, which are not themselves mitogenic agents. In contrast, hepatocyte growth factor (HGF) is itself fully mitogenic. The results of the study demonstrate that cAMP, EGF, HGF and PMA stimulate p70 ribosomal S6 kinase (p70 S6 kinase). However, insulin/IGF-1 also stimulate p70 S6 kinase. Thus stimulation of p70 S6 kinase might be necessary, but is certainly not sufficient, for the induction of DNA synthesis and is not specific for any stimulated pathway. In contrast, phosphatidylinositol 3-kinase (PI 3-kinase) and protein kinase B (PKB) activation by insulin and HGF is strong and sustained, whereas it is weak and transient with EGF and absent in the presence of TSH or PMA. These findings suggest that: (i) stimulation of PI 3-kinases and/or PKB is not involved in the cAMP-dependent pathways leading to thyrocyte proliferation, or in the action of PMA, (ii) the stimulation of the PI 3-kinase/PKB pathway may account for the permissive action of insulin/IGF-1 in the proliferation of these cells, and (iii) the stimulation of this pathway by HGF may explain why this agent does not require insulin or IGF-1 for its mitogenic action. PMID:10816429

  9. Growth of Normal Mouse Vaginal Epithelial Cells in and on Collagen Gels

    NASA Astrophysics Data System (ADS)

    Iguchi, Taisen; Uchima, Francis-Dean A.; Ostrander, Patricia L.; Bern, Howard A.

    1983-06-01

    Sustained growth in primary culture of vaginal epithelial cells from ovariectomized adult BALB/cCrg1 mice embedded within or seeded on collagen gel matrix was achieved in a serum-free medium composed of Ham's F-12 medium/Dulbecco's modified Eagle's medium, 1:1 (vol/vol), supplemented with insulin, bovine serum albumin fraction V, epidermal growth factor, cholera toxin, and transferrin. Three-dimensional growth of vaginal epithelial cells occurred inside the collagen gel matrix. Cell numbers increased 4- to 8-fold in collagen gel and about 4-fold on collagen gel after 9-10 days in culture. The effect of 17β -estradiol (0.00018-180 nM in gel or 0.018-180 nM on gel) and diethylstilbestrol (DES; 0.0186-186 nM in gel) on the growth of vaginal epithelial cells was examined. The addition of estrogen did not enhance the growth of vaginal epithelial cells during this time period either in the complete medium or in a suboptimal medium. Cultures on floating collagen gels in the serum-free medium are composed of 1-3 cell layers with superficial cornification. Estrogen does not appear to be a direct mitogen for vaginal epithelial cells, at least in this system.

  10. Long-term exposure of proximal tubular epithelial cells to glucose induces transforming growth factor-beta 1 synthesis via an autocrine PDGF loop.

    PubMed

    Fraser, Donald; Brunskill, Nigel; Ito, Takafumi; Phillips, Aled

    2003-12-01

    We have recently reported increased transforming growth factor (TGF)-beta1 gene transcription in proximal tubular cells within 12 hours of exposure to 25 mmol/L D-glucose, with a requirement for a second stimulus such as platelet-derived growth factor (PDGF) to increase its translation in short-term experiments. In the current study we investigated the effect on TGF-beta 1 production of prolonged exposure of proximal tubular cells to high glucose concentrations. Enzyme-linked immunosorbent assay of cell culture supernatant showed significant increase in latent TGF-beta 1 only after 7 days exposure to high glucose. Radiolabeling of glucose-stimulated cells with (3)H amino acids and subsequent immunoprecipitation of TGF-beta 1 demonstrated de novo synthesis from day 5 of high glucose exposure onwards. Similarly, polysome analysis showed enhanced translation of TGF-beta mRNA after 4 or more days of high glucose exposure. TGF-beta 1 synthesis, following addition of glucose, was inhibited by blockade of the PDGF-alpha receptor subunit. Glucose did not alter PDGF expression, nor expression of PDGF alpha-receptors. Activation of the receptor following addition of 25 mm D-glucose could be demonstrated suggesting increased sensitivity to endogenous PDGF. Exposure to glucose activated p38MAP kinase, and inhibition of this activation abrogated both glucose induced TGF-beta 1 transcriptional activation and TGF-beta 1 synthesis. Inhibition of p38MAP kinase did not influence the effect of exogenous PDGF when cells were stimulated sequentially by glucose and PDGF. We postulate that glucose induces an early increase in TGF-beta 1 transcription via activation of p38MAP kinase. In addition, glucose causes a late increase in PDGF-dependent TGF-beta 1 translation by enhancing cellular sensitivity to PDGF. This provides a potential explanation for the clinical observation that prolonged poor glycemic control may contribute to progression of diabetic nephropathy. PMID:14633628

  11. Interactions Between β-Catenin and Transforming Growth Factor-β Signaling Pathways Mediate Epithelial-Mesenchymal Transition and Are Dependent on the Transcriptional Co-activator cAMP-response Element-binding Protein (CREB)-binding Protein (CBP)*

    PubMed Central

    Zhou, Beiyun; Liu, Yixin; Kahn, Michael; Ann, David K.; Han, Arum; Wang, Hongjun; Nguyen, Cu; Flodby, Per; Zhong, Qian; Krishnaveni, Manda S.; Liebler, Janice M.; Minoo, Parviz; Crandall, Edward D.; Borok, Zea

    2012-01-01

    Interactions between transforming growth factor-β (TGF-β) and Wnt are crucial to many biological processes, although specific targets, rationale for divergent outcomes (differentiation versus block of epithelial proliferation versus epithelial-mesenchymal transition (EMT)) and precise mechanisms in many cases remain unknown. We investigated β-catenin-dependent and transforming growth factor-β1 (TGF-β1) interactions in pulmonary alveolar epithelial cells (AEC) in the context of EMT and pulmonary fibrosis. We previously demonstrated that ICG-001, a small molecule specific inhibitor of the β-catenin/CBP (but not β-catenin/p300) interaction, ameliorates and reverses pulmonary fibrosis and inhibits TGF-β1-mediated α-smooth muscle actin (α-SMA) and collagen induction in AEC. We now demonstrate that TGF-β1 induces LEF/TCF TOPFLASH reporter activation and nuclear β-catenin accumulation, while LiCl augments TGF-β-induced α-SMA expression, further confirming co-operation between β-catenin- and TGF-β-dependent signaling pathways. Inhibition and knockdown of Smad3, knockdown of β-catenin and overexpression of ICAT abrogated effects of TGF-β1 on α-SMA transcription/expression, indicating a requirement for β-catenin in these Smad3-dependent effects. Following TGF-β treatment, co-immunoprecipitation demonstrated direct interaction between endogenous Smad3 and β-catenin, while chromatin immunoprecipitation (ChIP)-re-ChIP identified spatial and temporal regulation of α-SMA via complex formation among Smad3, β-catenin, and CBP. ICG-001 inhibited α-SMA expression/transcription in response to TGF-β as well as α-SMA promoter occupancy by β-catenin and CBP, demonstrating a previously unknown requisite TGF-β1/β-catenin/CBP-mediated pro-EMT signaling pathway. Clinical relevance was shown by β-catenin/Smad3 co-localization and CBP expression in AEC of IPF patients. These findings suggest a new therapeutic approach to pulmonary fibrosis by specifically

  12. Activation of histamine H4 receptors decreases epithelial-to-mesenchymal transition progress by inhibiting transforming growth factor-β1 signalling pathway in non-small cell lung cancer.

    PubMed

    Cai, Wen-Ke; Hu, Jing; Li, Teng; Meng, Jing-Ru; Ma, Xue; Yin, Sun-Jun; Zhao, Can-Hu; He, Gong-Hao; Xu, Gui-Li

    2014-04-01

    Previous investigations found that epithelial-to-mesenchymal transition (EMT) was an important character of non-small cell lung cancer (NSCLC) and it was also suggested that histamine H4 receptors may have a role in preventing EMT progress in certain kind of tumours. However, the effect of H4 receptor activation on EMT progress of NSCLC and its potential mechanisms remain unclear. Therefore, we performed both in vitro and in vivo experiments to explore the effects of specific H4 receptor agonist 4-methylhistamine and antagonist JNJ7777120 on EMT progress. We showed the expression of H4 receptors in NSCLC and found that 4-methylhistamine increased the expression of the epithelial marker E-cadherin and decreased the expression of Vimentin, the mesenchymal marker, in both NSCLC cell lines and xenograft NSCLC tumours. Pretreatment with JNJ7777120 or H4 receptor gene silencing decreased while overexpression of H4 receptors facilitated this effect of 4-methylhistamine. Furthermore, we showed that down-regulation of cyclic adenosine monophosphate (cAMP) was the secondary signalling after H4 receptor activation, which in turn resulted in inactivation of transforming growth factor-β1 (TGF-β1) pathway and down-regulation of several important EMT inducing factors such as ZEB1, Snail and Slug. In conclusion, these findings revealed the anti-EMT effect of histamine H4 receptor activation in NSCLC, which provide novel insight into the development mechanism of NSCLC; and H4 receptors may be a new therapeutic target for NSCLC treatment. PMID:24447834

  13. Ispaghula (Plantago ovata) seed husk polysaccharides promote proliferation of human epithelial cells (skin keratinocytes and fibroblasts) via enhanced growth factor receptors and energy production.

    PubMed

    Deters, A M; Schröder, K R; Smiatek, T; Hensel, Andreas

    2005-01-01

    Endogenous carbohydrates, especially oligo- and polysaccharides, participate in the regulation of a broad range of biological activities, e. g., signal transduction, inflammation, fertilisation, cell-cell-adhesion and act as in vivo markers for the determination of cell types. In the present study, water-soluble (WS) and gel-forming polysaccharides (GF) of ispaghula seed husk (Plantago ovata Forsskal, Plantaginaceae) were characterised as neutral and acidic arabinoxylans and tested under in vitro conditions for regulating activities on cell physiology of human keratinocytes and human primary fibroblasts. Only water-soluble polysaccharides exhibited strong and significant effects on cell physiology of keratinocytes and fibroblasts. Proliferation of cells of the spontaneously immortalised keratinocyte cell line HaCaT was significantly up-regulated in a dose-independent manner. Analysis of activated signal pathways by RNA analysis proved an effect of the acidic arabinoxylan on the expression of keratinocyte growth factor (KGF) in HaCaT cells. Differentiation behaviour of normal human keratinocytes (NHK) determined by involucrin was slightly influenced, due to the enhanced cell proliferation, leading to a cell-cell-mediated indirect induction of early differentiation. WS did not influence late differentiation, as determined by keratin K1 and K10 titres. PMID:15678371

  14. Alveolar Epithelial Dynamics in Post-pneumonectomy Lung Growth

    PubMed Central

    Chamoto, Kenji; Gibney, Barry C.; Ackermann, Maximilian; Lee, Grace S.; Konerding, Moritz A.; Tsuda, Akira; Mentzer, Steven J.

    2013-01-01

    The intimate anatomic and functional relationship between epithelial cells and endothelial cells within the alveolus suggests the likelihood of a coordinated response during post-pneumonectomy lung growth. To define the population dynamics and potential contribution of alveolar epithelial cells to alveolar angiogenesis, we studied alveolar Type II and Type I cells during the 21 days after pneumonectomy. Alveolar Type II cells were defined and isolated by flow cytometry using a CD45−, MHC class II+, phosphine+ phenotype. These phenotypically defined alveolar Type II cells demonstrated an increase in cell number after pneumonectomy; the increase in cell number preceded the increase in Type I (T1α+) cells. Using a parabiotic wild type/GFP pneumonectomy model, less than 3% of the Type II cells and 1% of the Type I cells were positive for GFP—a finding consistent with the absence of a blood-borne contribution to alveolar epithelial cells. The CD45−, MHC class II+, phosphine+ Type II cells demonstrated the active transcription of angiogenesis-related genes both before and after pneumonectomy. When the Type II cells on day 7 after pneumonectomy were compared to non-surgical controls, 10 genes demonstrated significantly increased expression (p<.05). In contrast to the normal adult Type II cells, there was notable expression of inflammation-associated genes (Ccl2, Cxcl2, Ifng) as well as genes associated with epithelial growth (Ereg, Lep). Together, the data suggest an active contribution of local alveolar Type II cells to alveolar growth. PMID:23408540

  15. Insulin-Like Growth Factor I Receptor (IGF-IR) Ligands and BMI in Squamous Intra-Epithelial Lesion (SIL) of Cervix

    PubMed Central

    Batra, Swaraj; Saxena, Alpana

    2016-01-01

    Introduction Cancer cervix is the most common cancer in women in developing countries like India. Several studies have linked insulin-like growth factors-I & II (IGF-I and IGF-II) and IGF binding proteins-3 (IGFBP-3) with pathogenesis of Squamous Intraepithelial Lesion of cervix (SIL). To the best of our knowledge, no study has shown any correlation between circulating C-Peptide levels and SIL. Aim The present study has attempted to evaluate the correlation between SIL and IGF-IR ligands (IGF-I, IGF-II, C-Peptide), IGF binding protein (IGFBP-3) and Body Mass Index (BMI). Materials and Methods The present case-control study consisted of 31 histologically proven SIL cases and 31 age matched controls without evidence of SIL. A 10 ml blood sample was collected in heparinized vial. Plasma was separated immediately using centrifugation and was stored at -800 C till further analysis. Plasma levels of IGF-I, IGF-II, C-peptide and IGFBP3 were measured using commercially available Enzyme Linked Immunosorbent Assay (ELISA) kit. Height and weight was noted for calculation of BMI. Bio-effective molar ratio (BEMR) was calculated as 3.72 x {(0.25 x IGF-I) + (0.032 x IGF-II) + (0.0025 x C-peptide)} / {(1435 + IGFBP-3) – (2.79 x IGF-I) – (2.87 x IGF-II)}. Statistical analysis was performed using SPSS and Microsoft Excel software employing student t-test, Mann-Whitney and Chi-square test for trend while binary logistic regression was used to estimate the odds ratios (OR) and corresponding 95% Confidence Intervals (CI). Result IGF-I, IGF-II levels and BEMR were significantly increased in SIL compared to controls (p= 0.001, p <0.001, and p <0.001, respectively). C-Peptide levels were higher in controls than SIL (p = 0.04). IGFBP-3 & BMI in SIL were not significantly related when compared with controls. Risk of SIL in 4th quartile for BEMR, IGF-I, and IGF-II was 12.18(95% CI= 3.13-47.39), 3.94(95% CI = 1.24-12.56), and 4.57(95% CI = 1.42-14.7), respectively. Conclusion Elevated

  16. Human Epidermal Growth Factor Receptor 2 (HER-2/neu)-Directed Therapy for Rare Metastatic Epithelial Tumors with HER-2 Amplification

    PubMed Central

    Shin, Daniel Sanghoon; Sherry, Timothy; Kallen, Michael E.; Wong, Steven; Drakaki, Alexandra

    2016-01-01

    Case 1 A 67-year-old Asian female was diagnosed with locally advanced high-grade salivary duct carcinoma in June 2011. Molecular analysis revealed human epidermal growth factor receptor 2 (HER-2) amplification. She received adjuvant therapy with carboplatin/paclitaxel/ trastuzumab and maintenance of trastuzumab. Upon disease progression, trastuzumab could not be continued due to lack of financial coverage. Instead, she was treated with compassionate use of lapatinib from April 2013 and standard 5-fluorouracil. Her disease ultimately progressed and she expired later in 2013. Case 2 A 68-year-old Asian male was diagnosed with extramammary Paget's disease of the scrotum with HER-2 amplification in May 2011. He received 6 cycles of adjuvant trastuzumab/docetaxel/carboplatin followed by maintenance trastuzumab, which was changed to compassionate use of lapatinib as his insurance did not cover further administration of trastuzumab. He showed clinical benefits from single-agent lapatinib and a combination of lapatinib/capecitabine upon progression to the single-agent lapatinib. Ultimately, he was started on ado-trastuzumab emtansine, which was approved at that time by the FDA for HER-2-positive breast cancer progressed on trastuzumab. He is having clinical and radiographic complete response based on current imaging and normalization of his tumor markers. Conclusion HER-2-targeted therapy should be considered for tumors with HER-2 amplification. In our case series, we would like to emphasize this approach in other rare histologies. Specifically, our patient with extramammary Paget's disease of the scrotum represents the first reported case of a non-breast, non-gastric tumor with HER-2 overexpression with complete clinical and radiographic response to HER-2-targeted therapy PMID:27403128

  17. MicroRNA-27b reverses docetaxel resistance of non-small cell lung carcinoma cells via targeting epithelial growth factor receptor.

    PubMed

    Chen, Shi; Wang, Qian; Zhou, Xian-Mei; Zhu, Ji-Ping; Li, Tian; Huang, Mao

    2016-07-01

    MicroRNA (miR)‑27b has been reported to partici-pate in regulating the activity of non‑small cell lung carcinoma (NSCLC) cells. Additionally, when downregulated in NSCLC it promotes resistance to docetaxel; however, the underlying molecular mechanism remains largely unknown. Using reverse transcription‑quantitative polymerase chain reaction, the present study determined that the expression of miR‑27b was significantly reduced in NSCLC cells that were resistant to docetaxel. In addition, epidermal growth factor receptor (EGFR) was identified as a possible target of miR‑27b by searching the online miRNA database, TargetScan. A luciferase assay further validated EGFR as an effective target gene of miR‑27b. In addition, it was determined that in tumor tissue samples resistant to docetaxel miR‑27b was significantly downregulated, whilst EGFR was significantly upregulated. miR‑27b negatively regulated the expression of EGFR. This was evident as the transfection of miR‑27b mimics led to downregulation of the expression levels of EGFR, whilst miR‑27b inhibitors upregulated the expression levels of EGFR. Furthermore, it was demonstrated that the transfection of miR‑27b mimics significantly suppressed the apoptosis and promote the viability of A549 human lung carcinoma cells. In line with this, the introduction of miR‑27b inhibitors significantly induced apoptosis and inhibited the proliferation of A549 cells. These results indicate that miR‑27b may promote NSCLC cell viability and enhance resistance to docetaxel treatment through direct inhibition of EGFR expression. Additionally, miR‑27b may become a promising molecular target for improving the effectiveness of chemotherapy with docetaxel. PMID:27221512

  18. FGF growth factor analogs

    DOEpatents

    Zamora, Paul O.; Pena, Louis A.; Lin, Xinhua; Takahashi, Kazuyuki

    2012-07-24

    The present invention provides a fibroblast growth factor heparin-binding analog of the formula: ##STR00001## where R.sub.1, R.sub.2, R.sub.3, R.sub.4, R.sub.5, X, Y and Z are as defined, pharmaceutical compositions, coating compositions and medical devices including the fibroblast growth factor heparin-binding analog of the foregoing formula, and methods and uses thereof.

  19. Caffeic acid phenethyl ester reduces the secretion of vascular endothelial growth factor through the inhibition of the ROS, PI3K and HIF-1α signaling pathways in human retinal pigment epithelial cells under hypoxic conditions.

    PubMed

    Paeng, Sung Hwa; Jung, Won-Kyo; Park, Won Sun; Lee, Dae-Sung; Kim, Gi-Young; Choi, Yung Hyun; Seo, Su-Kil; Jang, Won Hee; Choi, Jung Sik; Lee, Young-Min; Park, Saegwang; Choi, Il-Whan

    2015-05-01

    Choroidal neovascularization (CNV) can lead to progressive and severe visual loss. Vascular endothelial growth factor (VEGF) promotes the development of CNV. Caffeic acid phenethyl ester (CAPE), a biologically active component of the honeybee (Apis mellifera) propolis, has been demonstrated to have several interesting biological regulatory properties. The objective of this study was to determine whether treatment with CAPE results in the inhibition of the production of vascular endothelial growth factor (VEGF) in retinal pigment epithelial cells (RPE cells) under hypoxic conditions and to explore the possible underlying mechanisms. An in vitro experimental model of hypoxia was used to mimic an ischemic microenvironment for the RPE cells. Human RPE cells (ARPE-19) were exposed to hypoxia with or without CAPE pre-treatment. ARPE-19 cells were used to investigate the pathway involved in the regulation of VEGF production under hypoxic conditions, based on western blot analysis, enzyme-linked immunosorbent assay (ELISA) and electrophoretic mobility shift assay (EMSA). The amount of VEGF released from the hypoxia-exposed cells was significantly higher than that of the normoxic controls. Pre-treatment with CAPE suppressed the hypoxia-induced production of VEGF in the ARPE-19 cells, and this effect was inhibited through the attenuation of reactive oxygen species (ROS) production, and the inhibition of phosphoinositide 3-kinase (PI3K)/AKT and hypoxia-inducible factor-1α (HIF-1α) expression. These in vitro findings suggest that CAPE may prove to be a novel anti-angiogenic agent for the treatment of diseases associated with CNV. PMID:25738890

  20. Hedgehog signaling in prostate epithelial-mesenchymal growth regulation

    PubMed Central

    Peng, Yu-Ching; Joyner, Alexandra L.

    2015-01-01

    The prostate gland plays an important role in male reproduction, and is also an organ prone to diseases such as benign prostatic hyperplasia (BPH) and prostate cancer. The prostate consists of ducts with an inner layer of epithelium surrounded by stroma. Reciprocal signaling between these two cell compartments is instrumental to normal prostatic development, homeostasis, regeneration, as well as tumor formation. Hedgehog (HH) signaling is a master regulator in numerous developmental processes. In many organs, HH plays a key role in epithelial-mesenchymal signaling that regulates organ growth and tissue differentiation, and abnormal HH signaling has been implicated in the progression of various epithelial carcinomas. In this review, we focus on recent studies exploring the multipotency of endogenous postnatal and adult epithelial and stromal stem cells and studies addressing the role of HH in prostate development and cancer. We discuss the implications of the results for a new understanding of prostate development and disease. Insight into the cellular and molecular mechanisms underlying epithelial-mesenchymal growth regulation should provide a basis for devising innovative therapies to combat diseases of the prostate. PMID:25641695

  1. Heterogeneity and stochastic growth regulation of biliary epithelial cells dictate dynamic epithelial tissue remodeling

    PubMed Central

    Kamimoto, Kenji; Kaneko, Kota; Kok, Cindy Yuet-Yin; Okada, Hajime; Miyajima, Atsushi; Itoh, Tohru

    2016-01-01

    Dynamic remodeling of the intrahepatic biliary epithelial tissue plays key roles in liver regeneration, yet the cellular basis for this process remains unclear. We took an unbiased approach based on in vivo clonal labeling and tracking of biliary epithelial cells in the three-dimensional landscape, in combination with mathematical simulation, to understand their mode of proliferation in a mouse liver injury model where the nascent biliary structure formed in a tissue-intrinsic manner. An apparent heterogeneity among biliary epithelial cells was observed: whereas most of the responders that entered the cell cycle upon injury exhibited a limited and tapering growth potential, a select population continued to proliferate, making a major contribution in sustaining the biliary expansion. Our study has highlighted a unique mode of epithelial tissue dynamics, which depends not on a hierarchical system driven by fixated stem cells, but rather, on a stochastically maintained progenitor population with persistent proliferative activity. DOI: http://dx.doi.org/10.7554/eLife.15034.001 PMID:27431614

  2. Osthole suppresses hepatocyte growth factor (HGF)-induced epithelial-mesenchymal transition via repression of the c-Met/Akt/mTOR pathway in human breast cancer cells.

    PubMed

    Hung, Chao-Ming; Kuo, Daih-Huang; Chou, Chun-Hung; Su, Yen-Chao; Ho, Chi-Tang; Way, Tzong-Der

    2011-09-14

    Substantial activation of the HGF/c-Met signaling pathway is involved in the progression of several types of cancers and associated with increased tumor invasion and metastatic potential. Underlying HGF-induced tumorigenesis, epithelial to mesenchymal transition (EMT) shows a positive correlation with progression in patients. We previously determined that osthole is a potent fatty acid synthase (FASN) inhibitor. FASN is implicated in cancer progression and may regulate lipid raft function. We therefore examined whether osthole could block HGF-induced tumorigenesis by disrupting lipid rafts. Here, we found that osthole could abrogate HGF-induced cell scattering, migration, and invasion in MCF-7 breast cancer cells. Osthole also effectively inhibited the HGF-induced decrease of E-cadherin and increase of vimentin via down-regulation of phosphorylated Akt and mTOR. Interestingly, osthole blocked HGF-induced c-Met phosphorylation and repressed the expression of total c-Met protein in MCF-7 cells. In addition, C75, a pharmacological inhibitor of FASN, repressed the expression of total c-Met protein in MCF-7 cells. Consistent with a role for FASN, loss of c-Met in cells treated with osthole was prevented by the exogenous addition of palmitate. Briefly, our result suggests a connection between FASN activity and c-Met protein expression and that osthole is a potential compound for breast cancer therapy by targeting the major pathway of HGF/c-Met-induced EMT. PMID:21806057

  3. Growth factors for nanobacteria

    NASA Astrophysics Data System (ADS)

    Ciftcioglu, Neva; Kajander, E. Olavi

    1999-12-01

    Nanobacteria are novel microorganisms recently isolated from fetal bovine serum and blood of cows and humans. These coccoid, gram negative bacteria in alpha-2 subgroup of Proteobacteria grow slowly under mammalian cell culture conditions but not in common media for microbes. Now we have found two different kinds of culture supplement preparations that improve their growth and make them culturable in the classical sense. These are supernatant fractions of conditioned media obtained from 1 - 3 months old nanobacteria cultures and from about a 2 weeks old Bacillus species culture. Both improved multiplication and particle yields and the latter increased their resistance to gentamicin. Nanobacteria cultured with any of the methods shared similar immunological property, structure and protein pattern. The growth supporting factors were heat-stabile and nondialyzable, and dialysis improved the growth promoting action. Nanobacteria formed stony colonies in a bacteriological medium supplemented with the growth factors. This is an implication that nanobacterial growth is influenced by pre-existing bacterial flora.

  4. Inhibition of protein tyrosine phosphatase activity mediates epidermal growth factor receptor signaling in human airway epithelial cells exposed to Zn{sup 2+}

    SciTech Connect

    Tal, T.L.; Graves, L.M.; Silbajoris, R.; Bromberg, P.A.; Wu, W.; Samet, J.M. . E-mail: samet.james@epa.gov

    2006-07-01

    Epidemiological studies have implicated zinc (Zn{sup 2+}) in the toxicity of ambient particulate matter (PM) inhalation. We previously showed that exposure to metal-laden PM inhibits protein tyrosine phosphatase (PTP) activity in human primary bronchial epithelial cells (HAEC) and leads to Src-dependent activation of EGFR signaling in B82 and A431 cells. In order to elucidate the mechanism of Zn{sup 2+}-induced EGFR activation in HAEC, we treated HAEC with 500 {mu}M ZnSO{sub 4} for 5-20 min and measured the state of activation of EGFR, c-Src and PTPs. Western blots revealed that exposure to Zn{sup 2+} results in increased phosphorylation at both trans- and autophosphorylation sites in the EGFR. Zn{sup 2+}-mediated EGFR phosphorylation did not require ligand binding and was ablated by the EGFR kinase inhibitor PD153035, but not by the Src kinase inhibitor PP2. Src activity was inhibited by Zn{sup 2+} treatment of HAEC, consistent with Src-independent EGFR transactivation in HAEC exposed to Zn{sup 2+}. The rate of exogenous EGFR dephosphorylation in lysates of HAEC exposed to Zn{sup 2+} or V{sup 4+} was significantly diminished. Moreover, exposure of HAEC to Zn{sup 2+} also resulted in a significant impairment of dephosphorylation of endogenous EGFR. These data show that Zn{sup 2+}-induced activation of EGFR in HAEC involves a loss of PTP activities whose function is to dephosphorylate EGFR in opposition to baseline EGFR kinase activity. These findings also suggest that there are marked cell-type-specific differences in the mechanism of EGFR activation induced by Zn{sup 2+} exposure.

  5. New microbial growth factor

    NASA Technical Reports Server (NTRS)

    Bok, S. H.; Casida, L. E., Jr.

    1977-01-01

    A screening procedure was used to isolate from soil a Penicillium sp., two bacterial isolates, and a Streptomyces sp. that produced a previously unknown microbial growth factor. This factor was an absolute growth requirement for three soil bacteria. The Penicillium sp. and one of the bacteria requiring the factor, an Arthrobacter sp., were selected for more extensive study concerning the production and characteristics of the growth factor. It did not seem to be related to the siderochromes. It was not present in soil extract, rumen fluid, or any other medium component tested. It appears to be a glycoprotein of high molecular weight and has high specific activity. When added to the diets for a meadow-vole mammalian test system, it caused an increased consumption of diet without a concurrent increase in rate of weight gain.

  6. Growth factors in haemopoiesis.

    PubMed

    Jones, A L; Millar, J L

    1989-01-01

    Haemopoietic growth factors have for over two decades allowed experimentalists to grow haemopoietic bone marrow cells in vitro. With refinements in technique and the discovery of novel growth factors, all of the known haemopoietic lineages can now be grown in vitro. This has allowed a much greater understanding of the complex process of haemopoiesis from the haemopoietic stem cell to the mature, functioning end-cell. The in vivo action of these growth factors has been harder to investigate. Although recombinant technology has afforded us the much greater quantities necessary for in vivo work, problems remain with administration because of effects on other tissues. Interpretation of results is difficult because of the complex inter-relationships which exist between factors. Some of these have been defined in vitro and it appears likely that they also operate in vivo. Erythropoietin is a physiological regulator of erythropoiesis. It has been detected in vivo with levels responding appropriately to stress (i.e. elevated in anaemia) and, when administered in pharmacological doses, has been shown to correct anaemia. Granulocyte/macrophage colony-stimulating factor (GM-CSF) has been detected in vivo and may influence the production and function of granulocytes and macrophages, although how it is regulated is unknown. Granulocyte colony-stimulating factor (G-CSF) and macrophage colony-stimulating factor are ore lineage-specific. Interleukin 3 (IL-3), although it has not been detected in vivo, may act on a primitive marrow precursor by expanding the population and making these cells more susceptible to other growth factors, such as GM-CSF. Interleukin 1 (IL-1) has been detected in vivo, does not appear to have any isolated action on bone marrow (except possibly radioprotection) but probably acts synergistically with other growth factors, such as G-CSF. Interleukins 2, 4, 5 and 6 have not been detected in vivo. All have effects on B-cells. In addition IL-2 is an essential

  7. Epithelial growth by rat vibrissae follicles in vitro requires mesenchymal contact via native extracellular matrix

    SciTech Connect

    Link, R.E.; Paus, R.; Stenn, K.S.; Kuklinska, E.; Moellmann, G. )

    1990-08-01

    An in vitro assay utilizing the rat vibrissa anagen follicle as a model for studying the epithelial-mesenchymal interactions (EMI) in hair growth is described. Through selective disruption of the epithelial-mesenchymal interface, we investigate whether the specialized extracellular matrix (ECM) of the dermal papilla and basement membrane zone (BMZ) serves a crucial function in hair follicle EMI. Epithelial bulbs incubated intact within their follicular sheaths incorporate thymidine primarily into cells of the hair matrix and outer root sheath, as shown by autoradiography. However, after removal of its mesenchymal associations (dermal papilla and extrabulbar connective tissue), the epithelial bulb showed no incorporation. Neither externally added collagen (type I or IV) nor the basement membrane components in Matrigel could substitute for the growth supporting influence of native surrounding stroma. Mechanical separation of the bulb from the dermal papilla in the basement membrane zone inhibited thymidine incorporation by the epithelium even though mesenchyme was still in close proximity. Enzymatic digestion of the dermal papilla ECM and the basal lamina by Dispase, a fibronectinase and type IV collagenase, also inhibited bulb growth without evidence of cytotoxicity. These experiments suggest that direct epithelial to mesenchymal contact is required for the support of follicular epithelial growth in vitro and that specific ECM components, possibly fibronectin and/or type IV collagen, rather than diffusable factors alone, play a crucial role in the mechanism of hair follicle EMI. The in vitro system described here provides an alternative to developmental EMI models and may serve as a valuable tool for studying EMI in the adult mammalian organism.

  8. Thyrotropin via cyclic AMP induces insulin receptor expression and insulin Co-stimulation of growth and amplifies insulin and insulin-like growth factor signaling pathways in dog thyroid epithelial cells.

    PubMed

    Burikhanov, R; Coulonval, K; Pirson, I; Lamy, F; Dumont, J E; Roger, P P

    1996-11-15

    Despite the similarity of their receptors and signal transduction pathways, insulin is regarded as a regulator of glucose, protein, and lipid metabolism, whereas insulin-like growth factors (IGF-I and IGF-II) mainly act as mitogenic hormones. In the dog thyroid primary culture model, the triggering of DNA synthesis by thyrotropin (TSH) through cAMP, or by cAMP-independent factors including epidermal growth factor, hepatocyte growth factor and phorbol esters, requires insulin or IGFs as comitogenic factors. In the present study, in TSH-treated cells, IGF-I receptors and insulin receptors were paradoxically equivalent in their capacity to elicit the comitogenic pathway, which, however, was mediated only by IGF-I receptors in dog thyroid cells stimulated by cAMP-independent mitogens. Moreover, prior cell exposure to TSH or forskolin increased their responsiveness to insulin, IGF-I, and IGF-II, as seen on DNA synthesis and activation of a common insulin/IGF signaling pathway. To understand these observations, binding characteristics and expression of insulin and IGF-I receptors were examined. To analyze IGF-I receptor characteristics, the unexpected interference of a huge presence of IGF-binding proteins at the cell membrane was avoided using labeled Long R3 IGF-I instead of IGF-I. Strikingly, TSH, through cAMP, time-dependently induced insulin binding and insulin receptor mRNA and protein accumulation without any effect on IGF-I receptors. These findings constitute a first example of an induction of insulin receptor gene expression by a cAMP-mediated hormone. In dog thyroid cells, this allows low physiological insulin concentrations to act as a comitogenic factor and might explain in part the enhanced responsiveness to IGFs in response to TSH. This raises the possibility that TSH-insulin interactions may play a role in the regulation of thyroid growth and function in vivo. PMID:8910605

  9. Transcriptional regulators transforming growth factor-beta 1 and estrogen-related receptor-alpha identified as putative mediators of calf rumen epithelial tissue development and function during weaning

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Molecular mechanisms controlling rumen epithelial development at weaning remain largely unknown. To identify gene networks and regulatory factors responsive to concentrate versus forage feeding at weaning, Holstein bull calves (n = 18) were fed commercial milk replacer only (MRO) until 42 d of age. ...

  10. Purification of scatter factor, a fibroblast-derived basic protein that modulates epithelial interactions and movement.

    PubMed Central

    Gherardi, E; Gray, J; Stoker, M; Perryman, M; Furlong, R

    1989-01-01

    Scatter factor is a fibroblast-derived protein that causes separation of contiguous epithelial cells and increased local mobility of unanchored cells. Highly purified scatter factor has been obtained by a combination of ion-exchange and reverse-phase chromatography from serum-free medium conditioned by a ras-transformed clone (D4) of mouse NIH 3T3 fibroblasts. Under nonreducing conditions scatter factor has a pI of approximately 9.5 and migrates in SDS/polyacrylamide gels as a single band at approximately 62 kDa from which epithelial scatter activity can be recovered. Treatment with reducing agents destroys biological activity and is associated with the appearance of two major bands at approximately 57 and approximately 30 kDa. Whether both the 57-kDa and 30-kDa polypeptides are required for biological activity remains to be established. All the activities observed in crude medium conditioned by cells producing scatter factor are retained by highly purified preparations of scatter factor. These include (i) increased local movement, modulation of morphology, and inhibition of junction formation by single epithelial cells and (ii) disruption of epithelial interactions and cell scattering from preformed epithelial sheets. These changes occur with picomolar concentrations of purified scatter factor and without an effect on cell growth. Images PMID:2527367

  11. Keratinocyte growth factor and androgen blockade work in concert to protect against conditioning regimen-induced thymic epithelial damage and enhance T-cell reconstitution after murine bone marrow transplantation

    PubMed Central

    Kelly, Ryan M.; Highfill, Steven L.; Panoskaltsis-Mortari, Angela; Taylor, Patricia A.; Boyd, Richard L.; Holländer, Georg A.

    2008-01-01

    Myeloablative conditioning results in thymic epithelial cell (TEC) injury, slow T-cell reconstitution, and a high risk of opportunistic infections. Keratinocyte growth factor (KGF) stimulates TEC proliferation and, when given preconditioning, reduces TEC injury. Thymocytes and TECs express androgen receptors, and exposure to androgen inhibits thymopoiesis. In this study, we have investigated whether TEC stimulation via preconditioning treatment with KGF and leuprolide acetate (Lupron), 2 clinically approved agents, given only before conditioning would circumvent the profound TEC and associated T-cell deficiency seen in allogeneic bone marrow transplant (BMT) recipients. Only combined treatment with KGF plus leuprolide acetate normalized TEC subset numbers and thymic architecture. Thymopoiesis and thymic output were supranormal, leading to the accelerated peripheral reconstitution of naive CD4 and CD8 T cells with a broad Vβ repertoire and decreased homeostatic T-cell proliferation. Combined therapy facilitated T:B cooperativity and enabled a B-cell humoral response to a CD4 T cell–dependent neoantigen challenge soon after BMT. In vivo antigen-specific CD8 T-cell responses and clearance of a live pathogen was superior with combined versus individual agent therapy. Thus, KGF combined with androgen blockade represents a novel approach to restore thymic function and facilitates the rapid recovery of peripheral T-cell function after allogeneic BMT. PMID:18334670

  12. Peptide growth factors, part B

    SciTech Connect

    Barnes, D.; Sirbasku, D.A.

    1987-01-01

    This book discusses the following topics: Platelet-Derived Growth Factor;Nerve and Glial Growth Factors;PC12 Pheochromocytoma Cells;Techniques for the Study of Growth Factor Activity;Genetic Approaches and Biological Effects.

  13. Stress-Derived Corticotropin Releasing Factor Breaches Epithelial Endotoxin Tolerance

    PubMed Central

    Yu, Yong; Geng, Xiao-Rui; Yang, Gui; Liu, Zhi-Gang; Zheng, Peng-Yuan; Yang, Ping-Chang

    2013-01-01

    Background and aims Loss of the endotoxin tolerance of intestinal epithelium contributes to a number of intestinal diseases. The etiology is not clear. Psychological stress is proposed to compromise the intestinal barrier function. The present study aims to elucidate the role of the stress-derived corticotropin releasing factor (CRF) in breaching the established intestinal epithelial endotoxin tolerance. Methods Epithelial cells of HT-29, T84 and MDCK were exposed to lipopolysaccharide to induce the endotoxin tolerance; the cells were then stimulated with CRF. The epithelial barrier function was determined using as indicators of the endotoxin tolerant status. A water-avoid stress mouse model was employed to test the role of CRF in breaching the established endotoxin tolerance in the intestine. Results The established endotoxin tolerance in the epithelial cell monolayers was broken down by a sequent exposure to CRF and LPS manifesting a marked drop of the transepithelial resistance (TER) and an increase in the permeability to a macromolecular tracer, horseradish peroxidase (HRP). The exposure to CRF also increased the expression of Cldn2 in the epithelial cells, which could be mimicked by over expression of TLR4 in epithelial cells. Over expression of Cldn2 resulted in low TER in epithelial monolayers and high permeability to HRP. After treating mice with the 10-day chronic stress, the intestinal epithelial barrier function was markedly compromised, which could be prevented by blocking either CRF, or TLR4, or Cldn2. Conclusions Psychological stress-derived CRF can breach the established endotoxin tolerance in the intestinal mucosa. PMID:23840363

  14. Regulation of Lysophosphatidic Acid-induced Epidermal Growth Factor Receptor Transactivation and Interleukin-8 Secretion in Human Bronchial Epithelial Cells by Protein Kinase Cδ, Lyn Kinase, and Matrix Metalloproteinases*

    PubMed Central

    Zhao, Yutong; He, Donghong; Saatian, Bahman; Watkins, Tonya; Spannhake, Ernst Wm.; Pyne, Nigel J.; Natarajan, Viswanathan

    2009-01-01

    We have demonstrated earlier that lysophosphatidic acid (LPA)-induced interleukin-8 (IL-8) secretion is regulated by protein kinase Cδ (PKCδ)-dependent NF-κB activation in human bronchial epithelial cells (HBEpCs). Here we provide evidence for signaling pathways that regulate LPA-mediated transactivation of epidermal growth factor receptor (EGFR) and the role of cross-talk between G-protein-coupled receptors and receptor-tyrosine kinases in IL-8 secretion in HBEpCs. Treatment of HBEpCs with LPA stimulated tyrosine phosphorylation of EGFR, which was attenuated by matrix metalloproteinase (MMP) inhibitor (GM6001), heparin binding (HB)-EGF inhibitor (CRM 197), and HB-EGF neutralizing antibody. Overexpression of dominant negative PKCδ or pretreatment with a PKCδ inhibitor (rottlerin) or Src kinase family inhibitor (PP2) partially blocked LPA-induced MMP activation, proHB-EGF shedding, and EGFR tyrosine phosphorylation. Down-regulation of Lyn kinase, but not Src kinase, by specific small interfering RNA mitigated LPA-induced MMP activation, proHB-EGF shedding, and EGFR phosphorylation. In addition, overexpression of dominant negative PKCδ blocked LPA-induced phosphorylation and translocation of Lyn kinase to the plasma membrane. Furthermore, down-regulation of EGFR by EGFR small interfering RNA or pretreatment of cells with EGFR inhibitors AG1478 and PD158780 almost completely blocked LPA-dependent EGFR phosphorylation and partially attenuated IL-8 secretion, respectively. These results demonstrate that LPA-induced IL-8 secretion is partly dependent on EGFR transactivation regulated by PKCδ-dependent activation of Lyn kinase and MMPs and proHB-EGF shedding, suggesting a novel mechanism of cross-talk and interaction between G-protein-coupled receptors and receptor-tyrosine kinases in HBEpCs. PMID:16687414

  15. Kaempferol Suppresses Transforming Growth Factor-β1-Induced Epithelial-to-Mesenchymal Transition and Migration of A549 Lung Cancer Cells by Inhibiting Akt1-Mediated Phosphorylation of Smad3 at Threonine-179.

    PubMed

    Jo, Eunji; Park, Seong Ji; Choi, Yu Sun; Jeon, Woo-Kwang; Kim, Byung-Chul

    2015-07-01

    Kaempferol, a natural dietary flavonoid, is well known to possess chemopreventive and therapeutic anticancer efficacy; however, its antimetastatic effects have not been mechanistically studied so far in any cancer model. This study was aimed to investigate the inhibitory effect and accompanying mechanisms of kaempferol on epithelial-to-mesenchymal transition (EMT) and cell migration induced by transforming growth factor-β1 (TGF-β1). In human A549 non-small lung cancer cells, kaempferol strongly blocked the enhancement of cell migration by TGF-β1-induced EMT through recovering the loss of E-cadherin and suppressing the induction of mesenchymal markers as well as the upregulation of TGF-β1-mediated matrix metalloproteinase-2 activity. Interestingly, kaempferol reversed TGF-β1-mediated Snail induction and E-cadherin repression by weakening Smad3 binding to the Snail promoter without affecting its C-terminus phosphorylation, complex formation with Smad4, and nuclear translocation under TGF-β1 stimulation. Mechanism study revealed that the phosphorylation of Smad3 linker region induced by TGF-β1 was required for the induction of EMT and cell migration, and selective downregulation of the phosphorylation of Smad3 at Thr179 residue (not Ser204, Ser208, and Ser213) in the linker region was responsible for the inhibition by kaempferol of TGF-β1-induced EMT and cell migration. Furthermore, Akt1 was required for TGF-β1-mediated induction of EMT and cell migration and directly phosphorylated Smad3 at Thr179, and kaempferol completely abolished TGF-β1-induced Akt1 phosphorylation. In summary, kaempferol blocks TGF-β1-induced EMT and migration of lung cancer cells by inhibiting Akt1-mediated phosphorylation of Smad3 at Thr179 residue, providing the first evidence of a molecular mechanism for the anticancer effect of kaempferol. PMID:26297431

  16. Kaempferol Suppresses Transforming Growth Factor-β1–Induced Epithelial-to-Mesenchymal Transition and Migration of A549 Lung Cancer Cells by Inhibiting Akt1-Mediated Phosphorylation of Smad3 at Threonine-1791

    PubMed Central

    Jo, Eunji; Park, Seong Ji; Choi, Yu Sun; Jeon, Woo-Kwang; Kim, Byung-Chul

    2015-01-01

    Kaempferol, a natural dietary flavonoid, is well known to possess chemopreventive and therapeutic anticancer efficacy; however, its antimetastatic effects have not been mechanistically studied so far in any cancer model. This study was aimed to investigate the inhibitory effect and accompanying mechanisms of kaempferol on epithelial-to-mesenchymal transition (EMT) and cell migration induced by transforming growth factor-β1 (TGF-β1). In human A549 non–small lung cancer cells, kaempferol strongly blocked the enhancement of cell migration by TGF-β1–induced EMT through recovering the loss of E-cadherin and suppressing the induction of mesenchymal markers as well as the upregulation of TGF-β1–mediated matrix metalloproteinase-2 activity. Interestingly, kaempferol reversed TGF-β1–mediated Snail induction and E-cadherin repression by weakening Smad3 binding to the Snail promoter without affecting its C-terminus phosphorylation, complex formation with Smad4, and nuclear translocation under TGF-β1 stimulation. Mechanism study revealed that the phosphorylation of Smad3 linker region induced by TGF-β1 was required for the induction of EMT and cell migration, and selective downregulation of the phosphorylation of Smad3 at Thr179 residue (not Ser204, Ser208, and Ser213) in the linker region was responsible for the inhibition by kaempferol of TGF-β1–induced EMT and cell migration. Furthermore, Akt1 was required for TGF-β1–mediated induction of EMT and cell migration and directly phosphorylated Smad3 at Thr179, and kaempferol completely abolished TGF-β1–induced Akt1 phosphorylation. In summary, kaempferol blocks TGF-β1–induced EMT and migration of lung cancer cells by inhibiting Akt1-mediated phosphorylation of Smad3 at Thr179 residue, providing the first evidence of a molecular mechanism for the anticancer effect of kaempferol. PMID:26297431

  17. An in vitro model of epithelial cell growth stimulation in the rodent mammary gland.

    PubMed

    Ehmann, U K; DeVries, J T; Chen, M S C; Adamos, A A; Guzman, R C; Omary, M B

    2003-08-01

    Mouse mammary epithelial cell cultures previously described bring about extensive proliferation and a cell population with the appropriate markers for luminal ductal epithelial cells, and also the ability to form normal tissue after implantation into mice. This success may result from a culture environment that resembles certain aspects of the environment in the mammary gland. Mouse mammary epithelial cells, whose proliferation is limited when plated alone, can be stimulated to multiply by contact with lethally irradiated cells of the LA7 rat mammary tumour line. Most of the proliferative stimulus is imparted by direct cell contact between LA7 and mouse mammary cells. Junctions, including adherens junctions, form among all cells in the culture, much as junctions form in the mammary gland. LA7 cells secrete TGFalpha and bFGF, factors found in the mammary gland, and factors to which mouse mammary cells respond in culture. Mouse mammary cells express keratins 8 and 18, markers for luminal cells of the mammary duct. LA7 cells express keratin 14 and vimentin, markers for myoepithelial cells. These facts, taken together, fit a model of cell replacement in an epithelial tissue and also imitate the relationship between luminal ductal cells and myoepithelial cells in the mammary gland. This method of culturing cells is useful, not only for in vitro-in vivo carcinogenesis studies, but also for the study of mechanisms by which growth signals are imparted from one cell to another. PMID:12950387

  18. Extracellular matrix and growth factors in branching morphogenesis

    NASA Technical Reports Server (NTRS)

    Hardman, P.; Spooner, B. S.

    1993-01-01

    The unifying hypothesis of the NSCORT in gravitational biology postulates that the ECM and growth factors are key interrelated components of a macromolecular regulatory system. The ECM is known to be important in growth and branching morphogenesis of embryonic organs. Growth factors have been detected in the developing embryo, and often the pattern of localization is associated with areas undergoing epithelial-mesenchymal interactions. Causal relationships between these components may be of fundamental importance in control of branching morphogenesis.

  19. Growth hormone, growth factors, and acromegaly

    SciTech Connect

    Ludecke, D.K.; Tolis, G.T.

    1987-01-01

    This book contains five sections, each consisting of several papers. The section headings are: Biochemistry and Physiology of GH and Growth Factors, Pathology of Acromegaly, Clinical Endocrinology of Acromegaly, Nonsurgical Therapy of Acromegaly, and Surgical Therapy of Acromegaly.

  20. The Rho guanine nucleotide exchange factor ARHGEF5 promotes tumor malignancy via epithelial-mesenchymal transition.

    PubMed

    Komiya, Y; Onodera, Y; Kuroiwa, M; Nomimura, S; Kubo, Y; Nam, J-M; Kajiwara, K; Nada, S; Oneyama, C; Sabe, H; Okada, M

    2016-01-01

    Epithelial tumor cells often acquire malignant properties, such as invasion/metastasis and uncontrolled cell growth, by undergoing epithelial-mesenchymal transition (EMT). However, the mechanisms by which EMT contributes to malignant progression remain elusive. Here we show that the Rho guanine nucleotide exchange factor (GEF) ARHGEF5 promotes tumor malignancy in a manner dependent on EMT status. We previously identified ARHGEF5, a member of the Dbl family of GEFs, as a multifunctional mediator of Src-induced cell invasion and tumor growth. In the present study, ARHGEF5 was upregulated during tumor growth factor-β-induced EMT in human epithelial MCF10A cells, and promoted cell migration by activating the Rho-ROCK pathway. ARHGEF5 was necessary for the invasive and in vivo metastatic activity of human colorectal cancer HCT116 cells. These findings underscore the crucial role of ARHGEF5 in cell migration and invasion/metastasis. An in vivo tumorigenesis assay revealed that ARHGEF5 had the potential to promote tumor growth via the phosphatidylinositol 3-kinase (PI3K) pathway. However, ARHGEF5 was not required for tumor growth in epithelial-like human colorectal cancer HCT116 and HT29 cells, whereas the growth of mesenchymal-like SW480 and SW620 cells depended on ARHGEF5. Induction of EMT by tumor necrosis factor-α or Slug in HCT116 cells resulted in the dependence of tumor growth on ARHGEF5. In these mesenchymal-like cells, Akt was activated via ARHGEF5 and its activity was required for tumor growth. Analysis of a transcriptome data set revealed that the combination of ARHGEF5 upregulation and E-cadherin downregulation or Snail upregulation was significantly correlated with poor prognosis in patients with colorectal cancers. Taken together, our findings suggest that EMT-induced ARHGEF5 activation contributes to the progression of tumor malignancy. ARHGEF5 may serve as a potential therapeutic target in a subset of malignant tumors that have undergone EMT. PMID

  1. The Roles of Growth Factors in Keratinocyte Migration

    PubMed Central

    Seeger, Mark A.; Paller, Amy S.

    2015-01-01

    Significance: The re-epithelialization of wounded skin requires the rapid and coordinated migration of keratinocytes (KC) into the wound bed. Almost immediately after wounding, cells present at or attracted to the wound site begin to secrete a complex milieu of growth factors. These growth factors exert mitogenic and motogenic effects on KCs, inducing the rapid proliferation and migration of KCs at the wound edge. Recent Advances: New roles for growth factors in KC biology are currently being discovered and investigated. This review will highlight the growth factors, particularly transforming growth factor-α (TGF-α), heparin-binding epidermal growth factor (HB-EGF), insulin-like growth factor 1 (IGF-1), fibroblast growth factor 7 (FGF-7), FGF-10, and hepatocyte growth factor (HGF), which have conclusively been shown to be the most motogenic for KCs. Critical Issues: The cellular and molecular heterogeneity of wounded tissue makes establishing direct relationships between specific growth factors and KC migration difficult in situ. The absence of this complexity in simplified in vitro experimental models of migration makes the clinical relevance of the results obtained from these in vitro studies ambiguous. Future Directions: Deciphering the relationship between growth factors and KC migration is critical for understanding the process of wound healing in normal and disease states. Insights into the basic science of the effects of growth factors on KC migration will hopefully lead to the development of new therapies to treat acute and chronic wounds. PMID:25945284

  2. Heparin Binding Epidermal Growth Factor Like Growth Factor Heals Chronic Tympanic Membrane Perforations With Advantage Over Fibroblast Growth Factor 2 and Epidermal Growth Factor in an Animal Model

    PubMed Central

    Santa Maria, Peter Luke; Weierich, Kendall; Kim, Sungwoo; Yang, Yunzhi Peter

    2016-01-01

    Hypothesis That heparin binding epidermal growth factor like growth factor (HB-EGF) heals chronic tympanic membrane (TM) perforations at higher rates than fibroblast growth factor 2 (FGF2) and epidermal growth factor (EGF) in an animal model. Background A non-surgical treatment for chronic TM perforation would benefit those unable to access surgery or those unable to have surgery, as well as reducing the cost of tympanoplasty. Growth factor (GF) treatments have been reported in the literature with variable success with the lack of a suitable animal providing a major obstacle. Methods The GFs were tested in a validated mouse model of chronic TM perforation. A bio absorbable hydrogel polymer was used to deliver the GF at a steady concentration as it dissolved over four weeks. A control (polymer only, n=18) was compared to polymer loaded with HB-EGF (5ug/ml, n=18), FGF2 (100ug/ml, n=19) and EGF (250ug/ml, n=19). Perforations were inspected at four weeks. Results The healing rates, as defined as one hundred percent perforation closure, were control (5/18, 27.8%), HB-EGF (15/18, 83.3%), FGF2 (6/19, 31.6%) and EGF (3/19, 15.8%). There were no differences between FGF2 (p=0.80) and EGF (p=0.31) with control healing rates. HB-EGF (p= 0.000001) showed a significant difference for healing. The HB-EGF healed TMs showed layers similar to a normal TM, whilst the other groups showed a lack of epithelial migration. Conclusion This study confirms the advantage of HB-EGF over two other commonly used growth factors and is a promising non-surgical treatment of chronic TM perforations. PMID:26075672

  3. A critical synopsis: Continuous growth of proximal tubular kidney epithelial cells in hormone-supplemented serum-free medium

    NASA Technical Reports Server (NTRS)

    Chuman, L. M.; FINE; COHEN; Saier, M. H.

    1985-01-01

    The kidney forms urine and reabsorbs electrolytes and water. Kidney cell lines and hormone supplemented serum free medium were used for growth. The hormones were insulin, transferrin, vasopressin, cholesterol, prostaglandins, hydrocortisone, and triidothyronine. Epithelial cell lines are polar and form hemicysts. The Madin-Darby canine kidney(MDCK) cell line used is distal tubulelike. LLC-PK sub 1 cells are derived from pig kidneys and have the properties of different kidney segments. The LLC-PK sub 1 cells with proximal tubule properties were maintained in hormone-supplemented serum free medium. Seven factors (the aforementioned homrones and selenium) were needed for growth. Hormone-defined medium supported LLC-PK sub 1 cell growth, allowed transport (as seen by hemicyst formation), and influenced cell morphology. Vasopressin (used for growth and morphology) could be partially replaced by isobutylmethylxanthine or dibutyryl cAMP. The defined medium was used to isolate rabbit proximal tubule kidney epithelial cells free of fibroblasts.

  4. Ciliary neurotrophic factor promotes the activation of corneal epithelial stem/progenitor cells and accelerates corneal epithelial wound healing.

    PubMed

    Zhou, Qingjun; Chen, Peng; Di, Guohu; Zhang, Yangyang; Wang, Yao; Qi, Xia; Duan, Haoyun; Xie, Lixin

    2015-05-01

    Ciliary neurotrophic factor (CNTF), a well-known neuroprotective cytokine, has been found to play an important role in neurogenesis and functional regulations of neural stem cells. As one of the most innervated tissue, however, the role of CNTF in cornea epithelium remains unclear. This study was to explore the roles and mechanisms of CNTF in the activation of corneal epithelial stem/progenitor cells and wound healing of both normal and diabetic mouse corneal epithelium. In mice subjecting to mechanical removal of corneal epithelium, the corneal epithelial stem/progenitor cell activation and wound healing were promoted by exogenous CNTF application, while delayed by CNTF neutralizing antibody. In cultured corneal epithelial stem/progenitor cells, CNTF enhanced the colony-forming efficiency, stimulated the mitogenic proliferation, and upregulated the expression levels of corneal epithelial stem/progenitor cell-associated transcription factors. Furthermore, the promotion of CNTF on the corneal epithelial stem/progenitor cell activation and wound healing was mediated by the activation of STAT3. Moreover, in diabetic mice, the content of CNTF in corneal epithelium decreased significantly when compared with that of normal mice, and the supplement of CNTF promoted the diabetic corneal epithelial wound healing, accompanied with the advanced activation of corneal epithelial stem/progenitor cells and the regeneration of corneal nerve fibers. Thus, the capability of expanding corneal epithelial stem/progenitor cells and promoting corneal epithelial wound healing and nerve regeneration indicates the potential application of CNTF in ameliorating limbal stem cell deficiency and treating diabetic keratopathy. PMID:25546438

  5. Electrical inhibition of lens epithelial cell proliferation: an additional factor in secondary cataract?

    PubMed Central

    Wang, Entong; Reid, Brian; Lois, Noemi; Forrester, John V.; McCaig, Colin D.; Zhao, Min

    2005-01-01

    Cataract is the most common cause of blindness but is at least curable by surgery. Unfortunately, many patients gradually develop the complication of posterior capsule opacification (PCO) or secondary cataract. This arises from stimulated cell growth within the lens capsule and can greatly impair vision. It is not fully understood why residual lens epithelial cell growth occurs after surgery. We propose and show that cataract surgery might remove an important inhibitory factor for lens cell growth, namely electric fields. The lens generates a unique pattern of electric currents constantly flowing out from the equator and entering the anterior and posterior poles. We show here that cutting and removing part of the anterior capsule as in cataract surgery significantly decreases the equatorial outward electric currents. Application of electric fields in culture inhibits proliferation of human lens epithelial cells. This inhibitory effect is likely to be mediated through a cell cycle control mechanism that decreases entry of cells into S phase from G1 phase by decreasing the G1-specific cell cycle protein cyclin E and increasing the cyclin-Cdk complex inhibitor p27kip1. Capsulorrhexis in vivo, which reduced endogenous lens electric fields, significantly increased LEC growth. This, together with our previous findings that electric fields have significant effects on the direction of lens cell migration, points to a controlling mechanism for the aberrant cell growth in posterior capsule opacification. A novel approach to control growth of lens epithelial cells using electric fields combined with other controlling mechanisms may be more effective in the prevention and treatment of this common complication of cataract surgery. PMID:15764648

  6. Transcriptional regulators transforming growth factor-β1 and estrogen-related receptor-α identified as putative mediators of calf rumen epithelial tissue development and function during weaning.

    PubMed

    Connor, E E; Baldwin, R L; Walker, M P; Ellis, S E; Li, C; Kahl, S; Chung, H; Li, R W

    2014-07-01

    Molecular mechanisms regulating rumen epithelial development remain largely unknown. To identify gene networks and regulatory factors controlling rumen development, Holstein bull calves (n=18) were fed milk replacer only (MRO) until 42 d of age. Three calves each were euthanized at 14 and 42 d of age for tissue collection to represent preweaning, and the remaining calves were provided diets of either milk replacer + orchard grass hay (MH; n=6) to initiate weaning without development of rumen papillae, or milk replacer + calf starter (MG; n=6) to initiate weaning and development of rumen papillae. At 56 and 70 d of age, 3 calves from the MH and MG groups were euthanized for collection of rumen epithelium. Total RNA and protein were extracted for microarray analysis and to validate detected changes in selected protein expression, respectively. As expected, calves fed MRO had no rumen papillae and development of papillae was greater in MG versus MH calves. Differentially expressed genes between the MRO diet at d 42 (preweaning) versus the MG or MH diets at d 56 (during weaning) were identified using permutation analysis of differential expression. Expression of 345 and 519 transcripts was uniquely responsive to MG and MH feeding, respectively. Ingenuity Pathway Analysis (Qiagen, Redwood City, CA) indicated that the top-ranked biological function affected by the MG diet was the cell cycle, and TFGB1, FBOX01, and PPARA were identified as key transcriptional regulators of genes responsive to the MG diet and associated with development of rumen papillae. Increased expressions of TGFB1 mRNA and protein in response to the MG diet were confirmed by subsequent analyses. The top-ranking biological function affected by the MH diet was energy production. Receptors for IGF-1 and insulin, ESRRA, and PPARD were identified by ingenuity pathway analysis as transcriptional regulators of genes responsive to the MH diet. Further analysis of TGFB1 and ESRRA mRNA expression in rumen

  7. Enhanced growth medium and method for culturing human mammary epithelial cells

    DOEpatents

    Stampfer, Martha R.; Smith, Helene S.; Hackett, Adeline J.

    1983-01-01

    Methods are disclosed for isolating and culturing human mammary epithelial cells of both normal and malignant origin. Tissue samples are digested with a mixture including the enzymes collagenase and hyaluronidase to produce clumps of cells substantially free from stroma and other undesired cellular material. Growing the clumps of cells in mass culture in an enriched medium containing particular growth factors allows for active cell proliferation and subculture. Clonal culture having plating efficiencies of up to 40% or greater may be obtained using individual cells derived from the mass culture by plating the cells on appropriate substrates in the enriched media. The clonal growth of cells so obtained is suitable for a quantitative assessment of the cytotoxicity of particular treatment. An exemplary assay for assessing the cytotoxicity of the drug adriamycin is presented.

  8. Mice overexpressing CD97 in intestinal epithelial cells provide a unique model for mammalian postnatal intestinal cylindrical growth.

    PubMed

    Aust, Gabriela; Kerner, Christiane; Gonsior, Susann; Sittig, Doreen; Schneider, Hartmut; Buske, Peter; Scholz, Markus; Dietrich, Norman; Oldenburg, Sindy; Karpus, Olga N; Galle, Jörg; Amasheh, Salah; Hamann, Jörg

    2013-07-01

    Postnatal enlargement of the mammalian intestine comprises cylindrical and luminal growth, associated with crypt fission and crypt/villus hyperplasia, respectively, which subsequently predominate before and after weaning. The bipartite adhesion G protein-coupled receptor CD97 shows an expression gradient along the crypt-villus axis in the normal human intestine. We here report that transgenic mice overexpressing CD97 in intestinal epithelial cells develop an upper megaintestine. Intestinal enlargement involves an increase in length and diameter but does not affect microscopic morphology, as typical for cylindrical growth. The megaintestine is acquired after birth and before weaning, independent of the genotype of the mother, excluding altered availability of milk constituents as driving factor. CD97 overexpression does not regulate intestinal growth factors, stem cell markers, and Wnt signaling, which contribute to epithelial differentiation and renewal, nor does it affect suckling-to-weaning transition. Consistent with augmented cylindrical growth, suckling but not adult transgenic mice show enlarged crypts and thus more crypt fissions caused by a transient increase of the crypt transit-amplifying zone. Intestinal enlargement by CD97 requires its seven-span transmembrane/cytoplasmic C-terminal fragment but not the N-terminal fragment binding partner CD55. In summary, ectopic expression of CD97 in intestinal epithelial cells provides a unique model for intestinal cylindrical growth occurring in breast-fed infants. PMID:23676664

  9. Growth Control in Colon Epithelial Cells: Gadolinium Enhances Calcium-Mediated Growth Regulation

    PubMed Central

    Attili, Durga; Jenkins, Brian; Aslam, Muhammad Nadeem; Dame, Michael K.

    2013-01-01

    Gadolinium, a member of the lanthanoid family of transition metals, interacts with calcium-binding sites on proteins and other biological molecules. The overall goal of the present investigation was to determine if gadolinium could enhance calcium-induced epithelial cell growth inhibition in the colon. Gadolinium at concentrations as low as 1–5 µM combined with calcium inhibits proliferation of human colonic epithelial cells more effectively than calcium alone. Gadolinium had no detectable effect on calcium-induced differentiation in the same cells based on change in cell morphology, induction of E-cadherin synthesis, and translocation of E-cadherin from the cytosol to the cell surface. When the colon epithelial cells were treated with gadolinium and then exposed to increased calcium concentrations, movement of extracellular calcium into the cell was suppressed. In contrast, gadolinium treatment had no effect on ionomycin-induced release of stored intracellular calcium into the cytoplasm. Whether these in vitro observations can be translated into an approach for reducing abnormal proliferation in the colonic mucosa (including polyp formation) is not known. These results do, however, provide an explanation for our recent findings that a multi-mineral supplement containing all of the naturally occurring lanthanoid metals including gadolinium are more effective than calcium alone in preventing colon polyp formation in mice on a high-fat diet. PMID:23008064

  10. The suppression of fibroblast growth factor 2/fibroblast growth factor 4-dependent tumour angiogenesis and growth by the anti-growth factor activity of dextran derivative (CMDB7).

    PubMed Central

    Bagheri-Yarmand, R.; Kourbali, Y.; Mabilat, C.; Morère, J. F.; Martin, A.; Lu, H.; Soria, C.; Jozefonvicz, J.; Crépin, M.

    1998-01-01

    Our previous studies showed that carboxymethyl benzylamide dextran (CMDB7) blocks basic fibroblast growth factor (FGF-2)-dependent cell proliferation of a human breast epithelial line (HBL100), suggesting its potential role as a potent antiangiogenic substance. The derived cell line (HH9), which was transformed with the hst/FGF4 gene, has been shown to be highly proliferative in vitro and to induce angiogenic tumours in nude mice. We show here that CMDB7 inhibits the mitogenic activities of the conditioned media from HBL 100 and HH9 cells in a dose-dependent manner. When HH9 cells were injected s.c. into nude mice, CMDB7 treatment (300 mg kg(-1) week(-1)) suppressed the tumour take and the tumour growth by about 50% and 80% respectively. Immunohistochemical analysis showed a highly significant decrease, by more than threefold, in the endothelial density of viable tumour regions, together with a significant increase in the necrosis area. This antiangiogenic activity of CMDB7 was further demonstrated by direct inhibition of calf pulmonary artery (CPAE) and human umbilical vein (HUVEC) endothelial cell proliferation and migration in vitro. In addition, we showed that CMDB7 inhibits specifically the mitogenic effects of the growth factors that bind to heparin such as FGF-2, FGF-4, platelet-derived growth factor (PDGF-BB) and transforming growth factor (TGF-beta1), but not those of epidermal growth factor (EGF) and insulin-like growth factor (IGF-1). These results demonstrate that CMDB7 inhibits FGF-2/FGF-4-dependent tumour growth and angiogenesis, most likely by disrupting the autocrine and paracrine effects of growth factors released from the tumour cells. Images Figure 4 PMID:9662260

  11. Polarity, cell division, and out-of-equilibrium dynamics control the growth of epithelial structures

    PubMed Central

    Cerruti, Benedetta; Puliafito, Alberto; Shewan, Annette M.; Yu, Wei; Combes, Alexander N.; Little, Melissa H.; Chianale, Federica; Primo, Luca; Serini, Guido; Mostov, Keith E.; Celani, Antonio

    2013-01-01

    The growth of a well-formed epithelial structure is governed by mechanical constraints, cellular apico-basal polarity, and spatially controlled cell division. Here we compared the predictions of a mathematical model of epithelial growth with the morphological analysis of 3D epithelial structures. In both in vitro cyst models and in developing epithelial structures in vivo, epithelial growth could take place close to or far from mechanical equilibrium, and was determined by the hierarchy of time-scales of cell division, cell–cell rearrangements, and lumen dynamics. Equilibrium properties could be inferred by the analysis of cell–cell contact topologies, and the nonequilibrium phenotype was altered by inhibiting ROCK activity. The occurrence of an aberrant multilumen phenotype was linked to fast nonequilibrium growth, even when geometric control of cell division was correctly enforced. We predicted and verified experimentally that slowing down cell division partially rescued a multilumen phenotype induced by altered polarity. These results improve our understanding of the development of epithelial organs and, ultimately, of carcinogenesis. PMID:24145168

  12. Risk factors for epithelial ovarian cancer in Beijing, China.

    PubMed

    Chen, Y; Wu, P C; Lang, J H; Ge, W J; Hartge, P; Brinton, L A

    1992-02-01

    A study in Beijing, China of 112 pathologically confirmed epithelial ovarian cancer cases and 224 age-matched community controls enabled evaluation of risk in relation to reproductive, medical, familial, and selected lifestyle factors. An inverse relationship was observed between the number of full-term pregnancies and ovarian cancer risk. Compared to nulliparous women, subjects with one, two, or three full-term pregnancies were at 50%, 70%, or 90% reduced risks, respectively (P for trend less than 0.01). A positive correlation was found between the number of ovulatory years and risk, with a 2.6-fold increased risk for women with 30 or more compared to less than 10 ovulatory years (P for trend less than 0.01). Infertility, as estimated in various ways, was also found to be an important risk factor. When parity was taken into account, age at first pregnancy was not related to ovarian cancer risk. No protective effect was associated with mumps virus infection. In contrast, risk increased significantly as serum mumps virus antibody titres increased (P for trend less than 0.01). An elevated risk was found in women with a history of long-term (greater than 3 months) application of talc-containing dusting powder to the lower abdomen and perineum (Relative risk 3.9, 95% confidence interval: 0.9-10.63). These findings suggest that Chinese women have risk factors similar to those of occidental women. PMID:1544753

  13. [Prognostic and predictive factors in epithelial ovarian cancer].

    PubMed

    Boudou-Rouquette, P; Pautier, P; Morice, P; Lhommé, C

    2009-04-01

    Even if prognosis of epithelial ovarian cancer remains very bad, survival and response to treatment are variable according to the patients. Determination of new prognostic markers helps us to adapt therapeutics for each patient and is necessary for the elaboration and the interpretation of clinical research studies. Many prognostic factors related to the tumor, the patient or the treatment, have been evaluated. The goal of this work is to review these parameters. So far, the most powerful variables are volume of residual disease after cytoreductive surgery, FIGO tumor stage, histologic type and grade of differentiation. The progress and accessibility to novel technologies applied to biology will make possible in the future the assessment of new prognostic profiles-based on genetic and/or proteomic tumor characteristics. The future also relies on the identification of predictive factors of response to treatment, but force is to note that on the last hundred publications testing predictive factors (p53, HER2, Topo-2-alpha, BRCA...), none have modified today our clinical practices. PMID:19357017

  14. Wnt-10b, uniquely among Wnts, promotes epithelial differentiation and shaft growth

    SciTech Connect

    Ouji, Yukiteru Yoshikawa, Masahide; Moriya, Kei; Nishiofuku, Mariko; Matsuda, Ryosuke; Ishizaka, Shigeaki

    2008-03-07

    Although Wnts are expressed in hair follicles throughout life from embryo to adult, and considered to be critical for their development and maturation, their roles remain largely unknown. In the present study, we investigated the effects of Wnts (Wnt-3a, Wnt-5a, Wnt-10b, and Wnt-11) on epithelial cell differentiation using adult mouse-derived primary skin epithelial cell (MPSEC) cultures and hair growth using hair follicle organ cultures. Only Wnt-10b showed evident promotion of epithelial cell differentiation and hair shaft growth, in contrast to Wnt-3a, 5a, and 11. Our results suggest that Wnt-10b is unique and plays an important role in differentiation of epithelial cells in the hair follicle.

  15. Interstitial fibrosis and growth factors.

    PubMed Central

    Lasky, J A; Brody, A R

    2000-01-01

    Interstitial pulmonary fibrosis (IPF) is scarring of the lung caused by a variety of inhaled agents including mineral particles, organic dusts, and oxidant gases. The disease afflicts millions of individuals worldwide, and there are no effective therapeutic approaches. A major reason for this lack of useful treatments is that few of the molecular mechanisms of disease have been defined sufficiently to design appropriate targets for therapy. Our laboratory has focused on the molecular mechanisms through which three selected peptide growth factors could play a role in the development of IPF. Hundreds of growth factors and cytokines could be involved in the complex disease process. We are studying platelet-derived growth factor because it is the most potent mesenchymal cell mitogen yet described, transforming growth factor beta because it is a powerful inducer of extracellular matrix (scar tissue) components by mesenchymal cells, and tumor necrosis factor alpha because it is a pleiotropic cytokine that we and others have shown is essential for the development of IPF in animal models. This review describes some of the evidence from studies in humans, in animal models, and in vitro, that supports the growth factor hypothesis. The use of modern molecular and transgenic technologies could elucidate those targets that will allow effective therapeutic approaches. Images Figure 1 Figure 2 PMID:10931794

  16. Growth factors in ischemic stroke

    PubMed Central

    Lanfranconi, S; Locatelli, F; Corti, S; Candelise, L; Comi, G P; Baron, P L; Strazzer, S; Bresolin, N; Bersano, A

    2011-01-01

    Abstract Data from pre-clinical and clinical studies provide evidence that colony-stimulating factors (CSFs) and other growth factors (GFs) can improve stroke outcome by reducing stroke damage through their anti-apoptotic and anti-inflammatory effects, and by promoting angiogenesis and neurogenesis. This review provides a critical and up-to-date literature review on CSF use in stroke. We searched for experimental and clinical studies on haemopoietic GFs such as granulocyte CSF, erythropoietin, granulocyte-macrophage colony-stimulating factor, stem cell factor (SCF), vascular endothelial GF, stromal cell-derived factor-1α and SCF in ischemic stroke. We also considered studies on insulin-like growth factor-1 and neurotrophins. Despite promising results from animal models, the lack of data in human beings hampers efficacy assessments of GFs on stroke outcome. We provide a comprehensive and critical view of the present knowledge about GFs and stroke, and an overview of ongoing and future prospects. PMID:20015202

  17. Growth and differentiation of human lens epithelial cells in vitro on matrix

    NASA Technical Reports Server (NTRS)

    Blakely, E. A.; Bjornstad, K. A.; Chang, P. Y.; McNamara, M. P.; Chang, E.; Aragon, G.; Lin, S. P.; Lui, G.; Polansky, J. R.

    2000-01-01

    PURPOSE: To characterize the growth and maturation of nonimmortalized human lens epithelial (HLE) cells grown in vitro. METHODS: HLE cells, established from 18-week prenatal lenses, were maintained on bovine corneal endothelial (BCE) extracellular matrix (ECM) in medium supplemented with basic fibroblast growth factor (FGF-2). The identity, growth, and differentiation of the cultures were characterized by karyotyping, cell morphology, and growth kinetics studies, reverse transcription-polymerase chain reaction (RT-PCR), immunofluorescence, and Western blot analysis. RESULTS: HLE cells had a male, human diploid (2N = 46) karyotype. The population-doubling time of exponentially growing cells was 24 hours. After 15 days in culture, cell morphology changed, and lentoid formation was evident. Reverse transcription-polymerase chain reaction (RT-PCR) indicated expression of alphaA- and betaB2-crystallin, fibroblast growth factor receptor 1 (FGFR1), and major intrinsic protein (MIP26) in exponential growth. Western analyses of protein extracts show positive expression of three immunologically distinct classes of crystallin proteins (alphaA-, alphaB-, and betaB2-crystallin) with time in culture. By Western blot analysis, expression of p57(KIP2), a known marker of terminally differentiated fiber cells, was detectable in exponential cultures, and levels increased after confluence. MIP26 and gamma-crystallin protein expression was detected in confluent cultures, by using immunofluorescence, but not in exponentially growing cells. CONCLUSIONS: HLE cells can be maintained for up to 4 months on ECM derived from BCE cells in medium containing FGF-2. With time in culture, the cells demonstrate morphologic characteristics of, and express protein markers for, lens fiber cell differentiation. This in vitro model will be useful for investigations of radiation-induced cataractogenesis and other studies of lens toxicity.

  18. Ivermectin Inhibits Growth of Chlamydia trachomatis in Epithelial Cells

    PubMed Central

    Pettengill, Matthew A.; Lam, Verissa W.; Ollawa, Ikechukwu; Marques-da-Silva, Camila; Ojcius, David M.

    2012-01-01

    Ivermectin is currently approved for treatment of both clinical and veterinary infections by nematodes, including Onchocerca cervicalis in horses and Onchocerca volvulus in humans. However, ivermectin has never been shown to be effective against bacterial pathogens. Here we show that ivermectin also inhibits infection of epithelial cells by the bacterial pathogen, Chlamydia trachomatis, at doses that could be envisioned clinically for sexually-transmitted or ocular infections by Chlamydia. PMID:23119027

  19. Ivermectin inhibits growth of Chlamydia trachomatis in epithelial cells.

    PubMed

    Pettengill, Matthew A; Lam, Verissa W; Ollawa, Ikechukwu; Marques-da-Silva, Camila; Ojcius, David M

    2012-01-01

    Ivermectin is currently approved for treatment of both clinical and veterinary infections by nematodes, including Onchocerca cervicalis in horses and Onchocerca volvulus in humans. However, ivermectin has never been shown to be effective against bacterial pathogens. Here we show that ivermectin also inhibits infection of epithelial cells by the bacterial pathogen, Chlamydia trachomatis, at doses that could be envisioned clinically for sexually-transmitted or ocular infections by Chlamydia. PMID:23119027

  20. Keratinocyte growth factor and hepatocyte growth factor/scatter factor are heparin-binding growth factors for alveolar type II cells in fibroblast-conditioned medium.

    PubMed Central

    Panos, R J; Rubin, J S; Csaky, K G; Aaronson, S A; Mason, R J

    1993-01-01

    Epithelial-mesenchymal interactions mediate aspects of normal lung growth and development and are important in the restoration of normal alveolar architecture after lung injury. To determine if fibroblasts are a source of soluble growth factors for alveolar type II cells, we investigated the effect of fibroblast-conditioned medium (CM) on alveolar type II cell DNA synthesis. Serum-free CM from confluent adult human lung fibroblasts was concentrated fivefold by lyophilization. Type II cells were isolated from adult rats by elastase dissociation and incubated with [3H]thymidine and varying dilutions of concentrated CM and serum from day 1 to 3 of culture. Stimulation of type II cell DNA synthesis by fibroblast-CM was maximal after 48 h of conditioning and required the presence of serum. The activity of the CM was eliminated by boiling and by treatment with trypsin, pepsin, or dithiothreitol and was additive with saturating concentrations of acidic fibroblast growth factor, epidermal growth factor, and insulin. The growth factor activity bound to heparin-Sepharose and was eluted with 0.6 and 1.0 M NaCl. Neutralizing antibody studies demonstrated that the primary mitogens isolated in the 0.6 and 1.0 M NaCl fractions were keratinocyte growth factor (KGF, fibroblast growth factor 7) and hepatocyte growth factor/scatter factor (HGF/SF), respectively. HGF/SF was demonstrated in the crude CM and KGF was detected in the 0.6 M NaCl eluent by immunoblotting. Northern blot analysis confirmed that the lung fibroblasts expressed both KGF and HGF/SF transcripts. Human recombinant KGF and HGF/SF induced a concentration- and serum-dependent increase in rat alveolar type II cell DNA synthesis. We conclude that adult human lung fibroblasts produce at least two soluble heparin-binding growth factors, KGF and HGF/SF, which promote DNA synthesis and proliferation of rat alveolar type II cells in primary culture. KGF and HGF/SF may be important stimuli for alveolar type II cell

  1. Growth factors in porcine full and partial thickness burn repair. Differing targets and effects of keratinocyte growth factor, platelet-derived growth factor-BB, epidermal growth factor, and neu differentiation factor.

    PubMed Central

    Danilenko, D. M.; Ring, B. D.; Tarpley, J. E.; Morris, B.; Van, G. Y.; Morawiecki, A.; Callahan, W.; Goldenberg, M.; Hershenson, S.; Pierce, G. F.

    1995-01-01

    The topical application of recombinant growth factors such as epidermal growth factor, platelet-derived growth factor-BB homodimer (rPDGF-BB), keratinocyte growth factor (rKGF), and neu differentiation factor has resulted in significant acceleration of healing in several animal models of wound repair. In this study, we established highly reproducible and quantifiable full and deep partial thickness porcine burn models in which burns were escharectomized 4 or 5 days postburn and covered with an occlusive dressing to replicate the standard treatment in human burn patients. We then applied these growth factors to assess their efficacy on several parameters of wound repair: extracellular matrix and granulation tissue production, percent reepithelialization, and new epithelial area. In full thickness burns, only rPDGF-BB and the combination of rPDGF-BB and rKGF induced significant changes in burn repair. rPDGF-BB induced marked extracellular matrix and granulation tissue production (P = 0.013) such that the burn defect was filled within several days of escharectomy, but had no effect on new epithelial area or reepithelialization. The combination of rPDGF-BB and rKGF in full thickness burns resulted in a highly significant increase in extracellular matrix and granulation tissue area (P = 0.0009) and a significant increase in new epithelial area (P = 0.007), but had no effect on reepithelialization. In deep partial thickness burns, rKGF induced the most consistent changes. Daily application of rKGF induced a highly significant increase in new epithelial area (P < 0.0001) but induced only a modest increase in reepithelialization (83.7% rKGF-treated versus 70.2% control; P = 0.016) 12 days postburn. rKGF also doubled the number of fully reepithelialized burns (P = 0.02) at 13 days postburn, at least partially because of marked stimulation of both epidermal and follicular proliferation as assessed by proliferating cell nuclear antigen expression. In situ hybridization for

  2. Control of growth and squamous differentiation in normal human bronchial epithelial cells by chemical and biological modifiers and transferred genes

    SciTech Connect

    Pfeifer, A.M.; Lechner, J.F.; Masui, T.; Reddel, R.R.; Mark, G.E.; Harris, C.C.

    1989-03-01

    The majority of human lung cancers arise from bronchial epithelial cells. The normal pseudostratified bronchial epithelium is composed of basal, mucous, and ciliated cells. This multi-differentiated epithelium usually responds to xenobiotics and physical injury by undergoing basal cell hyperplasia, mucous cell hyperplasia, and squamous metaplasia. One step of the multistage process of carcinogenesis is thought to involve aberrations in control of the squamous metaplastic processes. Decreased responsiveness to regulators of terminal squamous differentiation may confer a selective clonal expansion advantage to an initiated cell. We studied the effects of endogenous (e.g., transforming growth factor beta 1 (TGF-beta 1) and serum) and exogenous (e.g., 12-O-tetradecanoyl-13-phorbol-acetate (TPA), tobacco smoke condensate, and aldehydes) modifiers of normal human bronchial epithelial (NHBE) cell in a serum-free culture system. NHBE cells are growth inhibited by all of these compounds and induced to undergo squamous differentiation by TGF-beta 1 or TPA. In contrast, lung carcinoma cell lines are relatively resistant to inducers of terminal squamous differentiation which may provide them with a selective growth advantage. Chemical agents and activated protooncogenes (ras,raf,myc) altered the response to endogenous and exogenous inducers of squamous differentiation and caused extended cellular lifespan, aneuploidy, and/or tumorigenicity. The data suggest a close relationship between dysregulation of terminal differentiation pathways and neoplastic transformation of human bronchial epithelial cells.

  3. Long-term survival of a patient with lung cancer metastasis to the spine following surgical treatment combined with radiation and epithelial growth factor receptor inhibitor therapy: A case report

    PubMed Central

    XU, SONGFENG; YU, XIUCHUN; XU, MING

    2015-01-01

    The prognosis of patients with lung cancer metastasis to the spine is poor, and the choice of surgery is questionable based on the aggressiveness of the disease. The present study describes a case of a 56-year-old male with metastatic spinal cord compression. The patient underwent surgery for posterior decompression and internal fixation, in addition to receiving postoperative radiation and epidermal growth factor receptor (EGFR) inhibitor medication. After 24 months, positron emission tomography-computed tomography scan showed a reduction in the left upper lobe mass in the short axis and inactivation of the neoplasm in the left upper lobe and T9 vertebra. Based on these promising results, it is suggested that orthopedic oncologists consider the combination of radiation and EGFR inhibitor therapy with surgery for the treatment of lung cancer metastasis to the spine. PMID:25452785

  4. IglC and PdpA Are Important for Promoting Francisella Invasion and Intracellular Growth in Epithelial Cells

    PubMed Central

    Law, H. T.; Nix, Eli B.; Nano, Francis E.; Guttman, Julian Andrew

    2014-01-01

    The highly infectious bacteria, Francisella tularensis, colonize a variety of organs and replicate within both phagocytic as well as non-phagocytic cells, to cause the disease tularemia. These microbes contain a conserved cluster of important virulence genes referred to as the Francisella Pathogenicity Island (FPI). Two of the most characterized FPI genes, iglC and pdpA, play a central role in bacterial survival and proliferation within phagocytes, but do not influence bacterial internalization. Yet, their involvement in non-phagocytic epithelial cell infections remains unexplored. To examine the functions of IglC and PdpA on bacterial invasion and replication during epithelial cell infections, we infected liver and lung epithelial cells with F. novicida and F. tularensis ‘Type B’ Live Vaccine Strain (LVS) deletion mutants (ΔiglC and ΔpdpA) as well as their respective gene complements. We found that deletion of either gene significantly reduced their ability to invade and replicate in epithelial cells. Gene complementation of iglC and pdpA partially rescued bacterial invasion and intracellular growth. Additionally, substantial LAMP1-association with both deletion mutants was observed up to 12 h suggesting that the absence of IglC and PdpA caused deficiencies in their ability to dissociate from LAMP1-positive Francisella Containing Vacuoles (FCVs). This work provides the first evidence that IglC and PdpA are important pathogenic factors for invasion and intracellular growth of Francisella in epithelial cells, and further highlights the discrete mechanisms involved in Francisella infections between phagocytic and non-phagocytic cells. PMID:25115488

  5. Walking along the Fibroblast Growth Factor 10 Route: A Key Pathway to Understand the Control and Regulation of Epithelial and Mesenchymal Cell-Lineage Formation during Lung Development and Repair after Injury

    PubMed Central

    Bellusci, Saverio

    2014-01-01

    Basic research on embryonic lung development offers unique opportunities to make important discoveries that will impact human health. Developmental biologists interested in the molecular control of branching morphogenesis have intensively studied the developing lung, with its complex and seemingly stereotyped ramified structure. However, it is also an organ that is linked to a vast array of clinical problems in humans such as bronchopulmonary dysplasia in premature babies and emphysema, chronic obstructive pulmonary disease, fibrosis, and cancer in adults. Epithelial stem/progenitor cells reside in niches where they interact with specific extracellular matrices as well as with mesenchymal cells; the latter are still poorly characterized. Interactions of epithelial stem/progenitor cells with their microenvironments are usually instructive, controlling quiescence versus activation, proliferation, differentiation, and migration. During the past 18 years, Fgf10 has emerged not only as a marker for the distal lung mesenchyme during early lung development, but also as a key player in branching morphogenesis and a critical component of the niche for epithelial stem cells. In this paper, we will present the current knowledge regarding the lineage tree in the lung, with special emphasis on cell-lineage decisions in the lung mesenchyme and the role of Fgf10 in this context. PMID:25298902

  6. Walking along the Fibroblast Growth Factor 10 Route: A Key Pathway to Understand the Control and Regulation of Epithelial and Mesenchymal Cell-Lineage Formation during Lung Development and Repair after Injury.

    PubMed

    El Agha, Elie; Bellusci, Saverio

    2014-01-01

    Basic research on embryonic lung development offers unique opportunities to make important discoveries that will impact human health. Developmental biologists interested in the molecular control of branching morphogenesis have intensively studied the developing lung, with its complex and seemingly stereotyped ramified structure. However, it is also an organ that is linked to a vast array of clinical problems in humans such as bronchopulmonary dysplasia in premature babies and emphysema, chronic obstructive pulmonary disease, fibrosis, and cancer in adults. Epithelial stem/progenitor cells reside in niches where they interact with specific extracellular matrices as well as with mesenchymal cells; the latter are still poorly characterized. Interactions of epithelial stem/progenitor cells with their microenvironments are usually instructive, controlling quiescence versus activation, proliferation, differentiation, and migration. During the past 18 years, Fgf10 has emerged not only as a marker for the distal lung mesenchyme during early lung development, but also as a key player in branching morphogenesis and a critical component of the niche for epithelial stem cells. In this paper, we will present the current knowledge regarding the lineage tree in the lung, with special emphasis on cell-lineage decisions in the lung mesenchyme and the role of Fgf10 in this context. PMID:25298902

  7. Epidermal growth factor signaling in transformed cells

    PubMed Central

    Lindsey, Stephan; Langhans, Sigrid A.

    2016-01-01

    Members of the epidermal growth factor receptor (EGFR/ErbB) family play a critical role in normal cell growth and development. However, many ErbB family members, especially EGFR, are aberrantly expressed or deregulated in tumors and are thought to play crucial roles in cancer development and metastatic progression. In this chapter, we provide an overview of key mechanisms contributing to aberrant EGFR/ErbB signaling in transformed cells which results in many phenotypic changes associated with the earliest stages of tumor formation, including several hallmarks of epithelial-to-mesenchymal transition (EMT). These changes often occur through interaction with other major signaling pathways important to tumor progression resulting in a multitude of transcriptional changes that ultimately impact cell morphology, proliferation and adhesion, all of which are crucial for tumor progression. The resulting mesh of signaling networks will need to be taken into account as new regimens are designed for targeting EGFR for therapeutic intervention. As new insights into the molecular mechanisms of the cross-talk of EGFR signaling with other signaling pathways and their role in therapeutic resistance to anti-EGFR therapies are gained a continual reassessment of clinical therapeutic regimes and strategies will be required. Understanding the consequences and complexity of EGF signaling and how it relates to tumor progression is critical for the development of clinical compounds and establishing clinical protocols for the treatment of cancer. PMID:25619714

  8. Interdependent epidermal growth factor receptor signalling and trafficking.

    PubMed

    Jones, Sylwia; Rappoport, Joshua Z

    2014-06-01

    Epidermal growth factor (EGF) receptor (EGFR) signalling regulates diverse cellular functions, promoting cell proliferation, differentiation, migration, cell growth and survival. EGFR signalling is critical during embryogenesis, in particular in epithelial development, and disruption of the EGFR gene results in epithelial immaturity and perinatal death. EGFR signalling also functions during wound healing responses through accelerating wound re-epithelialisation, inducing cell migration, proliferation and angiogenesis. Upregulation of EGFR signalling is often observed in carcinomas and has been shown to promote uncontrolled cell proliferation and metastasis. Therefore aberrant EGFR signalling is a common target for anticancer therapies. Various reports indicate that EGFR signalling primarily occurs at the plasma membrane and EGFR degradation following endocytosis greatly attenuates signalling. Other studies argue that EGFR internalisation is essential for complete activation of downstream signalling cascades and that endosomes can serve as signalling platforms. The aim of this review is to discuss current understanding of intersection between EGFR signalling and trafficking. PMID:24681003

  9. Growth factors in orthopedic surgery

    PubMed Central

    Zaharia, C; Despa, N; Simionescu, M; Jinga, V; Fleseriu, I

    2010-01-01

    Growth factors have represented an essential issue of interest for the researchers and clinicians in orthopedics and trauma over the last 40 years. In the last 10 to 15 years, the advances registered in this field have permitted the identification of the most active cellular and humoral factors as well as the improvement of their use in the orthopedic and trauma surgery. Their domain of application has been continuously enlarged and the results have been visible from the beginning. The authors present their appreciation on the actual state of this subject as well as their experience with results and related conclusions. PMID:20302195

  10. Surface expression of CD74 by type II alveolar epithelial cells: a potential mechanism for macrophage migration inhibitory factor-induced epithelial repair.

    PubMed

    Marsh, Leigh M; Cakarova, Lidija; Kwapiszewska, Grazyna; von Wulffen, Werner; Herold, Susanne; Seeger, Werner; Lohmeyer, Juergen

    2009-03-01

    Macrophage migration inhibitory factor (MIF) is a pleiotropic proinflammatory cytokine involved in acute lung injury and other processes such as wound repair and tumor growth. MIF exerts pro-proliferative effects on a variety of cell types including monocytes/macrophages, B cells, and gastric epithelial cell lines through binding to the major histocompatibility complex type II-associated invariant chain, CD74. In acute lung injury, inflammatory damage of the alveolar epithelium leads to loss of type I alveolar epithelial cells (AEC-I), which are replaced by proliferation and differentiation of type II alveolar epithelial cells (AEC-II). In this study we have investigated the potential of MIF to contribute to alveolar repair by stimulating alveolar epithelial cell proliferation. We show that murine AEC-II, but not AEC-I, express high surface levels of CD74 in vivo. Culture of AEC-II in vitro resulted in decreased mRNA levels for CD74 and loss of surface CD74 expression, which correlated with a transition of AEC-II to an AEC-I-like phenotype. MIF stimulation of AEC-II induced rapid and prolonged phosphorylation of ERK1/2 and Akt, increased expression of cyclins D1 and E, as well as AEC-II proliferation. Corresponding MIF signaling and enhanced thymidine incorporation was observed after MIF stimulation of MLE-12 cells transfected to overexpress CD74. In contrast, MIF did not induce MAPK activation, gene transcription, or increased proliferation in differentiated AEC-I-like cells that lack CD74. These data suggest a previously unidentified role of MIF-CD74 interaction by inducing proliferation of AEC-II, which may contribute to alveolar repair. PMID:19136583

  11. Epithelial cells prime the immune response to an array of gut-derived commensals towards a tolerogenic phenotype through distinct actions of thymic stromal lymphopoietin and transforming growth factor

    PubMed Central

    Zeuthen, Louise Hjerrild; Fink, Lisbeth Nielsen; Frokiaer, Hanne

    2008-01-01

    Humans and other mammals coexist with a diverse array of microbes colonizing the intestine, termed the microflora. The relationship is symbiotic, with the microbes benefiting from a stable environment and nutrient supply, and the host gaining competitive exclusion of pathogens and continuously maintenance of the gut immune homeostasis. Here we report novel crosstalk mechanisms between the human enterocyte cell line, Caco2, and underlying human monocyte-derived DC in a transwell model where Gram-positive (G+) commensals prevent Toll-like receptor-4 (TLR4)-dependent Escherichia coli-induced semimaturation in a TLR2-dependent fashion. These findings add to our understanding of the hypo-responsiveness of the gut epithelium towards the microflora. Gut DC posses a more tolerogenic phenotype than conventional DC. Here we show that Caco2 spent medium (SM) induces tolerogenic DC with lower expression of maturation markers, interleukin (IL)-12p70, and tumour necrosis factor-α when matured with G+ and Gram-negative (G–) commensals, while IL-10 production is enhanced in DC upon encountering G+ commensals and reduced upon encountering G– bacteria. The Caco2 SM-induced tolerogenic phenotype is also seen in DC priming of naive T cells with elevated levels of transforming growth factor-β (TGF-β) and markedly reduced levels of bacteria-induced interferon-γ production. Caco2 cell production of IL-8, thymic stromal lymphopoietin (TSLP) and TGF-β increases upon microbial stimulation in a strain dependent manner. TSLP and TGF-β co-operate in inducing the tolerogenic DC phenotype but other mediators might be involved. PMID:17655740

  12. Both epidermal growth factor and insulin-like growth factor receptors are dispensable for structural intestinal adaptation

    PubMed Central

    Sun, Raphael C.; Diaz-Miron, Jose L.; Choi, Pamela M.; Sommovilla, Joshua; Guo, Jun; Erwin, Christopher R.; Warner, Brad W.

    2015-01-01

    Purpose Intestinal adaptation structurally represents increases in crypt depth and villus height in response to small bowel resection (SBR). Previously, we found that neither epidermal growth factor receptor (EGFR) nor insulin-like growth factor 1 receptor (IGF1R) function was individually required for normal adaptation. In this study, we sought to determine the effect of disrupting both EGFR and IGF1R expression on resection-induced adaptation. Methods Intestinal-specific EGFR and IGF1R double knockout mice (EGFR/IGF1R-IKO) (n=6) and wild-type (WT) control mice (n=7) underwent 50% proximal SBR. On postoperative day (POD) 7, structural adaptation was scored by measuring crypt depth and villus height. Rates of crypt cell proliferation, apoptosis, and submucosal capillary density were also compared. Results After 50% SBR, normal adaptation occurred in both WT and EGFR/IGF1R-IKO. Rates of proliferation and apoptosis were no different between the two groups. The angiogenic response was less in the EGFR/IGF1R-IKO compared to WT mice. Conclusion Disrupted expression of EGFR and IGF1R in the intestinal epithelial cells does not affect resection-induced structural adaptation but attenuates angiogenesis after SBR. These findings suggest that villus growth is driven by receptors and pathways that occur outside the epithelial cell component, while angiogenic responses may be influenced by epithelial-endothelial crosstalk. PMID:25818318

  13. Transforming growth factor-beta 1 and fibroblast growth factors in rat growth plate.

    PubMed

    Jingushi, S; Scully, S P; Joyce, M E; Sugioka, Y; Bolander, M E

    1995-09-01

    Chondrocytes in the growth plate progress in an orderly fashion from resting through proliferating to hypertrophic cells. In the region of hypertrophic chondrocytes, the cartilage is invaded by capillary loops and endochondral ossification is initiated. It is currently believed that growth factors may regulate the proliferation and maturation of chondrocytes and the synthesis of extracellular matrix in the growth plate. The ordered sequence of proliferation and differentiation observed in the growth plate provides a unique opportunity to study the role of acidic fibroblast growth factor, basic fibroblast growth factor, and transforming growth factor-beta 1 in the regulation of these processes. In this study, expression of the mRNA of these growth factors was examined using total RNA extracted from the physis and epiphysis of rat tibias. Transforming growth factor-beta 1 mRNA was detected by Northern hybridization. Expression of the genes encoding acidic and basic fibroblast growth factors was demonstrated by polymerase chain reaction amplification. In addition, using polyclonal antibodies against these growth factors, we localized them by immunohistochemical analysis. Strong intracellular staining with a predominantly nuclear pattern was observed in chondrocytes from the proliferating and upper hypertrophic zones. In contrast, chondrocytes in the resting zone stained only faintly for the presence of these growth factors. Some chondrocytes in the resting zone adjacent to the proliferating zone stained with these antibodies, and the antibodies also stained cells in the zone of Ranvier, which regulates latitudinal bone growth. Lastly, the location of transforming growth factor-beta 1 was examined further with use of a polyclonal antipeptide antibody specific for its extracellular epitope.(ABSTRACT TRUNCATED AT 250 WORDS) PMID:7472755

  14. Epidermal growth factor, from gene organization to bedside

    PubMed Central

    Zeng, Fenghua; Harris, Raymond C.

    2014-01-01

    In 1962, epidermal growth factor (EGF) was discovered by Dr. Stanley Cohen while studying nerve growth factor (NGF). It was soon recognized that EGF is the prototypical member of a family of peptide growth factors that activate the EGF receptors, and that the EGF/EGF receptor signaling pathway plays important roles in proliferation, differentiation and migration of a variety of cell types, especially in epithelial cells. After the basic characterization of EGF function in the first decade or so after its discovery, the studies related to EGF and its signaling pathway have extended to a broad range of investigations concerning its biological and pathophysiological roles in development and in human diseases. In this review, we briefly describe the gene organization and tissue distribution of EGF, with emphasis on its biological and pathological roles in human diseases. PMID:24513230

  15. CD40-induced growth inhibition in epithelial cells is mimicked by Epstein-Barr Virus-encoded LMP1: involvement of TRAF3 as a common mediator.

    PubMed

    Eliopoulos, A G; Dawson, C W; Mosialos, G; Floettmann, J E; Rowe, M; Armitage, R J; Dawson, J; Zapata, J M; Kerr, D J; Wakelam, M J; Reed, J C; Kieff, E; Young, L S

    1996-11-21

    CD40, a member of the tumour necrosis factor receptor family, is expressed on the surface of B lymphocytes where its ligation provides a potent survival signal. CD40 is also expressed in basal epithelial cells and in a number of different carcinomas where its function remains unknown. We observed that contrary to the studies in normal B cells, CD40 ligation in carcinoma cell lines and in normal primary epithelial cells resulted in growth inhibition and enhanced susceptibility to apoptosis induced by anti-neoplastic drugs, TNF-alpha, Fas and ceramide. This effect was also observed in CD40-transfected Rat-1 fibroblasts. The expression of Bcl-2 did not affect growth inhibition induced by CD40 ligation in epithelial cells but the Epstein - Barr Virus-encoded latent membrane protein 1 (LMP1) blocked the effect. Whilst transient expression of LMP-1 resulted in the inhibition of epithelial cell growth, this effect was not observed with a LMP1 mutant lacking the binding domain for TRAF3, a protein which may mediate signal transduction by interacting with the cytoplasmic domains of both CD40 and LMP1. Transient expression of TRAF3 also inhibited epithelial cell growth, whilst expression of a dominant-negative TRAF3 partially blocked the inhibitory effect of CD40 ligation and of transient LMP1 expression. These results suggest that CD40 regulates epithelial cell growth in a manner mimicked by LMP1 and implicate TRAF3 as a common mediator in the transduction of the growth inhibitory signals generated via the CD40 and LMP1 pathways. PMID:8950992

  16. Epidermal growth factor and growth in vivo

    SciTech Connect

    Rhodes, J.A.

    1986-01-01

    Epidermal growth factor (EGF) causes a dose-dependent thickening of the epidermis in suckling mice. The cellular mechanisms underlying this thickening were analyzed by measuring the effect of EGF on the cell-cycle. Neonatal mice were given daily injections of either 2ug EGF/g body weight/day or an equivalent volume of saline, and on the seventh day received a single injection of /sup 3/H-thymidine. At various times the mice were perfused with fixative; 1um sections of skin were stained with a modification of Harris' hematoxylin and were autoradiographed. The sections were analyzed using three methods based on the dependence on time after injection of /sup 3/H-thymidine of: frequency of labelled mitoses, labelling index, and reciprocal grains/nucleus. It was found that EGF caused a two-fold increase in the cell production rate. The effect of exogenous EGF on the morphology of gastric mucosa and incisors of suckling mice was also studied. The gastric mucosa appeared thicker in EGF-treated animals, but the effect was not statistically significant. In contrast to its effect on epidermis and gastric mucosa, EGF caused a significant, dose-dependent decrease in the size of the incisors. Because the mouse submandibular salivary gland is the major source of EGF the effect of sialoadenectomy on female reproductive functions was examined. Ablation of the submandibular gland had no effect on: length of estrus cycle, ability of the female to produce litters, length of the gestation period, litter size, and weight of the litter at birth. There was also no effect on survival of the offspring or on age at which the eyelids separated.

  17. Nerve growth factor enhances Clara cell proliferation after lung injury.

    PubMed

    Sonar, S S; Schwinge, D; Kilic, A; Yildirim, A O; Conrad, M L; Seidler, K; Müller, B; Renz, H; Nockher, W A

    2010-07-01

    The lung epithelia facilitate wound closure by secretion of various cytokines and growth factors. Nerve growth factor (NGF) has been well described in airway inflammation; however, its likely role in lung repair has not been examined thus far. To investigate the repair function of NGF, experiments were performed in vitro using cultured alveolar epithelial cells and in vivo using a naphthalene-induced model of Clara epithelial cell injury. Both in vitro and in vivo experiments revealed airway epithelial cell proliferation following injury to be dependent on NGF and the expression of its receptor, tropomyosin-receptor-kinase A. Additionally, NGF also augmented in vitro migration of alveolar type II cells. In vivo, transgenic mice over-expressing NGF in Clara cells (NGFtg) did not reveal any proliferation or alteration in Clara cell phenotype. However, following Clara cell specific injury, proliferation was increased in NGFtg and impaired upon inhibition of NGF. Furthermore, NGF also promoted the expression of collagen I and fibronectin in vitro and in vivo during repair, where significantly higher levels were measured in re-epithelialising NGFtg mice. Our study demonstrates that NGF promotes the proliferation of lung epithelium in vitro and the renewal of Clara cells following lung injury in vivo. PMID:20075049

  18. Connective tissue growth factor in tumor pathogenesis

    PubMed Central

    2012-01-01

    Key roles for connective tissue growth factor (CTGF/CCN2) are demonstrated in the wound repair process where it promotes myofibroblast differentiation and angiogenesis. Similar mechanisms are active in tumor-reactive stroma where CTGF is expressed. Other potential roles include prevention of hypoxia-induced apoptosis and promoting epithelial-mesenchymal transistion (EMT). CTGF expression in tumors has been associated to both tumor suppression and progression. For example, CTGF expression in acute lymphoblastic leukemia, breast, pancreas and gastric cancer correlates to worse prognosis whereas the opposite is true for colorectal, lung and ovarian cancer. This discrepancy is not yet understood. High expression of CTGF is a hallmark of ileal carcinoids, which are well-differentiated endocrine carcinomas with serotonin production originating from the small intestine and proximal colon. These tumors maintain a high grade of differentiation and low proliferation. Despite this, they are malignant and most patients have metastatic disease at diagnosis. These tumors demonstrate several phenotypes potentially related to CTGF function namely: cell migration, absent tumor cell apoptosis, as well as, reactive and well vascularised myofibroblast rich stroma and fibrosis development locally and in distal organs. The presence of CTGF in other endocrine tumors indicates a role in the progression of well-differentiated tumors. PMID:23259759

  19. Epidermal growth factor system regulates mucin production in airways

    PubMed Central

    Takeyama, Kiyoshi; Dabbagh, Karim; Lee, Heung-Man; Agustí, Carlos; Lausier, James A.; Ueki, Iris F.; Grattan, Kathleen M.; Nadel, Jay A.

    1999-01-01

    Goblet-cell hyperplasia is a critical pathological feature in hypersecretory diseases of airways. However, the underlying mechanisms are unknown, and no effective therapy exists. Here we show that stimulation of epidermal growth factor receptors (EGF-R) by its ligands, EGF and transforming growth factor α (TGFα), causes MUC5AC expression in airway epithelial cells both in in vitro and in vivo. We found that a MUC5AC-inducing epithelial cell line, NCI-H292, expresses EGF-R constitutively; EGF-R gene expression was stimulated further by tumor necrosis factor α (TNFα). EGF-R ligands increased the expression of MUC5AC at both gene and protein levels, and this effect was potentiated by TNFα. Selective EGF-R tyrosine kinase inhibitors blocked MUC5AC expression induced by EGF-R ligands. Pathogen-free rats expressed little EGF-R protein in airway epithelial cells; intratracheal instillation of TNFα induced EGF-R in airway epithelial cells, and subsequent instillation of EGF-R ligands increased the number of goblet cells, Alcian blue–periodic acid–Schiff staining (reflecting mucous glycoconjugates), and MUC5AC gene expression, whereas TNFα, EGF, or TGFα alone was without effect. In sensitized rats, three intratracheal instillations of ovalbumin resulted in EGF-R expression and goblet-cell production in airway epithelium. Pretreatment with EGF-R tyrosine kinase inhibitor, BIBX1522, prevented goblet-cell production both in rats stimulated by TNFα-EGF-R ligands and in an asthma model. These findings suggest potential roles for inhibitors of the EGF-R cascade in hypersecretory diseases of airways. PMID:10077640

  20. Plakoglobin Suppresses Epithelial Proliferation and Hair Growth in Vivo

    PubMed Central

    Charpentier, Emmanuelle; Lavker, Robert M.; Acquista, Elizabeth; Cowin, Pamela

    2000-01-01

    Plakoglobin regulates cell adhesion by providing a modulatable connection between both classical and desmosomal cadherins and their respective cytoskeletal linker proteins. Both plakoglobin and the related protein β-catenin are posttranscriptionally upregulated in response to Wnt-1 in cultured cells. Upregulation of β-catenin has been implicated in potentiating hyperproliferation and tumor formation. To investigate the role of plakoglobin in these functions we expressed a full-length (PG) and an NH2-terminally truncated form of plakoglobin (ΔN80PG) in mouse epidermis and hair follicles, tissues which undergo continuous and easily observed postnatal renewal and remodeling. Expression of these constructs results in stunted hair growth, a phenotype that has also been observed in transgenic mice expressing Wnt3 and Dvl2 (Millar et al. 1999). Hair follicles from PG and ΔN80PG mice show premature termination of the growth phase (anagen) of the hair cycle, an event that is regulated in part by FGF5 (Hebert et al. 1994). The proliferative rate of the epidermal cells was reduced and apoptotic changes, which are associated with entry into the regressive phase of the hair follicle cycle (catagen), occurred earlier than usual. PMID:10769039

  1. Epidermal growth factor receptor family in lung cancer and premalignancy.

    PubMed

    Franklin, Wilbur A; Veve, Robert; Hirsch, Fred R; Helfrich, Barbara A; Bunn, Paul A

    2002-02-01

    Lung cancer, like many other epithelial malignancies, is thought to be the outcome of genetic and epigenetic changes that result in a constellation of phenotypic abnormalities in bronchial epithelium. These include morphologic epithelial dysplasia, angiogenesis, increased proliferative rate, and changes in expression of cell surface proteins, particularly overexpression of epidermal growth factor receptor (EGFR) family proteins. The EFGR family is a group of four structurally similar tyrosine kinases (EGFR, HER2/neu, ErbB-3, and ErbB-4) that dimerize on binding with a number of ligands, including EGF and transforming growth factor alpha. Epidermal growth factor receptor overexpression is pronounced in virtually all squamous carcinomas and is also found in > or = 65% of large cell and adenocarcinomas. It is not expressed in situ by small cell lung carcinoma. Overexpression of EGFR is one of the earliest and most consistent abnormalities in bronchial epithelium of high-risk smokers. It is present at the stage of basal cell hyperplasia and persists through squamous metaplasia, dysplasia, and carcinoma in situ. Recent studies of the effect of inhibitors of receptor tyrosine kinases suggest that patterns of coexpression of multiple members of the EGFR family could be important in determining response. Intermediate endpoints of such trials could include monitoring of phosphorylation levels in signal transduction molecules downstream of the receptor dimers. These trials represent a new targeted approach to lung cancer treatment and chemoprevention that will require greater attention to molecular endpoints than required in past trials. PMID:11894009

  2. Growth Factor–dependent Activation of αvβ3 Integrin in Normal Epithelial Cells: Implications for Tumor Invasion

    PubMed Central

    Trusolino, Livio; Serini, Guido; Cecchini, Germana; Besati, Cristina; Ambesi-Impiombato, Francesco Saverio; Marchisio, Pier Carlo; De Filippi, Rosaria

    1998-01-01

    Integrin activation is a multifaceted phenomenon leading to increased affinity and avidity for matrix ligands. To investigate whether cytokines produced during stromal infiltration of carcinoma cells activate nonfunctional epithelial integrins, a cellular system of human thyroid clones derived from normal glands (HTU-5) and papillary carcinomas (HTU-34) was employed. In HTU-5 cells, αvβ3 integrin was diffused all over the membrane, disconnected from the cytoskeleton, and unable to mediate adhesion. Conversely, in HTU-34 cells, αvβ3 was clustered at focal contacts (FCs) and mediated firm attachment and spreading. αvβ3 recruitment at FCs and ligand-binding activity, essentially identical to those of HTU-34, occurred in HTU-5 cells upon treatment with hepatocyte growth factor/scatter factor (HGF/SF). The HTU-34 clone secreted HGF/SF and its receptor was constitutively tyrosine phosphorylated suggesting an autocrine loop responsible for αvβ3 activated state. Antibody-mediated inhibition of HGF/SF function in HTU-34 cells disrupted αvβ3 enrichment at FCs and impaired adhesion. Accordingly, activation of αvβ3 in normal cells was produced by HTU-34 conditioned medium on the basis of its content of HGF/SF. These results provide the first example of a growth factor–driven integrin activation mechanism in normal epithelial cells and uncover the importance of cytokine-based autocrine loops for the physiological control of integrin activation. PMID:9722624

  3. miR-494 suppresses tumor growth of epithelial ovarian carcinoma by targeting IGF1R.

    PubMed

    Li, Na; Zhao, Xiaosu; Wang, Lufei; Zhang, Shi; Cui, Manhua; He, Jin

    2016-06-01

    A growing body of evidence suggests that microRNA-494 (miR-494) could act as tumor-suppressive or oncogenic microRNAs (miRNAs) in different types of tumors. However, the biological roles and underlying mechanisms of miR-494 remain unknown in human epithelial ovarian carcinoma (EOC). Therefore, the aims of this study were to investigate the miR-494 expression and the significance of its clinical diagnosis in patients suffering EOC and to analyze its role and underlying molecular mechanism on the carcinogenesis of EOC. Here, we found that miR-494 was significantly decreased in EOC cell lines and tissues and its expression was negatively correlated with advanced International Federation of Gynecology and Obstetrics (FIGO) stage, high pathological grade, and lymph node metastasis (all P < 0.01). Functional studies showed that overexpression of miR-494 in EOC cells could remarkably inhibit proliferation, colony formation, migration, and invasion and induce cell apoptosis, G0/G1 phase arrest. An in vivo analysis revealed that the overexpression of miR-494 suppressed tumor growth in a nude mouse xenograft model system. Bioinformatic assay and dual-luciferase assay confirmed that insulin-like growth factor 1 receptor (IGF1R) was as a direct target of miR-494 in EOC cells. Western blot assay showed that overexpression of miR-494 inhibited IGF1R expression and its downstream signal protein expression. In addition, downregulation of IGF1R has similar effects with miR-494 overexpression on EOC cells and overexpression of IGF1R effectively rescued the inhibition of overexpressed miR-494 in EOC cells. These data suggested that miR-494 functions as a tumor suppressor in EOC by targeting IGF1R. PMID:26695144

  4. The expression and function of vascular endothelial growth factor in retinal pigment epithelial (RPE) cells is regulated by 4-hydroxynonenal (HNE) and glutathione S-transferaseA4-4

    SciTech Connect

    Vatsyayan, Rit; Lelsani, Poorna Chandra Rao; Chaudhary, Pankaj; Kumar, Sushil; Awasthi, Sanjay; Awasthi, Yogesh C.

    2012-01-06

    Highlights: Black-Right-Pointing-Pointer Low concentration of HNE (0.1-1.0 {mu}M) induced secretion of VEGF in RPE cells. Black-Right-Pointing-Pointer VEGF secreted medium of RPE cells promoted proliferation of endothelial cells. Black-Right-Pointing-Pointer VEGFR2 expression was attenuated with increasing concentrations of HNE. Black-Right-Pointing-Pointer These effects of HNE could be blocked by the over expression of GSTA4-4 in cells. -- Abstract: It is well established that 4-hydroxynonenal (HNE) plays a major role in oxidative stress-induced signaling and the toxicity of oxidants. Surprisingly our recent studies also demonstrate that low levels of HNE generated during oxidative stress promote cell survival mechanisms and proliferation. Since the expression and secretion of VEGF is known to be affected by Oxidative stress, during present studies, we have examined dose dependent effect of HNE on VEGF expression and secretion in a model of retinal pigment epithelial (RPE) cells in culture. Results of these studies showed that while inclusion of 0.1 {mu}M HNE in the medium caused increased secretion of VEGF, its secretion and expression was significantly suppressed in the presence of >5 {mu}M HNE in the media. These concentration dependent hormetic effects of HNE on VEGF secretion could be blocked by the over expression of GSTA4-4 indicating that these effects were specifically attributed to HNE and regulated by GSTA4-4. VEGF secreted into the media showed angiogenic properties as indicated by increased migration and tube formation of HUVEC in matrigel when grown in media from RPE cells treated with 1 {mu}M HNE. The corresponding media from GSTA4-4 over expressing RPE cells had no effect on migration and tube formation of HUVEC in matrigel. These results are consistent with earlier studies showing that at low concentrations, HNE promotes proliferative mechanisms and suggest that HNE induces VEGF secretion from RPE cells that acts in a paracrine fashion to induce

  5. Hepatocyte Nuclear Factor-1β Induces Redifferentiation of Dedifferentiated Tubular Epithelial Cells

    PubMed Central

    Omata, Mitsugu; Doke, Yukiko; Yamada, Chikaomi; Kawashima, Kayoko; Sho, Rumiko; Enomoto, Kei; Furuya, Mayumi; Inomata, Norio

    2016-01-01

    Tubular epithelial cells (TECs) can be dedifferentiated by repetitive insults, which activate scar-producing cells generated from interstitial cells such as fibroblasts, leading to the accumulation and deposition of extracellular matrix molecules. The dedifferentiated TECs play a crucial role in the development of renal fibrosis. Therefore, renal fibrosis may be attenuated if dedifferentiated TECs are converted back to their normal state (re-epithelialization). However, the mechanism underlying the re-epithelialization remains to be elucidated. In the present study, TGF-β1, a profibrotic cytokine, induced dedifferentiation of cultured TECs, and the dedifferentiated TECs were re-epithelialized by the removal of TGF-β1 stimulation. In the re-epithelialization process, transcription factor hepatocyte nuclear factor 1, beta (HNF-1β) was identified as a candidate molecule involved in inducing re-epithelialization by means of DNA microarray and biological network analysis. In functional validation studies, the re-epithelialization by TGF-β1 removal was abolished by HNF-1β knockdown. Furthermore, the ectopic expression of HNF-1β in the dedifferentiated TECs induced the re-epithelialization without the inhibition of TGF-β/Smad signaling, even in the presence of TGF-β1 stimulation. In mouse renal fibrosis model, unilateral ureteral obstruction model, HNF-1β expression in the TECs of the kidney was suppressed with fibrosis progression. Furthermore, the HNF-1β downregulated TECs resulted in dedifferentiation, which was characterized by expression of nestin. In conclusion, HNF-1β suppression in TECs is a crucial event for the dedifferentiation of TECs, and the upregulation of HNF-1β in TECs has a potential to restore the dedifferentiated TECs into their normal state, leading to the attenuation of renal fibrosis. PMID:27196561

  6. Feedback control of growth, differentiation, and morphogenesis of pancreatic endocrine progenitors in an epithelial plexus niche

    PubMed Central

    Bankaitis, Eric D.; Bechard, Matthew E.; Wright, Christopher V.E.

    2015-01-01

    In the mammalian pancreas, endocrine cells undergo lineage allocation upon emergence from a bipotent duct/endocrine progenitor pool, which resides in the “trunk epithelium.” Major questions remain regarding how niche environments are organized within this epithelium to coordinate endocrine differentiation with programs of epithelial growth, maturation, and morphogenesis. We used EdU pulse-chase and tissue-reconstruction approaches to analyze how endocrine progenitors and their differentiating progeny are assembled within the trunk as it undergoes remodeling from an irregular plexus of tubules to form the eventual mature, branched ductal arbor. The bulk of endocrine progenitors is maintained in an epithelial “plexus state,” which is a transient intermediate during epithelial maturation within which endocrine cell differentiation is continually robust and surprisingly long-lived. Within the plexus, local feedback effects derived from the differentiating and delaminating endocrine cells nonautonomously regulate the flux of endocrine cell birth as well as proliferative growth of the bipotent cell population using Notch-dependent and Notch-independent influences, respectively. These feedback effects in turn maintain the plexus state to ensure prolonged allocation of endocrine cells late into gestation. These findings begin to define a niche-like environment guiding the genesis of the endocrine pancreas and advance current models for how differentiation is coordinated with the growth and morphogenesis of the developing pancreatic epithelium. PMID:26494792

  7. Expression of transforming growth factor alpha and epidermal growth factor receptor in rat lung neoplasms induced by plutonium-239

    SciTech Connect

    Stegelmeier, B.L.; Gillett, N.A.; Hahn, F.F.; Kelly, G.; Rebar, A.H.

    1994-11-01

    Ninety-two rat lung proliferative lesions and neoplasms induced by inhaled {sup 239}PuO{sub 2} were evaluated for aberrant expression of transforming growth factor alpha (TGF-{alpha}) and epidermal growth factor receptor (EGFR). Expression of TGF-{alpha} protein, measured by immunohistochemistry, was higher in 94% of the squamous cell carcinomas and 87% of the foci of alveolar epithelial squamous metaplasia than that exhibited by the normal-appearing, adjacent lung parenchyma. In contrast, only 20% of adenocarcinomas and foci of epithelial hyperplasia expressed elevated levels of TGF-{alpha}. Many neoplasms expressing TGF-{alpha} also expressed excessive levels of EGFR mRNA. Southern and DNA slot blot analyses showed that the elevated EGFR expression was not due to amplification of the EGFR gene. These data suggest that increased amounts of TGF-{alpha} were early alterations in the progression of plutonium-induced squamous cell carcinoma, and these increases may occur in parallel with overexpression of the receptor for this growth factor. Together, these alterations create a potential autocrine loop for sustaining clonal expansion of cells initiated by high-LET radiation. 44 refs., 4 figs., 1 tab.

  8. Novel Therapy to Treat Corneal Epithelial Defects: A Hypothesis with Growth Hormone.

    PubMed

    Wirostko, Barbara; Rafii, MaryJane; Sullivan, David A; Morelli, Julia; Ding, Juan

    2015-07-01

    Impaired corneal wound healing that occurs with ocular surface disease, trauma, systemic disease, or surgical intervention can lead to persistent corneal epithelial defects (PCED), which result in corneal scarring, ulceration, opacification, corneal neovascularization, and, ultimately, visual compromise and vision loss. The current standard of care can include lubricants, ointments, bandage lenses, amniotic membranes, autologous serum eye drops, and corneal transplants. Various inherent problems exist with application and administration of these treatments, which often may not result in a completely healed surface. A topically applicable compound capable of promoting corneal epithelial cell proliferation and/or migration would be ideal to accelerate healing. We hypothesize that human growth hormone (HGH) is such a compound. In a recent study, HGH was shown to activate signal transducer and activators of transcription-5 (STAT5) signaling and promote corneal wound healing by enhancing corneal epithelial migration in a co-culture system of corneal epithelial cells and fibroblasts. These effects require an intact communication between corneal epithelia and fibroblasts. Further, HGH promotes corneal wound healing in a rabbit debridement model, thus demonstrating the effectiveness of HGH in vivo as well. In conclusion, HGH may represent an exciting and effective topical therapeutic to promote corneal wound healing. PMID:26045234

  9. Protection of photoreceptor cells from phototoxicity by transplanted retinal pigment epithelial cells expressing different neurotrophic factors.

    PubMed

    Abe, Toshiaki; Saigo, Yoko; Hojo, Masayoshi; Kano, Tetsuya; Wakusawa, Ryosuke; Tokita, Yumi; Tamai, Makoto

    2005-01-01

    Transplantation of cells or tissues and the intravitreal injection of neurotrophic factors are two methods that have been used to treat retinal diseases. The purpose of this study was to examine the effects of combining both methods: the transplantation of retinal pigment epithelial (RPE) cells expressing different neurotrophic factors. The neutrophic factors were Axokine, brain derived-neurotrophic factor (BDNF), and basic fibroblast growth factor (bFGF). The enhanced green fluorescence protein (eGFP) gene was used as a reporter gene. These genes were transduced into RPE cells by lipofection, selected by antibiotics, and transplanted into the subretinal space of 108 rats. The rats were examined at 1 week and 3 months after the transplantation to determine whether the transduced cells were present, were expressing the protein, and were able to protect photoreceptors against phototoxicity. The survival of the transplanted cells was monitored by the presence of eGFP. The degree of protection was determined by the thickness of the outer nuclear layer. Our results showed that the degree of photoreceptor protection was different for the different types of neurotrophic factors at 1 week. After 3 months, the number of surviving transplanted cell was markedly reduced, and protection was observed only with the BDNF-transduced RPE cells. A significant degree of rescue was also observed by BDNF-transduced RPE cells in the nontransplanted area of the retina at both the early and late times. Lymphocytic infiltration was not detected in the vitreous, retina, and choroid at any time. We conclude that the transplantation of BDNF-transduced RPE cells can reduce the photoreceptor damage induced by phototoxicity in the transplanted area and weakly in the nontransplanted area. PMID:16454354

  10. Small Is Beautiful: Insulin-Like Growth Factors and Their Role in Growth, Development, and Cancer

    PubMed Central

    Maki, Robert G.

    2010-01-01

    Insulin-like growth factors were discovered more than 50 years ago as mediators of growth hormone that effect growth and differentiation of bone and skeletal muscle. Interest of the role of insulin-like growth factors in cancer reached a peak in the 1990s, and then waned until the availability in the past 5 years of monoclonal antibodies and small molecules that block the insulin-like growth factor 1 receptor. In this article, we review the history of insulin-like growth factors and their role in growth, development, organism survival, and in cancer, both epithelial cancers and sarcomas. Recent developments regarding phase I to II clinical trials of such agents are discussed, as well as potential studies to consider in the future, given the lack of efficacy of one such monoclonal antibody in combination with cytotoxic chemotherapy in a first-line study in metastatic non–small-cell lung adenocarcinoma. Greater success with these agents clinically is expected when combining the agents with inhibitors of other cell signaling pathways in which cross-resistance has been observed. PMID:20975071

  11. Small is beautiful: insulin-like growth factors and their role in growth, development, and cancer.

    PubMed

    Maki, Robert G

    2010-11-20

    Insulin-like growth factors were discovered more than 50 years ago as mediators of growth hormone that effect growth and differentiation of bone and skeletal muscle. Interest of the role of insulin-like growth factors in cancer reached a peak in the 1990s, and then waned until the availability in the past 5 years of monoclonal antibodies and small molecules that block the insulin-like growth factor 1 receptor. In this article, we review the history of insulin-like growth factors and their role in growth, development, organism survival, and in cancer, both epithelial cancers and sarcomas. Recent developments regarding phase I to II clinical trials of such agents are discussed, as well as potential studies to consider in the future, given the lack of efficacy of one such monoclonal antibody in combination with cytotoxic chemotherapy in a first-line study in metastatic non-small-cell lung adenocarcinoma. Greater success with these agents clinically is expected when combining the agents with inhibitors of other cell signaling pathways in which cross-resistance has been observed. PMID:20975071

  12. TSC1 activates TGF-β-Smad2/3 signaling in growth arrest and epithelial-to-mesenchymal transition.

    PubMed

    Thien, Antje; Prentzell, Mirja Tamara; Holzwarth, Birgit; Kläsener, Kathrin; Kuper, Ineke; Boehlke, Christopher; Sonntag, Annika G; Ruf, Stefanie; Maerz, Lars; Nitschke, Roland; Grellscheid, Sushma-Nagaraja; Reth, Michael; Walz, Gerd; Baumeister, Ralf; Neumann-Haefelin, Elke; Thedieck, Kathrin

    2015-03-01

    The tuberous sclerosis proteins TSC1 and TSC2 are key integrators of growth factor signaling. They suppress cell growth and proliferation by acting in a heteromeric complex to inhibit the mammalian target of rapamycin complex 1 (mTORC1). In this study, we identify TSC1 as a component of the transforming growth factor β (TGF-β)-Smad2/3 pathway. Here, TSC1 functions independently of TSC2. TSC1 interacts with the TGF-β receptor complex and Smad2/3 and is required for their association with one another. TSC1 regulates TGF-β-induced Smad2/3 phosphorylation and target gene expression and controls TGF-β-induced growth arrest and epithelial-to-mesenchymal transition (EMT). Hyperactive Akt specifically activates TSC1-dependent cytostatic Smad signaling to induce growth arrest. Thus, TSC1 couples Akt activity to TGF-β-Smad2/3 signaling. This has implications for cancer treatments targeting phosphoinositide 3-kinases and Akt because they may impair tumor-suppressive cytostatic TGF-β signaling by inhibiting Akt- and TSC1-dependent Smad activation. PMID:25727005

  13. Serum-free growth of human mammary epithelial cells: rapid clonal growth in defined medium and extended serial passage with pituitary extract

    SciTech Connect

    Hammond, S.L.; Ham, R.G.; Stampfer, M.R.

    1984-09-01

    A serum-free medium with bovine pituitary extract as the only undefined supplement has been developed for long-term culture of human mammary epithelial cells. This medium supports serial subculture of normal cells for 10-20 passages (1:10 splits) without conditioning or special substrates, and it supports rapid clonal growth with plating efficiencies up to 35%. It consists of an optimized basal nutrient medium, (MCDB 170, supplemented with insulin, hydrocortisone, epidermal growth factor, ethanolamine, phosphoethanolamine, and bovine pituitary extract. Replacement of pituitary extract with prostaglandin E/sub 1/ and ovine prolactin yields a defined medium that supports rapid clonal growth and serial subculture for three of four passages. Cultures initiated in these media from normal reduction mammoplasty tissue remain diploid and maintain normal epithelia morphology, distribution of cell-associated fibronectin, expression of keratin fibrils, and a low level of expression of milk fat globule antigen. Large cell populations can now be generated and stored frozen, permitting multiple experiments over a period of time with cells from a single donor. These media greatly extend the range of experiments that can be performed both conveniently and reproducibly with cultured normal and tumor-derived human mammary epithelial cells. 31 references, 3 figures, 4 tables.

  14. Factors derived from Escherichia coli Nissle 1917, grown in different growth media, enhance cell death in a model of 5-fluorouracil-induced Caco-2 intestinal epithelial cell damage.

    PubMed

    Wang, Hanru; Bastian, Susan E P; Lawrence, Andrew; Howarth, Gordon S

    2015-01-01

    We evaluated supernatants (SNs) from Escherichia coli Nissle 1917 (EcN) grown in commonly used growth media for their capacity to affect the viability of Caco-2 colon cancer cells in the presence and absence of 5-Fluorouracil (5-FU) chemotherapy. EcN was grown in Luria-Bertani (LB), tryptone soya (TSB), Man Rogosa Sharpe (MRS), and M17 broth supplemented with 10% (v/v) lactose solution (M17). Human Caco-2 colon cancer cells were treated with DMEM (control), growth media alone (LB, TSB, MRS, and M17) or EcN SNs derived from these 4 media, in the presence and absence of 5-FU. Cell viability, reactive oxygen species (ROS), and cell monolayer permeability were determined. EcN SN in LB medium reduced Caco-2 cell viability significantly, to 51% at 48 h. The combination of this EcN SN and 5-FU further reduced cell viability to 37% at 48 h, compared to 5-FU control. MRS broth and EcN SN in MRS, together with 5-FU, generated significantly lower levels of ROS compared to 5-FU control. However, all 5-FU treatments significantly disrupted the Caco-2 cell barrier compared to control; with no significant differences observed among any of the 5-FU treatments. EcN SNs (LB+) was most effective at decreasing the viability of Caco-2 cells. This could indicate a potential role for this EcN SN in chemoprevention for colon cancer. PMID:25625670

  15. Spermidine Promotes Human Hair Growth and Is a Novel Modulator of Human Epithelial Stem Cell Functions

    PubMed Central

    Bíró, Tamás; Abu Bakar, Mohd Hilmi; Sugawara, Koji; Philpott, Michael P.; Harrison, Wesley; Pietilä, Marko; Paus, Ralf

    2011-01-01

    Background Rapidly regenerating tissues need sufficient polyamine synthesis. Since the hair follicle (HF) is a highly proliferative mini-organ, polyamines may also be important for normal hair growth. However, the role of polyamines in human HF biology and their effect on HF epithelial stem cells in situ remains largely unknown. Methods and Findings We have studied the effects of the prototypic polyamine, spermidine (0.1–1 µM), on human scalp HFs and human HF epithelial stem cells in serum-free organ culture. Under these conditions, spermidine promoted hair shaft elongation and prolonged hair growth (anagen). Spermidine also upregulated expression of the epithelial stem cell-associated keratins K15 and K19, and dose-dependently modulated K15 promoter activity in situ and the colony forming efficiency, proliferation and K15 expression of isolated human K15-GFP+ cells in vitro. Inhibiting the rate-limiting enzyme of polyamine synthesis, ornithine decarboyxlase (ODC), downregulated intrafollicular K15 expression. In primary human epidermal keratinocytes, spermidine slightly promoted entry into the S/G2-M phases of the cell cycle. By microarray analysis of human HF mRNA extracts, spermidine upregulated several key target genes implicated e.g. in the control of cell adherence and migration (POP3), or endoplasmic reticulum and mitochondrial functions (SYVN1, NACA and SLC25A3). Excess spermidine may restrict further intrafollicular polyamine synthesis by inhibiting ODC gene and protein expression in the HF's companion layer in situ. Conclusions These physiologically and clinically relevant data provide the first direct evidence that spermidine is a potent stimulator of human hair growth and a previously unknown modulator of human epithelial stem cell biology. PMID:21818338

  16. Diabetes, growth hormone-insulin-like growth factor pathways and association to benign prostatic hyperplasia.

    PubMed

    Wang, Zongwei; Olumi, Aria F

    2011-01-01

    Diabetes significantly increases the risk of benign prostatic hyperplasia (BPH) and low urinary tract symptoms (LUTS). The major endocrine aberration in connection with the metabolic syndrome is hyperinsulinemia. Insulin is an independent risk factor and a promoter of BPH. Insulin resistance may change the risk of BPH through several biological pathways. Hyperinsulinemia stimulates the liver to produce more insulin-like growth factor (IGF), another mitogen and an anti-apoptotic agent which binds insulin receptor/IGF receptor and stimulates prostate growth. The levels of IGFs and IGF binding proteins (IGFBPs) in prostate tissue and in blood are associated with BPH risk, with the regulation of circulating androgen and growth hormone. Stromal-epithelial interactions play a critical role in the development and growth of the prostate gland and BPH. Previously, we have shown that the expression of c-Jun in the fibroblastic stroma can promote secretion of IGF-I, which stimulates prostate epithelial cell proliferation through activating specific target genes. Here, we will review the epidemiologic, clinical, and molecular findings which have evaluated the relation between diabetes and development of BPH. PMID:21536370

  17. Loss of epithelial hypoxia-inducible factor prolyl hydroxylase 2 accelerates skin wound healing in mice.

    PubMed

    Kalucka, Joanna; Ettinger, Andreas; Franke, Kristin; Mamlouk, Soulafa; Singh, Rashim Pal; Farhat, Katja; Muschter, Antje; Olbrich, Susanne; Breier, Georg; Katschinski, Dörthe M; Huttner, Wieland; Weidemann, Alexander; Wielockx, Ben

    2013-09-01

    Skin wound healing in mammals is a complex, multicellular process that depends on the precise supply of oxygen. Hypoxia-inducible factor (HIF) prolyl hydroxylase 2 (PHD2) serves as a crucial oxygen sensor and may therefore play an important role during reepithelialization. Hence, this study was aimed at understanding the role of PHD2 in cutaneous wound healing using different lines of conditionally deficient mice specifically lacking PHD2 in inflammatory, vascular, or epidermal cells. Interestingly, PHD2 deficiency only in keratinocytes and not in myeloid or endothelial cells was found to lead to faster wound closure, which involved enhanced migration of the hyperproliferating epithelium. We demonstrate that this effect relies on the unique expression of β3-integrin in the keratinocytes around the tip of the migrating tongue in an HIF1α-dependent manner. Furthermore, we show enhanced proliferation of these cells in the stratum basale, which is directly related to their attenuated transforming growth factor β signaling. Thus, loss of the central oxygen sensor PHD2 in keratinocytes stimulates wound closure by prompting skin epithelial cells to migrate and proliferate. Inhibition of PHD2 could therefore offer novel therapeutic opportunities for the local treatment of cutaneous wounds. PMID:23798557

  18. Loss of Epithelial Hypoxia-Inducible Factor Prolyl Hydroxylase 2 Accelerates Skin Wound Healing in Mice

    PubMed Central

    Kalucka, Joanna; Ettinger, Andreas; Franke, Kristin; Mamlouk, Soulafa; Singh, Rashim Pal; Farhat, Katja; Muschter, Antje; Olbrich, Susanne; Breier, Georg; Katschinski, Dörthe M.; Huttner, Wieland; Weidemann, Alexander

    2013-01-01

    Skin wound healing in mammals is a complex, multicellular process that depends on the precise supply of oxygen. Hypoxia-inducible factor (HIF) prolyl hydroxylase 2 (PHD2) serves as a crucial oxygen sensor and may therefore play an important role during reepithelialization. Hence, this study was aimed at understanding the role of PHD2 in cutaneous wound healing using different lines of conditionally deficient mice specifically lacking PHD2 in inflammatory, vascular, or epidermal cells. Interestingly, PHD2 deficiency only in keratinocytes and not in myeloid or endothelial cells was found to lead to faster wound closure, which involved enhanced migration of the hyperproliferating epithelium. We demonstrate that this effect relies on the unique expression of β3-integrin in the keratinocytes around the tip of the migrating tongue in an HIF1α-dependent manner. Furthermore, we show enhanced proliferation of these cells in the stratum basale, which is directly related to their attenuated transforming growth factor β signaling. Thus, loss of the central oxygen sensor PHD2 in keratinocytes stimulates wound closure by prompting skin epithelial cells to migrate and proliferate. Inhibition of PHD2 could therefore offer novel therapeutic opportunities for the local treatment of cutaneous wounds. PMID:23798557

  19. Ets homologous factor regulates pathways controlling response to injury in airway epithelial cells.

    PubMed

    Fossum, Sara L; Mutolo, Michael J; Yang, Rui; Dang, Hong; O'Neal, Wanda K; Knowles, Michael R; Leir, Shih-Hsing; Harris, Ann

    2014-12-16

    Ets homologous factor (EHF) is an Ets family transcription factor expressed in many epithelial cell types including those lining the respiratory system. Disruption of the airway epithelium is central to many lung diseases, and a network of transcription factors coordinates its normal function. EHF can act as a transcriptional activator or a repressor, though its targets in lung epithelial cells are largely uncharacterized. Chromatin immunoprecipitation followed by deep sequencing (ChIP-seq), showed that the majority of EHF binding sites in lung epithelial cells are intergenic or intronic and coincide with putative enhancers, marked by specific histone modifications. EHF occupies many genomic sites that are close to genes involved in intercellular and cell-matrix adhesion. RNA-seq after EHF depletion or overexpression showed significant alterations in the expression of genes involved in response to wounding. EHF knockdown also targeted genes in pathways of epithelial development and differentiation and locomotory behavior. These changes in gene expression coincided with alterations in cellular phenotype including slowed wound closure and increased transepithelial resistance. Our data suggest that EHF regulates gene pathways critical for epithelial response to injury, including those involved in maintenance of barrier function, inflammation and efficient wound repair. PMID:25414352

  20. Growth Factor Mediated Signaling in Pancreatic Pathogenesis

    PubMed Central

    Nandy, Debashis; Mukhopadhyay, Debabrata

    2011-01-01

    Functionally, the pancreas consists of two types of tissues: exocrine and endocrine. Exocrine pancreatic disorders mainly involve acute and chronic pancreatitis. Acute pancreatitis typically is benign, while chronic pancreatitis is considered a risk factor for developing pancreatic cancer. Pancreatic carcinoma is the fourth leading cause of cancer related deaths worldwide. Most pancreatic cancers develop in the exocrine tissues. Endocrine pancreatic tumors are more uncommon, and typically are less aggressive than exocrine tumors. However, the endocrine pancreatic disorder, diabetes, is a dominant cause of morbidity and mortality. Importantly, different growth factors and their receptors play critical roles in pancreatic pathogenesis. Hence, an improved understanding of how various growth factors affect pancreatitis and pancreatic carcinoma is necessary to determine appropriate treatment. This chapter describes the role of different growth factors such as vascular endothelial growth factor (VEGF), insulin-like growth factor (IGF), platelet derived growth factor (PDGF), fibroblast growth factor (FGF), epidermal growth factor (EGF), and transforming growth factor (TGF) in various pancreatic pathophysiologies. Finally, the crosstalk between different growth factor axes and their respective signaling mechanisms, which are involved in pancreatitis and pancreatic carcinoma, are also discussed. PMID:24212642

  1. A systematic approach identifies FOXA1 as a key factor in the loss of epithelial traits during the epithelial-to-mesenchymal transition in lung cancer

    PubMed Central

    2013-01-01

    Background The epithelial-to-mesenchymal transition is an important mechanism in cancer metastasis. Although transcription factors including SNAIL, SLUG, and TWIST1 regulate the epithelial-to-mesenchymal transition, other unknown transcription factors could also be involved. Identification of the full complement of transcription factors is essential for a more complete understanding of gene regulation in this process. Chromatin immunoprecipitation-sequencing (ChIP-Seq) technologies have been used to detect genome-wide binding of transcription factors; here, we developed a systematic approach to integrate existing ChIP-Seq and transcriptome data. We scanned multiple transcription factors to investigate their functional impact on the epithelial-to-mesenchymal transition in the human A549 lung adenocarcinoma cell line. Results Among the transcription factors tested, impact scores identified the forkhead box protein A1 (FOXA1) as the most significant transcription factor in the epithelial-to-mesenchymal transition. FOXA1 physically associates with the promoters of its predicted target genes. Several critical epithelial-to-mesenchymal transition effectors involved in cellular adhesion and cellular communication were identified in the regulatory network of FOXA1, including FOXA2, FGA, FGB, FGG, and FGL1. The implication of FOXA1 in the epithelial-to-mesenchymal transition via its regulatory network indicates that FOXA1 may play an important role in the initiation of lung cancer metastasis. Conclusions We identified FOXA1 as a potentially important transcription factor and negative regulator in the initial stages of lung cancer metastasis. FOXA1 may modulate the epithelial-to-mesenchymal transition via its transcriptional regulatory network. Further, this study demonstrates how ChIP-Seq and expression data could be integrated to delineate the impact of transcription factors on a specific biological process. PMID:24093963

  2. Inhibitor of growth 4 suppresses colorectal cancer growth and invasion by inducing G1 arrest, inhibiting tumor angiogenesis and reversing epithelial-mesenchymal transition.

    PubMed

    Qu, Hui; Yin, Hong; Yan, Su; Tao, Min; Xie, Yufeng; Chen, Weichang

    2016-05-01

    Previous studies have found that inhibitor of growth 4 (ING4), a tumor suppressor, is reduced in human colorectal cancer (CRC), and is inversely correlated with clinical Dukes' stage, histological grade, lymph node metastasis and microvessel density (MVD). However, its underlying mechanism remains undetermined. In the present study, we analyzed ING4 expression in a panel of human CRC cells using low (LS174T and SW480) and high (LoVo and SW620) metastatic cell lines. We demonstrated that both the low and high metastatic CRC cells exhibited a lower level of ING4 compared to the level in normal human colorectal mucous epithelial FHC cells. Furthermore, ING4 expression in high metastatic CRC cells was less than that in low metastatic CRC cells. We then generated a lentivirus construct expressing ING4 and green fluorescent protein (GFP), established a ING4-stably transgenic LoVo CRC cell line, and investigated the effect of lentiviral-mediated ING4 expression on high metastatic LoVo CRC cells. Gain-of-function studies revealed that ING4 significantly inhibited LoVo CRC cell growth and invasion in vitro and induced cell cycle G1 phase arrest. Moreover, ING4 obviously suppressed LoVo CRC subcutaneously xenografted tumor growth and reduced tumor MVD in vivo in athymic BALB/c nude mice. Mechanistically, ING4 markedly upregulated P21 and E-cadherin but downregulated cyclin E, interleukin (IL)-6, IL-8, vascular endothelial growth factor (VEGF), Snail1, N-cadherin and vimentin in the LoVo CRC cells. Our data provide compelling evidence that i) ING4 suppresses CRC growth possibly via induction of G1 phase arrest through upregulation of P21 cyclin-dependent kinase (CDK) inhibitor and downregulation of cyclin E as well as inhibition of tumor angiogenesis through reduction of IL-6, IL-8 and VEGF proangiogenic factors; ii) ING4 inhibits CRC invasion and metastasis probably via a switch from mesenchymal marker N-cadherin to epithelial marker E-cadherin through

  3. The Ets transcription factor EHF as a regulator of cornea epithelial cell identity.

    PubMed

    Stephens, Denise N; Klein, Rachel Herndon; Salmans, Michael L; Gordon, William; Ho, Hsiang; Andersen, Bogi

    2013-11-29

    The cornea is the clear, outermost portion of the eye composed of three layers: an epithelium that provides a protective barrier while allowing transmission of light into the eye, a collagen-rich stroma, and an endothelium monolayer. How cornea development and aging is controlled is poorly understood. Here we characterize the mouse cornea transcriptome from early embryogenesis through aging and compare it with transcriptomes of other epithelial tissues, identifying cornea-enriched genes, pathways, and transcriptional regulators. Additionally, we profiled cornea epithelium and stroma, defining genes enriched in these layers. Over 10,000 genes are differentially regulated in the mouse cornea across the time course, showing dynamic expression during development and modest expression changes in fewer genes during aging. A striking transition time point for gene expression between postnatal days 14 and 28 corresponds with completion of cornea development at the transcriptional level. Clustering classifies co-expressed, and potentially co-regulated, genes into biologically informative categories, including groups that exhibit epithelial or stromal enriched expression. Based on these findings, and through loss of function studies and ChIP-seq, we show that the Ets transcription factor EHF promotes cornea epithelial fate through complementary gene activating and repressing activities. Furthermore, we identify potential interactions between EHF, KLF4, and KLF5 in promoting cornea epithelial differentiation. These data provide insights into the mechanisms underlying epithelial development and aging, identifying EHF as a regulator of cornea epithelial identity and pointing to interactions between Ets and KLF factors in promoting epithelial fate. Furthermore, this comprehensive gene expression data set for the cornea is a powerful tool for discovery of novel cornea regulators and pathways. PMID:24142692

  4. The Ets Transcription Factor EHF as a Regulator of Cornea Epithelial Cell Identity*

    PubMed Central

    Stephens, Denise N.; Klein, Rachel Herndon; Salmans, Michael L.; Gordon, William; Ho, Hsiang; Andersen, Bogi

    2013-01-01

    The cornea is the clear, outermost portion of the eye composed of three layers: an epithelium that provides a protective barrier while allowing transmission of light into the eye, a collagen-rich stroma, and an endothelium monolayer. How cornea development and aging is controlled is poorly understood. Here we characterize the mouse cornea transcriptome from early embryogenesis through aging and compare it with transcriptomes of other epithelial tissues, identifying cornea-enriched genes, pathways, and transcriptional regulators. Additionally, we profiled cornea epithelium and stroma, defining genes enriched in these layers. Over 10,000 genes are differentially regulated in the mouse cornea across the time course, showing dynamic expression during development and modest expression changes in fewer genes during aging. A striking transition time point for gene expression between postnatal days 14 and 28 corresponds with completion of cornea development at the transcriptional level. Clustering classifies co-expressed, and potentially co-regulated, genes into biologically informative categories, including groups that exhibit epithelial or stromal enriched expression. Based on these findings, and through loss of function studies and ChIP-seq, we show that the Ets transcription factor EHF promotes cornea epithelial fate through complementary gene activating and repressing activities. Furthermore, we identify potential interactions between EHF, KLF4, and KLF5 in promoting cornea epithelial differentiation. These data provide insights into the mechanisms underlying epithelial development and aging, identifying EHF as a regulator of cornea epithelial identity and pointing to interactions between Ets and KLF factors in promoting epithelial fate. Furthermore, this comprehensive gene expression data set for the cornea is a powerful tool for discovery of novel cornea regulators and pathways. PMID:24142692

  5. Nerve growth factor and asthma.

    PubMed

    Bonini, S; Lambiase, A; Lapucci, G; Properzi, F; Bresciani, M; Bracci Laudiero, M L; Mancini, M J; Procoli, A; Micera, A; Sacerdoti, G; Bonini, S; Levi-Schaffer, F; Rasi, G; Aloe, L

    2002-01-01

    An increasing body of evidence shows that nerve growth factor (NGF) exerts biological activity not only on the central and peripheral nervous system, but also on the immune system thereby influencing allergic diseases and asthma. (1) NGF circulating levels are increased in patients with allergic diseases and asthma, and are related to the severity of the inflammatory process and disease. In vernal keratoconjunctivitis, NGF plasma levels correlate with the number of mast cells infiltrating the conjunctiva, and NGF mRNA is increased in nasal mucosal scrapings of patients with allergic rhinitis who have high levels of NGF in serum and nasal fluids; NGF is further increased in nasal fluids after specific allergen challenge. (2) NGF is produced and released by several modulatory and effector cells of allergic inflammation and asthma, for example T-helper 2 lymphocytes, mast cells and eosinophils. (3) NGF receptors are expressed on the conjunctival epithelium of patients with allergic conjunctivitis and the number of NGF-receptor positive cells is increased in the conjunctiva of these patients. Indeed, local administration of NGF induces fibroblast activation and healing processes of human corneal ulcers, which suggests that NGF plays a role in tissue remodelling processes occurring in asthma. (4) NGF increases airway hyperreactivity to histamine in an animal model of asthma, while anti-NGF treatment reduces airway hyperreactivity induced by ovalbumin topical challenge in the sensitized mouse. PMID:12144547

  6. Synthetic heparin-binding growth factor analogs

    DOEpatents

    Pena, Louis A.; Zamora, Paul; Lin, Xinhua; Glass, John D.

    2007-01-23

    The invention provides synthetic heparin-binding growth factor analogs having at least one peptide chain that binds a heparin-binding growth factor receptor, covalently bound to a hydrophobic linker, which is in turn covalently bound to a non-signaling peptide that includes a heparin-binding domain. The synthetic heparin-binding growth factor analogs are useful as soluble biologics or as surface coatings for medical devices.

  7. Role and prognostic significance of the epithelial-mesenchymal transition factor ZEB2 in ovarian cancer

    PubMed Central

    Prislei, Silvia; Martinelli, Enrica; Zannoni, Gian Franco; Petrillo, Marco; Filippetti, Flavia; Mariani, Marisa; Mozzetti, Simona; Raspaglio, Giuseppina; Scambia, Giovanni; Ferlini, Cristiano

    2015-01-01

    ZEB2 is a key factor in epithelial-mesenchymal transition (EMT), a program controlling cell migration in embryonic development and adult tissue homeostasis. We demonstrated a role of ZEB2 in migration and anchorage-independent cell growth in ovarian cancer, as shown by ZEB2 silencing. We found that the RNA-binding protein HuR bound the 3′UTR of ZEB2 mRNA, acting as a positive regulator of ZEB2 protein expression. In Hey ovarian cell line, HuR silencing decreased ZEB2 and ZEB1 nuclear expression and impaired migration. In hypoglycemic conditions ZEB2 expression decreased, along with ZEB1, vimentin and cytoplasmic HuR, and a reduced cellular migration ability was observed. Analysis of ZEB2 and HuR expression in ovarian cancers revealed that nuclear ZEB2 is localized in tumor leading edge and co-localizes with cytoplasmic HuR. In a series of 143 ovarian cancer patients high expression of ZEB2 mRNA significantly correlated with a poor prognosis in term of both overall survival and progression- free survival. Moreover, at immunohistochemical evaluation, we found that prognostic significance of ZEB2 protein relies on its nuclear expression and co-localization with cytoplasmic HuR. In conclusion our findings indicated that nuclear ZEB2 may enhance progression of EMT transition and acquisition of an aggressive phenotype in ovarian cancer. PMID:26136338

  8. Autocrine growth factors and solid tumor malignancy.

    PubMed Central

    Walsh, J. H.; Karnes, W. E.; Cuttitta, F.; Walker, A.

    1991-01-01

    The ability of malignant cells to escape the constraint that normally regulate cell growth and differentiation has been a primary focus of attention for investigators of cancer cell biology. An outcome of this attention has been the discovery that the protein products of oncogenes play a role in the activation of growth signal pathways. A second outcome, possibly related to abnormal oncogene expression, has been the discovery that malignant cells frequently show an ability to regulate their own growth by the release of autocrine growth modulatory substances. Most important, the growth of certain malignant cell types has been shown to depend on autocrine growth circuits. A malignant tumor whose continued growth depends on the release of an autocrine growth factor may be vulnerable to treatment with specific receptor antagonists or immunoneutralizing antibodies designed to break the autocrine circuit. Information is rapidly emerging concerning autocrine growth factors in selected human solid tissue malignancy. Images PMID:1926844

  9. Transcription Factors OVOL1 and OVOL2 Induce the Mesenchymal to Epithelial Transition in Human Cancer

    PubMed Central

    Roca, Hernan; Hernandez, James; Weidner, Savannah; McEachin, Richard C.; Fuller, David; Sud, Sudha; Schumann, Taibriana; Wilkinson, John E.; Zaslavsky, Alexander; Li, Hangwen; Maher, Christopher A.; Daignault-Newton, Stephanie; Healy, Patrick N.; Pienta, Kenneth J.

    2013-01-01

    Cell plasticity regulated by the balance between the mesenchymal to epithelial transition (MET) and the opposite program, EMT, is critical in the metastatic cascade. Several transcription factors (TFs) are known to regulate EMT, though the mechanisms of MET remain unclear. We demonstrate a novel function of two TFs, OVOL1 and OVOL2, as critical inducers of MET in human cancers. Our findings indicate that the OVOL-TFs control MET through a regulatory feedback loop with EMT-inducing TF ZEB1, and the regulation of mRNA splicing by inducing Epithelial Splicing Regulatory Protein 1 (ESRP1). Using mouse prostate tumor models we show that expression of OVOL-TFs in mesenchymal prostate cancer cells attenuates their metastatic potential. The role of OVOL-TFs as inducers of MET is further supported by expression analyses in 917 cancer cell lines, suggesting their role as crucial regulators of epithelial-mesenchymal cell plasticity in cancer. PMID:24124593

  10. Recent Advancements in Prognostic Factors of Epithelial Ovarian Carcinoma

    PubMed Central

    Ezzati, Mohammad; Abdullah, Amer; Shariftabrizi, Ahmad; Hou, June; Kopf, Michael; Stedman, Jennifer K.; Samuelson, Robert; Shahabi, Shohreh

    2014-01-01

    Ovarian cancer remains the most common cause of gynecologic cancer-related death among women in developed countries. Nevertheless, subgroups of ovarian cancer patients experience relatively longer survival. Efforts to identify prognostic factors that characterize such patients are ongoing, with investigational areas including tumor characteristics, surgical management, inheritance patterns, immunologic factors, and genomic patterns. This review discusses various demographic, clinical, and molecular factors implicating longevity and ovarian cancer survival. Continued efforts at identifying these prognosticators may result in invaluable adjuncts to the treatment of ovarian cancer, with the ultimate goal of advancing patient care.

  11. Roles for Growth Factors in Cancer Progression

    PubMed Central

    Witsch, Esther; Sela, Michael; Yarden, Yosef

    2011-01-01

    Under physiological conditions, cells receive fate-determining signals from their tissue surroundings, primarily in the form of polypeptide growth factors. Integration of these extracellular signals underlies tissue homeostasis. Although departure from homeostasis and tumor initiation are instigated by oncogenic mutations rather than by growth factors, the latter are the major regulators of all subsequent steps of tumor progression, namely clonal expansion, invasion across tissue barriers, angiogenesis, and colonization of distant niches. Here, we discuss the relevant growth factor families, their roles in tumor biology, as well as the respective downstream signaling pathways. Importantly, cancer-associated activating mutations that impinge on these pathways often relieve, in part, the reliance of tumors on growth factors. On the other hand, growth factors are frequently involved in evolvement of resistance to therapeutic regimens, which extends the roles for polypeptide factors to very late phases of tumor progression and offers opportunities for cancer therapy. PMID:20430953

  12. Scatter factor influences the formation of prostate epithelial cell colonies on bone marrow stroma in vitro.

    PubMed

    Lang, S H; Clarke, N W; George, N J; Testa, N G

    1999-06-01

    Prostate cancer metastases form selectively in the bone marrow. Previously we demonstrated motility was important for the formation of primary prostatic epithelial cell colonies in bone marrow stroma (BMS) co-culture. In this study we looked at the influence of motility factors on the colony formation of epithelial cells derived from benign (bPEC) or malignant (mPEC) prostate tissue. After 7 days co-culture we found that anti-scatter factor consistently inhibited prostate epithelial cell colony formation on BMS (7/7 mPEC and 4/7 bPEC samples showed significant inhibition). Antibodies against bFGF and 5T4 did not significantly affect colony formation. Addition of fibroblast conditioned media (derived from benign prostates) to co-cultures stimulated the colony formation of bPEC (170%) and mPEC (252%). This stimulation was eliminated by depletion of SF from the conditioned media. Immunohistochemical staining found c-Met expression in 5/6 bPEC cultures and 7/9 mPEC cultures. When grown in BMS co-culture expression of c-Met was positive in 3/6 bPEC and 2/7 mPEC samples. In conclusion, scatter factor influences the in vitro formation of prostate epithelial cell colonies on BMS co-culture. PMID:10545020

  13. An epithelial marker promoter induction screen identifies histone deacetylase inhibitors to restore epithelial differentiation and abolishes anchorage independence growth in cancers.

    PubMed

    Tang, H M; Kuay, K T; Koh, P F; Asad, M; Tan, T Z; Chung, V Y; Lee, S C; Thiery, J P; Huang, Ry-J

    2016-01-01

    Epithelial-mesenchymal transition (EMT), a crucial mechanism in development, mediates aggressiveness during carcinoma progression and therapeutic refractoriness. The reversibility of EMT makes it an attractive strategy in designing novel therapeutic approaches. Therefore, drug discovery pipelines for EMT reversal are in need to discover emerging classes of compounds. Here, we outline a pre-clinical drug screening platform for EMT reversal that consists of three phases of drug discovery and validation. From the Phase 1 epithelial marker promoter induction (EpI) screen on a library consisting of compounds being approved by Food and Drug Administration (FDA), Vorinostat (SAHA), a histone deacetylase inhibitor (HDACi), is identified to exert EMT reversal effects by restoring the expression of an epithelial marker, E-cadherin. An expanded screen on 41 HDACi further identifies 28 compounds, such as class I-specific HDACi Mocetinosat, Entinostat and CI994, to restore E-cadherin and ErbB3 expressions in ovarian, pancreatic and bladder carcinoma cells. Mocetinostat is the most potent HDACi to restore epithelial differentiation with the lowest concentration required for 50% induction of epithelial promoter activity (EpIC-50).The HDACi exerts paradoxical effects on EMT transcriptional factors such as SNAI and ZEB family and the effects are context-dependent in epithelial- and mesenchymal-like cells. In vitro functional studies further show that HDACi induced significant increase in anoikis and decrease in spheroid formation in ovarian and bladder carcinoma cells with mesenchymal features. This study demonstrates a robust drug screening pipeline for the discovery of compounds capable of restoring epithelial differentiation that lead to significant functional lethality. PMID:27551531

  14. Vascular Endothelial Growth Factor Acts Primarily via Platelet-Derived Growth Factor Receptor α to Promote Proliferative Vitreoretinopathy

    PubMed Central

    Pennock, Steven; Haddock, Luis J.; Mukai, Shizuo; Kazlauskas, Andrius

    2015-01-01

    Proliferative vitreoretinopathy (PVR) is a nonneovascular blinding disease and the leading cause for failure in surgical repair of rhegmatogenous retinal detachments. Once formed, PVR is difficult to treat. Hence, there is an acute interest in developing approaches to prevent PVR. Of the many growth factors and cytokines that accumulate in vitreous as PVR develops, neutralizing vascular endothelial growth factor (VEGF) A has recently been found to prevent PVR in at least one animal model. The goal of this study was to test if Food and Drug Administration–approved agents could protect the eye from PVR in multiple animal models and to further investigate the underlying mechanisms. Neutralizing VEGF with aflibercept (VEGF Trap-Eye) safely and effectively protected rabbits from PVR in multiple models of disease. Furthermore, aflibercept reduced the bioactivity of both experimental and clinical PVR vitreous. Finally, although VEGF could promote some PVR-associated cellular responses via VEGF receptors expressed on the retinal pigment epithelial cells that drive this disease, VEGF's major contribution to vitreal bioactivity occurred via platelet-derived growth factor receptor α. Thus, VEGF promotes PVR by a noncanonical ability to engage platelet-derived growth factor receptor α. These findings indicate that VEGF contributes to nonangiogenic diseases and that anti–VEGF-based therapies may be effective on a wider spectrum of diseases than previously appreciated. PMID:25261788

  15. Growth factor involvement in tension-induced skeletal muscle growth

    NASA Technical Reports Server (NTRS)

    Vandenburgh, Herman H.

    1993-01-01

    Long-term manned space travel will require a better understanding of skeletal muscle atrophy which results from microgravity. Astronaut strength and dexterity must be maintained for normal mission operations and for emergency situations. Although exercise in space slows the rate of muscle loss, it does not prevent it. A biochemical understanding of how gravity/tension/exercise help to maintain muscle size by altering protein synthesis and/or degradation rate should ultimately allow pharmacological intervention to prevent muscle atrophy in microgravity. The overall objective is to examine some of the basic biochemical processes involved in tension-induced muscle growth. With an experimental in vitro system, the role of exogenous and endogenous muscle growth factors in mechanically stimulated muscle growth are examined. Differentiated avian skeletal myofibers can be 'exercised' in tissue culture using a newly developed dynamic mechanical cell stimulator device which simulates different muscle activity patterns. Patterns of mechanical activity which significantly affect muscle growth and metabolic characteristics were found. Both exogenous and endogenous growth factors are essential for tension-induced muscle growth. Exogenous growth factors found in serum, such as insulin, insulin-like growth factors, and steroids, are important regulators of muscle protein turnover rates and mechanically-induced muscle growth. Endogenous growth factors are synthesized and released into the culture medium when muscle cells are mechanically stimulated. At least one family of mechanically induced endogenous factors, the prostaglandins, help to regulate the rates of protein turnover in muscle cells. Endogenously synthesized IGF-1 is another. The interaction of muscle mechanical activity and these growth factors in the regulation of muscle protein turnover rates with our in vitro model system is studied.

  16. The emerging role of hepatocyte growth factor in renal diseases.

    PubMed

    Mao, Song; Zhang, Jianhua

    2016-06-01

    Hepatocyte growth factor (HGF), a kringle-containing polypeptide, acts on various epithelial cells to regulate cell growth, cell motility, and morphogenesis. HGF also accelerates tissue regeneration of injured organs and is regarded as a key molecule in organ regeneration. Besides the regeneration of the liver, HGF also plays a role in the renal regeneration. In addition, an adaptive alteration of HGF status in various renal diseases occurs. However, the precise role of HGF in various renal diseases remains elusive. The signaling pathways of HGF may be associated with renal diseases. In this review, we will try to provide an in-depth understanding of the underlying role of HGF and its possible interactions with other molecules in renal diseases. PMID:26460681

  17. Poly(rC)-Binding Protein 2 Regulates Hippo Signaling To Control Growth in Breast Epithelial Cells.

    PubMed

    Li, Fengmin; Bullough, Kimberly Z; Vashisht, Ajay A; Wohlschlegel, James A; Philpott, Caroline C

    2016-08-15

    Poly(rC)-binding proteins (PCBPs) are multifunctional adapters that mediate interactions between nucleic acids, iron cofactors, and other proteins, affecting the fates and activities of the components of these interactions. Here, we show that PCBP2 forms a complex with the Hippo pathway components Salvador (Sav1), Mst1, Mst2, and Lats1 in human cells and mouse tissues. Hippo is a kinase cascade that functions to phosphorylate and inactivate the transcriptional coactivators YAP and TAZ, which control cell growth and proliferation. PCBP2 specifically interacts with the scaffold protein Sav1 and prevents proteolytic cleavage of the Mst1 kinase, resulting in increased signaling through Hippo and suppressed activity of YAP and TAZ. Human breast epithelial cells lacking PCBP2 exhibit impaired proteasomal degradation of TAZ. They accumulate TAZ in both the nucleus and the cytosol, increase expression of YAP and TAZ connective tissue growth factor (CTGF) and Cyr61 target genes, and exhibit anchorage-independent growth. Thus, PCBP2 can function as a component of the Hippo complex, enhancing signaling, suppressing activity of YAP and TAZ, and altering the growth characteristics of cells. PMID:27215387

  18. Doxycycline Inducible Kruppel-Like Factor 4 Lentiviral Vector Mediates Mesenchymal to Epithelial Transition in Ovarian Cancer Cells

    PubMed Central

    Chen, Zixuan; Wang, Yinan; Liu, Wen; Zhao, Guannan; Lee, Suechin; Balogh, Andrea; Zou, Yanan; Guo, Yuqi; Zhang, Zhan; Gu, Weiwang; Li, Chengyao; Tigyi, Gabor; Yue, Junming

    2014-01-01

    Ovarian cancer presents therapeutic challenges due to its typically late detection, aggressive metastasis, and therapeutic resistance. The transcription factor Krüppel-like factor 4 (KLF4) has been implicated in human cancers as a tumor suppressor or oncogene, although its role depends greatly on the cellular context. The role of KLF4 in ovarian cancer has not been elucidated in mechanistic detail. In this study, we investigated the role of KLF4 in ovarian cancer cells by transducing the ovarian cancer cell lines SKOV3 and OVCAR3 with a doxycycline-inducible KLF4 lentiviral vector. Overexpression of KLF4 reduced cell proliferation, migration, and invasion. The epithelial cell marker gene E-cadherin was significantly upregulated, whereas the mesenchymal cell marker genes vimentin, twist1and snail2 (slug) were downregulated in both KLF4-expressing SKOV3 and OVCAR3 cells. KLF4 inhibited the transforming growth factor β (TGFβ)-induced epithelial to mesenchymal transition (EMT) in ovarian cancer cells. Taken together, our data demonstrate that KLF4 functions as a tumor suppressor gene in ovarian cancer cells by inhibiting TGFβ-induced EMT. PMID:25137052

  19. Direct and in vitro observation of growth hormone receptor molecules in A549 human lung epithelial cells by nanodiamond labeling

    NASA Astrophysics Data System (ADS)

    Cheng, C.-Y.; Perevedentseva, E.; Tu, J.-S.; Chung, P.-H.; Cheng, C.-L.; Liu, K.-K.; Chao, J.-I.; Chen, P.-H.; Chang, C.-C.

    2007-04-01

    This letter presents direct observation of growth hormone receptor in one single cancer cell using nanodiamond-growth hormone complex as a specific probe. The interaction of surface growth hormone receptor of A549 human lung epithelial cells with growth hormone was observed using nanodiamond's unique spectroscopic signal via confocal Raman mapping. The growth hormone molecules were covalent conjugated to 100nm diameter carboxylated nanodiamonds, which can be recognized specifically by the growth hormone receptors of A549 cell. The Raman spectroscopic signal of diamond provides direct and in vitro observation of growth hormone receptors in physiology condition in a single cell level.

  20. Neuropilin 1 is an entry factor that promotes EBV infection of nasopharyngeal epithelial cells.

    PubMed

    Wang, Hong-Bo; Zhang, Hua; Zhang, Jing-Ping; Li, Yan; Zhao, Bo; Feng, Guo-Kai; Du, Yong; Xiong, Dan; Zhong, Qian; Liu, Wan-Li; Du, Huamao; Li, Man-Zhi; Huang, Wen-Lin; Tsao, Sai Wah; Hutt-Fletcher, Lindsey; Zeng, Yi-Xin; Kieff, Elliott; Zeng, Mu-Sheng

    2015-01-01

    Epstein-Barr virus (EBV) is implicated as an aetiological factor in B lymphomas and nasopharyngeal carcinoma. The mechanisms of cell-free EBV infection of nasopharyngeal epithelial cells remain elusive. EBV glycoprotein B (gB) is the critical fusion protein for infection of both B and epithelial cells, and determines EBV susceptibility of non-B cells. Here we show that neuropilin 1 (NRP1) directly interacts with EBV gB(23-431). Either knockdown of NRP1 or pretreatment of EBV with soluble NRP1 suppresses EBV infection. Upregulation of NRP1 by overexpression or EGF treatment enhances EBV infection. However, NRP2, the homologue of NRP1, impairs EBV infection. EBV enters nasopharyngeal epithelial cells through NRP1-facilitated internalization and fusion, and through macropinocytosis and lipid raft-dependent endocytosis. NRP1 partially mediates EBV-activated EGFR/RAS/ERK signalling, and NRP1-dependent receptor tyrosine kinase (RTK) signalling promotes EBV infection. Taken together, NRP1 is identified as an EBV entry factor that cooperatively activates RTK signalling, which subsequently promotes EBV infection in nasopharyngeal epithelial cells. PMID:25670642

  1. Neuropilin 1 is an entry factor that promotes EBV infection of nasopharyngeal epithelial cells

    PubMed Central

    Wang, Hong-Bo; Zhang, Hua; Zhang, Jing-Ping; Li, Yan; Zhao, Bo; Feng, Guo-Kai; Du, Yong; Xiong, Dan; Zhong, Qian; Liu, Wan-Li; Du, Huamao; Li, Man-Zhi; Huang, Wen-Lin; Tsao, Sai Wah; Hutt-Fletcher, Lindsey; Zeng, Yi-Xin; Kieff, Elliott; Zeng, Mu-Sheng

    2015-01-01

    Epstein–Barr virus (EBV) is implicated as an aetiological factor in B lymphomas and nasopharyngeal carcinoma. The mechanisms of cell-free EBV infection of nasopharyngeal epithelial cells remain elusive. EBV glycoprotein B (gB) is the critical fusion protein for infection of both B and epithelial cells, and determines EBV susceptibility of non-B cells. Here we show that neuropilin 1 (NRP1) directly interacts with EBV gB23–431. Either knockdown of NRP1 or pretreatment of EBV with soluble NRP1 suppresses EBV infection. Upregulation of NRP1 by overexpression or EGF treatment enhances EBV infection. However, NRP2, the homologue of NRP1, impairs EBV infection. EBV enters nasopharyngeal epithelial cells through NRP1-facilitated internalization and fusion, and through macropinocytosis and lipid raft-dependent endocytosis. NRP1 partially mediates EBV-activated EGFR/RAS/ERK signalling, and NRP1-dependent receptor tyrosine kinase (RTK) signalling promotes EBV infection. Taken together, NRP1 is identified as an EBV entry factor that cooperatively activates RTK signalling, which subsequently promotes EBV infection in nasopharyngeal epithelial cells. PMID:25670642

  2. Hepatocyte nuclear factor 1 coordinates multiple processes in a model of intestinal epithelial cell function.

    PubMed

    Yang, Rui; Kerschner, Jenny L; Harris, Ann

    2016-04-01

    Mutations in hepatocyte nuclear factor 1 transcription factors (HNF1α/β) are associated with diabetes. These factors are well studied in the liver, pancreas and kidney, where they direct tissue-specific gene regulation. However, they also have an important role in the biology of many other tissues, including the intestine. We investigated the transcriptional network governed by HNF1 in an intestinal epithelial cell line (Caco2). We used chromatin immunoprecipitation followed by direct sequencing (ChIP-seq) to identify HNF1 binding sites genome-wide. Direct targets of HNF1 were validated using conventional ChIP assays and confirmed by siRNA-mediated depletion of HNF1, followed by RT-qPCR. Gene ontology process enrichment analysis of the HNF1 targets identified multiple processes with a role in intestinal epithelial cell function, including properties of the cell membrane, cellular response to hormones, and regulation of biosynthetic processes. Approximately 50% of HNF1 binding sites were also occupied by other members of the intestinal transcriptional network, including hepatocyte nuclear factor 4A (HNF4A), caudal type homeobox 2 (CDX2), and forkhead box A2 (FOXA2). Depletion of HNF1 in Caco2 cells increases FOXA2 abundance and decreases levels of CDX2, illustrating the coordinated activities of the network. These data suggest that HNF1 plays an important role in regulating intestinal epithelial cell function, both directly and through interactions with other intestinal transcription factors. PMID:26855178

  3. Growth factors from genes to clinical application

    SciTech Connect

    Sara, V.R. ); Hall, K.; Low, H. )

    1990-01-01

    The last decade has witnessed an explosion in the identification of growth factors and their receptors. This has been greatly facilitated by recombinant DNA technology, which has provided the tools not only to identify these proteins at the gene level but also to produce recombinant proteins for evaluating their biological activities. With the help of such techniques, we are moving toward an understanding of the biosynthesis of growth factors and their receptors, structure-function relationships, as well as mechanisms for intracellular signal transmission. The possibility of modifying these factors has opened new fields of clinical application. In this paper, four major areas of growth factor research are presented: the characterization of growth factor genes and their protein products, growth factor receptors and signal transduction by the receptors to mediate biological action, the biological actions of the various growth factors, and the role of growth factors in health and disease and their possible clinical application. Some of the topics covered include: structure of the IGFs and their variants; isoforms of PDGF receptor types; tyrosine kinase activation; structure of G-proteins in biological membranes; possible therapeutic application of NGF in the treatment of Parkinson's and Alzheimer's diseases; PDGF's possible role in the development of several fibroproliferative diseases and its therapeutic application in wound healing; and the possible use of angiogenic inhibitors in tumor treatment.

  4. Inverse regulation of human ERBB2 and epidermal growth factor receptors by tumor necrosis factor alpha.

    PubMed Central

    Kalthoff, H; Roeder, C; Gieseking, J; Humburg, I; Schmiegel, W

    1993-01-01

    Recombinant human tumor necrosis factor (TNF) alpha decreased the expression of ERBB2 mRNA by stimulating p55 TNF receptors of pancreatic tumor cells. This decrease contrasts with an increase in epidermal growth factor receptor (EGFR) mRNA. Both effects were selectively achieved by TNF-alpha or -beta, whereas interferon alpha or gamma or transforming growth factor beta showed no such effects. The inverse regulatory effects of TNF on ERBB2 and EGFR mRNA levels were evoked by different signaling pathways of p55 TNF receptors. The TNF-mediated ERBB2 mRNA decrease was followed by a reduction in protein. Four of five pancreatic tumor cell lines exhibited this down-regulation. This decrease of ERBB2 is a singular example of a modulation of this growth factor receptor by TNF. Overexpression of ERBB2 has been reported to cause resistance to TNF and other cytotoxic cytokines. In our study we show that the TNF-mediated down-regulation of ERBB2 in pancreatic tumor cells is accompanied by an increase in growth inhibition at low doses of TNF. The simultaneous alteration of the ERBB2/EGFR balance by TNF represents a striking model of cytokine receptor transregulation in the growth control of malignant pancreatic epithelial cells. Images Fig. 1 Fig. 2 Fig. 3 Fig. 4 Fig. 5 PMID:8105469

  5. Cytokines and Prognostic Factors in Epithelial Ovarian Cancer

    PubMed Central

    Jammal, Millena Prata; Martins-Filho, Agrimaldo; Silveira, Thales Parenti; Murta, Eddie Fernando Candido; Nomelini, Rosekeila Simões

    2016-01-01

    INTRODUCTION Ovarian cancer has a high mortality and delayed diagnosis. Inflammation is a risk factor for ovarian cancer, and the inflammatory response is involved in almost all stages of tumor development. Immunohistochemical staining in stroma and epithelium of a panel of cytokines in benign and malignant ovarian neoplasm was evaluated. In addition, immunostaining was related to prognostic factors in malignant tumors. METHOD The study group comprised 28 ovarian benign neoplasias and 28 ovarian malignant neoplasms. A panel of cytokines was evaluated by immunohistochemistry (Th1: IL-2 and IL-8; Th2: IL-5, IL-6, and IL-10; and TNFR1). Chi-square test with Yates’ correction was used, which was considered significant if less than 0.05. RESULTS TNFR1, IL-5, and IL-10 had more frequent immunostaining 2/3 in benign neoplasms compared with malignant tumors. Malignant tumors had more frequent immunostaining 2/3 for IL-2 in relation to benign tumors. The immunostaining 0/1 of IL 8 was more frequent in the stroma of benign neoplasms compared with malignant neoplasms. Evaluation of the ovarian cancer stroma showed that histological grade 3 was significantly correlated with staining 2/3 for IL-2 (P = 0.004). Women whose disease-free survival was less than 2.5 years had TNFR1 stromal staining 2/3 (P = 0.03) more frequently. CONCLUSION IL-2 and TNFR1 stromal immunostaining are related prognostic factors in ovarian cancer and can be the target of new therapeutic strategies. PMID:27512342

  6. Oral administration of transforming growth factor-β1 (TGF-β1) protects the immature gut from injury via Smad protein-dependent suppression of epithelial nuclear factor κB (NF-κB) signaling and proinflammatory cytokine production.

    PubMed

    Shiou, Sheng-Ru; Yu, Yueyue; Guo, Yuee; Westerhoff, Maria; Lu, Lei; Petrof, Elaine O; Sun, Jun; Claud, Erika C

    2013-11-29

    Inflammatory immune responses play an important role in mucosal homeostasis and gut diseases. Nuclear factor κB (NF-κB), central to the proinflammatory cascade, is activated in necrotizing enterocolitis (NEC), a devastating condition of intestinal injury with extensive inflammation in premature infants. TGF-β is a strong immune suppressor and a factor in breast milk, which has been shown to be protective against NEC. In an NEC animal model, oral administration of the isoform TGF-β1 activated the downstream effector Smad2 in intestine and significantly reduced NEC incidence. In addition, TGF-β1 suppressed NF-κB activation, maintained levels of the NF-κB inhibitor IκBα in the intestinal epithelium, and systemically decreased serum levels of IL-6 and IFN-γ. The immature human fetal intestinal epithelial cell line H4 was used as a reductionistic model of the immature enterocyte to investigate mechanism. TGF-β1 pretreatment inhibited the TNF-α-induced IκBα phosphorylation that targets the IκBα protein for degradation and inhibited NF-κB activation. Chromatin immunoprecipitation (ChIP) assays demonstrated decreased NF-κB binding to the promoters of IL-6, IL-8, and IκBα in response to TNF-α with TGF-β1 pretreatment. These TGF-β1 effects appear to be mediated through the canonical Smad pathway as silencing of the TGF-β central mediator Smad4 resulted in loss of the TGF-β1 effects. Thus, TGF-β1 is capable of eliciting anti-inflammatory effects by inhibiting NF-κB specifically in the intestinal epithelium as well as by decreasing systemic IL-6 and IFN-γ levels. Oral administration of TGF-β1 therefore can potentially be used to protect against gastrointestinal diseases. PMID:24129565

  7. Growth factors for the treatment of ischemic brain injury (growth factor treatment).

    PubMed

    Larpthaveesarp, Amara; Ferriero, Donna M; Gonzalez, Fernando F

    2015-01-01

    In recent years, growth factor therapy has emerged as a potential treatment for ischemic brain injury. The efficacy of therapies that either directly introduce or stimulate local production of growth factors and their receptors in damaged brain tissue has been tested in a multitude of models for different Central Nervous System (CNS) diseases. These growth factors include erythropoietin (EPO), vascular endothelial growth factor (VEGF), brain-derived neurotrophic factor (BDNF), and insulin-like growth factor (IGF-1), among others. Despite the promise shown in animal models, the particular growth factors that should be used to maximize both brain protection and repair, and the therapeutic critical period, are not well defined. We will review current pre-clinical and clinical evidence for growth factor therapies in treating different causes of brain injury, as well as issues to be addressed prior to application in humans. PMID:25942688

  8. An epithelial marker promoter induction screen identifies histone deacetylase inhibitors to restore epithelial differentiation and abolishes anchorage independence growth in cancers

    PubMed Central

    Tang, H M; Kuay, K T; Koh, P F; Asad, M; Tan, T Z; Chung, V Y; Lee, S C; Thiery, J P; Huang, RY-J

    2016-01-01

    Epithelial–mesenchymal transition (EMT), a crucial mechanism in development, mediates aggressiveness during carcinoma progression and therapeutic refractoriness. The reversibility of EMT makes it an attractive strategy in designing novel therapeutic approaches. Therefore, drug discovery pipelines for EMT reversal are in need to discover emerging classes of compounds. Here, we outline a pre-clinical drug screening platform for EMT reversal that consists of three phases of drug discovery and validation. From the Phase 1 epithelial marker promoter induction (EpI) screen on a library consisting of compounds being approved by Food and Drug Administration (FDA), Vorinostat (SAHA), a histone deacetylase inhibitor (HDACi), is identified to exert EMT reversal effects by restoring the expression of an epithelial marker, E-cadherin. An expanded screen on 41 HDACi further identifies 28 compounds, such as class I-specific HDACi Mocetinosat, Entinostat and CI994, to restore E-cadherin and ErbB3 expressions in ovarian, pancreatic and bladder carcinoma cells. Mocetinostat is the most potent HDACi to restore epithelial differentiation with the lowest concentration required for 50% induction of epithelial promoter activity (EpIC-50).The HDACi exerts paradoxical effects on EMT transcriptional factors such as SNAI and ZEB family and the effects are context-dependent in epithelial- and mesenchymal-like cells. In vitro functional studies further show that HDACi induced significant increase in anoikis and decrease in spheroid formation in ovarian and bladder carcinoma cells with mesenchymal features. This study demonstrates a robust drug screening pipeline for the discovery of compounds capable of restoring epithelial differentiation that lead to significant functional lethality. PMID:27551531

  9. The microRNA bantam functions in epithelial cells to regulate scaling growth of dendrite arbors in Drosophila sensory neurons

    PubMed Central

    Parrish, Jay Z.; Xu, Peizhang; Kim, Charles C.; Jan, Lily Yeh; Jan, Yuh Nung

    2009-01-01

    Summary In addition to establishing dendritic coverage of the receptive field, neurons need to adjust their dendritic arbors to match changes of the receptive field. Here we show that dendrite arborization (da) sensory neurons establish dendritic coverage of the body wall early in Drosophila larval development and then grow in precise proportion to their substrate, the underlying body wall epithelium, as the larva more than triples in length. This phenomenon, referred to as scaling growth of dendrites, requires the function of the microRNA (miRNA) bantam (ban) in the epithelial cells rather than the da neurons themselves. We further show that ban in epithelial cells dampens Akt kinase activity in adjacent neurons to influence dendrite growth. This signaling between epithelial cells and neurons receiving sensory input from the body wall synchronizes their growth to ensure proper dendritic coverage of the receptive field. PMID:19778508

  10. The CDX2 transcription factor regulates furin expression during intestinal epithelial cell differentiation.

    PubMed

    Gendron, Fernand-Pierre; Mongrain, Sébastien; Laprise, Patrick; McMahon, Stéphanie; Dubois, Claire M; Blais, Mylène; Asselin, Claude; Rivard, Nathalie

    2006-02-01

    CDX2, a member of the caudal family of transcription factors, is involved in enterocyte lineage specification. CDX2 activates many intestine-specific genes, such as sucrase-isomaltase and lactase-phlorizin hydrolase (LPH), and adhesion proteins, namely, LI-cadherin and claudin-2. In this study, we show that the proprotein convertase furin, involved in proteolytic maturation of proprotein substrates including LPH and cell surface proteins, is a CDX2 target. Indeed, expression of the rat furin homolog was induced 1.5-fold, as determined by microarray experiments that compared control with CDX2-expressing intestinal epithelial cells (IEC-6). As determined by transient transfection assays in Caco-2/15 cells, the furin P1 promoter 1.3-kb fragment between SacI and NheI was essential for CDX2 transcriptional activation. Electrophoretic mobility shift/supershift assays followed by site-specific mutagenesis and chromatin immunoprecipitation identified the CDX DNA-binding site (CBS)2 sequence from nt -1827 to -1821 as the major CBS involved in furin P1 promoter activation. Increased furin mRNA and protein expression correlated with both CDX2 expression and intestinal epithelial cell differentiation. In addition, furin mRNAs were detected predominantly in differentiated epithelial cells of the villus, as determined by in situ hybridization. Treatment of Caco-2/15 cells with a furin inhibitor led to inhibition of LPH activity. Morphological differentiation of enterocyte-like features in Caco-2/15 such as epithelial cell polarity and brush-border formation were strongly attenuated by furin inhibition. These results suggest that CDX2 regulates furin expression in intestinal epithelial cells. Furin may be important in modulating the maturation and/or activation of key factors involved in enterocyte differentiation. PMID:16239403

  11. Matriptase activation connects tissue factor-dependent coagulation initiation to epithelial proteolysis and signaling.

    PubMed

    Le Gall, Sylvain M; Szabo, Roman; Lee, Melody; Kirchhofer, Daniel; Craik, Charles S; Bugge, Thomas H; Camerer, Eric

    2016-06-23

    The coagulation cascade is designed to sense tissue injury by physical separation of the membrane-anchored cofactor tissue factor (TF) from inactive precursors of coagulation proteases circulating in plasma. Once TF on epithelial and other extravascular cells is exposed to plasma, sequential activation of coagulation proteases coordinates hemostasis and contributes to host defense and tissue repair. Membrane-anchored serine proteases (MASPs) play critical roles in the development and homeostasis of epithelial barrier tissues; how MASPs are activated in mature epithelia is unknown. We here report that proteases of the extrinsic pathway of blood coagulation transactivate the MASP matriptase, thus connecting coagulation initiation to epithelial proteolysis and signaling. Exposure of TF-expressing cells to factors (F) VIIa and Xa triggered the conversion of latent pro-matriptase to an active protease, which in turn cleaved the pericellular substrates protease-activated receptor-2 (PAR2) and pro-urokinase. An activation pathway-selective PAR2 mutant resistant to direct cleavage by TF:FVIIa and FXa was activated by these proteases when cells co-expressed pro-matriptase, and matriptase transactivation was necessary for efficient cleavage and activation of wild-type PAR2 by physiological concentrations of TF:FVIIa and FXa. The coagulation initiation complex induced rapid and prolonged enhancement of the barrier function of epithelial monolayers that was dependent on matriptase transactivation and PAR2 signaling. These observations suggest that the coagulation cascade engages matriptase to help coordinate epithelial defense and repair programs after injury or infection, and that matriptase may contribute to TF-driven pathogenesis in cancer and inflammation. PMID:27114461

  12. Dendritic cell-derived tumor necrosis factor α modifies airway epithelial cell responses.

    PubMed

    Lutfi, R; Ledford, J R; Zhou, P; Lewkowich, I P; Page, K

    2012-01-01

    Mucosal dendritic cells (DC) are intimately associated with the airway epithelium and thus are ideally situated to be first responders to pathogens. We hypothesize that DC drive innate immune responses through early release of tumor necrosis factor (TNF) α, which drives airway epithelial cell responses. In a mouse model, TNFα release was significantly increased following a single exposure to German cockroach (GC) frass, an event independent of neutrophil recruitment into the airways. While lung epithelial cells and alveolar macrophages failed to release TNFα following GC frass exposure, bone marrow-derived DC (BMDC) produced substantial amounts of TNFα suggesting their importance as early responding cells. This was confirmed by flow cytometry of pulmonary myeloid DC. Addition of GC frass-pulsed BMDC or conditioned media from GC frass-pulsed BMDC to primary mouse tracheal epithelial cells (MTEC) or MLE-15 cells induced chemokine (C-C) motif ligand (CCL) 20 and granulocyte macrophage (GM) colony-stimulating factor (CSF), both of which are important for DC recruitment, survival and differentiation. Importantly, DC do not produce CCL20 or GM-CSF following allergen exposure. Blocking TNFα receptor 1 (TNFR1) completely abolished chemokine production, suggesting that BMDC-derived TNFα induced airway epithelial cell activation and enhancement of the innate immune response. Lastly, blocking TNFR1 in vivo resulted in significantly decreased CCL20 and GM-CSF production in the lungs of mice. Together, our data strongly suggest that DC-derived TNFα plays a crucial role in the initiation of innate immune responses through the modification of airway epithelial cell responses. PMID:22517116

  13. Growth factor involvement in tension-induced skeletal muscle growth

    NASA Technical Reports Server (NTRS)

    Vandenburgh, H. H.

    1987-01-01

    Muscle tissue culture techniques were developed to grow skeletal myofibers which differentiate into more adult-like myofibers. Mechanical simulation studies of these muscle cells in a newly developed mechanical cell simulator can now be performed to study growth processes in skeletal muscle. Conditions in the mechanical cell simulator were defined where mechanical activity can either prevent muscle wasting or stimulate muscle growth. The role of endogenous and exogenous growth factors in tension-induced muscle growth is being investigated under the defined conditions of tissue culture.

  14. Tumor necrosis factor-alpha is expressed by glomerular visceral epithelial cells in human membranous nephropathy.

    PubMed Central

    Neale, T. J.; Rüger, B. M.; Macaulay, H.; Dunbar, P. R.; Hasan, Q.; Bourke, A.; Murray-McIntosh, R. P.; Kitching, A. R.

    1995-01-01

    The role of tumor necrosis factor alpha (TNF-alpha) was examined in biopsy-proven glomerulonephritis by immunohistochemistry, in situ hybridization, immunogold electron microscopy, immunoassay in serum and urine, and urinary immunoblot. Striking glomerular capillary wall and visceral glomerular epithelial cell TNF-alpha protein staining was observed in all cases of membranous nephropathy and membranous lupus nephropathy. Staining was less frequently observed in crescentic glomerulonephritis and in isolated cases of other histological subtypes of glomerulonephritis, usually in association with glomerular macrophages. By immunogold electron microscopy TNF-alpha was localized in membranous nephropathy within the visceral glomerular epithelial cells, and also in the glomerular basement membrane, especially in relation to immune deposits. In situ hybridization localized TNF-alpha mRNA exclusively to glomerular epithelial cells in all biopsies with membranous morphology but not in other histological subtypes. Concentrations of TNF-alpha were significantly increased compared with normal controls in the urine of patients with membranous nephropathy and with crescentic glomerulonephritis. The expression of TNF-alpha by glomerular epithelial cells exclusively and universally in biopsies showing a membranous morphology strongly suggests this cytokine has a role in the pathogenesis of membranous nephropathy. Images Figure 1 Figure 2 Figure 3 Figure 5 PMID:7778683

  15. Mesenchymal to Epithelial Transition Induced by Reprogramming Factors Attenuates the Malignancy of Cancer Cells

    PubMed Central

    Takaishi, Mikiro; Tarutani, Masahito; Takeda, Junji; Sano, Shigetoshi

    2016-01-01

    Epithelial to mesenchymal transition (EMT) is a biological process of metastatic cancer. However, an effective anticancer therapy that directly targets the EMT program has not yet been discovered. Recent studies have indicated that mesenchymal to epithelial transition (MET), the reverse phenomenon of EMT, is observed in fibroblasts during the generation of induced pluripotent stem cells. In the present study, we investigated the effects of reprogramming factors (RFs) on squamous cell carcinoma (SCC) cells. RFs-introduced cancer cells (RICs) demonstrated the enhanced epithelial characteristics in morphology with altered expression of mRNA and microRNAs. The motility and invasive activities of RICs in vitro were significantly reduced. Furthermore, xenografts of RICs exhibited no lymph node metastasis, whereas metastasis was detected in parental SCC-inoculated mice. Thus, we concluded that RICs regained epithelial properties through MET and showed reduced cancer malignancy in vitro and in vivo. Therefore, the understanding of the MET process in cancer cells by introduction of RFs may lead to the designing of a novel anticancer strategy. PMID:27258152

  16. Environmental factors influencing growth and pubertal development.

    PubMed Central

    Delemarre-van de Waal, H A

    1993-01-01

    Postnatal growth is based on hereditary signals and environmental factors in a complex regulatory network. Each factor must be in an optimal state for normal growth of the child. Fetal conditions may also have consequences on postnatal height. Intrauterine growth retardation can be recovered postnatally, although postnatal growth remains depressed in about one-third of cases. After birth, the environment may exert either a positive or negative effect on growth. In underdeveloped countries, malnutrition plays a major role in inhibiting the growth process. Children from families of higher socioeconomic classes are taller than their coevals in the lower socioeconomic groups. Urbanization also has a positive effect on growth. Better child care is supported by sufficient food supply, appropriate health and sanitation services, and a higher level of education. Over the last century, these factors have induced a taller stature and a more rapid maturity in Europe, North America, and Australia; a phenomenon which has been referred to as "the secular trend" in growth. Recently, a secular trend has also been reported in some developing countries. Although urbanization in general appears to be associated with better conditions of living, this is not the case in the slums of South America or in Africa where rural children are better off than children living in the poor cities. This paper describes in more detail the different hereditary and environmental factors that act during the fetal period and postnatally, and which play a role in human growth and pubertal development. PMID:8243404

  17. Vascular growth factors in neuropsychiatry

    PubMed Central

    Newton, Samuel S.; Fournier, Neil M.; Duman, Ronald S.

    2014-01-01

    Recent advances in understanding the cellular and molecular basis of psychiatric illnesses have shed light on the important role played by trophic factors in modulating functional parameters associated with disease causality and drug action. Disease mechanisms are now thought to involve multiple cell types, including neurons and endothelial cells. These functionally distinct but interactively coupled cell types engage in cellular cross talk via shared and common signaling molecules. Dysregulation in their cellular signaling pathways influences brain function and alters behavioral performance. Multifunctional trophic factors such as VEGF and EPO that possess both neurotrophic and angiogenic actions are of particular interest due to their ability to rescue structural and plasticity deficits in neurons and vasculature. Obtaining insight into the behavioral, cellular and molecular actions of multi-functional trophic factors has the potential to open new and transformative therapeutic approaches. PMID:23475069

  18. Inhibition of hypoxia inducible factor-1α downregulates the expression of epithelial to mesenchymal transition early marker proteins without undermining cell survival in hypoxic lens epithelial cells

    PubMed Central

    Neelam, Sudha; Brooks, Morgan M.

    2015-01-01

    Purpose The purpose of this study was to identify potential therapeutic strategies to slow down or prevent the expression of early-onset epithelial to mesenchymal transition (EMT) marker proteins (fibronectin and alpha smooth muscle actin, α-SMA) without sacrificing the synthesis and accumulation of the prosurvival protein vascular endothelial growth factor (VEGF) in cultured virally transformed human lens epithelial (HLE) cells. Methods HLE-B3 cells, maintained in a continuous hypoxic environment (1% oxygen), were treated with SB216763, a specific inhibitor of glycogen synthase kinase-3β (GSK-3β) catalytic activity. Western blot analysis was employed to detect the cytoplasmic and nuclear levels of β-catenin, as well as the total lysate content of fibronectin and α-SMA. Enzyme-linked immunosorbent assay (ELISA) was used to measure the levels of VEGF in cell culture medium. A hypoxia-inducible factor-1α (HIF-1α) translation inhibitor and an HIF-2α translation inhibitor were independently employed to evaluate the effect of hypoxia inducible factor inhibition on EMT marker protein and VEGF expression. XAV932 was used to assess the suppression of nuclear β-catenin and its downstream effect on EMT marker proteins and VEGF expression. Results SB216763-treated HLE-B3 cells caused marked inhibition of GSK-3β activity prompting a significant increase in the translocation of cytoplasmic β-catenin to the nucleus. The enhancement of nuclear β-catenin looked as if it positively correlated with a significant increase in the basal expression of VEGF as well as increased expression of fibronectin and α-SMA. In conjunction with SB216763, coadministration of an HIF-1α translation inhibitor, but not an HIF-2α translation inhibitor, markedly suppressed the expression of fibronectin and α-SMA without affecting VEGF levels. Treatment with XAV932 significantly reduced the level of nuclear β-catenin, but the levels of neither the EMT marker proteins nor VEGF were changed

  19. New Clue Found to Growth Factor Action.

    ERIC Educational Resources Information Center

    Hoffman, Michelle

    1991-01-01

    Discussed is the discovery which may help to explain epidermal growth factor effects on the cell skeleton. The role of a protein called profilin in the regulation of the microfilament system is described. (CW)

  20. Expression of growth factors in Dictyostelium discoideum.

    PubMed

    Asgari, S; Arun, S; Slade, M B; Marshall, J; Williams, K L; Wheldrake, J F

    2001-07-01

    Growth factors and their binding proteins are important proteins regulating mammalian cell proliferation and differentiation so there is considerable interest in producing them as recombinant proteins, especially in hosts that do not already produce a complex mixture of growth factors. Many growth factors require post-translational modifications making them unsuitable for production in Escherichia coli or other prokaryotes. Since several expression vector systems have been recently developed for foreign protein production in the cellular slime mould, Dictyostelium discoideum, we attempted to use two of these systems to express human insulin-like growth factor binding protein 6 (hIGFBP6) and bovine beta-cellulin (bBTC) as secreted proteins. Although both proteins were successfully produced in stably transformed amoebae, no secretion was detected in spite of several attempts to facilitate this occurring. PMID:11361083

  1. [T-LYMPHOCYTES AND TISSUE GROWTH FACTORS].

    PubMed

    Tishevskaya, N V; Gevorkyan, N M; Kozlova, N I

    2015-08-01

    Lympnoici regulation, in aciaition to ensuring tne protection of tne antigen, is aimecl at maintaining a qualitative, quantitative, structural and functional integrity of the body. T-lymphocytes and growth factors are involved in cell proliferation, differentiation, and tissue and organ regeneration. Lymphocyte's, sensitivity to homeostasis changes and their morphogenetic function are connected with a large number of receptors to bioactive substances and with their ability to syn- thesize and secrete hormones and tissue growth factors. At the same time tissue growth factors are involved in the development of thymocytes, in the differentiation of T helper and cytotoxic lymphocytes. Growth factors modulate the functions of Thl, Th2, Treg, Thl7, Th9. The important aspects of the interaction of T cells and EGF, TGF-P, FGF, VEGF, PlGF, HGF/SF in normal and pathological conditions are shown in this review. PMID:26591583

  2. The pH-Responsive PacC Transcription Factor of Aspergillus fumigatus Governs Epithelial Entry and Tissue Invasion during Pulmonary Aspergillosis

    PubMed Central

    Alcazar-Fuoli, Laura; Cairns, Timothy C.; Muñoz, Alberto; Walker, Louise A.; Herbst, Susanne; Safari, Maryam; Cheverton, Angela M.; Chen, Dan; Liu, Hong; Saijo, Shinobu; Fedorova, Natalie D.; Armstrong-James, Darius; Munro, Carol A.; Read, Nick D.; Filler, Scott G.; Espeso, Eduardo A.; Nierman, William C.; Haas, Hubertus; Bignell, Elaine M.

    2014-01-01

    Destruction of the pulmonary epithelium is a major feature of lung diseases caused by the mould pathogen Aspergillus fumigatus. Although it is widely postulated that tissue invasion is governed by fungal proteases, A. fumigatus mutants lacking individual or multiple enzymes remain fully invasive, suggesting a concomitant requirement for other pathogenic activities during host invasion. In this study we discovered, and exploited, a novel, tissue non-invasive, phenotype in A. fumigatus mutants lacking the pH-responsive transcription factor PacC. Our study revealed a novel mode of epithelial entry, occurring in a cell wall-dependent manner prior to protease production, and via the Dectin-1 β-glucan receptor. ΔpacC mutants are defective in both contact-mediated epithelial entry and protease expression, and significantly attenuated for pathogenicity in leukopenic mice. We combined murine infection modelling, in vivo transcriptomics, and in vitro infections of human alveolar epithelia, to delineate two major, and sequentially acting, PacC-dependent processes impacting epithelial integrity in vitro and tissue invasion in the whole animal. We demonstrate that A. fumigatus spores and germlings are internalised by epithelial cells in a contact-, actin-, cell wall- and Dectin-1 dependent manner and ΔpacC mutants, which aberrantly remodel the cell wall during germinative growth, are unable to gain entry into epithelial cells, both in vitro and in vivo. We further show that PacC acts as a global transcriptional regulator of secreted molecules during growth in the leukopenic mammalian lung, and profile the full cohort of secreted gene products expressed during invasive infection. Our study reveals a combinatorial mode of tissue entry dependent upon sequential, and mechanistically distinct, perturbations of the pulmonary epithelium and demonstrates, for the first time a protective role for Dectin-1 blockade in epithelial defences. Infecting ΔpacC mutants are hypersensitive to

  3. Predictive factors for intrauterine growth restriction

    PubMed Central

    Albu, AR; Anca, AF; Horhoianu, VV; Horhoianu, IA

    2014-01-01

    Abstract Reduced fetal growth is seen in about 10% of the pregnancies but only a minority has a pathological background and is known as intrauterine growth restriction or fetal growth restriction (IUGR / FGR). Increased fetal and neonatal mortality and morbidity as well as adult pathologic conditions are often associated to IUGR. Risk factors for IUGR are easy to assess but have poor predictive value. For the diagnostic purpose, biochemical serum markers, ultrasound and Doppler study of uterine and spiral arteries, placental volume and vascularization, first trimester growth pattern are object of assessment today. Modern evaluations propose combined algorithms using these strategies, all with the goal of a better prediction of risk pregnancies. Abbreviations: SGA = small for gestational age; IUGR = intrauterine growth restriction; FGR = fetal growth restriction; IUFD = intrauterine fetal demise; HIV = human immunodeficiency virus; PAPP-A = pregnancy associated plasmatic protein A; β-hCG = beta human chorionic gonadotropin; MoM = multiple of median; ADAM-12 = A-disintegrin and metalloprotease 12; PP-13 = placental protein 13; VEGF = vascular endothelial growth factor; PlGF = placental growth factor; sFlt-1 = soluble fms-like tyrosine kinase-1; UAD = uterine arteries Doppler ultrasound; RI = resistence index; PI = pulsatility index; VOCAL = Virtual Organ Computer–Aided Analysis software; VI = vascularization index; FI = flow index; VFI = vascularization flow index; PQ = placental quotient PMID:25408721

  4. Organic growth factor requirements of some yeasts.

    PubMed

    Madan, M; Gulati, N

    1980-01-01

    Some sporogenous yeasts (Brettanomyces bruxellensis, Debaryomyces hansenii, Hansenula ciferrii, Hansenula polymorpha, Pichia polymorpha, Saccharomycopsis guttulata, and Saccharomyces chevalieri), isolated from various fruits have been examined for their organic growth factor requisites. H. ciferrii was completely deficient in thiamine, biotin, inositol, riboflavin, niacin, and partially deficient in pantothenic acid. It required an external supply of 0.1-1.0 ppm thiamine, 0.01-0.1 ppm biotin, 10.0 ppm inositol, 0.10 ppm niacin and riboflavin for its optimum growth. H. polymorpha showed partial deficiency only in xanthine. P. polymorpha gave indications of partial deficiencies in thiamine and biotin. S. guttulata was completely deficient in biotin, and partially deficient in adenine sulphate. It required 0.01 ppm biotin for optimum growth. S chevalieri was completely deficient in pyridoxine and partially deficient in thiamine. It required 0.1 ppm pyridoxine for maximum growth. D. hansenii and B bruxellensis were auxoautotrophic for the various growth factors studied. PMID:7242379

  5. Effects of steroids on the secretion of immunoregulatory factors by thymic epithelial cell cultures.

    PubMed Central

    Stimson, W H; Crilly, P J

    1981-01-01

    Rat thymic epithelial cells were cultured for 39 days in the presence of various concentrations of oestradiol, testosterone, progesterone and corticosterone and the supernatants assessed for effects on the stimulation of cells from the thymus, bone marrow, lymph nodes and spleen, with several agents. All the steroids, except progesterone, were found to significantly regulate the secretion of immunoregulatory factors by the epithelial cells at physiological levels but the effects were dose dependent. Fractionation of active supernatants indicated that the capacity to enhance or depress cellular proliferation was mainly associated with substances having molecular weights greater than 30,000 or less than 1000, respectively. This study supports the idea that certain steroids can influence the immune response indirectly through the thymus. PMID:7298074

  6. Tumour necrosis factor-alpha up-regulates decay-accelerating factor gene expression in human intestinal epithelial cells.

    PubMed Central

    Andoh, A; Fujiyama, Y; Sumiyoshi, K; Sakumoto, H; Okabe, H; Bamba, T

    1997-01-01

    The increased expression of decay-accelerating factor (DAF) has been detected in intestinal epithelial cells at the inflamed mucosa. In this study, we examined the effects of tumour necrosis factor (TNF)-alpha on DAF expression in three intestinal epithelial cell lines. DAF mRNA expression was evaluated by Northern blot analysis, and DAF protein expression was analysed by biotin labelling and immunoprecipitation. TNF-alpha induced a marked increase in DAF mRNA and protein expression in HT-29, T84 and Caco-2 cells. In HT-29 cells, the effects of TNF-a on DAF mRNA accumulation were observed in a dose-dependent manner; DAF mRNA accumulation reached a maximum at 3-6 hr, and then gradually decreased. These effects of TNF-alpha required de novo protein synthesis. Messenger RNA stability studies suggested that TNF-alpha partially regulated DAF gene expression by a posttranscriptional mechanism. Moreover, the combination of TNF-alpha and interleukin (IL)-4 induced an additive increase in DAF mRNA accumulation in HT-29 and T84 cells. In human intestinal epithelial cells, TNF-alpha acts as a potent inducer of DAF mRNA expression, indicating an important role for TNF-alpha in the regulation of DAF expression at the inflamed mucosa. Images Figure 1 Figure 2 Figure 3 Figure 4 Figure 6 Figure 7 Figure 8 PMID:9155641

  7. Dpysl4 Is Involved in Tooth Germ Morphogenesis through Growth Regulation, Polarization and Differentiation of Dental Epithelial Cells

    PubMed Central

    Yasukawa, Masato; Ishida, Kentaro; Yuge, Yohei; Hanaoka, Mai; Minami, Yoko; Ogawa, Miho; Sasaki, Takashi; Saito, Masahiro; Tsuji, Takashi

    2013-01-01

    Dihydropyrimidinase-related protein 4 (Dpysl4) is a known regulator of hippocampal neuron development. Here, we report that Dpysl4 is involved in growth regulation, polarization and differentiation of dental epithelial cells during tooth germ morphogenesis. A reduction in Dpysl4 gene expression in the tooth germ produced a loss of ameloblasts, resulting in the decrease of synthesis and secretion of enamel. The inhibition of Dpysl4 gene expression led to promotion of cell proliferation of inner enamel epithelial cells and inhibition of the differentiation of these cells into pre-ameloblasts, which was confirmed by analyzing cell polarization, columnar cell structure formation and the expression of ameloblast marker genes. By contrast, overexpression of Dpysl4 in dental epithelial cells induces inhibition of growth and increases the expression of the inner enamel epithelial cell marker gene, Msx2. These findings suggest that Dpysl4 plays essential roles in tooth germ morphogenesis through the regulation of dental epithelial cell proliferation, cell polarization and differentiation. PMID:23630450

  8. The Oct1 transcription factor and epithelial malignancies: Old protein learns new tricks.

    PubMed

    Vázquez-Arreguín, Karina; Tantin, Dean

    2016-06-01

    The metazoan-specific POU domain transcription factor family comprises activities underpinning developmental processes such as embryonic pluripotency and neuronal specification. Some POU family proteins efficiently bind an 8-bp DNA element known as the octamer motif. These proteins are known as Oct transcription factors. Oct1/POU2F1 is the only widely expressed POU factor. Unlike other POU factors it controls no specific developmental or organ system. Oct1 was originally described to operate at target genes associated with proliferation and immune modulation, but more recent results additionally identify targets associated with oxidative and cytotoxic stress resistance, metabolic regulation, stem cell function and other unexpected processes. Oct1 is pro-oncogenic in multiple contexts, and several recent reports provide broad evidence that Oct1 has prognostic and therapeutic value in multiple epithelial tumor settings. This review focuses on established and emerging roles of Oct1 in epithelial tumors, with an emphasis on mechanisms of transcription regulation by Oct1 that may underpin these findings. This article is part of a Special Issue entitled: The Oct Transcription Factor Family, edited by Dr. Dean Tantin. PMID:26877236

  9. ATP differentially upregulates fibroblast growth factor 2 and transforming growth factor α in neonatal and adult mice: effect on neuroproliferation.

    PubMed

    Jia, C; Cussen, A R; Hegg, C C

    2011-03-17

    Multiple neurotrophic factors play a role in proliferation, differentiation and survival in the olfactory epithelium (OE); however, the signaling cascade has not been fully elucidated. We tested the hypotheses that ATP induces the synthesis and secretion of two neurotrophic factors, fibroblast growth factor 2 (FGF2) and transforming growth factor alpha (TGFα), and that these neurotrophic factors have a role in inducing proliferation. Protein levels of FGF2 and TGFα were increased 20 h post-intranasal instillation of ATP compared to vehicle control in adult Swiss Webster mice. Pre-intranasal treatment with purinergic receptor antagonist pyridoxal-phosphate-6-azophenyl-20,40-disulfonic acid (PPADS) significantly blocked this ATP-induced increase, indicating that upregulation of FGF2 and TGFα expression is mediated by purinergic receptor activation. However, in neonatal mouse, intranasal instillation of ATP significantly increased the protein levels of FGF2, but not TGFα. Likewise, ATP evoked the secretion of FGF2, but not TGFα, from neonatal mouse olfactory epithelial slices and PPADS significantly blocked ATP-evoked FGF2 release. To determine the role of FGF2 and TGFα in inducing proliferation, 5-bromo-2-deoxyuridine (BrdU) incorporation was examined in adult olfactory epithelium. Intranasal treatment with FGF receptor inhibitor PD173074 or epidermal growth factor receptor inhibitor AG1478 following ATP instillation significantly blocked ATP-induced BrdU incorporation. Collectively, these data demonstrate that ATP induces proliferation in adult mouse olfactory epithelium by promoting FGF2 and TGFα synthesis and activation of their receptors. These data suggest that different mechanisms regulate neurogenesis in neonatal and adult OE, and FGF2 and TGFα may have different roles throughout development. PMID:21187124

  10. Placenta Growth Factor in Diabetic Wound Healing

    PubMed Central

    Cianfarani, Francesca; Zambruno, Giovanna; Brogelli, Laura; Sera, Francesco; Lacal, Pedro Miguel; Pesce, Maurizio; Capogrossi, Maurizio C.; Failla, Cristina Maria; Napolitano, Monica; Odorisio, Teresa

    2006-01-01

    Reduced microcirculation and diminished expression of growth factors contribute to wound healing impairment in diabetes. Placenta growth factor (PlGF), an angiogenic mediator promoting pathophysiological neovascularization, is expressed during cutaneous wound healing and improves wound closure by enhancing angiogenesis. By using streptozotocin-induced diabetic mice, we here demonstrate that PlGF induction is strongly reduced in diabetic wounds. Diabetic transgenic mice overexpressing PlGF in the skin displayed accelerated wound closure compared with diabetic wild-type littermates. Moreover, diabetic wound treatment with an adenovirus vector expressing the human PlGF gene (AdCMV.PlGF) significantly accelerated the healing process compared with wounds treated with a control vector. The analysis of treated wounds showed that PlGF gene transfer improved granulation tissue formation, maturation, and vascularization, as well as monocytes/macrophages local recruitment. Platelet-derived growth factor, fibroblast growth factor-2, and vascular endothelial growth factor mRNA levels were increased in AdCMV.PlGF-treated wounds, possibly enhancing PlGF-mediated effects. Finally, PlGF treatment stimulated cultured dermal fibroblast migration, pointing to a direct role of PlGF in accelerating granulation tissue maturation. In conclusion, our data indicate that reduced PlGF expression contributes to impaired wound healing in diabetes and that PlGF gene transfer to diabetic wounds exerts therapeutic activity by promoting different aspects of the repair process. PMID:17003476

  11. Mortality factor 4 like 1 protein mediates epithelial cell death in a mouse model of pneumonia.

    PubMed

    Zou, Chunbin; Li, Jin; Xiong, Sheng; Chen, Yan; Wu, Qin; Li, Xiuying; Weathington, Nathaniel M; Han, SeungHye; Snavely, Courtney; Chen, Bill B; Mallampalli, Rama K

    2015-10-28

    Unchecked epithelial cell death is fundamental to the pathogenesis of pneumonia. The recognition of unique signaling pathways that preserve epithelial cell viability may present new opportunities for interventional strategies. We describe that mortality factor 4 like 1 (Morf4l1), a protein involved in chromatin remodeling, is constitutively expressed at low levels in the lung because of its continuous degradation mediated by an orphan ubiquitin E3 ligase subunit, Fbxl18. Expression of Morf4l1 increases in humans with pneumonia and is up-regulated in lung epithelia after exposure to Pseudomonas aeruginosa or lipopolysaccharide. In a mouse model of pneumonia induced by P. aeruginosa, Morf4l1 is stabilized by acetylation that protects it from Fbxl18-mediated degradation. After P. aeruginosa infection of mice, overexpression of Morf4l1 resulted in lung epithelial cell death, whereas its depletion restored cell viability. Using in silico modeling and drug-target interaction studies, we identified that the U.S. Food and Drug Administration-approved thrombin inhibitor argatroban is a Morf4l1 antagonist. Argatroban inhibited Morf4l1-dependent histone acetylation, reduced its cytotoxicity, and improved survival of mice with experimental lung injury at doses that had no anticoagulant activity. These studies uncover a previously unrecognized biological mechanism whereby pathogens subvert cell viability by extending the life span of a cytotoxic host protein. Morf4l1 may be a potential molecular target for non-antibiotic pharmacotherapy during severe pulmonary infection. PMID:26511508

  12. IL-13 Augments Compressive Stress-Induced Tissue Factor Expression in Human Airway Epithelial Cells.

    PubMed

    Mitchel, Jennifer A; Antoniak, Silvio; Lee, Joo-Hyeon; Kim, Sae-Hoon; McGill, Maureen; Kasahara, David I; Randell, Scott H; Israel, Elliot; Shore, Stephanie A; Mackman, Nigel; Park, Jin-Ah

    2016-04-01

    Tissue factor (TF) is best known as a cellular initiator of coagulation, but it is also a multifunctional protein that has been implicated in multiple pathophysiologic conditions, including asthma. In the lung, airway epithelial cells express TF, but it is unknown how TF expression is regulated by asthma-associated mediators. We investigated the role of IL-13, a type 2 cytokine, alone and in combination with compressive stress, which mimics asthmatic bronchoconstriction, on TF expression and release of TF-positive extracellular vesicles from primary normal human bronchial epithelial cells. Well-differentiated normal human bronchial epithelial cells were treated with IL-13 and compressive stress, alone and in combination. TF mRNA, protein and activity were measured in the cells and conditioned media. TF was also measured in the bronchoalveolar lavage (BAL) fluid of allergen-challenged mice and patients with asthma. IL-13 and compressive stress increased TF expression, but only compressive stress induced TF-positive extracellular vesicle release. Pretreatment with IL-13 augmented compressive stress-induced TF expression and release. TF protein and activity in BAL fluid were increased in allergen-sensitized and -challenged mice. TF was elevated in the BAL fluid of patients with mild asthma after an allergen challenge. Our in vitro and in vivo data indicate close cooperation between mechanical and inflammatory stimuli on TF expression and release of TF-positive extracellular vesicles in the lungs, which may contribute to pathophysiology of asthma. PMID:26407210

  13. Mortality factor 4 like 1 protein mediates epithelial cell death in a mouse model of pneumonia

    PubMed Central

    Zou, Chunbin; Li, Jin; Xiong, Sheng; Chen, Yan; Wu, Qin; Li, Xiuying; Weathington, Nathaniel M.; Han, Seung Hye; Snavely, Courtney; Chen, Bill B.; Mallampalli, Rama K.

    2016-01-01

    Unchecked epithelial cell death is fundamental to the pathogenesis of pneumonia. The recognition of unique signaling pathways that preserve epithelial cell viability may present new opportunities for interventional strategies. We describe that mortality factor 4 like 1 (Morf4l1), a protein involved in chromatin remodeling, is constitutively expressed at low levels in the lung because of its continuous degradation mediated by an orphan ubiquitin E3 ligase subunit, Fbxl18. Expression of Morf4l1 increases in humans with pneumonia and is up-regulated in lung epithelia after exposure to Pseudomonas aeruginosa or lipopolysaccharide. In a mouse model of pneumonia induced by P. aeruginosa, Morf4l1 is stabilized by acetylation that protects it from Fbxl18-mediated degradation. After P. aeruginosa infection of mice, overexpression of Morf4l1 resulted in lung epithelial cell death, whereas its depletion restored cell viability. Using in silico modeling and drug-target interaction studies, we identified that the U.S. Food and Drug Administration–approved thrombin inhibitor argatroban is a Morf4l1 antagonist. Argatroban inhibited Morf4l1-dependent histone acetylation, reduced its cytotoxicity, and improved survival of mice with experimental lung injury at doses that had no anticoagulant activity. These studies uncover a previously unrecognized biological mechanism whereby pathogens subvert cell viability by extending the life span of a cytotoxic host protein. Morf4l1 may be a potential molecular target for non-antibiotic pharmacotherapy during severe pulmonary infection. PMID:26511508

  14. In vitro small intestinal epithelial cell growth on a nanocomposite polycaprolactone scaffold

    PubMed Central

    Gupta, Ashish; Vara, Dina S.; Punshon, Geoffrey; Sales, Kevin M.; Winslet, Marc C.; Seifalian, Alexander M.

    2009-01-01

    Tissue engineering of the small intestine remains experimental despite worldwide attempts to develop a functional substitute for short bowel syndrome. Most published studies have reported predominant use of PLLA (poly-L-lactide acid)/PGA (polyglycolic acid) copolymer as the scaffold material, and studies have been limited by in vivo experiments. This lack of progress has inspired a fresh perspective and provoked further investigation and development in this field of tissue engineering. In the present paper, we exploit a relatively new nanocomposite of POSS (polyhedral oligomeric silsesquioxane) and PCL [poly(caprolactone-urea)urethane] as a material to develop porous scaffolds using a solvent casting/particulate leaching technique to fabricate porous scaffolds in different pore sizes and porosities. Scaffolds were characterized for pore morphology and porosity using scanning electron microscopy and micro-computed tomography. Rat intestinal epithelial cells were then seeded on to the polymer scaffolds for an in vitro study of cell compatibility and proliferation, which was assessed by Alamar Blue™ and lactate dehydrogenase assays performed for 21 days post-seeding. The results obtained demonstrate that POSS–PCL nanocomposite was produced as a macroporous scaffold with porosity over the range of 40–80% and pore size over the range of 150–250 μm. This scaffold was shown to support epithelial cell proliferation and growth. In conclusion, as a further step in investigating small intestinal tissue engineering, the nanocomposite employed in this study may prove to be a useful alternative to poly(lactic-co-glycolic acid) in the future. PMID:19860739

  15. The Endogenous Cell-Fate Factor Dachshund restrains Prostate Epithelial Cell Migration via Repression of Cytokine Secretion via a CXCL Signaling Module

    PubMed Central

    Chen, Ke; Wu, Kongming; Jiao, Xuanmao; Wang, Liping; Ju, Xiaoming; Wang, Min; Di Sante, Gabriele; Xu, Shaohua; Wang, Qiong; Li, Kevin; Sun, Xin; Xu, Congwen; Li, Zhiping; Casimiro, Mathew C.; Ertel, Adam; Addya, Sankar; McCue, Peter; Lisanti, Michael P.; Wang, Chenguang; Davis, Richard J.; Mardon, Graeme; Pestell, Richard G.

    2015-01-01

    Prostate cancer (PCa) is the second leading form of cancer death in men. In a subset of PCa patients increased chemokine signaling IL-8 and IL-6 correlates with androgen therapy-resistant prostate cancer (CRPC). IL-8 and IL-6 are produced by prostate epithelial cells and promote PCa cell invasion, however the mechanisms restraining prostate epithelial cell cytokine secretion are poorly understood. Herein the cell-fate determinant factor DACH1 inhibited androgen therapy-resistant prostate cancer (CRPC) tumor growth in mice. Using Dach1fl/fl/Probasin-Cre bi-transgenic mice, we show IL-8 and IL-6 secretion was altered ~1000 fold by endogenous Dach1. Endogenous Dach1 is shown to serve as a key endogenous restraint to prostate epithelial cell growth and restrains migration via CXCL signaling. DACH1 inhibited expression, transcription and secretion of the CXCL genes (IL-8, IL-6) by binding to their promoter regulatory regions in chromatin. DACH1 is thus a newly defined determinant of benign and malignant prostate epithelium cellular growth, migration and cytokine abundance in vivo. PMID:25769723

  16. Upregulation of epidermal growth factor receptor 4 in oral leukoplakia

    PubMed Central

    Kobayashi, Hiroshi; Kumagai, Kenichi; Gotoh, Akito; Eguchi, Takanori; Yamada, Hiroyuki; Hamada, Yoshiki; Suzuki, Satsuki; Suzuki, Ryuji

    2013-01-01

    In the present study, we investigate the expression profile of the epidermal growth factor receptor family, which comprises EGFR/ErbB1, HER2/ErbB2, HER3/ErbB3 and HER4/ErbB4 in oral leukoplakia (LP). The expression of four epidermal growth factor receptor (EGFR) family genes and their ligands were measured in LP tissues from 14 patients and compared with levels in 10 patients with oral lichen planus (OLP) and normal oral mucosa (NOM) from 14 healthy donors by real-time polymerase chain reaction (PCR) and immunohistochemistry. Synchronous mRNA coexpression of ErbB1, ErbB2, ErbB3 and ErbB4 was detected in LP lesions. Out of the receptors, only ErbB4 mRNA and protein was more highly expressed in LP compared with NOM tissues. These were strongly expressed by epithelial keratinocytes in LP lesions, as shown by immunohistochemistry. Regarding the ligands, the mRNA of Neuregulin2 and 4 were more highly expressed in OLP compared with NOM tissues. Therefore, enhanced ErbB4 on the keratinocytes and synchronous modulation of EGFR family genes may contribute to the pathogenesis and carcinogenesis of LP. PMID:23492901

  17. Colony Stimulating Factor-1 exerts direct effects on the proliferation and invasiveness of endometrial epithelial cells

    PubMed Central

    Aligeti, Sabitha; Kirma, Nameer B.; Binkely, Peter A; Schenken, Robert S.; Tekmal, Rajeshwar Rao

    2011-01-01

    Although macrophage colony stimulating factor (CSF-1) has been suggested to play a role in part in maintaining the chronic inflammatory response in endometriosis, our previous data suggest that CSF-1 may also play a role in early endometriosis lesion formation. In the current studies, we have examined this possibility and have shown that CSF-1, in an autocrine fashion, has a direct effect on endometrial epithelial cell proliferation and attachment to peritoneal mesothelial cells, early steps in endometriosis lesion formation on the peritoneum. PMID:21481374

  18. Autocrine and paracrine actions of intestinal fibroblast-derived insulin-like growth factors.

    PubMed

    Simmons, J G; Pucilowska, J B; Lund, P K

    1999-04-01

    Paracrine and autocrine actions of the insulin-like growth factors (IGFs) are inferred by local expression within the bowel. CCD-18Co cells, IEC-6 cells, and immunoneutralization were used to analyze whether IGFs have direct autocrine or paracrine effects on proliferation of cultured intestinal fibroblasts and epithelial cells. Growth factor expression was analyzed by ribonuclease protection assay and RT-PCR. Extracellular matrix (ECM) was analyzed for effects on cell proliferation. CCD-18Co cells express IGF-II mRNAs and low levels of IGF-I mRNA. Conditioned medium from CCD-18Co cells (CCD-CM) stimulated proliferation of IEC-6 and CCD-18Co cells. Neutralization of IGF immunoreactivity in CCD-CM reduced but did not abolish this effect. RT-PCR and immunoneutralization demonstrated that other growth factors contribute to mitogenic activity of CCD-CM. Preincubation of CCD-CM with ECM prepared from IEC-6 or CCD-18Co cells reduced its mitogenic activity. ECM from CCD-18Co cells enhanced growth factor-dependent proliferation of IEC-6 cells. IEC-6 cell ECM inhibited IGF-I action on CCD-18Co cells. We conclude that IGF-II is a potent autocrine mitogen for intestinal fibroblasts. IGF-II interacts with other fibroblast-derived growth factors and ECM to stimulate proliferation of intestinal epithelial cells in a paracrine manner. PMID:10198323

  19. Induction of reactive oxygen species generation inhibits epithelial-mesenchymal transition and promotes growth arrest in prostate cancer cells.

    PubMed

    Das, Trinath P; Suman, Suman; Damodaran, Chendil

    2014-07-01

    Oxidative stress is one causative factor of the pathogenesis and aggressiveness of most of the cancer types, including prostate cancer (CaP). A moderate increase in reactive oxygen species (ROS) induces cell proliferation whereas excessive amounts of ROS promote apoptosis. In this study, we explored the pro-oxidant property of 3,9-dihydroxy-2-prenylcoumestan (psoralidin [pso]), a dietary agent, on CaP (PC-3 and C4-2B) cells. Pso greatly induced ROS generation (more than 20-fold) that resulted in the growth inhibition of CaP cells. Overexpression of anti-oxidant enzymes superoxide dismutase 1 (SOD1), SOD2, and catalase, or pretreatment with the pharmacological inhibitor N-acetylcysteine (NAC) significantly attenuated both pso-mediated ROS generation and pso-mediated growth inhibition in CaP cells. Furthermore, pso administration significantly inhibited the migratory and invasive property of CaP cells by decreasing the transcription of β-catenin, and slug, which promote epithelial-mesenchymal transition (EMT), and by concurrently inducing E-cadherin expression in CaP cells. Pso-induced ROS generation in CaP cells resulted in loss of mitochondrial membrane potential, cytochrome-c release, and activation of caspase-3 and -9 and poly (ADP-ribose) polymerase (PARP), which led to apoptosis. On the other hand, overexpression of anti-oxidants rescued pso-mediated effects on CaP cells. These findings suggest that increasing the threshold of intracellular ROS could prevent or treat CaP growth and metastasis. PMID:23475579

  20. Foxn1 Transcription Factor Regulates Wound Healing of Skin through Promoting Epithelial-Mesenchymal Transition.

    PubMed

    Gawronska-Kozak, Barbara; Grabowska, Anna; Kur-Piotrowska, Anna; Kopcewicz, Marta

    2016-01-01

    Transcription factors are key molecules that finely tune gene expression in response to injury. We focused on the role of a transcription factor, Foxn1, whose expression is limited to the skin and thymus epithelium. Our previous studies showed that Foxn1 inactivity in nude mice creates a pro-regenerative environment during skin wound healing. To explore the mechanistic role of Foxn1 in the skin wound healing process, we analyzed post-injured skin tissues from Foxn1::Egfp transgenic and C57BL/6 mice with Western Blotting, qRT-PCR, immunofluorescence and flow cytometric assays. Foxn1 expression in non-injured skin localized to the epidermis and hair follicles. Post-injured skin tissues showed an intense Foxn1-eGFP signal at the wound margin and in leading epithelial tongue, where it co-localized with keratin 16, a marker of activated keratinocytes. This data support the concept that suprabasal keratinocytes, expressing Foxn1, are key cells in the process of re-epithelialization. The occurrence of an epithelial-mesenchymal transition (EMT) was confirmed by high levels of Snail1 and Mmp-9 expression as well as through co-localization of vimentin/E-cadherin-positive cells in dermis tissue at four days post-wounding. Involvement of Foxn1 in the EMT process was verified by co-localization of Foxn1-eGFP cells with Snail1 in histological sections. Flow cytometric analysis showed the increase of double positive E-cadherin/N-cadherin cells within Foxn1-eGFP population of post-wounded skin cells isolates, which corroborated histological and gene expression analyses. Together, our findings indicate that Foxn1 acts as regulator of the skin wound healing process through engagement in re-epithelization and possible involvement in scar formation due to Foxn1 activity during the EMT process. PMID:26938103

  1. Foxn1 Transcription Factor Regulates Wound Healing of Skin through Promoting Epithelial-Mesenchymal Transition

    PubMed Central

    Gawronska-Kozak, Barbara; Grabowska, Anna; Kur-Piotrowska, Anna; Kopcewicz, Marta

    2016-01-01

    Transcription factors are key molecules that finely tune gene expression in response to injury. We focused on the role of a transcription factor, Foxn1, whose expression is limited to the skin and thymus epithelium. Our previous studies showed that Foxn1 inactivity in nude mice creates a pro-regenerative environment during skin wound healing. To explore the mechanistic role of Foxn1 in the skin wound healing process, we analyzed post-injured skin tissues from Foxn1::Egfp transgenic and C57BL/6 mice with Western Blotting, qRT-PCR, immunofluorescence and flow cytometric assays. Foxn1 expression in non-injured skin localized to the epidermis and hair follicles. Post-injured skin tissues showed an intense Foxn1-eGFP signal at the wound margin and in leading epithelial tongue, where it co-localized with keratin 16, a marker of activated keratinocytes. This data support the concept that suprabasal keratinocytes, expressing Foxn1, are key cells in the process of re-epithelialization. The occurrence of an epithelial-mesenchymal transition (EMT) was confirmed by high levels of Snail1 and Mmp-9 expression as well as through co-localization of vimentin/E-cadherin-positive cells in dermis tissue at four days post-wounding. Involvement of Foxn1 in the EMT process was verified by co-localization of Foxn1-eGFP cells with Snail1 in histological sections. Flow cytometric analysis showed the increase of double positive E-cadherin/N-cadherin cells within Foxn1-eGFP population of post-wounded skin cells isolates, which corroborated histological and gene expression analyses. Together, our findings indicate that Foxn1 acts as regulator of the skin wound healing process through engagement in re-epithelization and possible involvement in scar formation due to Foxn1 activity during the EMT process. PMID:26938103

  2. Epidermal Growth Factor and Intestinal Barrier Function.

    PubMed

    Tang, Xiaopeng; Liu, Hu; Yang, Shufen; Li, Zuohua; Zhong, Jinfeng; Fang, Rejun

    2016-01-01

    Epidermal growth factor (EGF) is a 53-amino acid peptide that plays an important role in regulating cell growth, survival, migration, apoptosis, proliferation, and differentiation. In addition, EGF has been established to be an effective intestinal regulator helping to protect intestinal barrier integrity, which was essential for the absorption of nutrients and health in humans and animals. Several researches have demonstrated that EGF via binding to the EGF receptor and subsequent activation of Ras/MAPK, PI3K/AKT, PLC-γ/PKC, and STATS signal pathways regulates intestinal barrier function. In this review, the relationship between epidermal growth factor and intestinal development and intestinal barrier is described, to provide a better understanding of the effects of EGF on intestine development and health. PMID:27524860

  3. Epidermal Growth Factor and Intestinal Barrier Function

    PubMed Central

    Liu, Hu; Yang, Shufen; Li, Zuohua; Zhong, Jinfeng

    2016-01-01

    Epidermal growth factor (EGF) is a 53-amino acid peptide that plays an important role in regulating cell growth, survival, migration, apoptosis, proliferation, and differentiation. In addition, EGF has been established to be an effective intestinal regulator helping to protect intestinal barrier integrity, which was essential for the absorption of nutrients and health in humans and animals. Several researches have demonstrated that EGF via binding to the EGF receptor and subsequent activation of Ras/MAPK, PI3K/AKT, PLC-γ/PKC, and STATS signal pathways regulates intestinal barrier function. In this review, the relationship between epidermal growth factor and intestinal development and intestinal barrier is described, to provide a better understanding of the effects of EGF on intestine development and health. PMID:27524860

  4. Biochemical characterization of the Drosophila dpp protein, a member of the transforming growth factor beta family of growth factors.

    PubMed Central

    Panganiban, G E; Rashka, K E; Neitzel, M D; Hoffmann, F M

    1990-01-01

    The decapentaplegic (dpp) gene of Drosophila melanogaster is required for pattern formation in the embryo and for viability of the epithelial cells in the imaginal disks. The dpp protein product predicted from the DNA sequence is similar to members of a family of growth factors that includes transforming growth factor beta (TGF-beta). We have produced polyclonal antibodies to a recombinant dpp protein made in bacteria and used a metallothionein promoter to express a dpp cDNA in Drosophila S2 cells. Similar to other proteins in the TGF-beta family, the dpp protein produced by the Drosophila cells was proteolytically cleaved, and both portions of the protein were secreted from the cells. The amino-terminal 47-kilodalton (kDa) peptide was found in the medium and in the proteins adhering to the plastic petri dish. The carboxy-terminal peptide, the region with sequence similarity to the active ligand portion of TGF-beta, was found extracellularly as a 30-kDa homodimer. Most of the 30-kDa homodimer was in the S2 cell protein adsorbed onto the surface of the plastic dish. The dpp protein could be released into solution by increased salt concentration and nonionic detergent. Under these conditions, the amino-terminal and carboxy-terminal portions of dpp were not associated in a stable complex. Images PMID:1692958

  5. Hypoxia-induced alveolar epithelial-mesenchymal transition requires mitochondrial ROS and hypoxia-inducible factor 1

    PubMed Central

    Zhou, Guofei; Dada, Laura A.; Wu, Minghua; Kelly, Aileen; Trejo, Humberto; Zhou, Qiyuan; Varga, John

    2009-01-01

    Patients with acute lung injury develop hypoxia, which may lead to lung dysfunction and aberrant tissue repair. Recent studies have suggested that epithelial-mesenchymal transition (EMT) contributes to pulmonary fibrosis. We sought to determine whether hypoxia induces EMT in alveolar epithelial cells (AEC). We found that hypoxia induced the expression of α-smooth muscle actin (α-SMA) and vimentin and decreased the expression of E-cadherin in transformed and primary human, rat, and mouse AEC, suggesting that hypoxia induces EMT in AEC. Both severe hypoxia and moderate hypoxia induced EMT. The reactive oxygen species (ROS) scavenger Euk-134 prevented hypoxia-induced EMT. Moreover, hypoxia-induced expression of α-SMA and vimentin was prevented in mitochondria-deficient ρ0 cells, which are incapable of ROS production during hypoxia. CoCl2 and dimethyloxaloylglycine, two compounds that stabilize hypoxia-inducible factor (HIF)-α under normoxia, failed to induce α-SMA expression in AEC. Furthermore, overexpression of constitutively active HIF-1α did not induce α-SMA. However, loss of HIF-1α or HIF-2α abolished induction of α-SMA mRNA during hypoxia. Hypoxia increased the levels of transforming growth factor (TGF)-β1, and preincubation of AEC with SB431542, an inhibitor of the TGF-β1 type I receptor kinase, prevented the hypoxia-induced EMT, suggesting that the process was TGF-β1 dependent. Furthermore, both ROS and HIF-α were necessary for hypoxia-induced TGF-β1 upregulation. Accordingly, we have provided evidence that hypoxia induces EMT of AEC through mitochondrial ROS, HIF, and endogenous TGF-β1 signaling. PMID:19801454

  6. Tissue Engineering Using Transfected Growth-Factor Genes

    NASA Technical Reports Server (NTRS)

    Madry, Henning; Langer, Robert S.; Freed, Lisa E.; Trippel, Stephen; Vunjak-Novakovic, Gordana

    2005-01-01

    A method of growing bioengineered tissues includes, as a major component, the use of mammalian cells that have been transfected with genes for secretion of regulator and growth-factor substances. In a typical application, one either seeds the cells onto an artificial matrix made of a synthetic or natural biocompatible material, or else one cultures the cells until they secrete a desired amount of an extracellular matrix. If such a bioengineered tissue construct is to be used for surgical replacement of injured tissue, then the cells should preferably be the patient s own cells or, if not, at least cells matched to the patient s cells according to a human-leucocyteantigen (HLA) test. The bioengineered tissue construct is typically implanted in the patient's injured natural tissue, wherein the growth-factor genes enhance metabolic functions that promote the in vitro development of functional tissue constructs and their integration with native tissues. If the matrix is biodegradable, then one of the results of metabolism could be absorption of the matrix and replacement of the matrix with tissue formed at least partly by the transfected cells. The method was developed for articular chondrocytes but can (at least in principle) be extended to a variety of cell types and biocompatible matrix materials, including ones that have been exploited in prior tissue-engineering methods. Examples of cell types include chondrocytes, hepatocytes, islet cells, nerve cells, muscle cells, other organ cells, bone- and cartilage-forming cells, epithelial and endothelial cells, connective- tissue stem cells, mesodermal stem cells, and cells of the liver and the pancreas. Cells can be obtained from cell-line cultures, biopsies, and tissue banks. Genes, molecules, or nucleic acids that secrete factors that influence the growth of cells, the production of extracellular matrix material, and other cell functions can be inserted in cells by any of a variety of standard transfection techniques.

  7. A Homeobox Gene Related to Drosophila Distal-Less Promotes Ovarian Tumorigenicity by Inducing Expression of Vascular Endothelial Growth Factor and Fibroblast Growth Factor-2

    PubMed Central

    Hara, Fumikata; Samuel, Shaija; Liu, Jinsong; Rosen, Daniel; Langley, Robert R.; Naora, Honami

    2007-01-01

    Homeobox genes control developmental patterning and are increasingly being found to be deregulated in tumors. The DLX4 homeobox gene maps to the 17q21.3-q22 region that is amplified in some epithelial ovarian cancers. Because amplification of this region correlates with poor prognosis, we investigated whether DLX4 overexpression contributes to aggressive behavior of this disease. DLX4 was not detected in normal ovary and cystadenomas, whereas its expression in ovarian carcinomas was strongly associated with high tumor grade and advanced disease stage. Overexpression of DLX4 in ovarian cancer cells promoted growth in low serum and colony formation. Imaging of mice bearing intraperitoneal tumors revealed that DLX4 overexpression substantially increased tumor burden. Tumors that overexpressed DLX4 were more vascularized than vector-control tumors. Conditioned medium of DLX4-overexpressing tumor cells was more effective than medium conditioned by vector-control cells in stimulating endothelial cell growth. These observations were associated with the ability of DLX4 to induce expression of vascular endothelial growth factor as well as intracellular and secreted isoforms of fibroblast growth factor-2. Moreover, increased levels of these fibroblast growth factor-2 isoforms induced vascular endothelial growth factor expression in tumor cells. This study reveals a novel role for a homeobox gene in ovarian tumorigenicity by its induction of a proangiogenic, growth-stimulatory molecular program. PMID:17456765

  8. A homeobox gene related to Drosophila distal-less promotes ovarian tumorigenicity by inducing expression of vascular endothelial growth factor and fibroblast growth factor-2.

    PubMed

    Hara, Fumikata; Samuel, Shaija; Liu, Jinsong; Rosen, Daniel; Langley, Robert R; Naora, Honami

    2007-05-01

    Homeobox genes control developmental patterning and are increasingly being found to be deregulated in tumors. The DLX4 homeobox gene maps to the 17q21.3-q22 region that is amplified in some epithelial ovarian cancers. Because amplification of this region correlates with poor prognosis, we investigated whether DLX4 overexpression contributes to aggressive behavior of this disease. DLX4 was not detected in normal ovary and cystadenomas, whereas its expression in ovarian carcinomas was strongly associated with high tumor grade and advanced disease stage. Overexpression of DLX4 in ovarian cancer cells promoted growth in low serum and colony formation. Imaging of mice bearing intraperitoneal tumors revealed that DLX4 overexpression substantially increased tumor burden. Tumors that overexpressed DLX4 were more vascularized than vector-control tumors. Conditioned medium of DLX4-overexpressing tumor cells was more effective than medium conditioned by vector-control cells in stimulating endothelial cell growth. These observations were associated with the ability of DLX4 to induce expression of vascular endothelial growth factor as well as intracellular and secreted isoforms of fibroblast growth factor-2. Moreover, increased levels of these fibroblast growth factor-2 isoforms induced vascular endothelial growth factor expression in tumor cells. This study reveals a novel role for a homeobox gene in ovarian tumorigenicity by its induction of a proangiogenic, growth-stimulatory molecular program. PMID:17456765

  9. RasGRP1 opposes proliferative EGFR–SOS1–Ras signals and restricts intestinal epithelial cell growth

    PubMed Central

    Depeille, Philippe; Henricks, Linda M.; van de Ven, Robert A. H.; Lemmens, Ed; Wang, Chih-Yang; Matli, Mary; Werb, Zena; Haigis, Kevin M.; Donner, David; Warren, Robert; Roose, Jeroen P.

    2015-01-01

    The character of EGFR signals can influence cell fate but mechanistic insights into intestinal EGFR-Ras signalling are limited. Here we show that two distinct Ras nucleotide exchange factors, RasGRP1 and SOS1, lie downstream of EGFR but act in functional opposition. RasGRP1 is expressed in intestinal crypts where it restricts epithelial growth. High RasGRP1 expression in colorectal cancer (CRC) patient samples correlates with a better clinical outcome. Biochemically, we find that RasGRP1 creates a negative feedback loop that limits proliferative EGFR–SOS1–Ras signals in CRC cells. Genetic Rasgrp1 depletion from mice with either an activating mutation in KRas or with aberrant Wnt signalling due to a mutation in Apc resulted in both cases in exacerbated Ras–ERK signalling and cell proliferation. The unexpected opposing cell biological effects of EGFR–RasGRP1 and EGFR–SOS1 signals in the same cell shed light on the intricacy of EGFR-Ras signalling in normal epithelium and carcinoma. PMID:26005835

  10. [The basic and applied study on the epidermal growth factor].

    PubMed

    Huang, B R; Cai, L W; Xiang, X Z

    2001-04-01

    This article reviews the results of the basic research about epidermal growth factor and its receptor, and the development of the novel drug, EGF eyedrop, that containing chemically synthesized EGF gene, the construction of EGF expression vector, the transformation of the host cells, the purification of the recombinant protein EGF, the preparation of three batches of the EGF product and identification, the preclinical and clinical trials. Relevant studies show that recombinant EGF consisting of 51 amino acids can be secreted into the medium under the control of the alpha factor leading sequence in the yeast cells. The EGF can accelerate the growth of corneal-limbal epithelial cells and the healing of an alkali burned corneal. The EGF can be used in curing oral cavity ulcer and skin burned wound. And it has the preventive effects on experimental duodenal ulcer of rat. The antiserum was made for test of the concentration of blood EGF and urine EGF by RIA. Data from studies demonstrate the inhibition effect of EGF on the growth of tumor cells, such as A431 and BT325 cells in the presence of high EGF concentration (> 10 ng/ml). The expression of EGFR and DNA ploidy in renal carcinoma has clinical significance. Crystallization and preliminary x-ray diffraction studies of the EGF has been made. The MW of the EGF product is 6000, and the pI is about 4.6 and it has correct N-terminal amino acids sequences, immunogenicity and biological activity. There is no vestige of the DNA of the yeast cells. Animal experiments reveal that there is no cumulation of the EGF in the body, and EGF can promote corneal epithelial healing. There is no toxicological effect during cornea wound healing of rabbit. A randomized, double-blind, placebo-controlled, multi-center clinical trial was conducted in four hospitals to assess safety, ocular tolerance and efficacy of an ophthalmic solution of EGF for 200 cases of cornea transplantation and 247 cases of nebulae. Unequivocal results were obtained

  11. Nerve Growth Factor and Diabetic Neuropathy

    PubMed Central

    Vinik, Aaron

    2003-01-01

    Neuropathy is one of the most debilitating complications of both type 1 and type 2 diabetes, with estimates of prevalence between 50–90% depending on the means of detection. Diabetic neuropathies are heterogeneous and there is variable involvement of large myelinated fibers and small, thinly myelinated fibers. Many of the neuronal abnormalities in diabetes can be duplicated by experimental depletion of specific neurotrophic factors, their receptors or their binding proteins. In experimental models of diabetes there is a reduction in the availability of these growth factors, which may be a consequence of metabolic abnormalities, or may be independent of glycemic control. These neurotrophic factors are required for the maintenance of the neurons, the ability to resist apoptosis and regenerative capacity. The best studied of the neurotrophic factors is nerve growth factor (NGF) and the related members of the neurotrophin family of peptides. There is increasing evidence that there is a deficiency of NGF in diabetes, as well as the dependent neuropeptides substance P (SP) and calcitonin gene-related peptide (CGRP) that may also contribute to the clinical symptoms resulting from small fiber dysfunction. Similarly, NT3 appears to be important for large fiber and IGFs for autonomic neuropathy. Whether the observed growth factor deficiencies are due to decreased synthesis, or functional, e.g. an inability to bind to their receptor, and/or abnormalities in nerve transport and processing, remains to be established. Although early studies in humans on the role of neurotrophic factors as a therapy for diabetic neuropathy have been unsuccessful, newer agents and the possibilities uncovered by further studies should fuel clinical trials for several generations. It seems reasonable to anticipate that neurotrophic factor therapy, specifically targeted at different nerve fiber populations, might enter the therapeutic armamentarium. PMID:14668049

  12. Transforming growth factor alpha and epidermal growth factor levels in bladder cancer and their relationship to epidermal growth factor receptor.

    PubMed Central

    Mellon, J. K.; Cook, S.; Chambers, P.; Neal, D. E.

    1996-01-01

    We have examined levels of epidermal growth factor (EGF) and transforming growth factor alpha (TGF-alpha) in neoplastic and non-neoplastic bladder tissue using a standard radioimmunoassay technique. Tumour samples had much higher TGF-alpha levels compared with EGF and TGF-alpha levels in malignant tissue were significantly higher than in benign bladder samples. There was, in addition, a difference in mean EGF levels from 'normal' bladder samples from non-tumour bearing areas of bladder in patients with bladder cancer compared with 'normal' bladder tissue obtained at the time of organ retrieval surgery. Levels of EGF and TGF-alpha did not correlate with levels of EGF receptor (EGFR) as determined by a radioligand binding method but levels of TGF-alpha > 10 ng gm-1 of tumour tissue did correlate with EGFR positivity defined using immunohistochemistry. These data suggest that TGF-alpha is the likely ligand for EGFR in bladder tumours. PMID:8605103

  13. Metformin Inhibits the IL-6-Induced Epithelial-Mesenchymal Transition and Lung Adenocarcinoma Growth and Metastasis

    PubMed Central

    Wang, Yubo; Han, Rui; Li, Li; Xiang, Tong; He, Luhang; Long, Haixia; Zhu, Bo; He, Yong

    2014-01-01

    Objective Epithelial-mesenchymal transition (EMT) plays an important role in cancer tumorigenesis. However, the underlying mechanisms of EMT in lung adenocarcinoma, and how this process might be inhibited, remain to be explored. This study investigated the role of IL-6 in lung adenocarcinoma cell EMT and explored the potential effects of metformin on this process. Methods Invasion assay and MTT assay was performed to determine cell invasion and cell proliferation. Western blotting, immunofluorescence, real-time PCR, ELISA, and immunohistochemistry were performed to detect the expression of IL-6, E-cadherin, Vimentin, and p-STAT3. Results We discovered that IL-6, via STAT3 phosphorylation, could promote lung adenocarcinoma cell invasion via EMT in vitro. This was supported by the inverse correlation between E-cadherin and IL-6 expression, positive correlation between IL-6 and vimentin mRNA expression and between STAT3 phosphorylation and IL-6 expression in tumor tissues. Importantly, metformin inhibited tumor growth and distant metastases in tumor-bearing nude mice and reversed IL-6-induced EMT both in vitro and in vivo. Furthermore, we found that blockade of STAT3 phosphorylation might be the underlying mechanism of metformin inhibition of IL-6-induced EMT. Conclusions Collectively, our present results show that enhanced IL-6 expression, via STAT3 phosphorylation, is a mechanism of EMT in lung adenocarcinoma. We found that metformin could inhibit IL-6-induced EMT possibly by blocking STAT3 phosphorylation. PMID:24789104

  14. Epithelial Wnt ligand secretion is required for adult hair follicle growth and regeneration

    PubMed Central

    Myung, Peggy; Takeo, Makoto; Ito, Mayumi; Atit, Radhika

    2012-01-01

    β-catenin, a key transducer molecule of Wnt signaling, is required for adult hair follicle growth and regeneration. However, the cellular source of Wnt ligands required for Wnt/β-catenin activation during anagen induction is unknown. In this study, we genetically deleted Wntless, a gene required for Wnt ligand secretion by Wnt-producing cells, specifically in the hair follicle epithelium during telogen phase. We show that epithelial Wnt ligands are required for anagen, as loss of Wntless in the follicular epithelium resulted in a profound hair cycle arrest. Both the follicular epithelium and dermal papilla showed markedly decreased Wnt/β-catenin signaling during anagen induction compared to control hair follicles. Surprisingly, hair follicle stem cells that are responsible for hair regeneration maintained expression of stem cell markers but exhibited significantly reduced proliferation. Finally, we demonstrate that epidermal Wnt ligands are critical for adult wound-induced de novo hair formation. Collectively, these data show that Wnt ligands secreted by the hair follicle epithelium are required for adult hair follicle regeneration and provide new insight into potential cellular targets for the treatment of hair disorders such as alopecia. PMID:22810306

  15. Rv3351c, a Mycobacterium tuberculosis gene that affects bacterial growth and alveolar epithelial cell viability.

    PubMed

    Pavlicek, Rebecca L; Fine-Coulson, Kari; Gupta, Tuhina; Quinn, Frederick D; Posey, James E; Willby, Melisa; Castro-Garza, Jorge; Karls, Russell K

    2015-12-01

    Despite the interactions known to occur between various lower respiratory tract pathogens and alveolar epithelial cells (AECs), few reports examine factors influencing the interplay between Mycobacterium tuberculosis bacilli and AECs during infection. Importantly, in vitro studies have demonstrated that the M. tuberculosis hbha and esxA gene products HBHA and ESAT6 directly or indirectly influence AEC survival. In this report, we identify Rv3351c as another M. tuberculosis gene that impacts the fate of both the pathogen and AEC host. Intracellular replication of an Rv3351c mutant in the human AEC type II pneumocyte cell line A549 was markedly reduced relative to the complemented mutant and parent strain. Deletion of Rv3351c diminished the release of lactate dehydrogenase and decreased uptake of trypan blue vital stain by host cells infected with M. tuberculosis bacilli, suggesting attenuated cytotoxic effects. Interestingly, an isogenic hbha mutant displayed reductions in AEC killing similar to those observed for the Rv3351c mutant. This opens the possibility that multiple M. tuberculosis gene products interact with AECs. We also observed that Rv3351c aids intracellular replication and survival of M. tuberculosis in macrophages. This places Rv3351c in the same standing as HBHA and ESAT6, which are important factors in AECs and macrophages. Defining the mechanism(s) by which Rv3351c functions to aid pathogen survival within the host may lead to new drug or vaccine targets. PMID:26492080

  16. Sustained-release genistein from nanostructured lipid carrier suppresses human lens epithelial cell growth

    PubMed Central

    Liu, Jin-Lu; Zhang, Wen-Ji; Li, Xue-Dong; Yang, Na; Pan, Wei-San; Kong, Jun; Zhang, Jin-Song

    2016-01-01

    AIM To design and investigate the efficacy of a modified nanostructured lipid carrier loaded with genistein (Gen-NLC) to inhibit human lens epithelial cells (HLECs) proliferation. METHODS Gen-NLC was made by melt emulsification method. The morphology, particle size (PS), zeta potentials (ZP), encapsulation efficiency (EE) and in vitro release were characterized. The inhibition effect of nanostructured lipid carrier (NLC), genistein (Gen) and Gen-NLC on HLECs proliferation was evaluated by cell counting kit-8 (CCK-8) assay, gene and protein expression of the proliferation marker Ki67 were evaluated with real-time quantitative polymerase chain reaction (RT-qPCR) and immunofluorescence analyses. RESULTS The mean PS of Gen-NLC was 80.12±1.55 nm with a mean polydispersity index of 0.11±0.02. The mean ZP was -7.14±0.38 mV and the EE of Gen in the nanoparticles was 92.3%±0.73%. Transmission electron microscopy showed that Gen-NLC displayed spherical-shaped particles covered by an outer-layer structure. In vitro release experiments demonstrated a prolonged drug release for 72h. The CCK-8 assay results showed the NLC had no inhibitory effect on HLECs and Gen-NLC displayed a much more prominent inhibitory effect on cellular growth compared to Gen of the same concentration. The mRNA and protein expression of Ki67 in LECs decreased significantly in Gen-NLC group. CONCLUSION Sustained drug release by Gen-NLCs may impede HLEC growth. PMID:27275415

  17. Growth Factors and Tension-Induced Skeletal Muscle Growth

    NASA Technical Reports Server (NTRS)

    Vandenburgh, Herman H.

    1994-01-01

    The project investigated biochemical mechanisms to enhance skeletal muscle growth, and developed a computer based mechanical cell stimulator system. The biochemicals investigated in this study were insulin/(Insulin like Growth Factor) IGF-1 and Steroids. In order to analyze which growth factors are essential for stretch-induced muscle growth in vitro, we developed a defined, serum-free medium in which the differentiated, cultured avian muscle fibers could be maintained for extended periods of time. The defined medium (muscle maintenance medium, MM medium) maintains the nitrogen balance of the myofibers for 3 to 7 days, based on myofiber diameter measurements and myosin heavy chain content. Insulin and IGF-1, but not IGF-2, induced pronounced myofiber hypertrophy when added to this medium. In 5 to 7 days, muscle fiber diameters increase by 71 % to 98% compared to untreated controls. Mechanical stimulation of the avian muscle fibers in MM medium increased the sensitivity of the cells to insulin and IGF-1, based on a leftward shift of the insulin dose/response curve for protein synthesis rates. (54). We developed a ligand binding assay for IGF-1 binding proteins and found that the avian skeletal muscle cultures produced three major species of 31, 36 and 43 kD molecular weight (54) Stretch of the myofibers was found to have no significant effect on the efflux of IGF-1 binding proteins, but addition of exogenous collagen stimulated IGF-1 binding protein production 1.5 to 5 fold. Steroid hormones have a profound effect on muscle protein turnover rates in vivo, with the stress-related glucocorticoids inducing rapid skeletal muscle atrophy while androgenic steroids induce skeletal muscle growth. Exercise in humans and animals reduces the catabolic effects of glucocorticoids and may enhance the anabolic effects of androgenic steroids on skeletal muscle. In our continuing work on the involvement of exogenrus growth factors in stretch-induced avian skeletal muscle growth, we

  18. Growth hormone, insulin-like growth factor system and carcinogenesis.

    PubMed

    Boguszewski, Cesar Luiz; Boguszewski, Margaret Cristina da Silva; Kopchick, John J

    2016-01-01

    The growth hormone (GH) and insulin-like growth factor (IGF) system plays an important role in the regulation of cell proliferation, differentiation, apoptosis, and angiogenesis. In terms of cell cycle regulation, the GH-IGF system induces signalling pathways for cell growth that compete with other signalling systems that result in cell death; thus the final effect of these opposed forces is critical for normal and abnormal cell growth. The association of the GH-IGF system with carcinogenesis has long been hypothesised, mainly based on in vitro studies and the use of a variety of animal models of human cancer, and also on epidemiological and clinical evidence in humans. While ample experimental evidence supports a role of the GH-IGF system in tumour promotion and progression, with several of its components being currently tested as central targets for cancer therapy, the strength of evidence from patients with acromegaly, GH deficiency, or treated with GH is much weaker. In this review, we will attempt to consolidate this data. (Endokrynol Pol 2016; 67 (4): 414-426). PMID:27387246

  19. Growth hormone-insulinlike growth factor I and immune function.

    PubMed

    Gelato, M C

    1993-04-01

    Growth hormone (GH) and insulinlike growth factor I (IGF-I) may be part of a neuroendocrine immune axis that stimulates cellular proliferation of primary lymphoid organs (bone marrow, thymus) as well as stimulates activation of peripheral lymphocytes and macrophages to enhance specific immune responses. GH can also stimulate production of thymic hormones and cytokines, and in this way impact on immune function. It is not clear whether GH and IGF-I act independently or whether the action of GH is mediated by local production of IGF-I by lymphocytes. Both GH and IGF-I and their receptors are present in lymphocytes. Thus, cells of the immune system may be important targets of the GH-IGF-I axis. PMID:18407143

  20. Growth Factors and Astrocytes Metabolism: Possible Roles for Platelet Derived Growth Factor.

    PubMed

    Cabezas, Ricardo; Avila-Rodriguez, Marco; Vega-Vela, Nelson E; Echeverria, Valentina; González, Janneth; Hidalgo, Oscar A; Santos, Altair B; Aliev, Gjumrakch; Barreto, George E

    2016-01-01

    Astrocytes exert multiple functions in the brain such as the development of blood-brain barrier characteristics, the promotion of neurovascular coupling, attraction of cells through the release of chemokines, clearance of toxic substances and generation of antioxidant molecules and growth factors. In this aspect, astrocytes secrete several growth factors (BDNF, GDNF, NGF, and others) that are fundamental for cell viability, oxidant protection, genetic expression and modulation of metabolic functions. The platelet derived growth factor (PDGF), which is expressed by many SNC cells, including astrocytes, is an important molecule that has shown neuroprotective potential, improvement of wound healing, regulation of calcium metabolism and mitochondrial function. Here we explore some of these astrocyte-driven functions of growth factors and their possible therapeutic uses in the context of neurodegeneration. PMID:26477707

  1. Targeting Insulin-Like Growth Factor 1 Receptor Inhibits Pancreatic Cancer Growth and Metastasis

    PubMed Central

    Subramani, Ramadevi; Lopez-Valdez, Rebecca; Arumugam, Arunkumar; Nandy, Sushmita; Boopalan, Thiyagarajan; Lakshmanaswamy, Rajkumar

    2014-01-01

    Pancreatic cancer is one of the most lethal cancers. Increasing incidence and mortality indicates that there is still much lacking in detection and management of the disease. This is partly due to a lack of specific symptoms during early stages of the disease. Several growth factor receptors have been associated with pancreatic cancer. Here, we have investigated if an RNA interference approach targeted to IGF-IR could be effective and efficient against pancreatic cancer growth and metastasis. For that, we evaluated the effects of IGF-1R inhibition using small interfering RNA (siRNAs) on tumor growth and metastasis in HPAC and PANC-1 pancreatic cancer cell lines. We found that silencing IGF-1R inhibits pancreatic cancer growth and metastasis by blocking key signaling pathways such AKT/PI3K, MAPK, JAK/STAT and EMT. Silencing IGF-1R resulted in an anti-proliferative effect in PANC-1 and HPAC pancreatic cancer cell lines. Matrigel invasion, transwell migration and wound healing assays also revealed a role for IGF-1R in metastatic properties of pancreatic cancer. These results were further confirmed using Western blotting analysis of key intermediates involved in proliferation, epithelial mesenchymal transition, migration, and invasion. In addition, soft agar assays showed that silencing IGF-1R also blocks the colony forming capabilities of pancreatic cancer cells in vitro. Western blots, as well as, flow cytometric analysis revealed the induction of apoptosis in IGF-1R silenced cells. Interestingly, silencing IGF-1R also suppressed the expression of insulin receptor β. All these effects together significantly control pancreatic cancer cell growth and metastasis. To conclude, our results demonstrate the significance of IGF-1R in pancreatic cancer. PMID:24809702

  2. Platelet-derived growth factor and transforming growth factor-beta in invertebrate immune and neuroendocrine interactions: another sign of conservation in evolution.

    PubMed

    Ottaviani, E; Franchini, A; Kletsas, D

    2001-08-01

    Growth factor-like molecules have been found in various invertebrate species. In particular, we have reported the presence of platelet-derived growth factor (PDGF)-AB and transforming growth factor-beta (TGF-beta)1 immunoreactive molecules in molluscs, insects and annelids. Moreover, PDGF-AB and TGF-beta1 affect the main immune functions, such as phagocytosis, chemotaxis and cell motility. Changes in cell shape are induced via interactions of growth factors with their respective specific receptors. The extracellular signals are transduced by the activation of classical signal transduction pathways, such as those involving PKA and PKC, and pivotal transcription regulators, i.e. the Fos, Jun and SMAD proteins. The two growth factors intervene in stress responses by activating the CRH-ACTH-biogenic amine axis. Exogenous administration of PDGF-AB and TGF-beta1 in a molluscan wound provokes an accelerated migration of immunocytes and fibroblasts to the injured area, stimulating granulation tissue formation and wound re-epithelialization. These findings suggest that these molecules are ancestral and that their function is well conserved and crucial in the maintenance of invertebrate homeostasis. PMID:11489427

  3. Cytokine regulation of granulocyte-macrophage colony-stimulating factor (GM-CSF) production by human retinal pigment epithelial cells

    PubMed Central

    Crane, I J; Kuppner, M C; Mckillop-Smith, S; Wallace, C A; Forrester, J V

    1999-01-01

    GM-CSF is an important regulator of macrophage, granulocyte and dendritic cell behaviour and function. These cell types have been implicated in the retinal damage characteristic of endogenous posterior uveitis. Dendritic cells in the choroid have access to retinal antigens processed by the retinal pigment epithelial (RPE) cells of the blood–retinal barrier and are thought to be candidates for the presentation of antigen in uveoretinitis. We therefore investigated the production of GM-CSF and its regulation in human RPE cells. IL-1β, tumour necrosis factor-alpha (TNF-α) and transforming growth factor-beta (TGF-β) all stimulated GM-CSF production by RPE cells and a combination of these cytokines increased GM-CSF production over five-fold compared with that with the individual cytokines alone. Interferon-gamma (IFN-γ) rapidly down-regulated these responses. IFN-γ did not appear to be acting directly on IL-1β or via the synthesis of another protein. GM-CSF mRNA expression showed the same pattern of response to these cytokines, indicating transcriptional or pre-transcriptional regulation, and there was no evidence that IFN-γ was acting by destabilizing GM-CSF mRNA. These results are generally important in understanding the ways in which cytokine regulation differs between different cell types and also more specifically for determining ways in which a cytokine with a significant role in the development of autoimmune uveoretinitis may be manipulated. PMID:9933455

  4. Transcriptome profiling reveals novel gene expression signatures and regulating transcription factors of TGFβ-induced epithelial-to-mesenchymal transition.

    PubMed

    Du, Liutao; Yamamoto, Shota; Burnette, Barry L; Huang, Danshang; Gao, Kun; Jamshidi, Neema; Kuo, Michael D

    2016-08-01

    Dysregulated epithelial to mesenchymal transition (EMT) in cancer cells endows invasive and metastatic properties upon cancer cells that favor successful colonization of distal target organs and therefore play a critical role in transforming early-stage carcinomas into invasive malignancies. EMT has also been associated with tumor recurrence and drug resistance and cancer stem cell initiation. Therefore, better understanding of the mechanisms behind EMT could ultimately contribute to the development of novel prognostic approaches and individualized therapies that specifically target EMT processes. As an effort to characterize the central transcriptome changes during EMT, we have developed a Transforming growth factor (TGF)-beta-based in vitro EMT model and used it to profile EMT-related gene transcriptional changes in two different cell lines, a non-small cell lung cancer cell line H358, and a breast cell line MCF10a. After 7 days of TGF-beta/Oncostatin M (OSM) treatment, changes in cell morphology to a mesenchymal phenotype were observed as well as concordant EMT-associated changes in mRNA and protein expression. Further, increased motility was noted and flow cytometry confirmed enrichment in cancer stem cell-like populations. Microarray-based differential expression analysis identified an EMT-associated gene expression signature which was confirmed by RT-qPCR and which significantly overlapped with a previously published EMT core signature. Finally, two novel EMT-regulating transcription factors, IRF5 and LMCD1, were identified and independently validated. PMID:27318801

  5. Functional characteristics of the growth factor receptor family and some ligands in the oropharyngeal cavity of the Chukar partridge (Alectoris chukar).

    PubMed

    Saruhan, Berna G; Sağsöz, Hakan; Akbalik, Mehmet E; Erdoğan, Serkan

    2015-01-01

    1. The aim of the present study is to describe, immunohistochemically, the expression and cell type localisation of growth factor receptors and some of their ligands in the oropharyngeal organs of the Chukar partridge. 2. The tissue samples from 10 healthy adult partridges were dissected under ether anaesthesia and then embedded in paraffin following routine histological procedures. The immunoreaction for receptors and ligands of the epidermal growth factor receptor (EGFR)/ligand system was localised in the cell membrane, nucleus and cytoplasm of the luminal and glandular epithelial cells, stromal and striated muscle cells, and vascular endothelial and smooth muscle cells. 3. Variations were observed in the avian oropharyngeal organs. The immunostaining for the erbB1/HER1 (human epidermal growth factor receptor 1) and the EGF (epidermal growth factor) and AREG (Amphiregulin) ligands in the luminal epithelial cells was higher than in the glandular epithelial, stromal and striated muscle cells. However, the immunostaining for erbB3/HER3 (human epidermal growth factor receptor 3) and erbB4/HER4 (human epidermal growth factor receptor 4) were similar in the luminal epithelium, stromal and striated muscle cells. 4. Growth factor receptors and some of their ligands were localised in different cell types in the oropharyngeal organs. We suggest that erbB/HERs (human epidermal growth factor receptors) and their ligands play an important role in proliferation, differentiation, growth, survival and migration of the cells. PMID:26569385

  6. Growth factor signaling in lung morphogenetic centers: automaticity, stereotypy and symmetry

    PubMed Central

    Warburton, David; Bellusci, Saverio; Del Moral, Pierre-Marie; Kaartinen, Vesa; Lee, Matt; Tefft, Denise; Shi, Wei

    2003-01-01

    Lung morphogenesis is stereotypic, both for lobation and for the first several generations of airways, implying mechanistic control by a well conserved, genetically hardwired developmental program. This program is not only directed by transcriptional factors and peptide growth factor signaling, but also co-opts and is modulated by physical forces. Peptide growth factors signal within repeating epithelial-mesenchymal temporospatial patterns that constitute morphogenetic centers, automatically directing millions of repetitive events during both stereotypic branching and nonstereotypic branching as well as alveolar surface expansion phases of lung development. Transduction of peptide growth factor signaling within these centers is finely regulated at multiple levels. These may include ligand expression, proteolytic activation of latent ligand, ligand bioavailability, ligand binding proteins and receptor affinity and presentation, receptor complex assembly and kinase activation, phosphorylation and activation of adapter and messenger protein complexes as well as downstream events and cross-talk both inside and outside the nucleus. Herein we review the critical Sonic Hedgehog, Fibroblast Growth Factor, Bone Morphogenetic Protein, Vascular Endothelial Growth Factor and Transforming Growth Factorβ signaling pathways and propose how they may be functionally coordinated within compound, highly regulated morphogenetic gradients that drive first stereotypic and then non-stereotypic, automatically repetitive, symmetrical as well as asymmetrical branching events in the lung. PMID:12818006

  7. Growth Hormone and Insulin-Like Growth Factor-1.

    PubMed

    Nicholls, Adam R; Holt, Richard I G

    2016-01-01

    Human growth hormone (GH) was first isolated from the human pituitary gland in 1945 and found to promote the growth of children with hypopituitarism. Since the formation of the World Anti-Doping Association, human GH has appeared on the list of forbidden substances. There is a significant amount of anecdotal evidence that human GH is misused by athletes to enhance performance, and there have been a number of high-profile cases of GH use in professional sport. GH secretagogues (GH-Ss), which increase GH secretion, and insulin-like growth factor (IGF-1), which mediates many of the effects of GH, are also misused, although there is less evidence for this. The effectiveness of GH, IGF-1, and GH-Ss as performance-enhancing drugs remains unclear. Evidence from studies of GH use in people with hypopituitarism show several desirable outcomes, including increased lean body mass, increased strength, and increased exercise capacity. These anabolic and metabolic properties, coupled with the difficulty in detecting them, make them attractive as agents of misuse. Studies in healthy young adults have also demonstrated a performance benefit with GH and IGF-1. PMID:27347885

  8. Intraepithelial ischemia is a principal factor promoting cancerization of the covering epithelial tissues.

    PubMed

    Karseladze, A I

    2016-09-01

    Prominent angiogenesis, which is a hallmark of invasive cancer is preceded at the precancerous stage by marked ischemia. Our hypothesis proposes a structural mechanism responsible for altering blood flow in the covering epithelium and leading to marked reduction of vascularization in the foci of dysplasia. This mechanism varies from one type of epithelium to another. In squamous epithelium only basal cells are in direct contact with stromal vessels. To supply nutrients to the rest of the cells located at different levels, the subjacent stroma forms excrescences which penetrate upward together with blood capillaries. As soon as precancerous dysplastic alterations start and progress the number of intraepithelial blood vessels simultaneously decreases, thus leading to ischemia which precedes or promotes malignization of the covering squamous epithelium. To compensate for the deficit in blood supply, the dysplastic cells penetrate deeper into the underlying stroma, commencing invasion. Thus, the cells destroy the subjacent stroma not because they are initially "malignant", but due to ischemia which provokes the search for nutrients. Comparing squamous epithelium with glandular respiratory epithelium shows that the latter contains no blood capillaries at all. However, unlike squamous epithelial coverings, in respiratory epithelial covering, each cell is attached directly to the basal membrane and has ample access to the blood supply. Covering respiratory epithelium itself seldom gives rise directly to malignant growth. Cancerization of this type of epithelium occurs in the foci of squamous metaplasia. The latter are not supplied by a sufficient amount of blood vessels and in the majority of cases remain fragile and vulnerable structures, easily prone to malignization. Further study of these phenomenon should include the clarification of the influence of carcinogenic agents on the mechanism of adequate vascularization at the precancerous stage. PMID:27515223

  9. Waste management - cytokines, growth factors and cachexia.

    PubMed

    Saini, Amarjit; Al-Shanti, Nasser; Nasser, Al-Shanti; Stewart, Claire E H

    2006-12-01

    Muscle damage with a lack of regeneration, manifests itself in several life-threatening diseases, including cancer cachexia, congestive heart failure, AIDS and sepsis. Often misdiagnosed as a condition simply of weight loss, cachexia is actually a highly complex metabolic disorder involving features of anorexia, anaemia, lipolysis and insulin resistance. A significant loss of lean body mass arises from such conditions, resulting in wasting of skeletal muscle. Unlike starvation, the weight loss seen in chronic illnesses arises equally from loss of muscle and of fat. The cachectic state is particularly problematic in cancer, typifying poor prognosis and often lowering responses to chemotherapy and radiation treatment. More than half of cancer patients suffer from cachexia, and strikingly, nearly one-third of cancer deaths are related to cachexia rather than the tumour burden. In considering this disorder, we are faced with a conundrum; how is it possible for uncontrolled growth to prevail in the tumour, in the face of unrestrained tissue loss in our muscles? Consistently, the catabolic state has been associated with a shift in the homeostatic balance between muscle synthesis and degradation mediated by the actions of growth factors and cytokines. Indeed, tumour necrosis factor-alpha (TNF-alpha) levels are raised in several animal models of cachectic muscle wasting, whereas the insulin-like growth factor (IGF) system acts potently to regulate muscle development, hypertrophy and maintenance. This concept of skeletal muscle homeostasis, often viewed as the net balance between two separate processes of protein synthesis and degradation has however changed. More recently, the view is that these two biochemical processes are not occurring independently of each other but in fact are finely co-ordinated by a web of intricate signalling networks. This review, therefore, aims to discuss data currently available regarding the mechanisms of degeneration and regeneration with

  10. Apoptosis Factors of Lens Epithelial Cells Responsible for Cataractogenesis in Vitrectomized Eyes with Silicone Oil Tamponade

    PubMed Central

    Zhu, Lili; Zhao, Ke; Lou, Dinghua

    2016-01-01

    Background The aim of this study was to determine the expression of apoptotic factors Bax, Bcl-2, and Caspase-3 in lens epithelial cells (LECs) from cataracts secondary to pars plana vitrectomy with silicone oil (SO) tamponade. We also investigated the impact of SO emulsification on the expression of apoptotic factors. Material/Methods Anterior capsulotomy specimens of 20 eyes in 20 patients with cataract secondary to SO tamponade (Group 2), were collected. Another 20 eyes of 20 patients with age-related cataract (Group 1) were recruited as controls. The anterior capsule specimens were obtained from the patients during cataract surgery, frozen and later analyzed with respect to immunohistochemical stains of Bax, Bcl-2, and Caspase-3 using a confocal microscope. Results Age, sex, and laterality did not show any difference between the 2 groups. There was a greater increase in Bax and Caspase-3 expression in LEC in Group 2 than in Group 1 (PBax<0.0001, PCaspase-3<0.0001). The Bcl-2 expression decreased in Group 2, although the difference was not significant (P=0.616). The changes of apoptosis factors are not associated to SO emulsification (PBax=0.354, PBcl-2=0.728, PCaspase-3=0.939). Conclusions The caspase-3-dependent apoptosis of LECs increased in complicated cataract patients with SO endotamponade. The Bax played a critical role in regulating apoptosis of LECs in vitrectomized eyes with SO tamponade. The SO emulsification had no significant impact on the expression of apoptosis factors. PMID:26956740

  11. Innate immunity in HIV-1 infection: epithelial and non-specific host factors of mucosal immunity- a workshop report.

    PubMed

    Nittayananta, W; Weinberg, A; Malamud, D; Moyes, D; Webster-Cyriaque, J; Ghosh, S

    2016-04-01

    The interplay between HIV-1 and epithelial cells represents a critical aspect in mucosal HIV-1 transmission. Epithelial cells lining the oral cavity cover subepithelial tissues, which contain virus-susceptible host cells including CD4(+) T lymphocytes, monocytes/macrophages, and dendritic cells. Oral epithelia are among the sites of first exposure to both cell-free and cell-associated virus HIV-1 through breast-feeding and oral-genital contact. However, oral mucosa is considered to be naturally resistant to HIV-1 transmission. Oral epithelial cells have been shown to play a crucial role in innate host defense. Nevertheless, it is not clear to what degree these local innate immune factors contribute to HIV-1 resistance of the oral mucosa. This review paper addressed the following issues that were discussed at the 7th World Workshop on Oral Health and Disease in AIDS held in Hyderabad, India, during November 6-9, 2014: (i) What is the fate of HIV-1 after interactions with oral epithelial cells?; (ii) What are the keratinocyte and other anti-HIV effector oral factors, and how do they contribute to mucosal protection?; (iii) How can HIV-1 interactions with oral epithelium affect activation and populations of local immune cells?; (iv) How can HIV-1 interactions alter functions of oral epithelial cells? PMID:27109285

  12. Bacteria Regulate Intestinal Epithelial Cell Differentiation Factors Both In Vitro and In Vivo

    PubMed Central

    Becker, Svetlana; Oelschlaeger, Tobias A.; Wullaert, Andy; Pasparakis, Manolis; Wehkamp, Jan; Stange, Eduard F.; Gersemann, Michael

    2013-01-01

    Background The human colon harbours a plethora of bacteria known to broadly impact on mucosal metabolism and function and thought to be involved in inflammatory bowel disease pathogenesis and colon cancer development. In this report, we investigated the effect of colonic bacteria on epithelial cell differentiation factors in vitro and in vivo. As key transcription factors we focused on Hes1, known to direct towards an absorptive cell fate, Hath1 and KLF4, which govern goblet cell. Methods Expression of the transcription factors Hes1, Hath1 and KLF4, the mucins Muc1 and Muc2 and the defensin HBD2 were measured by real-time PCR in LS174T cells following incubation with several heat-inactivated E. coli strains, including the probiotic E. coli Nissle 1917+/− flagellin, Lactobacilli and Bifidobacteria. For protein detection Western blot experiments and chamber-slide immunostaining were performed. Finally, mRNA and protein expression of these factors was evaluated in the colon of germfree vs. specific pathogen free vs. conventionalized mice and colonic goblet cells were counted. Results Expression of Hes1 and Hath1, and to a minor degree also of KLF4, was reduced by E. coli K-12 and E. coli Nissle 1917. In contrast, Muc1 and HBD2 expression were significantly enhanced, independent of the Notch signalling pathway. Probiotic E. coli Nissle 1917 regulated Hes1, Hath1, Muc1 and HBD2 through flagellin. In vivo experiments confirmed the observed in vitro effects of bacteria by a diminished colonic expression of Hath1 and KLF4 in specific pathogen free and conventionalized mice as compared to germ free mice whereas the number of goblet cells was unchanged in these mice. Conclusions Intestinal bacteria influence the intestinal epithelial differentiation factors Hes1, Hath1 and KLF4, as well as Muc1 and HBD2, in vitro and in vivo. The induction of Muc1 and HBD2 seems to be triggered directly by bacteria and not by Notch. PMID:23418447

  13. Regulation of the actin cytoskeleton in Helicobacter pylori-induced migration and invasive growth of gastric epithelial cells

    PubMed Central

    2011-01-01

    Dynamic rearrangement of the actin cytoskeleton is a significant hallmark of Helicobacter pylori (H. pylori) infected gastric epithelial cells leading to cell migration and invasive growth. Considering the cellular mechanisms, the type IV secretion system (T4SS) and the effector protein cytotoxin-associated gene A (CagA) of H. pylori are well-studied initiators of distinct signal transduction pathways in host cells targeting kinases, adaptor proteins, GTPases, actin binding and other proteins involved in the regulation of the actin lattice. In this review, we summarize recent findings of how H. pylori functionally interacts with the complex signaling network that controls the actin cytoskeleton of motile and invasive gastric epithelial cells. PMID:22044652

  14. Expression of transforming growth factor-β (TGF-β) in chronic idiopathic cough

    PubMed Central

    Xie, Shaoping; Macedo, Patricia; Hew, Mark; Nassenstein, Christina; Lee, Kang-Yun; Chung, Kian Fan

    2009-01-01

    In patients with chronic idiopathic cough, there is a chronic inflammatory response together with evidence of airway wall remodelling and an increase in airway epithelial nerves expressing TRPV-1. We hypothesised that these changes could result from an increase in growth factors such as TGFβ and neurotrophins. We recruited 13 patients with persistent non-asthmatic cough despite specific treatment of associated primary cause(s), or without associated primary cause, and 19 normal non-coughing volunteers without cough as controls, who underwent fiberoptic bronchoscopy with bronchoalveolar lavage (BAL) and bronchial biopsies. There was a significant increase in the levels of TGFβ in BAL fluid, but not of nerve growth factor(NGF) and brain-derived nerve growth factor(BDNF) compared to normal volunteers. Levels of TFGβ gene and protein expression were assessed in bronchial biopsies. mRNA expression for TGFβ was observed in laser-captured airway smooth muscle and epithelial cells, and protein expression by immunohistochemistry was increased in ASM cells in chronic cough patients, associated with an increase in nuclear expression of the transcription factor, smad 2/3. Subbasement membrane thickness was significantly higher in cough patients compared to normal subjects and there was a positive correlation between TGF-β levels in BAL and basement membrane thickening. TGFβ in the airways may be important in the airway remodelling changes observed in chronic idiopathic cough patients, that could in turn lead to activation of the cough reflex. PMID:19463161

  15. [BETA-III TUBULIN AS A POTENTIAL TARGET FOR BLOCKING INVASIVE GROWTH OF MALIGNANT EPITHELIAL TUMORS].

    PubMed

    Portyanko, A S; Akalovich, S T; Doroshenko, T M

    2015-01-01

    Invasive growth is the first step of metastatic cascade in the growth of malignant tumors. The mobility of cells, which is a necessary factor of the invasive growth of malignant tumors, is closely linked to the dynamic structure of cytoskeleton. An important role in cell motility is played by microtubules and actin microfilaments. Microtubules consist of tubulin--a heterodimer comprising α and β subunits, which can be represented by different isotypes. The appearance of beta-III tubulin in a tumor is essential for chemoresistance and prognosis of some tumors in humans. This study focuses on determining the possibility of using beta-III tubulin as a target molecule for the suppression of invasive growth. It is established that blocking of the beta-III tubulin expression in colorectal cancer cells does not affect their viability, but reduces the cell adhesion to the extracellular matrix by 40% for HT-29 (∂ = 0.0044) and by 15% (p = 0.0436) for HCT 116) and produces a four-fold decrease in the invasive activity (p = 0.0000 and 0.0001, respectively). These facts allow considering beta-III tubulin as a target molecule in the development of antitumor drugs. PMID:26591579

  16. Cytotoxic Potential of Bacillus cereus Strains ATCC 11778 and 14579 Against Human Lung Epithelial Cells Under Microaerobic Growth Conditions.

    PubMed

    Kilcullen, Kathleen; Teunis, Allison; Popova, Taissia G; Popov, Serguei G

    2016-01-01

    Bacillus cereus, a food poisoning bacterium closely related to Bacillus anthracis, secretes a multitude of virulence factors including enterotoxins, hemolysins, and phospholipases. However, the majority of the in vitro experiments evaluating the cytotoxic potential of B. cereus were carried out in the conditions of aeration, and the impact of the oxygen limitation in conditions encountered by the microbe in natural environment such as gastrointestinal tract remains poorly understood. This research reports comparative analysis of ATCC strains 11778 (BC1) and 14579 (BC2) in aerobic and microaerobic (static) cultures with regard to their toxicity for human lung epithelial cells. We showed that BC1 increased its toxicity upon oxygen limitation while BC2 was highly cytotoxic in both growth conditions. The combined effect of the pore-forming, cholesterol-dependent hemolysin, cereolysin O (CLO), and metabolic product(s) such as succinate produced in microaerobic conditions provided substantial contribution to the toxicity of BC1 but not BC2 which relied mainly on other toxins. This mechanism is shared between CB1 and B. anthracis. It involves the permeabilization of the cell membrane which facilitates transport of toxic bacterial metabolites into the cell. The toxicity of BC1 was potentiated in the presence of bovine serum albumin which appeared to serve as reservoir for bacteria-derived nitric oxide participating in the downstream production of reactive oxidizing species with the properties of peroxynitrite. In agreement with this the BC1 cultures demonstrated the increased oxidation of the indicator dye Amplex Red catalyzed by peroxidase as well as the increased toxicity in the presence of externally added ascorbic acid. PMID:26870026

  17. Cytotoxic Potential of Bacillus cereus Strains ATCC 11778 and 14579 Against Human Lung Epithelial Cells Under Microaerobic Growth Conditions

    PubMed Central

    Kilcullen, Kathleen; Teunis, Allison; Popova, Taissia G.; Popov, Serguei G.

    2016-01-01

    Bacillus cereus, a food poisoning bacterium closely related to Bacillus anthracis, secretes a multitude of virulence factors including enterotoxins, hemolysins, and phospholipases. However, the majority of the in vitro experiments evaluating the cytotoxic potential of B. cereus were carried out in the conditions of aeration, and the impact of the oxygen limitation in conditions encountered by the microbe in natural environment such as gastrointestinal tract remains poorly understood. This research reports comparative analysis of ATCC strains 11778 (BC1) and 14579 (BC2) in aerobic and microaerobic (static) cultures with regard to their toxicity for human lung epithelial cells. We showed that BC1 increased its toxicity upon oxygen limitation while BC2 was highly cytotoxic in both growth conditions. The combined effect of the pore-forming, cholesterol-dependent hemolysin, cereolysin O (CLO), and metabolic product(s) such as succinate produced in microaerobic conditions provided substantial contribution to the toxicity of BC1 but not BC2 which relied mainly on other toxins. This mechanism is shared between CB1 and B. anthracis. It involves the permeabilization of the cell membrane which facilitates transport of toxic bacterial metabolites into the cell. The toxicity of BC1 was potentiated in the presence of bovine serum albumin which appeared to serve as reservoir for bacteria-derived nitric oxide participating in the downstream production of reactive oxidizing species with the properties of peroxynitrite. In agreement with this the BC1 cultures demonstrated the increased oxidation of the indicator dye Amplex Red catalyzed by peroxidase as well as the increased toxicity in the presence of externally added ascorbic acid. PMID:26870026

  18. Architectural Transcription Factor HMGI(Y) Promotes Tumor Progression and Mesenchymal Transition of Human Epithelial Cells

    PubMed Central

    Reeves, Raymond; Edberg, Dale D.; Li, Ying

    2001-01-01

    Numerous studies have demonstrated that overexpression or aberrant expression of the HMGI(Y) family of architectural transcription factors is frequently associated with both neoplastic transformation of cells and metastatic tumor progression. Little is known, however, about the molecular roles played by the HMGI(Y) proteins in these events. Here we report that human breast epithelial cells harboring tetracycline-regulated HMGI(Y) transgenes acquire the ability to form both primary and metastatic tumors in nude mice only when the transgenes are actively expressed. Unexpectedly, the HMG-Y, rather than the HMG-I, isoform of these proteins is the most effective elicitor of both neoplastic transformation and metastatic progression in vivo. Furthermore, expression of either antisense or dominant-negative HMGI(Y) constructs inhibits both the rate of proliferation of tumor cells and their ability to grow anchorage independently in soft agar. Array analysis of transcription profiles demonstrates that the HMG-I and HMG-Y isoform proteins each modulate the expression of distinctive constellations of genes known to be involved in signal transduction, cell proliferation, tumor initiation, invasion, migration, induction of angiogenesis, and colonization. Immunohistochemical analyses of tumors formed in nude mice indicate that many have undergone an epithelial-mesenchymal transition in vivo. Together, these findings demonstrate that overexpression of the HMGI(Y) proteins, more specifically, the HMG-Y isoform protein, is causally associated with both neoplastic transformation and metastatic progression and suggest that induction of integrins and their signaling pathways may play significant molecular roles in these biological events. PMID:11134344

  19. Architectural transcription factor HMGI(Y) promotes tumor progression and mesenchymal transition of human epithelial cells.

    PubMed

    Reeves, R; Edberg, D D; Li, Y

    2001-01-01

    Numerous studies have demonstrated that overexpression or aberrant expression of the HMGI(Y) family of architectural transcription factors is frequently associated with both neoplastic transformation of cells and metastatic tumor progression. Little is known, however, about the molecular roles played by the HMGI(Y) proteins in these events. Here we report that human breast epithelial cells harboring tetracycline-regulated HMGI(Y) transgenes acquire the ability to form both primary and metastatic tumors in nude mice only when the transgenes are actively expressed. Unexpectedly, the HMG-Y, rather than the HMG-I, isoform of these proteins is the most effective elicitor of both neoplastic transformation and metastatic progression in vivo. Furthermore, expression of either antisense or dominant-negative HMGI(Y) constructs inhibits both the rate of proliferation of tumor cells and their ability to grow anchorage independently in soft agar. Array analysis of transcription profiles demonstrates that the HMG-I and HMG-Y isoform proteins each modulate the expression of distinctive constellations of genes known to be involved in signal transduction, cell proliferation, tumor initiation, invasion, migration, induction of angiogenesis, and colonization. Immunohistochemical analyses of tumors formed in nude mice indicate that many have undergone an epithelial-mesenchymal transition in vivo. Together, these findings demonstrate that overexpression of the HMGI(Y) proteins, more specifically, the HMG-Y isoform protein, is causally associated with both neoplastic transformation and metastatic progression and suggest that induction of integrins and their signaling pathways may play significant molecular roles in these biological events. PMID:11134344

  20. Decorin: A Growth Factor Antagonist for Tumor Growth Inhibition

    PubMed Central

    Järvinen, Tero A. H.; Prince, Stuart

    2015-01-01

    Decorin (DCN) is the best characterized member of the extracellular small leucine-rich proteoglycan family present in connective tissues, typically in association with or “decorating” collagen fibrils. It has substantial interest to clinical medicine owing to its antifibrotic, anti-inflammatory, and anticancer effects. Studies on DCN knockout mice have established that a lack of DCN is permissive for tumor development and it is regarded as a tumor suppressor gene. A reduced expression or a total disappearance of DCN has been reported to take place in various forms of human cancers during tumor progression. Furthermore, when used as a therapeutic molecule, DCN has been shown to inhibit tumor progression and metastases in experimental cancer models. DCN affects the biology of various types of cancer by targeting a number of crucial signaling molecules involved in cell growth, survival, metastasis, and angiogenesis. The active sites for the neutralization of different growth factors all reside in different parts of the DCN molecule. An emerging concept that multiple proteases, especially those produced by inflammatory cells, are capable of cleaving DCN suggests that native DCN could be inactivated in a number of pathological inflammatory conditions. In this paper, we review the role of DCN in cancer. PMID:26697491

  1. Human Genome-Wide RNAi Screen for Host Factors That Modulate Intracellular Salmonella Growth

    PubMed Central

    Thornbrough, Joshua M.; Hundley, Tom; Valdivia, Raphael; Worley, Micah J.

    2012-01-01

    Salmonella enterica is a bacterial pathogen of humans that can proliferate within epithelial cells as well as professional phagocytes of the immune system. While much has been learned about the microbial genes that influence the infectious process through decades of intensive research, relatively little is known about the host factors that affect infection. We performed a genome-wide siRNA screen to identify host genes that Salmonella enterica serovar Typhimurium (S. typhimurium) utilizes to facilitate growth within human epithelial cells. In this screen, with siRNAs targeting every predicted gene in the human genome, we identified 252 new human-host-susceptibility factors (HSFs) for S. typhimurium. We also identified 39 genes whose silencing results in increased intracellular growth of S. typhimurium. The HSFs identified are regulated most centrally by NFκB and associate with each other through an extremely dense network of interactions that center around a group of kinases. Most genes identified were not previously appreciated as playing roles in the intracellular lifecycle of S. enterica. Numerous HSFs identified with interesting characteristics that could play plausible roles in mediating intracellular microbial growth are discussed. Importantly, this study reveals significant overlap between the host network that supports S. typhimurium growth within human epithelial cells and the one that promotes the growth of Mycobacterium tuberculosis within human macrophages. In addition to providing much new information about the molecular mechanisms underlying S. enterica-host cell interplay, all 252 HSFs identified are candidates for new anti-microbial targets for controlling S. enterica infections, and some may provide broad-spectrum anti-microbial activity. PMID:22701604

  2. Blocking M2 muscarinic receptor signaling inhibits tumor growth and reverses epithelial-mesenchymal transition (EMT) in non-small cell lung cancer (NSCLC)

    PubMed Central

    Zhao, Qingnan; Gu, Xiajing; Zhang, Chun; Lu, Qin; Chen, Hongzhuan; Xu, Lu

    2015-01-01

    Lung cancers express non-neuronal, cholinergic autoparacrine loop, which facilitates tumor growth. Interruption of M3 muscarinic cholinergic signaling has been reported to inhibit small cell lung cancer (SCLC) growth. The purpose of this study is to investigate if blocking autoparacrine muscarinic cholinergic signaling could inhibit non-small cell lung cancer (NSCLC) growth and possible underlying mechanisms. Our results showed that PC9 and A549 cells expressed all 5 subtypes of muscarinic receptor (mAChR) and blocking M2 mAChR (M2R) signaling using selective antagonist methoctramine or short hairpin RNA (shRNA) inhibited tumor cell proliferation in vitro and in vivo. Consistent with AChR agonists stimulating p44/42 MAPK (Erk1/2) and Akt phosphorylation, blocking M2R signaling decreased MAPK and Akt phosphorylation, indicating that non-neuronal ACh functions as an autoparacrine growth factor signaling in part through activation of M2R and downstream MAPK and Akt pathways. Importantly, further studies revealed that blocking M2R signaling also reversed epithelial-mesenchymal transition (EMT) in vitro and in vivo, indicating that non-neuronal ACh promotes EMT partially through activation of M2R. These findings demonstrate that M2R plays a role in the growth and progression of NSCLC and suggest M2R antagonists may be an efficacious adjuvant therapy for NSCLC. PMID:25778781

  3. In vitro inhibition of lens epithelial cell growth by continuous wave Nd:YAG laser

    SciTech Connect

    Miyake, K.; Iwata, S.; Ando, F.; Daikuzono, N.; Federman, J.L.

    1989-04-01

    Bovine lens epithelial cells were suspended in MEM medium and subjected to continuous wave, low power, pulsed neodymium:yttrium-aluminum-garnet (Nd:YAG) laser irradiation. The temperature of each suspension was maintained at 36 degrees C. Laser applications ranged from 1 to 10 watts and from 100 to 2000 seconds, but the total dose to each of the epithelial cell suspension was 2000 J. Six to thirty-nine percent of the cells were dead immediately after irradiation. Surviving cells, cultured for 15 days, showed decreased attachment and failed to grow. These preliminary results suggest that the Nd:YAG laser may be used during cataract surgery to prevent subsequent lens epithelial cell proliferation and the resulting vision reduction and glare.

  4. Keratinocyte growth factor accelerates wound closure in airway epithelium during cyclic mechanical strain.

    PubMed

    Waters, C M; Savla, U

    1999-12-01

    The airway epithelium may be damaged by inhalation of noxious agents, in response to pathogens, or during endotracheal intubation and mechanical ventilation. Maintenance of an intact epithelium is important for lung fluid balance, and the loss of epithelium may stimulate inflammatory responses. Epithelial repair in the airways following injury must occur on a substrate that undergoes cyclic elongation and compression during respiration. We have previously shown that cyclic mechanical strain inhibits wound closure in the airway epithelium (Savla and Waters, 1998b). In this study, we investigated the stimulation of epithelial wound closure by keratinocyte growth factor (KGF) in vitro and the mechanisms by which KGF overcomes the inhibition due to mechanical strain. Primary cultures of normal human bronchial epithelial cells (NHBE) and a cell line of human airway epithelial cells, Calu 3, were grown on Silastic membranes, and a wound was scraped across the well. The wells were then exposed to cyclic strain using the Flexercell Strain Unit, and wound closure was measured. While cyclic elongation (20% maximum) and cyclic compression (approximately 2%) both inhibited wound closure in untreated wells, treatment with KGF (50 ng/ml) significantly accelerated wound closure and overcame the inhibition due to cyclic strain. Since wound closure involves cell spreading, migration, and proliferation, we investigated the effect of cyclic strain on cell area, cell-cell distance, and cell velocity at the wound edge. While the cell area increased in unstretched monolayers, the cell area of monolayers in compressed regions decreased significantly. Treatment with KGF increased the cell area in both cyclically elongated and compressed cells. Also, when cells were treated with KGF, cell velocity was significantly increased in both static and cyclically strained monolayers, and cyclic strain did not inhibit cell migration. These results suggest that KGF is an important factor in

  5. Expression of pigment epithelium‐derived factor and thrombospondin‐1 regulate proliferation and migration of retinal pigment epithelial cells

    PubMed Central

    Farnoodian, Mitra; Kinter, James B.; Yadranji Aghdam, Saeed; Zaitoun, Ismail; Sorenson, Christine M.; Sheibani, Nader

    2015-01-01

    Abstract Age‐related macular degeneration (AMD) is the leading cause of vision loss among elderly. Although the pathogenesis of AMD is associated with retinal pigmented epithelium (RPE) dysfunction and abnormal neovascularization the detailed mechanisms remain unresolved. RPE is a specialized monolayer of epithelial cells with important functions in ocular homeostasis. Pathological RPE damage contributes to major ocular conditions including retinal degeneration and irreversible loss of vision in AMD. RPE cells also assist in the maintenance of the ocular angiogenic balance by production of positive and negative regulatory factors including vascular endothelial growth factor (VEGF), thrombospondin‐1 (TSP1), and pigment epithelium‐derived factor (PEDF). The altered production of PEDF and TSP1, as endogenous inhibitors of angiogenesis and inflammation, by RPE cells have been linked to pathogenesis of AMD and choroidal and retinal neovascularization. However, lack of simple methods for isolation and culture of mouse RPE cells has resulted in limited knowledge regarding the cell autonomous role of TSP1 and PEDF in RPE cell function. Here, we describe a method for routine isolation and propagation of RPE cells from wild‐type, TSP1, and PEDF‐deficient mice, and have investigated their impact on RPE cell function. We showed that expression of TSP1 and PEDF significantly impacted RPE cell proliferation, migration, adhesion, oxidative state, and phagocytic activity with minimal effect on their basal rate of apoptosis. Together, our results indicated that the expression of PEDF and TSP1 by RPE cells play crucial roles not only in regulation of ocular vascular homeostasis but also have significant impact on their cellular function. PMID:25602019

  6. Antiviral factors and type I/III interferon expression associated with regulatory factors in the oral epithelial cells from HIV-1-serodiscordant couples.

    PubMed

    Cervantes, Cesar A C; Oliveira, Luanda M S; Manfrere, Kelly C G; Lima, Josenilson F; Pereira, Natalli Z; Duarte, Alberto J S; Sato, Maria N

    2016-01-01

    Individuals who remain HIV-seronegative despite repeated unprotected exposure to the virus are defined as exposed seronegative (ESN) individuals. Innate and adaptive immunity, as well as genetic factors, provide ESNs with important advantages that allow for low infection susceptibility. The majority of HIV-1-infected individuals undergo antiretroviral therapy, which can decrease the level of HIV-1 exposure in ESNs. We analyzed type I interferon (IFN)-related antiviral and regulatory factors in peripheral blood mononuclear cells (PBMCs) and oral epithelial cells from serodiscordant couples. Our findings revealed that ESNs did not induce the expression of antiviral factors (APOBEC-3G, TRIM5-α, SAMDH1, STING, TBk1) or regulatory factors (Trex, Foxo3, Socs3, IL-10) in PBMCs, unlike their HIV-1-infected partners. In contrast, ESNs upregulated APOBEC-3G and type I/III IFNs (IFNs-α,-β/-λ) in oral mucosal epithelial cells similar to their HIV-infected partners. The serodiscordant groups exhibited an increased expression of type I IFN-induced regulators, such as Trex and Foxo3, in oral epithelial cells. TLR7, TLR8 and TLR9 were expressed in oral epithelial cells of both ESNs and HIV-1-infected subjects. These findings revealed evidence of antiviral factors, type I/III interferon and regulatory factor expression only in the oral mucosal compartment of ESNs, while HIV-1-infected partners systemically and oral mucosal expressed the antiviral profile. PMID:27168019

  7. Antiviral factors and type I/III interferon expression associated with regulatory factors in the oral epithelial cells from HIV-1-serodiscordant couples

    PubMed Central

    Cervantes, Cesar A. C.; Oliveira, Luanda M. S.; Manfrere, Kelly C. G.; Lima, Josenilson F.; Pereira, Natalli Z.; Duarte, Alberto J. S.; Sato, Maria N.

    2016-01-01

    Individuals who remain HIV-seronegative despite repeated unprotected exposure to the virus are defined as exposed seronegative (ESN) individuals. Innate and adaptive immunity, as well as genetic factors, provide ESNs with important advantages that allow for low infection susceptibility. The majority of HIV-1-infected individuals undergo antiretroviral therapy, which can decrease the level of HIV-1 exposure in ESNs. We analyzed type I interferon (IFN)-related antiviral and regulatory factors in peripheral blood mononuclear cells (PBMCs) and oral epithelial cells from serodiscordant couples. Our findings revealed that ESNs did not induce the expression of antiviral factors (APOBEC-3G, TRIM5-α, SAMDH1, STING, TBk1) or regulatory factors (Trex, Foxo3, Socs3, IL-10) in PBMCs, unlike their HIV-1-infected partners. In contrast, ESNs upregulated APOBEC-3G and type I/III IFNs (IFNs-α,-β/-λ) in oral mucosal epithelial cells similar to their HIV-infected partners. The serodiscordant groups exhibited an increased expression of type I IFN-induced regulators, such as Trex and Foxo3, in oral epithelial cells. TLR7, TLR8 and TLR9 were expressed in oral epithelial cells of both ESNs and HIV-1-infected subjects. These findings revealed evidence of antiviral factors, type I/III interferon and regulatory factor expression only in the oral mucosal compartment of ESNs, while HIV-1-infected partners systemically and oral mucosal expressed the antiviral profile. PMID:27168019

  8. Epidermal Growth Factor Receptor Inhibitors: A Review of Cutaneous Adverse Events and Management

    PubMed Central

    Chanprapaph, K.; Vachiramon, V.; Rattanakaemakorn, P.

    2014-01-01

    Epidermal growth factor inhibitors (EGFRI), the first targeted cancer therapy, are currently an essential treatment for many advance-stage epithelial cancers. These agents have the superior ability to target cancers cells and better safety profile compared to conventional chemotherapies. However, cutaneous adverse events are common due to the interference of epidermal growth factor receptor (EGFR) signaling in the skin. Cutaneous toxicities lead to poor compliance, drug cessation, and psychosocial discomfort. This paper summarizes the current knowledge concerning the presentation and management of skin toxicity from EGFRI. The common dermatologic adverse events are papulopustules and xerosis. Less common findings are paronychia, regulatory abnormalities of hair growth, maculopapular rash, mucositis, and postinflammatory hyperpigmentation. Radiation enhances EGFRI rash due to synergistic toxicity. There is a positive correlation between the occurrence and severity of cutaneous adverse effects and tumor response. To date, prophylactic systemic tetracycline and tetracycline class antibiotics have proven to be the most effective treatment regime. PMID:24723942

  9. The pleiotropic roles of transforming growth factor beta inhomeostasis and carcinogenesis of endocrine organs.

    SciTech Connect

    Fleisch, Markus C.; Maxwell, Christopher A.; Barcellos-Hoff,Mary-Helen

    2006-01-13

    Transforming growth factor beta (TGF-beta) is a ubiquitous cytokine that plays a critical role in numerous pathways regulating cellular and tissue homeostasis. TGF-beta is regulated by hormones and is a primary mediator of hormone response in uterus, prostate and mammary gland. This review will address the role of TGF-beta in regulating hormone dependent proliferation and morphogenesis. The subversion of TGF-beta regulation during the processes of carcinogenesis, with particular emphasis on its effects on genetic stability and epithelial to mesenchymal transition (EMT), will also be examined. An understanding of the multiple and complex mechanisms of TGF-beta regulation of epithelial function, and the ultimate loss of TGF-beta function during carcinogenesis, will be critical in the design of novel therapeutic interventions for endocrine-related cancers.

  10. Serum deprivation response inhibits breast cancer progression by blocking transforming growth factor-β signaling.

    PubMed

    Tian, Yao; Yu, Yue; Hou, Li-Kun; Chi, Jiang-Rui; Mao, Jie-Fei; Xia, Li; Wang, Xin; Wang, Ping; Cao, Xu-Chen

    2016-03-01

    Serum deprivation response (SDPR), a key substrate for protein kinase C, play a critical role in inducing membrane curvature and participate in the formation of caveolae. However, the function of SDPR in cancer development and progression is still not clear. Here, we found that SDPR is downregulated in human breast cancer. Overexpression of SDPR suppresses cell proliferation and invasion in MDA-MB-231 cells, while depletion of SDPR promotes cell proliferation and invasion in MCF10A cells. Subsequently, SDPR depletion induces epithelial-mesenchymal transition (EMT)-like phenotype. Finally, knockdown of SDPR activates transforming growth factor-β (TGF-β) signaling by upregulation of TGF-β1 expression. In conclusion, our results showed that SDPR inhibits breast cancer progression by blocking TGF-β signaling. Serum deprivation response suppresses cell proliferation and invasion in breast cancer cells. SDPR depletion induces epithelial-mesenchymal transition by activation of TGF-β signaling. PMID:26749136

  11. Synergistic action of master transcription factors controls epithelial-to-mesenchymal transition.

    PubMed

    Chang, Hongyuan; Liu, Yuwei; Xue, Mengzhu; Liu, Haiyue; Du, Shaowei; Zhang, Liwen; Wang, Peng

    2016-04-01

    Epithelial-to-mesenchymal transition (EMT) is a complex multistep process in which phenotype switches are mediated by a network of transcription factors (TFs). Systematic characterization of all dynamic TFs controlling EMT state transitions, especially for the intermediate partial-EMT state, represents a highly relevant yet largely unexplored task. Here, we performed a computational analysis that integrated time-course EMT transcriptomic data with public cistromic data and identified three synergistic master TFs (ETS2, HNF4A and JUNB) that regulate the transition through the partial-EMT state. Overexpression of these regulators predicted a poor clinical outcome, and their elimination readily abolished TGF-β-induced EMT. Importantly, these factors utilized a clique motif, physically interact and their cumulative binding generally characterized EMT-associated genes. Furthermore, analyses of H3K27ac ChIP-seq data revealed that ETS2, HNF4A and JUNB are associated with super-enhancers and the administration of BRD4 inhibitor readily abolished TGF-β-induced EMT. These findings have implications for systematic discovery of master EMT regulators and super-enhancers as novel targets for controlling metastasis. PMID:26926107

  12. Synergistic action of master transcription factors controls epithelial-to-mesenchymal transition

    PubMed Central

    Chang, Hongyuan; Liu, Yuwei; Xue, Mengzhu; Liu, Haiyue; Du, Shaowei; Zhang, Liwen; Wang, Peng

    2016-01-01

    Epithelial-to-mesenchymal transition (EMT) is a complex multistep process in which phenotype switches are mediated by a network of transcription factors (TFs). Systematic characterization of all dynamic TFs controlling EMT state transitions, especially for the intermediate partial-EMT state, represents a highly relevant yet largely unexplored task. Here, we performed a computational analysis that integrated time-course EMT transcriptomic data with public cistromic data and identified three synergistic master TFs (ETS2, HNF4A and JUNB) that regulate the transition through the partial-EMT state. Overexpression of these regulators predicted a poor clinical outcome, and their elimination readily abolished TGF-β-induced EMT. Importantly, these factors utilized a clique motif, physically interact and their cumulative binding generally characterized EMT-associated genes. Furthermore, analyses of H3K27ac ChIP-seq data revealed that ETS2, HNF4A and JUNB are associated with super-enhancers and the administration of BRD4 inhibitor readily abolished TGF-β-induced EMT. These findings have implications for systematic discovery of master EMT regulators and super-enhancers as novel targets for controlling metastasis. PMID:26926107

  13. Development of an Autonomous, Dual Chamber Bioreactor for the Growth of 3-Dimensional Epithelial-Stromal Tissues in Microgravity

    NASA Technical Reports Server (NTRS)

    Patel, Zarana S.; Wettergreen, Matthew A.; Huff, Janice L.

    2014-01-01

    We are developing a novel, autonomous bioreactor that can provide for the growth and maintenance in microgravity of 3-D organotypic epithelial-stromal cultures that require an air-liquid interface. These complex 3-D tissue models accurately represent the morphological features, differentiation markers, and growth characteristics observed in normal human epithelial tissues, including the skin, esophagus, lung, breast, pancreas, and colon. However, because of their precise and complex culture requirements, including that of an air-liquid interface, these 3-D models have yet to be utilized for life sciences research aboard the International Space Station. The development of a bioreactor for these cultures will provide the capability to perform biological research on the ISS using these realistic, tissue-like human epithelial-stromal cell models and will contribute significantly to advances in fundamental space biology research on questions regarding microgravity effects on normal tissue development, aging, cancer, and other disease processes. It will also allow for the study of how combined stressors, such as microgravity with radiation and nutritional deficiencies, affect multiple biological processes and will provide a platform for conducting countermeasure investigations on the ISS without the use of animal models. The technology will be autonomous and consist of a cell culture chamber that provides for air-liquid, liquid-liquid, and liquid-air exchanges within the chambers while maintaining the growth and development of the biological samples. The bioreactor will support multiple tissue types and its modular design will provide for incorporation of add-on capabilities such as microfluidics drug delivery, media sampling, and in situ biomarker analysis. Preliminary flight testing of the hardware will be conducted on a parabolic platform through NASA's Flight Opportunities Program.

  14. Characterization of insulin-like growth factor I and epidermal growth factor receptors in meningioma

    SciTech Connect

    Kurihara, M.; Tokunaga, Y.; Tsutsumi, K.; Kawaguchi, T.; Shigematsu, K.; Niwa, M.; Mori, K. )

    1989-10-01

    Receptors for insulin-like growth factor I (IGF-I) and epidermal growth factor (EGF) were localized and characterized in eight samples of human meningioma (four fibrous, two meningothelial, and two angioblastic types), using quantitative autoradiographic techniques. Effects of both growth factors on deoxyribonucleic acid (DNA) synthesis in the cultured meningioma cells were examined. High numbers of specific binding sites for both IGF-I and EGF were homogeneously present in tissue sections derived from fibrous and meningothelial types of meningiomas, whereas binding sites for these growth factors were not detectable in adjacent leptomeninges. While relatively large numbers of IGF-I binding sites were located in the wall of the intratumoral vasculature, the number of binding sites in the stromal component was lower in angioblastic-type meningiomas, including a low number of EGF binding sites detected only in the stromal portion. Scatchard analysis revealed the presence of a single class of high-affinity binding sites for both IGF-I and EGF in the meningiomas examined (dissociation constant (Kd) = 0.6 to 2.9 nM, and the maximum number of binding sites (Bmax) = 16 to 80 fmol/mg for IGF-I; and Kd = 0.6 to 4.0 nM, Bmax = 3 to 39 fmol/mg for EGF). Both growth factors increased the synthesis of DNA, in a dose-dependent manner, as measured by 3H-thymidine incorporation. The combination of IGF-I and EGF synergistically stimulated the synthesis of DNA, and the effects seen with 10% fetal bovine serum could be reproduced at a concentration of 10(-10) M. These observations can be interpreted to mean that both IGF-I and EGF may be involved in the growth modulation of meningiomas, possibly through paracrine or autocrine mechanisms.

  15. Vasoactive-intestinal-Peptide (vip) modulates the growth fraction of epithelial skin cells.

    PubMed

    Wollina, U; Bonnekoh, B; Mahrle, G

    1992-06-01

    Using the human keratinocyte cell line HaCaT, modifications of the growth fraction due to vasoactive intestinal peptide (VIP) were determined by immunostaining with monoclonal antibody Ki67. In addition, the expression of VIP receptor and epidermal growth factor (EGF) receptor have been analysed. VIP (10-(7) to 10-(11) M) produced an almost doubling of the total number of Ki67-positive cells in cultures with 2% fetal calf serum (FCS), wheras it was ineffective in FCS-free and 10% FCS cultures. The nuclear Ki67-staining patterns were classified into four categories. In FCS-free cultures VIP induced a shift from type III (light nucleus, staining nuclei) to type II (multiple, intensely stained spots). In cultures with 2% FCS, VIP induced a shift from type II to type III. VIP receptor expression was facilitated by VIP, when cells were grown in a medium supplemented with 10% FCS. VIP increased EGF receptor expression in FCS-free cultures but decreased the number EGF receptor-positive cells in experiments with 2% FCS. In conclusion, VIP is capable to modulate the growth fraction and receptor expression of HaCaT cells in vitro. The effects are dependent on the concentration of FCS within the culture medium. The findings might be of interest for keratinocyte pathology in general and dermatooncology in particular. PMID:21584504

  16. Human breast cancer cells and normal mammary epithelial cells: retinol metabolism and growth inhibition by the retinol metabolite 4-oxoretinol.

    PubMed

    Chen, A C; Guo, X; Derguini, F; Gudas, L J

    1997-10-15

    To understand the signaling and growth-inhibitory effects of retinoids, we have examined the metabolism of [3H]retinol in a number of estrogen receptor-positive (ER+) and estrogen receptor-negative (ER-) human breast cancer cell lines. We have also assayed the metabolism of [3H]retinol in normal human mammary epithelial cells. The ER+ breast cancer cell lines MCF-7 and T47D produce [3H]4-oxoretinol from [3H]retinol; the production of [3H]4-oxoretinol is increased by initial culture in the presence of nonradiolabeled retinoic acid (RA) or N-(4-hydroxyphenyl)retinamide, indicating that these drugs enhance [3H]retinol metabolism to [3H]4-oxoretinol. No metabolism of [3H]retinol to [3H]RA in these ER+ tumor lines was detected. ER- breast cancer lines MDA-MB-231, MDA-MB-468, and BT20 do not metabolize [3H]retinol to [3H]4-oxoretinol. In the ER- tumor lines, most of the [3H]retinol remains unmetabolized during the 24-h culture period; MDA-MB-468 and BT20 metabolize some [3H]retinol to [3H]RA. Unlike the majority of the tumor lines, the normal human breast epithelial cell strains AD074 and MCF10A rapidly metabolize [3H]retinol to [3H]retinyl esters. No detectable [3H]RA is produced from [3H]retinol in AD074 and MCF10A cells. Thus, the normal breast epithelial strains, the ER+ tumor lines and the ER- tumor lines differ greatly in their pathways of [3H]retinol metabolism. The levels of cellular retinol binding protein-I mRNA expression are not correlated with the levels or types of various retinol metabolites. Whereas the normal breast epithelial cells and the ER+ tumor lines are growth inhibited by RA, N-(4-hydroxyphenyl)retinamide, and 4-oxoretinol, only the 4-oxoretinol is growth inhibitory in the ER- tumor lines. The cellular retinoic acid-binding protein II mRNA levels are not correlated with the growth inhibition by RA or 4-oxoretinol in the normal and tumor lines. PMID:9377581

  17. Up-regulation of early growth response gene 1 (EGR-1) via ERK1/2 signals attenuates sulindac sulfide-mediated cytotoxicity in the human intestinal epithelial cells

    SciTech Connect

    Moon, Yuseok Yang, Hyun; Kim, Yung Bu

    2007-09-01

    Non-steroidal anti-inflammatory drugs (NSAIDs) are used to relieve pain and inflammation and have also received considerable attention because of their preventive effects against human cancer. However, the drug application is sometimes limited by the severe gastrointestinal ulcers and mucosal complications. In the present study, NSAID sulindac sulfide was investigated for the cytotoxic injury in the intestinal epithelial cells in association with an immediate inducible factor, early growth response gene 1 (EGR-1). Previously we reported that sulindac sulfide can suppress tumor cell invasion by inducing EGR-1. Extending the previous study, EGR-1 induction by sulindac sulfide was observed both in the non-transformed and transformed human intestinal epithelial cell lines. In terms of signaling pathway, ERK1/2 MAP kinases and its substrate Elk-1 transcription factor were involved in the sulindac sulfide-induced EGR-1 gene expression. Moreover, sulindac sulfide stimulated the nuclear translocation of the transcription factor EGR-1, which was also mediated by ERK1/2 signaling pathway. The roles of EGR-1 signals in the apoptotic cell death were assessed in the intestinal epithelial cells. Suppression of EGR-1 expression retarded cellular growth and colony forming activity in the intestinal epithelial cells. Moreover, induced EGR-1 ameliorated sulindac sulfide-mediated apoptotic cell death and enhanced the cellular survival. Taken all together, sulindac sulfide activated ERK1/2 MAP kinases which then mediated EGR-1 induction and nuclear translocation, all of which played important roles in the cellular survival from NSAID-mediated cytotoxicity in the human intestinal epithelial cells, implicating the protective roles of EGR-1 in the NSAID-mediated mucosal injuries.

  18. Role of Insulin-like Growth Factor Binding Protein-3 in Allergic Airway Remodeling

    PubMed Central

    Veraldi, Kristen L.; Gibson, Bethany T.; Yasuoka, Hidekata; Myerburg, Michael M.; Kelly, Elizabeth A.; Balzar, Silvana; Jarjour, Nizar N.; Pilewski, Joseph M.; Wenzel, Sally E.; Feghali-Bostwick, Carol A.

    2009-01-01

    Rationale: The hallmarks of allergic asthma are airway inflammation, obstruction, and remodeling. Airway remodeling may lead to irreversible airflow obstruction with increased morbidity and mortality. Despite advances in the treatment of asthma, the mechanisms underlying airway remodeling are still poorly understood. We reported that insulin-like growth factor (IGF) binding proteins (IGFBPs) contribute to extracellular matrix deposition in idiopathic pulmonary fibrosis; however, their contribution to airway remodeling in asthma has not been established. Objectives: We hypothesized that IGFBP-3 is overexpressed in asthma and contributes to airway remodeling. Methods: We evaluated levels of IGFBP-3 in tissues and bronchoalveolar lavage fluid from patients with asthma at baseline and 48 hours after allergen challenge, in reparative epithelium in an in vitro wounding assay, and in conditioned media from cytokine- and growth factor–stimulated primary epithelial cells. Measurements and Main Results: IGFBP-3 levels and distribution were evaluated by Western blot, ELISA, and immunofluorescence. IGFBP-3 is increased in vivo in the airway epithelium of patients with asthma compared with normal control subjects. The concentration of IGFBP-3 is increased in the bronchoalveolar lavage fluid of patients with asthma after allergen challenge, its levels are increased in reparative epithelium in an in vitro wounding assay and in the conditioned medium of primary airway epithelial cell cultures stimulated with IGF-I. Conclusions: Our results suggest that one mechanism of allergic airway remodeling is through the secretion of the profibrotic IGFBP-3 from IGF-I–stimulated airway epithelial cells during allergic inflammation. PMID:19608721

  19. Regulation of insulin-like growth factor (IGF)-binding protein expression by growth factors and cytokines alters IGF-mediated proliferation of postnatal lung fibroblasts.

    PubMed

    Price, Wayne A

    2004-06-01

    Postnatal day 5 is the beginning of septation and the peak of postnatal fibroblast proliferation. The author and colleagues studied fibroblasts from this developmental time period to determine factors that regulate cell proliferation. Exposure of cells to insulin-like growth factor (IGF)-I for 48 hours increased cell number whereas exposure to epithelial growth factor (EGF), platelet-derived growth factor (PDGF)-BB, fibroblast growth factor (FGF)-7, FGF-2, tumor necrosis factor-alpha (TNF-alpha), or interleukin (L)-1beta did not alter cell number. Long[R3]IGF-I (a synthetic IGF analog with reduced affinity for IGF-binding proteins [IGFBPs]) was more potent than IGF-I, with half-maximal stimulation at a dose of 0.6 nM for long[R3]IGF-I compared to 1.5 nM for IGF-I, suggesting that IGFBPs in the conditioned medium (CM) inhibit IGF activity. Addition of exogenous IGFBP-3 inhibited the IGF-stimulated increase in cell number. Addition of IGFBP-4 did not alter IGF activity because IGF-I stimulated proteolysis of IGFBP-4. The expression of mRNA for PAPP-A (a known IGFBP-4 protease) suggests that the clearance of IGFBP-4 is mediated by pregnancy-associated plasma protein (PAPP)-A. Exposure of cells to TNF-alpha or IL-1beta increased IGFBP-3 mRNA abundance and IGFBP-3 protein in CM. PDGF-BB and IL-1beta increased IGFBP-4 protein abundance and PDGF-BB and dibutyryl cAMP increased IGFBP-4 mRNA. The increase in CM IGFBP-3 following TNF-alpha exposure blocked IGF-mediated cell proliferation, suggesting that the growth factor- and cytokine-mediated changes in IGFBP abundance regulate postnatal fibroblast cell proliferation. PMID:15204833

  20. Fibroblast growth factor 23 and bone mineralisation

    PubMed Central

    Guo, Yu-Chen; Yuan, Quan

    2015-01-01

    Fibroblast growth factor 23 (FGF23) is a hormone that is mainly secreted by osteocytes and osteoblasts in bone. The critical role of FGF23 in mineral ion homeostasis was first identified in human genetic and acquired rachitic diseases and has been further characterised in animal models. Recent studies have revealed that the levels of FGF23 increase significantly at the very early stages of chronic kidney disease (CKD) and may play a critical role in mineral ion disorders and bone metabolism in these patients. Our recent publications have also shown that FGF23 and its cofactor, Klotho, may play an independent role in directly regulating bone mineralisation instead of producing a systematic effect. In this review, we will discuss the new role of FGF23 in bone mineralisation and the pathophysiology of CKD-related bone disorders. PMID:25655009

  1. Neuropeptides as lung cancer growth factors.

    PubMed

    Moody, Terry W; Moreno, Paola; Jensen, Robert T

    2015-10-01

    This manuscript is written in honor of the Festschrift for Abba Kastin. I met Abba at a Society for Neuroscience meeting and learned that he was Editor-in-Chief of the Journal Peptides. I submitted manuscripts to the journal on "Neuropeptides as Growth Factors in Cancer" and subsequently was named to the Editorial Advisory Board. Over the past 30 years I have published dozens of manuscripts in Peptides and reviewed hundreds of submitted manuscripts. It was always rewarding to interact with Abba, a consummate professional. When I attended meetings in New Orleans I would sometimes go out to dinner with him at the restaurant "Commanders Palace". When I chaired the Summer Neuropeptide Conference we were honored to have him receive the Fleur Strand Award one year in Israel. I think that his biggest editorial contribution has been the "Handbook of Biologically Active Peptides." I served as a Section Editor on "Cancer/Anticancer Peptides" and again found that it was a pleasure working with him. This review focuses on the mechanisms by which bombesin-like peptides, neurotensin and vasoactive intestinal peptide regulate the growth of lung cancer. PMID:25836991

  2. The Fibroblast Growth Factor signaling pathway

    PubMed Central

    Ornitz, David M; Itoh, Nobuyuki

    2015-01-01

    The signaling component of the mammalian Fibroblast Growth Factor (FGF) family is comprised of eighteen secreted proteins that interact with four signaling tyrosine kinase FGF receptors (FGFRs). Interaction of FGF ligands with their signaling receptors is regulated by protein or proteoglycan cofactors and by extracellular binding proteins. Activated FGFRs phosphorylate specific tyrosine residues that mediate interaction with cytosolic adaptor proteins and the RAS-MAPK, PI3K-AKT, PLCγ, and STAT intracellular signaling pathways. Four structurally related intracellular non-signaling FGFs interact with and regulate the family of voltage gated sodium channels. Members of the FGF family function in the earliest stages of embryonic development and during organogenesis to maintain progenitor cells and mediate their growth, differentiation, survival, and patterning. FGFs also have roles in adult tissues where they mediate metabolic functions, tissue repair, and regeneration, often by reactivating developmental signaling pathways. Consistent with the presence of FGFs in almost all tissues and organs, aberrant activity of the pathway is associated with developmental defects that disrupt organogenesis, impair the response to injury, and result in metabolic disorders, and cancer. © 2015 Wiley Periodicals, Inc. PMID:25772309

  3. Platelet-activating factor induces collagenase expression in corneal epithelial cells.

    PubMed Central

    Bazan, H E; Tao, Y; Bazan, N G

    1993-01-01

    Platelet-activating factor (PAF), a potent lipid mediator involved in inflammatory and immune responses, accumulates rapidly in response to injury in a variety of tissues, including the corneal epithelium. However, the precise role of this compound in the cascade of events following insult has not been defined. Here we examined the effect of PAF on gene expression in the epithelial cells of rabbit corneas in organ culture. We found that incubation with 100 nM methylcarbamoyl PAF, a nonhydrolyzable analog of PAF, produced rapid transient 2.8- and 3.5-fold increases in the expression of c-fos and c-jun, respectively, at 1 hr, followed by increased expression of the collagenase type I gene beginning at 3 hr and peaking at 14-fold by 8 hr. Addition of the protein-synthesis-inhibitor cycloheximide superinduced c-fos and c-jun, strongly potentiating the PAF effect, but inhibited the induction of collagenase type I expression, suggesting the existence of a transcriptional factor linking the two events. BN-50730, a selective antagonist of intracellular PAF-binding sites, blocked the expression of the immediate-early genes as well as the increase in collagenase type I mRNA. Our results suggest that one of the functions of PAF may be to enhance the breakdown of the extracellular matrix as a part of the remodeling process during corneal wound healing after injury. Pathologically, a PAF-induced overproduction of collagenase may be a factor in the development of corneal ulcers, as well as other pathophysiological conditions such as cartilage destruction in arthritis. If so, inhibitors of this signal-transduction pathway may be useful as tools for further investigation and, eventually, as therapeutic agents to treat such disorders. Images Fig. 1 Fig. 2 PMID:8378347

  4. INTRODUCTION OF A HA-RAS ONCOGENE INTO RAT LIVER EPITHELIAL CELLS AND PARENCHYMAL HEPATACYTES CONFERS REISTANCE TO THE GROWTH INHIBITORY EFFECTS OF TGF-B

    EPA Science Inventory

    Growth of rat liver epithelial cells (RLEC) and primary cultures of parenchymal hepatocytes is potentially inhibited by TGF-b. Transfection of a mutaged Ha-ras oncogene, but not a human c-myc oncogene, into RLEC resulted in cell lines ressitant to growth inhibition by TGF-b under...

  5. Regulation of tumorigenesis in oral epithelial cells by defined reprogramming factors Oct4 and Sox2.

    PubMed

    Cai, Jinghua; He, Baoxia; Li, Xinming; Sun, Minglei; Lam, Alfred King-Yin; Qiao, Bin; Qiu, Weiliu

    2016-08-01

    Oct4 and Sox2 are pluripotent stem cell factors but the interplay between them in tumorigenesis is unclear. The aim of the present study was to investigate the roles of Oct4 and Sox2 in the reprogramming of oral cancer stem cells. One or both Oct4 and Sox2 were overexpressed in immortalized oral epithelial (hTERT+-OME) cells by lentivirus transduction. In addition, Oct4 and Sox2 proteins in two oral squamous cell carcinoma cell (OSCC) lines (Cal27 and primary cultured OSCC from a T2N2M0 patient) were individually or combinedly knocked down by shRNA. The results showed that the doubly transduced (Oct4+Sox2+) cells could trigger neoplasms in immunodeficient mice after lentivirus transduction, but single transduced (Oct4+ or Sox2+) cells had no tumor formation ability. The knockdown Sox2low and knockdown Oct4lowSox2low cells resulted in decreased tumor size in the immunodeficient mice but the single knockdown Oct4low cancer cells acquired more aggressive xenografts. Our findings suggest that Oct4+Sox2+ cells may be reprogrammed cancer stem cells inducing oral carcinogenesis. PMID:27279579

  6. Octamer transcription factor 1 mediates epithelial-mesenchymal transition in colorectal cancer.

    PubMed

    Li, Yuji; Dong, Ming; Kong, Fanmin; Zhou, Jianping

    2015-12-01

    Colorectal cancer (CRC) is one of the most common cancer types worldwide. Octamer transcription factor 1 (OCT1) is associated with tumor progression and a poor patient survival rate. However, little is known regarding the effect of OCT1 in CRC. Moreover, because the epithelial-to-mesenchymal transition (EMT) is a key player in metastasis, whether OCT1 induces EMT in CRC remains unclear. In the present study, we investigate the role of OCT1 in CRC and its expression pattern and clinical significance. The expression of OCT1 in CRC tissues and the adjacent noncancerous tissues was detected using quantitative real-time PCR (QRT-PCR), Western blot, and immunohistochemistry analyses. In addition, silencing of OCT1 with small interfering RNA (siRNA) was performed in CRC cell lines, and the impact on proliferation, migration, and the EMT marker of CRC was analyzed. Our results found that OCT1 levels were significant higher in CRC tissues compared with the adjacent noncancerous tissues. Furthermore, OCT1 siRNA significantly reduced the proliferation rate of SW620 and LoVo cells, inhibited the migration and invasion, and could reverse EMT in these two CRC cells, indicating that OCT1 plays a critical role in CRC progression. PMID:26178483

  7. The hypoxia factor Hif-1α controls neural crest chemotaxis and epithelial to mesenchymal transition

    PubMed Central

    Barriga, Elias H.; Maxwell, Patrick H.

    2013-01-01

    One of the most important mechanisms that promotes metastasis is the stabilization of Hif-1 (hypoxia-inducible transcription factor 1). We decided to test whether Hif-1α also was required for early embryonic development. We focused our attention on the development of the neural crest, a highly migratory embryonic cell population whose behavior has been likened to cancer metastasis. Inhibition of Hif-1α by antisense morpholinos in Xenopus laevis or zebrafish embryos led to complete inhibition of neural crest migration. We show that Hif-1α controls the expression of Twist, which in turn represses E-cadherin during epithelial to mesenchymal transition (EMT) of neural crest cells. Thus, Hif-1α allows cells to initiate migration by promoting the release of cell–cell adhesions. Additionally, Hif-1α controls chemotaxis toward the chemokine SDF-1 by regulating expression of its receptor Cxcr4. Our results point to Hif-1α as a novel and key regulator that integrates EMT and chemotaxis during migration of neural crest cells. PMID:23712262

  8. Decay-Accelerating Factor Binding Determines the Entry Route of Echovirus 11 in Polarized Epithelial Cells▿

    PubMed Central

    Sobo, Komla; Rubbia-Brandt, Laura; Brown, T. David K.; Stuart, Amanda D.; McKee, Thomas A.

    2011-01-01

    The interaction between echovirus 11 strain 207 (EV11-207) and decay-accelerating factor (DAF or CD55) at the apical surface of polarized Caco-2 cells results in rapid transport of the virus to tight junctions and in its subsequent uptake. A virus mutant (EV11-207R) which differs at 6 amino acids and whose affinity for DAF is apparently significantly lower remains at the apical surface, from where its uptake occurs. Binding of EV11-207 to DAF and its transport to tight junctions result in a loss of function of the junctions. In contrast, the mutant virus EV11-207R is not transferred to tight junctions, nor does it impair the integrity of these junctions. Cholesterol depletion from the apical membrane leads to DAF aggregation and, presumably, internalization and inhibits infection by EV11-207. However, infection by EV11-207R is significantly less sensitive to cholesterol depletion than infection by EV11-207, confirming the DAF requirement for EV11-207, but not EV11-207R, to infect cells. These data strongly indicate that in the case of infection of polarized epithelial cells by echovirus 11, DAF binding appears be a key determinant in the choice of entry pathway, at least in cell culture. PMID:21917947

  9. Regulation of tumorigenesis in oral epithelial cells by defined reprogramming factors Oct4 and Sox2

    PubMed Central

    Cai, Jinghua; He, Baoxia; Li, Xinming; Sun, Minglei; Lam, Alfred King-Yin; Qiao, Bin; Qiu, Weiliu

    2016-01-01

    Oct4 and Sox2 are pluripotent stem cell factors but the interplay between them in tumorigenesis is unclear. The aim of the present study was to investigate the roles of Oct4 and Sox2 in the reprogramming of oral cancer stem cells. One or both Oct4 and Sox2 were overexpressed in immortalized oral epithelial (hTERT+-OME) cells by lentivirus transduction. In addition, Oct4 and Sox2 proteins in two oral squamous cell carcinoma cell (OSCC) lines (Cal27 and primary cultured OSCC from a T2N2M0 patient) were individually or combinedly knocked down by shRNA. The results showed that the doubly transduced (Oct4+Sox2+) cells could trigger neoplasms in immunodeficient mice after lentivirus transduction, but single transduced (Oct4+ or Sox2+) cells had no tumor formation ability. The knockdown Sox2low and knockdown Oct4lowSox2low cells resulted in decreased tumor size in the immunodeficient mice but the single knockdown Oct4low cancer cells acquired more aggressive xenografts. Our findings suggest that Oct4+Sox2+ cells may be reprogrammed cancer stem cells inducing oral carcinogenesis. PMID:27279579

  10. Polyelectrolyte Complex for Heparin Binding Domain Osteogenic Growth Factor Delivery.

    PubMed

    Wing Moon Lam, Raymond; Abbah, Sunny Akogwu; Ming, Wang; Naidu, Mathanapriya; Ng, Felly; Tao, Hu; Goh Cho Hong, James; Ting, Kang; Hee Kit, Wong

    2016-01-01

    During reconstructive bone surgeries, supraphysiological amounts of growth factors are empirically loaded onto scaffolds to promote successful bone fusion. Large doses of highly potent biological agents are required due to growth factor instability as a result of rapid enzymatic degradation as well as carrier inefficiencies in localizing sufficient amounts of growth factor at implant sites. Hence, strategies that prolong the stability of growth factors such as BMP-2/NELL-1, and control their release could actually lower their efficacious dose and thus reduce the need for larger doses during future bone regeneration surgeries. This in turn will reduce side effects and growth factor costs. Self-assembled PECs have been fabricated to provide better control of BMP-2/NELL-1 delivery via heparin binding and further potentiate growth factor bioactivity by enhancing in vivo stability. Here we illustrate the simplicity of PEC fabrication which aids in the delivery of a variety of growth factors during reconstructive bone surgeries. PMID:27585207

  11. Design of Growth Factor Sequestering Biomaterials

    PubMed Central

    Belair, David G.; Le, Ngoc Nhi; Murphy, William L.

    2014-01-01

    Growth factors (GFs) are major regulatory proteins that can govern cell fate, migration, and organization. Numerous aspects of the cell milieu can modulate cell responses to GFs, and GF regulation is often achieved by the native extracellular matrix (ECM). For example, the ECM can sequester GFs and thereby control GF bioavailability. In addition, GFs can exert distinct effects depending on whether they are sequestered in solution, at two-dimensional interfaces, or within three-dimensional matrices. Understanding how the context of GF sequestering impacts cell function in the native ECM can instruct the design of soluble or insoluble GF sequestering moieties, which can then be used in a variety of bioengineering applications. This Feature Article provides an overview of the natural mechanisms of GF sequestering in the cell milieu, and reviews the recent bioengineering approaches that have sequestered GFs to modulate cell function. Results to date demonstrate that the cell response to GF sequestering depends on the affinity of the sequestering interaction, the spatial proximity of sequestering in relation to cells, the source of the GF (supplemented or endogenous), and the phase of the sequestering moiety (soluble or insoluble). We highlight the importance of context for the future design of biomaterials that can leverage endogenous molecules in the cell milieu and mitigate the need for supplemented factors. PMID:25182455

  12. Stanniocalcin-1 is induced by hypoxia inducible factor in rat alveolar epithelial cells.

    PubMed

    Ito, Yoko; Zemans, Rachel; Correll, Kelly; Yang, Ivana V; Ahmad, Aftab; Gao, Bifeng; Mason, Robert J

    2014-10-01

    Alveolar type II (ATII) cells remain differentiated and express surfactant proteins when cultured at an air-liquid (A/L) interface. When cultured under submerged conditions, ATII cells dedifferentiate and change their gene expression profile. We have previously shown that gene expression under submerged conditions is regulated by hypoxia inducible factor (HIF) signaling due to focal hypoxia resulting from ATII cell metabolism. Herein, we sought to further define gene expression changes in ATII cells cultured under submerged conditions. We performed a genome wide microarray on RNA extracted from rat ATII cells cultured under submerged conditions for 24-48h after switching from an A/L interface. We found significant alterations in gene expression, including upregulation of the HIF target genes stanniocalcin-1 (STC1), tyrosine hydroxylase (Th), enolase (Eno) 2, and matrix metalloproteinase (MMP) 13, and we verified upregulation of these genes by RT-PCR. Because STC1, a highly evolutionarily conserved glycoprotein with anti-inflammatory, anti-apoptotic, anti-oxidant, and wound healing properties, is widely expressed in the lung, we further explored the potential functions of STC1 in the alveolar epithelium. We found that STC1 was induced by hypoxia and HIF in rat ATII cells, and this induction occurred rapidly and reversibly. We also showed that recombinant human STC1 (rhSTC1) enhanced cell motility with extended lamellipodia formation in alveolar epithelial cell (AEC) monolayers but did not inhibit the oxidative damage induced by LPS. We also confirmed that STC1 was upregulated by hypoxia and HIF in human lung epithelial cells. In this study, we have found that several HIF target genes including STC1 are upregulated in AECs by a submerged condition, that STC1 is regulated by hypoxia and HIF, that this regulation is rapidly and reversibly, and that STC1 enhances wound healing moderately in AEC monolayers. However, STC1 did not inhibit oxidative damage in rat AECs

  13. Distinction Between Cell Proliferation and Apoptosis Signals Regulated by Brain-Derived Neurotrophic Factor in Human Periodontal Ligament Cells and Gingival Epithelial Cells.

    PubMed

    Kashiwai, Kei; Kajiya, Mikihito; Matsuda, Shinji; Ouhara, Kazuhisa; Takeda, Katsuhiro; Takata, Takashi; Kitagawa, Masae; Fujita, Tsuyoshi; Shiba, Hideki; Kurihara, Hidemi

    2016-07-01

    Previously, we reported that brain-derived neurotrophic factor (BDNF) enhances periodontal tissue regeneration by inducing periodontal ligament cell proliferation in vivo. In addition, the down growth of gingival epithelial cells, which comprises a major obstacle to the regeneration, was not observed. However, the underlying molecular mechanism is still unclear. Therefore, this study aimed to investigate the effect of BDNF on cell proliferation and apoptosis in human periodontal ligament (HPL) cells and human gingival epithelial cells (OBA9 cells) and to explore the molecular mechanism in vitro. HPL cells dominantly expressed a BDNF receptor, TrkB, and BDNF increased cell proliferation and ERK phosphorylation. However, its proliferative effect was diminished by a MEK1/2 inhibitor (U0126) and TrkB siRNA transfection. Otherwise, OBA9 cells showed a higher expression level of p75, which is a pan-neurotrophin receptor, than that of HPL cells. BDNF facilitated not cell proliferation but cell apoptosis and JNK phosphorylation in OBA9 cells. A JNK inhibitor (SP600125) and p75 siRNA transfection attenuated the BDNF-induced cell apoptosis. Moreover, OBA9 cells pretreated with SP600125 or p75 siRNA showed cell proliferation by BDNF stimulation, though it was reduced by U0126 and TrkB siRNA. Interestingly, overexpression of p75 in HPL cells upregulated cell apoptosis and JNK phosphorylation by BDNF treatment. These results indicated that TrkB-ERK signaling regulates BDNF-induced cell proliferation, whereas p75-JNK signaling plays roles in cell apoptotic and cytostatic effect of BDNF. Overall, BDNF activates periodontal ligament cells proliferation and inhibits the gingival epithelial cells growth via the distinct pathway. J. Cell. Biochem. 117: 1543-1555, 2016. © 2015 Wiley Periodicals, Inc. PMID:26581032

  14. Interaction of E-cadherin and PTEN regulates morphogenesis and growth arrest in human mammary epithelial cells

    SciTech Connect

    Fournier, Marcia V.; Fata, Jimmie E.; Martin, Katherine J.; Yaswen, Paul; Bissell, Mina J.

    2009-06-03

    PTEN is a dual function phosphatase with tumor suppressor function compromised in a wide spectrum of cancers. Because tissue polarity and architecture are crucial modulators of normal and malignant behavior, we postulated that PTEN may play a role in maintenance of tissue integrity. We used two non-malignant human mammary epithelial cell lines (HMECs) that form polarized, growth-arrested structures (acini) when cultured in 3-dimensional laminin-rich extracellular matrix gels (3D lrECM). As acini begin to form, PTEN accumulates in both the cytoplasm, and at cell-cell contacts where it colocalizes with E-cadherin/{beta}-catenin complex. Reduction of PTEN levels by shRNA in lrECM prevents formation of organized breast acini and disrupts growth arrest. Importantly, disruption of acinar polarity and cell-cell contact by E-cadherin function-blocking antibodies reduces endogenous PTEN protein levels and inhibits its accumulation at cell-cell contacts. Conversely, in SKBR3 breast cancer cells lacking endogenous E-cadherin expression, exogenous introduction of E-cadherin gene causes induction of PTEN expression and its accumulation at sites of cell interactions. These studies provide evidence that E-cadherin regulates both the PTEN protein levels and its recruitment to cell-cell junctions in 3D lrECM indicating a dynamic reciprocity between architectural integrity and the levels and localization of PTEN. This interaction thus appears to be a critical integrator of proliferative and morphogenetic signaling in breast epithelial cells.

  15. Enforced epithelial expression of IGF-1 causes hyperplastic prostate growth while negative selection is requisite for spontaneous metastogenesis

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The insulin-like growth factor-1 (IGF-1) signaling axis is important for cell growth, differentiation, and survival, and increased serum IGF is a risk factor for prostate and other cancers. To study IGF-1 action on the prostate, we created transgenic (PB-Des) mice that specifically express human IGF...

  16. Inhibition of fibroblast growth factor receptor 3-dependent lung adenocarcinoma with a human monoclonal antibody.

    PubMed

    Yin, Yongjun; Ren, Xiaodi; Smith, Craig; Guo, Qianxu; Malabunga, Maria; Guernah, Ilhem; Zhang, Yiwei; Shen, Juqun; Sun, Haijun; Chehab, Nabil; Loizos, Nick; Ludwig, Dale L; Ornitz, David M

    2016-05-01

    Activating mutations in fibroblast growth factor receptor 3 (FGFR3) have been identified in multiple types of human cancer and in congenital birth defects. In human lung cancer, fibroblast growth factor 9 (FGF9), a high-affinity ligand for FGFR3, is overexpressed in 10% of primary resected non-small cell lung cancer (NSCLC) specimens. Furthermore, in a mouse model where FGF9 can be induced in lung epithelial cells, epithelial proliferation and ensuing tumorigenesis is dependent on FGFR3. To develop new customized therapies for cancers that are dependent on FGFR3 activation, we have used this mouse model to evaluate a human monoclonal antibody (D11) with specificity for the extracellular ligand-binding domain of FGFR3, that recognizes both human and mouse forms of the receptor. Here, we show that D11 effectively inhibits signaling through FGFR3 in vitro, inhibits the growth of FGFR3-dependent FGF9-induced lung adenocarcinoma in mice, and reduces tumor-associated morbidity. Given the potency of FGF9 in this mouse model and the absolute requirement for signaling through FGFR3, this study validates the D11 antibody as a potentially useful and effective reagent for treating human cancers or other pathologies that are dependent on activation of FGFR3. PMID:27056048

  17. Inhibition of fibroblast growth factor receptor 3-dependent lung adenocarcinoma with a human monoclonal antibody

    PubMed Central

    Yin, Yongjun; Ren, Xiaodi; Smith, Craig; Guo, Qianxu; Malabunga, Maria; Guernah, Ilhem; Zhang, Yiwei; Shen, Juqun; Sun, Haijun; Chehab, Nabil; Loizos, Nick; Ludwig, Dale L.; Ornitz, David M.

    2016-01-01

    ABSTRACT Activating mutations in fibroblast growth factor receptor 3 (FGFR3) have been identified in multiple types of human cancer and in congenital birth defects. In human lung cancer, fibroblast growth factor 9 (FGF9), a high-affinity ligand for FGFR3, is overexpressed in 10% of primary resected non-small cell lung cancer (NSCLC) specimens. Furthermore, in a mouse model where FGF9 can be induced in lung epithelial cells, epithelial proliferation and ensuing tumorigenesis is dependent on FGFR3. To develop new customized therapies for cancers that are dependent on FGFR3 activation, we have used this mouse model to evaluate a human monoclonal antibody (D11) with specificity for the extracellular ligand-binding domain of FGFR3, that recognizes both human and mouse forms of the receptor. Here, we show that D11 effectively inhibits signaling through FGFR3 in vitro, inhibits the growth of FGFR3-dependent FGF9-induced lung adenocarcinoma in mice, and reduces tumor-associated morbidity. Given the potency of FGF9 in this mouse model and the absolute requirement for signaling through FGFR3, this study validates the D11 antibody as a potentially useful and effective reagent for treating human cancers or other pathologies that are dependent on activation of FGFR3. PMID:27056048

  18. Temporal and Spatial Expression of Transforming Growth Factor-β after Airway Remodeling to Tobacco Smoke in Rats.

    PubMed

    Hoang, Laura L; Nguyen, Yen P; Aspeé, Rayza; Bolton, Sarah J; Shen, Yi-Hsin; Wang, Lei; Kenyon, Nicholas J; Smiley-Jewell, Suzette; Pinkerton, Kent E

    2016-06-01

    Airway remodeling is strongly correlated with the progression of chronic obstructive pulmonary disease (COPD). In this study, our goal was to characterize progressive structural changes in site-specific airways, along with the temporal and spatial expression of transforming growth factor (TGF)-β in the lungs of male spontaneously hypertensive rats exposed to tobacco smoke (TS). Our studies demonstrated that TS-induced changes of the airways is dependent on airway generation and exposure duration for proximal, midlevel, and distal airways. Stratified squamous epithelial cell metaplasia was evident in the most proximal airways after 4 and 12 weeks but with minimal levels of TGF-β-positive epithelial cells after only 4 weeks of exposure. In contrast, epithelial cells in midlevel and distal airways were strongly TGF-β positive at both 4 and 12 weeks of TS exposure. Airway smooth muscle volume increased significantly at 4 and 12 weeks in midlevel airways. Immunohistochemistry of TGF-β was also found to be significantly increased at 4 and 12 weeks in lymphoid tissues and alveolar macrophages. ELISA of whole-lung homogenate demonstrated that TGF-β2 was increased after 4 and 12 weeks of TS exposure, whereas TGF-β1 was decreased at 12 weeks of TS exposure. Airway levels of messenger RNA for TGF-β2, as well as platelet-derived growth factor-A, granulocyte-macrophage colony-stimulating factor, and vascular endothelial growth factor-α, growth factors regulated by TGF-β, were significantly decreased in animals after 12 weeks of TS exposure. Our data indicate that TS increases TGF-β in epithelial and inflammatory cells in connection with airway remodeling, although the specific role of each TGF-β isoform remains to be defined in TS-induced airway injury and disease. PMID:26637070

  19. A novel function of colony-stimulating factor 1 receptor in hTERT immortalization of human epithelial cells.

    PubMed

    Li, N F; Kocher, H M; Salako, M A; Obermueller, E; Sandle, J; Balkwill, F

    2009-02-01

    The receptor for macrophage colony-stimulating factor 1 receptor (CSF1R) is a product of the proto-oncogene c-fms and a member of the class III transmembrane tyrosine kinase receptor family. Earlier, we described increased mRNA expression of CSF1R in human telomerase reverse transcriptase (hTERT) immortalized human ovarian surface epithelial (IOSE) cell lines derived from a single donor. Here, we further describe that CSF1R is upregulated at both the mRNA and protein level in hTERT immortalized human normal OSE cells from two different donors and in hTERT immortalized human pancreatic ductal epithelial cells. CSF1R was not upregulated in hTERT immortalized epithelial clones that subsequently underwent senescence or in immortalized fibroblasts. Upon stimulation by the CSF1R ligand CSF1, the immortalized epithelial cell lines showed rapid internalization of CSF1R with concomitant down-modulation and colocalization of phosphorylated NFkappaBp65 with hTERT protein, hTERT translocation into the nucleus and the binding of c-Myc to the hTERT promoter region. Reducing the expression of CSF1R using short hairpin interfering RNA abolished these effects and also decreased cell survival and the number of population doublings under suboptimal culture conditions. The telomerase inhibitor GRN163L confirmed a role for telomerase in the cleavage of the intracellular domain of CSF1R. On the basis of these findings, we suggest that CSF1R may be a critical factor facilitating hTERT immortalization of epithelial cells. PMID:18997822

  20. Canonical pathways, networks and transcriptional factor regulation by clinical strains of Mycobacterium tuberculosis in pulmonary alveolar epithelial cells.

    PubMed

    Mvubu, Nontobeko E; Pillay, Balakrishna; Gamieldien, Junaid; Bishai, William; Pillay, Manormoney

    2016-03-01

    Limited knowledge exists on pathways, networks and transcriptional factors regulated within epithelial cells by diverse Mycobacterium tuberculosis genotypes. This study aimed to elucidate these mechanisms induced in A549 epithelial cells by dominant clinical strains in KwaZulu-Natal, South Africa. RNA for sequencing was extracted from epithelial cells at 48 h post-infection with 5 strains at a multiplicity of infection of approximately 10:1. Bioinformatics analysis performed with the RNA-Seq Tuxedo pipeline identified differentially expressed genes. Changes in pathways, networks and transcriptional factors were identified using Ingenuity Pathway Analysis (IPA). The interferon signalling and hepatic fibrosis/hepatic stellate cell activation pathways were among the top 5 canonical pathways in all strains. Hierarchical clustering for enrichment of cholesterol biosynthesis and immune associated pathways revealed similar patterns for Beijing and Unique; F15/LAM4/KZN and F11; and, F28 and H37Rv strains, respectively. However, the induction of top scoring networks varied among the strains. Among the transcriptional factors, only EHL, IRF7, PML, STAT1, STAT2 and VDR were induced by all clinical strains. Activation of the different pathways, networks and transcriptional factors revealed in the current study may be an underlying mechanism that results in the differential host response by clinical strains of M. tuberculosis. PMID:26980499

  1. Expression and activation of erbB-2 and epidermal growth factor receptor in lung adenocarcinomas.

    PubMed Central

    Rachwal, W. J.; Bongiorno, P. F.; Orringer, M. B.; Whyte, R. I.; Ethier, S. P.; Beer, D. G.

    1995-01-01

    ErbB-2 and EGFR (epidermal growth factor receptor) are expressed in lung adenocarcinomas and associated with a poor prognosis. Immunocytochemical analysis revealed erbB-2 and EGFR coexperession as a characteristic feature of most lung adenocarcinomas, and at levels of receptor expression present in bronchial epithelial cells. In primary lung tumours and cell lines, erbB-2 detected using Western blot analysis demonstrated low-level phosphotyrosine staining of the 185 kDa band, as compared with breast cancer cell lines. A549 and A427 lung adenocarcinoma cells treated with neu differentiation factor (NDF) showed increased erbB-2 phosphotyrosine staining, but to a much lesser extent than breast cancer cells. The lung cells were examined for expression of the potential autocrine growth factors NDF and transforming growth factor alpha (TGF-alpha) by Northern blot analysis. Both NDF and TFG-alpha mRNA were abundantly expressed in the A549 cells. NDF mRNA was highest during active cell proliferation and decreased in confluent cells or after treatment with the growth-inhibitory steroid dexamethasone. Primary tumours and cell lines expressed EGFR, showing higher basal level phosphotyrosine staining than erbB-2. Treatment with NDF and EGF (epidermal growth factor) stimulated cell growth, and in A549 cells the presence of both factors provided an additive increase in cell growth. The growth stimulus that ligand-activated erbB-2 and EGFR provides to lung adenocarcinoma cells may establish a background of continued cell proliferation over which other critical transforming events may occur. Images Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 PMID:7599067

  2. Endorsement of Growth Factors in Experiential Training Groups

    ERIC Educational Resources Information Center

    Kiweewa, John; Gilbride, Dennis; Luke, Melissa; Seward, Derek

    2013-01-01

    The purpose of this study was to identify student growth factors during a semester long Master's level group counseling class. Results indicated that 12 growth factors accounted for 86% of the total number of critical incidents that participants reported as influencing their personal growth and awareness during the group experience. Two other…

  3. Gene Expression of Growth Factors and Growth Factor Receptors for Potential Targeted Therapy of Canine Hepatocellular Carcinoma

    PubMed Central

    IIDA, Gentoku; ASANO, Kazushi; SEKI, Mamiko; SAKAI, Manabu; KUTARA, Kenji; ISHIGAKI, Kumiko; KAGAWA, Yumiko; YOSHIDA, Orie; TESHIMA, Kenji; EDAMURA, Kazuya; WATARI, Toshihiro

    2013-01-01

    ABSTRACT The purpose of this study was to evaluate the gene expression of growth factors and growth factor receptors of primary hepatic masses, including hepatocellular carcinoma (HCC) and nodular hyperplasia (NH), in dogs. Quantitative real-time reverse transcriptase-polymerase chain reaction was performed to measure the expression of 18 genes in 18 HCCs, 10 NHs, 11 surrounding non-cancerous liver tissues and 4 healthy control liver tissues. Platelet-derived growth factor-B (PDGF-B), transforming growth factor-α, epidermal growth factor receptor, epidermal growth factor and hepatocyte growth factor were found to be differentially expressed in HCC compared with NH and the surrounding non-cancerous and healthy control liver tissues. PDGF-B is suggested to have the potential to become a valuable ancillary target for the treatment of canine HCC. PMID:24189579

  4. Molecular cloning and expression of an additional epidermal growth factor receptor-related gene.

    PubMed Central

    Plowman, G D; Whitney, G S; Neubauer, M G; Green, J M; McDonald, V L; Todaro, G J; Shoyab, M

    1990-01-01

    Epidermal growth factor (EGF), transforming growth factor alpha (TGF-alpha), and amphiregulin are structurally and functionally related growth regulatory proteins. These secreted polypeptides all bind to the 170-kDa cell-surface EGF receptor, activating its intrinsic kinase activity. However, amphiregulin exhibits different activities than EGF and TGF-alpha in a number of biological assays. Amphiregulin only partially competes with EGF for binding EGF receptor, and amphiregulin does not induce anchorage-independent growth of normal rat kidney cells (NRK) in the presence of TGF-beta. Amphiregulin also appears to abrogate the stimulatory effect of TGF-alpha on the growth of several aggressive epithelial carcinomas that overexpress EGF receptor. These findings suggest that amphiregulin may interact with a separate receptor in certain cell types. Here we report the cloning of another member of the human EGF receptor (HER) family of receptor tyrosine kinases, which we have named "HER3/ERRB3." The cDNA was isolated from a human carcinoma cell line, and its 6-kilobase transcript was identified in various human tissues. We have generated peptide-specific antisera that recognizes the 160-kDa HER3 protein when transiently expressed in COS cells. These reagents will allow us to determine whether HER3 binds amphiregulin or other growth regulatory proteins and what role HER3 protein plays in the regulation of cell growth. Images PMID:2164210

  5. Growth factor involvement in tension-induced skeletal muscle growth

    NASA Technical Reports Server (NTRS)

    Vandenburgh, Herman W.

    1987-01-01

    New muscle tissue culture techniques were developed to grow embryonic skeletal myofibers which are able to differentiate into more adultlike myofibers. Studies on mechanical simulation of cultured muscle cell growth will now be more directly applicable to mechanically-induced growth in adult muscle, and lead to better models for understanding muscle tissue atrophy caused by disuse in the microgravity of space.

  6. Chemical synthesis of a gene for human epidermal growth factor urogastrone and its expression in yeast.

    PubMed Central

    Urdea, M S; Merryweather, J P; Mullenbach, G T; Coit, D; Heberlein, U; Valenzuela, P; Barr, P J

    1983-01-01

    We have chemically synthesized and expressed in yeast a gene coding for human epidermal growth factor (urogastrone), a 53-amino-acid polypeptide that has been shown to promote epithelial cell proliferation and to inhibit gastric acid secretion. The synthetic gene, consisting of 170 base pairs, was designed with yeast-preferred codons and assembled by enzymatic ligation of synthetic fragments produced by phosphoramidite chemistry. The DNA synthesis protocol used allows for facile synthesis of oligonucleotides larger than 50 bases. Yeast cells were transformed with plasmids containing the synthetic gene under control of a yeast glyceraldehyde-3-phosphate dehydrogenase gene promoter and were shown to synthesize a biologically active human epidermal growth factor. Images PMID:6369317

  7. Cowden disease: gene marker studies and measurements of epidermal growth factor.

    PubMed Central

    Carlson, H E; Burns, T W; Davenport, S L; Luger, A M; Spence, M A; Sparkes, R S; Orth, D N

    1986-01-01

    Cowden disease (CD) is a familial syndrome characterized by tumors of the skin, oral mucosa, breast, thyroid, and intestinal epithelium. Since the syndrome is inherited as an autosomal dominant, we examined a battery of gene markers in a family with CD to detect linkage between the CD gene and known marker genes. There was no positive evidence for linkage of a CD locus with any of the markers; other investigators can add to our data to confirm and extend these findings. Additionally, we measured epidermal growth factor (EGF) in body fluids from CD patients and controls to determine if elevated EGF levels might be responsible for the widespread epithelial proliferation in CD. EGF levels in saliva, serum, plasma, and urine were similar in CD patients and control subjects. Although alterations in growth factors or their receptors may play a role in CD, excess circulating EGF is not responsible for the manifestations of the syndrome. Images Fig. 2 PMID:3487976

  8. Can we protect the gut in critical illness? The role of growth factors and other novel approaches.

    PubMed

    Dominguez, Jessica A; Coopersmith, Craig M

    2010-07-01

    The intestine plays a central role in the pathophysiology of critical illness and is frequently called the "motor" of the systemic inflammatory response. Perturbations to the intestinal barrier can lead to distant organ damage and multiple organ failure. Therefore, identifying ways to preserve intestinal integrity may be of paramount importance. Growth factors and other peptides have emerged as potential tools for modulation of intestinal inflammation and repair due to their roles in cellular proliferation, differentiation, migration, and survival. This review examines the involvement of growth factors and other peptides in intestinal epithelial repair during critical illness and their potential use as therapeutic targets. PMID:20643306

  9. Can We Protect the Gut in Critical Illness: The Role of Growth Factors and Other Novel Approaches

    PubMed Central

    Dominguez, Jessica A.; Coopersmith, Craig M.

    2010-01-01

    Synopsis The intestine plays a central role in the pathophysiology of critical illness and is frequently called the “motor” of the systemic inflammatory response. Perturbations to the intestinal barrier can lead to distant organ damage and multiple organ failure. Therefore, identifying ways to preserve intestinal integrity may be of paramount importance. Growth factors and other peptides have emerged as potential tools for modulation of intestinal inflammation and repair due to their roles in cellular proliferation, differentiation, migration, and survival. In this review, we will examine the involvement of growth factors and other peptides in intestinal epithelial repair during critical illness and their potential use as therapeutic targets. PMID:20643306

  10. Isolated placental vessel response to vascular endothelial growth factor and placenta growth factor in normal and growth-restricted pregnancy.

    PubMed

    Szukiewicz, Dariusz; Szewczyk, Grzegorz; Watroba, Mateusz; Kurowska, Ewa; Maslinski, Slawomir

    2005-01-01

    Vascular endothelial growth factor (VEGF) and placenta growth factor (PlGF) cause vasodilation. We examined the vasomotor response of isolated placental vessels to VEGF and PlGF in normal (group I) and intrauterine growth retardation (IUGR)-complicated pregnancy (group II). Rings of vessels were prepared in vitro and mounted on the vessel myograph plunged in tissue bath. The magnitude of dilation to increased doses of VEGF and PlGF has been studied. VEGF is a more potent vasodilator than PlGF. Both, VEGF- and PlGF-induced vasorelaxation was diminished in the IUGR (group II) nearly by half, compared to control (group I). Relative placental nitric oxide deficiency, or decreased sensitivity to VEGF and PlGF may contribute to the development of high impedance fetoplacental circulation. PMID:15591804

  11. The Role of Vascular Endothelial Growth Factors and Fibroblast Growth Factors in Angiogenesis during Otitis Media

    PubMed Central

    Husseman, Jacob; Palacios, Sean D.; Rivkin, Alexander Z.; Oehl, Heinz; Ryan, Allen F.

    2012-01-01

    The middle ear response to otitis media includes transformation and hyperplasia of the mucosal epithelium and subepithelial connective tissue. Significant neovascularization is also noted, which occurs both to support the hypertrophied mucosa and to mediate the increased trafficking of leukocytes. We investigated the role of two known potent angiogenic growth factor families, the fibroblast growth factors (FGFs) and vascular endothelial growth factors (VEGFs), in middle ear mucosal angiogenesis. DNA microarrays were used to evaluate the expression of FGFs and VEGFs, as well as their receptors and unique signaling proteins, in the middle ears of mice undergoing a complete course of acute bacterial otitis media. In addition, a member of each family was introduced to the middle ear submucosal compartment of the normal middle ears of guinea pigs, by a continuous-release osmotic minipump system over 1 week. During the course of bacterial otitis media, a significant regulation of a number of genes important for angiogenesis was identified. Histologic evaluation of middle ear mucosa following micropump infusion of both FGF1 and VEGF-A showed significant angiogenesis at the site of infusion in comparison to control saline infusion. These results support a role for FGFs and VEGFs in the neovascularization of the middle ear mucosa during otitis media, and offer a potential avenue for therapeutic intervention. PMID:22104377

  12. Direct binding of hepatocyte growth factor and vascular endothelial growth factor to CD44v6

    PubMed Central

    Volz, Yvonne; Koschut, David; Matzke-Ogi, Alexandra; Dietz, Marina S.; Karathanasis, Christos; Richert, Ludovic; Wagner, Moritz G.; Mély, Yves; Heilemann, Mike; Niemann, Hartmut H.; Orian-Rousseau, Véronique

    2015-01-01

    CD44v6, a member of the CD44 family of transmembrane glycoproteins is a co-receptor for two receptor tyrosine kinases (RTKs), Met and VEGFR-2 (vascular endothelial growth factor receptor 2). CD44v6 is not only required for the activation of these RTKs but also for signalling. In order to understand the role of CD44v6 in Met and VEGFR-2 activation and signalling we tested whether CD44v6 binds to their ligands, HGF (hepatocyte growth factor) and VEGF (vascular endothelial growth factor), respectively. FACS analysis and cellular ELISA showed binding of HGF and VEGF only to cells expressing CD44v6. Direct binding of CD44v6 to HGF and VEGF was demonstrated in pull-down assays and the binding affinities were determined using MicroScale Thermophoresis, fluorescence correlation spectroscopy and fluorescence anisotropy. The binding affinity of CD44v6 to HGF is in the micromolar range in contrast with the high-affinity binding measured in the case of VEGF and CD44v6, which is in the nanomolar range. These data reveal a heparan sulfate-independent direct binding of CD44v6 to the ligands of Met and VEGFR-2 and suggest different roles of CD44v6 for these RTKs. PMID:26181364

  13. Vascular Endothelial Growth Factor in Eye Disease

    PubMed Central

    Penn, J.S.; Madan, A.; Caldwell, R.B.; Bartoli, M.; Caldwell, R.W.; Hartnett, M.E.

    2012-01-01

    Collectively, angiogenic ocular conditions represent the leading cause of irreversible vision loss in developed countries. In the U.S., for example, retinopathy of prematurity, diabetic retinopathy and age-related macular degeneration are the principal causes of blindness in the infant, working age and elderly populations, respectively. Evidence suggests that vascular endothelial growth factor (VEGF), a 40 kDa dimeric glycoprotein, promotes angiogenesis in each of these conditions, making it a highly significant therapeutic target. However, VEGF is pleiotropic, affecting a broad spectrum of endothelial, neuronal and glial behaviors, and confounding the validity of anti-VEGF strategies, particularly under chronic disease conditions. In fact, among other functions VEGF can influence cell proliferation, cell migration, proteolysis, cell survival and vessel permeability in a wide variety of biological contexts. This article will describe the roles played by VEGF in the pathogenesis of retinopathy of prematurity, diabetic retinopathy and age-related macular degeneration. The potential disadvantages of inhibiting VEGF will be discussed, as will the rationales for targeting other VEGF-related modulators of angiogenesis. PMID:18653375

  14. [Epidermal growth factor, innovation and safety].

    PubMed

    Esquirol Caussa, Jordi; Herrero Vila, Elisabeth

    2015-10-01

    Bioidentical recombinant human epidermal growth factor (rhEGF) is available in concentrations and purity suitable for therapeutic use in long time stable formulations. Beneficial effects in several skin pathologies and lesions have been reported (traumatic and surgical wound healing, laser induced wounds, abnormal scars, keloids, radiation or chemotherapy induced dermatitis, post inflammatory hyperpigmentation or for skin aging damage repairing) and also may be considered for the treatment of several oropharingeal and high gastroesophageal tract mucosa diseases (mouth sores, pharyngeal fistulas, ulcers), and several corneal or conjunctive mucosa lesions. rhEGF has not shown any important side or collateral effects in humans or in laboratory experimentation animals, showing optimal tolerability and safety with continuous use for months. Compounding gives advantages of versatility, individualization, personalization, molecular stability, safety and effectiveness in ideal conditions, showing good tissue penetration, both on intact skin and skin lesions that expose the lower planes to the surface. rhEGF compounds can be considered for prevention or as a treatment of diverse skin and mucosa diseases and conditions through compounding preparations. PMID:25433777

  15. Overexpression of angiotensin II type 1 receptor in breast cancer cells induces epithelial-mesenchymal transition and promotes tumor growth and angiogenesis.

    PubMed

    Oh, Eunhye; Kim, Ji Young; Cho, Youngkwan; An, Hyunsook; Lee, Nahyun; Jo, Hunho; Ban, Changill; Seo, Jae Hong

    2016-06-01

    The angiotensin II type I receptor (AGTR1) has been implicated in diverse aspects of human disease, from the regulation of blood pressure and cardiovascular homeostasis to cancer progression. We sought to investigate the role of AGTR1 in cell proliferation, epithelial-mesenchymal transition (EMT), migration, invasion, angiogenesis and tumor growth in the breast cancer cell line MCF7. Stable overexpression of AGTR1 was associated with accelerated cell proliferation, concomitant with increased expression of survival factors including poly(ADP-ribose) polymerase (PARP) and X-linked inhibitor of apoptosis (XIAP), as well as extracellular signal-regulated kinase (ERK) activation. AGTR1-overexpressing MCF7 cells were more aggressive than their parent line, with significantly increased activity in migration and invasion assays. These observations were associated with changes in EMT markers, including reduced E-cadherin expression and increased p-Smad3, Smad4 and Snail levels. Treatment with the AGTR1 antagonist losartan attenuated these effects. AGTR1 overexpression also accelerated tumor growth and increased Ki-67 expression in a xenograft model. This was associated with increased tumor angiogenesis, as evidenced by a significant increase in microvessels in the intratumoral and peritumoral areas, and enhanced tumor invasion, with the latter response associated with increased EMT marker expression and matrix metallopeptidase 9 (MMP-9) upregulation. In vivo administration of losartan significantly reduced both tumor growth and angiogenesis. Our findings suggest that AGTR1 plays a significant role in tumor aggressiveness, and its inhibition may have therapeutic implications. PMID:26975580

  16. Effect of Aflatoxin B1 on Growth of Bovine Mammary Epithelial Cells in 3D and Monolayer Culture System

    PubMed Central

    Forouharmehr, Ali; Harkinezhad, Taher; Qasemi-Panahi, Babak

    2013-01-01

    Purpose: Many studies have been showed transfer of aflatoxins, toxins produced by Aspergillus flvaus and Aspergillus parasiticus fungi, into milk. These toxins are transferred into the milk through digestive system by eating contaminated food. Due to the toxicity of these materials, it seems that it has side effects on the growth of mammary cells. Therefore, the present work aimed to investigate possible toxic effects of aflatoxin B1 (AFB1) on bovine mammary epithelial cells in monolayer and three-dimensional cultures. Methods: Specimens of the mammary tissue of bovine were sized out in size 2×2 cm in slaughterhouse. After disinfection and washing in sterile PBS, primary cell culture was performed by enzymatic digestion of tissue with collagenase. When proper numbers of cells were achieved in monolayer culture, cells were seeded in a 24-well culture plate for three-dimensional (3D) culture in Matrigel matrix. After 21 days of 3D culture and reaching the required number of cells, the concentrations of 15, 25 and 35 µL of AFB1 were added to the culture in quadruplicate and incubated for 8 hours. Cellular cytotoxicity was examined using standard colorimetric assay and finally, any change in the morphology of the cells was studied by microscopic technique. Results: Microscopic investigations showed necrosis of the AFB1-exposed cells compared to the control cells. Also, bovine mammary epithelial cells were significantly affected by AFB1 in dose and time dependent manner in cell viability assays. Conclusion: According to the results, it seems that AFB1 can induce cytotoxicity and necrosis in bovine mammary epithelial cells. PMID:24312827

  17. Skeletal Muscle Depletion and Markers for Cancer Cachexia Are Strong Prognostic Factors in Epithelial Ovarian Cancer

    PubMed Central

    Aust, Stefanie; Knogler, Thomas; Pils, Dietmar; Obermayr, Eva; Reinthaller, Alexander; Zahn, Lisa; Radlgruber, Ilja; Mayerhoefer, Marius Erik; Grimm, Christoph; Polterauer, Stephan

    2015-01-01

    Objective Tumor cachexia is an important prognostic parameter in epithelial ovarian cancer (EOC). Tumor cachexia is characterized by metabolic and inflammatory disturbances. These conditions might be reflected by body composition measurements (BCMs) ascertained by pre-operative computed tomography (CT). Thus, we aimed to identify the prognostically most relevant BCMs assessed by pre-operative CT in EOC patients. Methods We evaluated muscle BCMs and well established markers of nutritional and inflammatory status, as well as clinical-pathological parameters in 140 consecutive patients with EOC. Furthermore, a multiplexed inflammatory marker panel of 25 cytokines was used to determine the relationship of BCMs with inflammatory markers and patient’s outcome. All relevant parameters were evaluated in uni- and multivariate survival analysis. Results Muscle attenuation (MA)—a well established BCM parameter—is an independent prognostic factor for survival in multivariate analysis (HR 2.25; p = 0.028). Low MA—reflecting a state of cachexia—is also associated with residual tumor after cytoreductive surgery (p = 0.046) and with an unfavorable performance status (p = 0.015). Moreover, MA is associated with Eotaxin and IL-10 out of the 25 cytokine multiplex marker panel in multivariate linear regression analysis (p = 0.021 and p = 0.047, respectively). Conclusion MA—ascertained by routine pre-operative CT—is an independent prognostic parameter in EOC patients. Low MA is associated with the inflammatory, as well as the nutritional component of cachexia. Therefore, the clinical value of pre-operative CT could be enhanced by the assessment of MA. PMID:26457674

  18. Fibroblast Growth Factors and Vascular Endothelial Growth Factor Promote Cardiac Reprogramming under Defined Conditions

    PubMed Central

    Yamakawa, Hiroyuki; Muraoka, Naoto; Miyamoto, Kazutaka; Sadahiro, Taketaro; Isomi, Mari; Haginiwa, Sho; Kojima, Hidenori; Umei, Tomohiko; Akiyama, Mizuha; Kuishi, Yuki; Kurokawa, Junko; Furukawa, Tetsushi; Fukuda, Keiichi; Ieda, Masaki

    2015-01-01

    Summary Fibroblasts can be directly reprogrammed into cardiomyocyte-like cells (iCMs) by overexpression of cardiac transcription factors, including Gata4, Mef2c, and Tbx5; however, this process is inefficient under serum-based culture conditions, in which conversion of partially reprogrammed cells into fully reprogrammed functional iCMs has been a major hurdle. Here, we report that a combination of fibroblast growth factor (FGF) 2, FGF10, and vascular endothelial growth factor (VEGF), termed FFV, promoted cardiac reprogramming under defined serum-free conditions, increasing spontaneously beating iCMs by 100-fold compared with those under conventional serum-based conditions. Mechanistically, FFV activated multiple cardiac transcriptional regulators and converted partially reprogrammed cells into functional iCMs through the p38 mitogen-activated protein kinase and phosphoinositol 3-kinase/AKT pathways. Moreover, FFV enabled cardiac reprogramming with only Mef2c and Tbx5 through the induction of cardiac reprogramming factors, including Gata4. Thus, defined culture conditions promoted the quality of cardiac reprogramming, and this finding provides new insight into the mechanism of cardiac reprogramming. PMID:26626177

  19. NME genes in epithelial morphogenesis

    PubMed Central

    2012-01-01

    The NME family of genes encodes highly conserved multifunctional proteins that have been shown to participate in nucleic acid metabolism, energy homeostasis, cell signaling, and cancer progression. Some family members, particularly isoforms 1 and 2, have attracted extensive interests because of their potential anti-metastasis activity. Unfortunately, there have been few consensus mechanistic explanations for this critical function because of the numerous molecular functions ascribed to these proteins, including nucleoside diphosphate kinase, protein kinase, nuclease, transcription factor, growth factor, among others. In addition, different studies showed contradictory prognostic correlations between NME expression levels and tumor progression in clinical samples. Thus, analyses using pliable in vivo systems have become critical for unraveling at least some aspects of the complex functions of this family of genes. Recent works using the Drosophila genetic system have suggested a role for NME in regulating epithelial cell motility and morphogenesis, which has also been demonstrated in mammalian epithelial cell culture. This function is mediated by promoting internalization of growth factor receptors in motile epithelial cells, and the adherens junction components such as E-cadherin and β-catenin in epithelia that form the tissue linings. Interestingly, NME genes in epithelial cells appear to function in a defined range of expression levels. Either down-regulation or over-expression can perturb epithelial integrity, resulting in different aspects of epithelial abnormality. Such biphasic functions provide a plausible explanation for the documented anti-metastatic activity and the suspected oncogenic function. This review summarizes these recent findings and discusses their implications. PMID:21336542

  20. Androgen Receptor-Mediated Growth Suppression of HPr-1AR and PC3-Lenti-AR Prostate Epithelial Cells

    PubMed Central

    Bolton, Eric C.

    2015-01-01

    The androgen receptor (AR) mediates the developmental, physiologic, and pathologic effects of androgens including 5α-dihydrotestosterone (DHT). However, the mechanisms whereby AR regulates growth suppression and differentiation of luminal epithelial cells in the prostate gland and proliferation of malignant versions of these cells are not well understood, though they are central to prostate development, homeostasis, and neoplasia. Here, we identify androgen-responsive genes that restrain cell cycle progression and proliferation of human prostate epithelial cell lines (HPr-1AR and PC3-Lenti-AR), and we investigate the mechanisms through which AR regulates their expression. DHT inhibited proliferation of HPr-1AR and PC3-Lenti-AR, and cell cycle analysis revealed a prolonged G1 interval. In the cell cycle, the G1/S-phase transition is initiated by the activity of cyclin D and cyclin-dependent kinase (CDK) complexes, which relieve growth suppression. In HPr-1AR, cyclin D1/2 and CDK4/6 mRNAs were androgen-repressed, whereas CDK inhibitor, CDKN1A, mRNA was androgen-induced. The regulation of these transcripts was AR-dependent, and involved multiple mechanisms. Similar AR-mediated down-regulation of CDK4/6 mRNAs and up-regulation of CDKN1A mRNA occurred in PC3-Lenti-AR. Further, CDK4/6 overexpression suppressed DHT-inhibited cell cycle progression and proliferation of HPr-1AR and PC3-Lenti-AR, whereas CDKN1A overexpression induced cell cycle arrest. We therefore propose that AR-mediated growth suppression of HPr-1AR involves cyclin D1 mRNA decay, transcriptional repression of cyclin D2 and CDK4/6, and transcriptional activation of CDKN1A, which serve to decrease CDK4/6 activity. AR-mediated inhibition of PC3-Lenti-AR proliferation occurs through a similar mechanism, albeit without down-regulation of cyclin D. Our findings provide insight into AR-mediated regulation of prostate epithelial cell proliferation. PMID:26372468

  1. IL-6 promotes growth and epithelial-mesenchymal transition of CD133+ cells of non-small cell lung cancer

    PubMed Central

    Lee, Soo Ok; Yang, Xiaodong; Duan, Shanzhou; Tsai, Ying; Strojny, Laura R.; Keng, Peter; Chen, Yuhchyau

    2016-01-01

    We examined IL-6 effects on growth, epithelial-mesenchymal transition (EMT) process, and metastatic ability of CD133+ and CD133– cell subpopulations isolated from three non-small cell lung cancer (NSCLC) cell lines: A549, H157, and H1299. We developed IL-6 knocked-down and scramble (sc) control cells of A549 and H157 cell lines by lentiviral infection system, isolated CD133+ and CD133– sub-populations, and investigated the IL-6 role in self-renewal/growth of these cells. IL-6 showed either an inhibitory or lack of effect in modulating growth of CD133– cells depending on intracellular IL-6 levels, but there was higher self-renewal ability of IL-6 expressing CD133+ cells than IL-6 knocked down cells, confirming the promoter role of IL-6 in CD133+ cells growth. We then examined tumor growth of xenografts developed from CD133+ cells of A549IL-6si vs. A549sc cell lines. Consistently, there was retarded growth of tumors developed from A549IL-6si, CD133+ cells compared to tumors originating from A549sc, CD133+ cells. The effect of IL-6 in promoting CD133+ self-renewal was due to hedgehog (Hhg) and Erk signaling pathway activation and higher Bcl-2/Bcl-xL expression. We also investigated whether IL-6 regulates the EMT process of CD133− and CD133+ cells differently. Expression of the EMT/metastasis-associated molecules in IL-6 expressing cells was higher than in IL-6 knocked down cells. Together, we demonstrated dual roles of IL-6 in regulating growth of CD133– and CD133+ subpopulations of lung cancer cells and significant regulation of IL-6 on EMT/metastasis increase in CD133+ cells, not in CD133– cells. PMID:26675547

  2. Transforming growth factor-beta. Murine glomerular receptors and responses of isolated glomerular cells.

    PubMed Central

    MacKay, K; Striker, L J; Stauffer, J W; Doi, T; Agodoa, L Y; Striker, G E

    1989-01-01

    Proliferation of resident glomerular cells and the accumulation of mesangial matrix are histologic abnormalities which are observed in the course of many progressive glomerular diseases. We explored the potential regulatory effects of transforming growth factor-beta (TGF-beta) on these processes. We found that cultured mouse glomerular endothelial, mesangial, and epithelial cells as well as isolated intact rat glomeruli possess high-affinity receptors for TGF-beta. We also found that, although TGF-beta consistently inhibited the proliferation of glomerular endothelial and epithelial cells, it acted as a bifunctional regulator of mesangial cell proliferation. TGF-beta significantly increased the production of collagen and fibronectin by glomerular mesangial cells whereas only fibronectin production was augmented in glomerular epithelial cells. The presence of TGF-beta receptors on intact glomeruli and on each glomerular cell type and the demonstrated responsiveness of these cells to TGF-beta combine to suggest that potentially important interactions may occur between resident glomerular cells and TGF-beta in vivo. Images PMID:2539392

  3. Interleukin-17 is a potent immuno-modulator and regulator of normal human intestinal epithelial cell growth

    SciTech Connect

    Schwartz, S.; Beaulieu, J.F. . E-mail: frank.ruemmele@nck.ap-hop-paris.fr

    2005-11-18

    Upregulation of the T-cell derived cytokine interleukin (IL-17) was reported in the inflamed intestinal mucosa of patients with inflammatory bowel disorders. In this study, we analyzed the effect of IL-17 on human intestinal epithelial cell (HIEC) turnover and functions. Proliferation and apoptosis in response to IL-17 was monitored in HIEC (cell counts, [{sup 3}H]thymidine incorporation method, and annexinV-PI-apoptosis assay). Signalling pathways were analyzed by Western blots, electromobility shift assay, and immunofluorescence studies. IL-17 proved to be a potent inhibitor of HIEC proliferation without any pro-apoptotic/necrotic effect. The growth inhibitory effect of IL-17 was mediated via the p38 stress kinase. Consequently, the p38-SAPkinase-inhibitor SB203580 abrogated this anti-mitotic effect. In parallel, IL-17 provoked the degradation of I{kappa}B{alpha}, allowing nuclear translocation of the p65 NF-{kappa}B subunit and induction of the NF-{kappa}B-controlled genes IL-6 and -8. IL-17 potently blocks epithelial cell turnover while at the same time amplifying an inflammatory response in a positive feedback manner.

  4. Vascular Growth Factors and Glomerular Disease.

    PubMed

    Bartlett, Christina S; Jeansson, Marie; Quaggin, Susan E

    2016-01-01

    The glomerulus is a highly specialized microvascular bed that filters blood to form primary urinary filtrate. It contains four cell types: fenestrated endothelial cells, specialized vascular support cells termed podocytes, perivascular mesangial cells, and parietal epithelial cells. Glomerular cell-cell communication is critical for the development and maintenance of the glomerular filtration barrier. VEGF, ANGPT, EGF, SEMA3A, TGF-β, and CXCL12 signal in paracrine fashions between the podocytes, endothelium, and mesangium associated with the glomerular capillary bed to maintain filtration barrier function. In this review, we summarize the current understanding of these signaling pathways in the development and maintenance of the glomerulus and the progression of disease. PMID:26863327

  5. Fibroblast Growth Factor Signaling in Metabolic Regulation

    PubMed Central

    Nies, Vera J. M.; Sancar, Gencer; Liu, Weilin; van Zutphen, Tim; Struik, Dicky; Yu, Ruth T.; Atkins, Annette R.; Evans, Ronald M.; Jonker, Johan W.; Downes, Michael Robert

    2016-01-01

    The prevalence of obesity is a growing health problem. Obesity is strongly associated with several comorbidities, such as non-alcoholic fatty liver disease, certain cancers, insulin resistance, and type 2 diabetes, which all reduce life expectancy and life quality. Several drugs have been put forward in order to treat these diseases, but many of them have detrimental side effects. The unexpected role of the family of fibroblast growth factors in the regulation of energy metabolism provides new approaches to the treatment of metabolic diseases and offers a valuable tool to gain more insight into metabolic regulation. The known beneficial effects of FGF19 and FGF21 on metabolism, together with recently discovered similar effects of FGF1 suggest that FGFs and their derivatives carry great potential as novel therapeutics to treat metabolic conditions. To facilitate the development of new therapies with improved targeting and minimal side effects, a better understanding of the molecular mechanism of action of FGFs is needed. In this review, we will discuss what is currently known about the physiological roles of FGF signaling in tissues important for metabolic homeostasis. In addition, we will discuss current concepts regarding their pharmacological properties and effector tissues in the context of metabolic disease. Also, the recent progress in the development of FGF variants will be reviewed. Our goal is to provide a comprehensive overview of the current concepts and consensuses regarding FGF signaling in metabolic health and disease and to provide starting points for the development of FGF-based therapies against metabolic conditions. PMID:26834701

  6. Treatment of Silk Fibroin with Poly(ethylene glycol) for the Enhancement of Corneal Epithelial Cell Growth

    PubMed Central

    Suzuki, Shuko; Dawson, Rebecca A.; Chirila, Traian V.; Shadforth, Audra M. A.; Hogerheyde, Thomas A.; Edwards, Grant A.; Harkin, Damien G.

    2015-01-01

    in the performance of these materials as a substratum for corneal epithelial cell growth. The reduced mechanical stability of freestanding PEG-treated membranes makes them a less viable choice than the non-treated membranes. PMID:26034883

  7. Novel Drosophila receptor that binds multiple growth factors

    SciTech Connect

    Rosner, M.R.; Thompson, K.L.; Garcia, V.; Decker, S.J.

    1986-05-01

    The authors have recently reported the identification of a novel growth factor receptor from Drosophila cell cultures that has dual binding specificity for both insulin and epidermal growth factor (EGF). This 100 kDa protein is also antigenically related to the cytoplasmic region of the mammalian EGF receptor-tyrosine kinase. They now report that this protein binds to mammalian nerve growth factor and human transforming growth factor alpha as well as insulin and EGF with apparent dissociation constants ranging from 10/sup -6/ to 10/sup -8/ M. The 100 kDa protein can be affinity-labeled with these /sup 125/I-labeled growth factors after immunoprecipitation with anti-EGF receptor antiserum. These four growth factors appear to share a common binding site, as evidenced by their ability to block affinity labelling by /sup 125/I-insulin. No significant binding to the 100 kDa protein was observed with platelet-derived growth factor, transforming growth factor beta, or glucagon. The 100 kDa Drosophila protein has a unique ligand-binding spectrum with no direct counterpart in mammalian cells and may represent an evolutionary precursor of the mammalian receptors for these growth factors.

  8. Growth factor array fabrication using a color ink jet printer.

    PubMed

    Watanabe, Kohei; Miyazaki, Takeshi; Matsuda, Ryoichi

    2003-04-01

    We have developed a novel method for growth factor analysis using a commercial color ink jet printer to fabricate substrata patterned with growth factors. We prepared substrata with insulin printed in a simple pattern or containing multiple areas of varying quantities of printed insulin. When we cultured the mouse myoblast cell line, C2C12, on the insulin-patterned substrata, the cells were grown in the same pattern with the insulin-printed pattern. Cell culture with the latter substrata demonstrated that quantity control of insulin deposition by a color ink jet printer is possible. For further applications, we developed substrata with insulin-like growth factor-I (IGF-I) and basic fibroblast growth factor (bFGF) spotted in 16 different areas in varying combinations and concentrations (growth factor array). With this growth factor array, C2C12 cells were cultured, and the onset of muscle cell differentiation was monitored for the expression of the myogenic regulator myogenin. The ratio of cells expressing myogenin varied with the doses of IGF-I and bFGF in the sections, demonstrating a feasibility of growth factor array fabrication by a color ink jet printer. Since a printer manipulates several colors, this method can be easily applied to multivariate analyses of growth factors and attachment factors affecting cell growth and differentiation. This method may provide a powerful tool for cell biology and tissue engineering, especially for stem cell research in investigating unknown conditions for differentiation. PMID:12719645

  9. Tumor-promoting functions of transforming growth factor-β in progression of cancer

    PubMed Central

    2012-01-01

    Transforming growth factor-β (TGF-β) elicits both tumor-suppressive and tumor-promoting functions during cancer progression. Here, we describe the tumor-promoting functions of TGF-β and how these functions play a role in cancer progression. Normal epithelial cells undergo epithelial-mesenchymal transition (EMT) through the action of TGF-β, while treatment with TGF-β and fibroblast growth factor (FGF)-2 results in transdifferentiation into activated fibroblastic cells that are highly migratory, thereby facilitating cancer invasion and metastasis. TGF-β also induces EMT in tumor cells, which can be regulated by oncogenic and anti-oncogenic signals. In addition to EMT promotion, invasion and metastasis of cancer are facilitated by TGF-β through other mechanisms, such as regulation of cell survival, angiogenesis, and vascular integrity, and interaction with the tumor microenvironment. TGF-β also plays a critical role in regulating the cancer-initiating properties of certain types of cells, including glioma-initiating cells. These findings thus may be useful for establishing treatment strategies for advanced cancer by inhibiting TGF-β signaling. PMID:22111550

  10. Infection of human fallopian tube epithelial cells with Neisseria gonorrhoeae protects cells from tumor necrosis factor alpha-induced apoptosis.

    PubMed

    Morales, Priscilla; Reyes, Paz; Vargas, Macarena; Rios, Miguel; Imarai, Mónica; Cardenas, Hugo; Croxatto, Horacio; Orihuela, Pedro; Vargas, Renato; Fuhrer, Juan; Heckels, John E; Christodoulides, Myron; Velasquez, Luis

    2006-06-01

    Following infection with Neisseria gonorrhoeae, bacteria may ascend into the Fallopian tubes (FT) and induce salpingitis, a major cause of infertility. In the FT, interactions between mucosal epithelial cells and gonococci are pivotal events in the pathogen's infection cycle and the inflammatory response. In the current study, primary FT epithelial cells were infected in vitro with different multiplicities of infection (MOI) of Pil+ Opa+ gonococci. Bacteria showed a dose-dependent association with cells and induced the secretion of tumor necrosis factor alpha (TNF-alpha). A significant finding was that gonococcal infection (MOI = 1) induced apoptosis in approximately 30% of cells, whereas increasing numbers of bacteria (MOI = 10 to 100) did not induce apoptosis. Apoptosis was observed in only 11% of cells with associated bacteria, whereas >84% of cells with no adherent bacteria were apoptotic. TNF-alpha was a key contributor to apoptosis, since (i) culture supernatants from cells infected with gonococci (MOI = 1) induced apoptosis in naïve cultures, suggesting that a soluble factor was responsible; (ii) gonococcal infection-induced apoptosis was inhibited with anti-TNF-alpha antibodies; and (iii) the addition of exogenous TNF-alpha induced apoptosis, which was inhibited by the presence of increasing numbers of bacteria (MOI = 10 to 100). These data suggest that TNF-alpha-mediated apoptosis of FT epithelial cells is likely a primary host defense mechanism to prevent pathogen colonization. However, epithelial cell-associated gonococci have evolved a mechanism to protect the cells from undergoing TNF-alpha-mediated apoptosis, and this modulation of the host innate response may contribute to establishment of infection. Understanding the antiapoptotic mechanisms used by Neisseria gonorrhoeae will inform the pathogenesis of salpingitis and could suggest new intervention strategies for prevention and treatment of the disease. PMID:16714596

  11. Glial cell line-derived neurotrophic factor promotes barrier maturation and wound healing in intestinal epithelial cells in vitro.

    PubMed

    Meir, Michael; Flemming, Sven; Burkard, Natalie; Bergauer, Lisa; Metzger, Marco; Germer, Christoph-Thomas; Schlegel, Nicolas

    2015-10-15

    Recent data suggest that neurotrophic factors from the enteric nervous system are involved in intestinal epithelial barrier regulation. In this context the glial cell line-derived neurotrophic factor (GDNF) was shown to affect gut barrier properties in vivo directly or indirectly by largely undefined processes in a model of inflammatory bowel disease (IBD). We further investigated the potential role and mechanisms of GDNF in the regulation of intestinal barrier functions. Immunostaining of human gut specimen showed positive GDNF staining in enteric neuronal plexus and in enterocytes. In Western blots of the intestinal epithelial cell lines Caco2 and HT29B6, significant amounts of GDNF were detected, suggesting that enterocytes represent an additional source of GDNF. Application of recombinant GDNF on Caco2 and HT29B6 cells for 24 h resulted in significant epithelial barrier stabilization in monolayers with immature barrier functions. Wound-healing assays showed a significantly faster closure of the wounded areas after GDNF application. GDNF augmented cAMP levels and led to significant inactivation of p38 MAPK in immature cells. Activation of p38 MAPK signaling by SB-202190 mimicked GDNF-induced barrier maturation, whereas the p38 MAPK activator anisomycin blocked GDNF-induced effects. Increasing cAMP levels had adverse effects on barrier maturation, as revealed by permeability measurements. However, increased cAMP augmented the proliferation rate in Caco2 cells, and GDNF-induced proliferation of epithelial cells was abrogated by the PKA inhibitor H89. Our data show that enterocytes represent an additional source of GDNF synthesis. GDNF contributes to wound healing in a cAMP/PKA-dependent manner and promotes barrier maturation in immature enterocytes cells by inactivation of p38 MAPK signaling. PMID:26294673

  12. Pharmacological effects of epidermal growth factor (EGF) with focus on the urinary and gastrointestinal tracts.

    PubMed

    Vinter-Jensen, L

    1999-01-01

    . The species of higher order were most sensitive to treatment with EGF. EGF did not consistently change the total body weight despite EGF consistently reduced circulating levels of insulin-like growth factor I (IGF-I) in Goettingen minipigs as well as in rats. Low circulating levels of IGF-I are usually associated with retarded growth. This review mostly focuses on the organs which appeared to be most sensitive to EGF, the urinary and gastrointestinal tracts including the liver and the pancreas. The histopathological changes consisted mainly of epithelial proliferations in the gastrointestinal, urinary and respiratory tracts. These findings match the knowledge obtained from animals overexpressing the EGF agonist, transforming growth factor alpha (TGF alpha), and the mice with a knock out of the gene encoding for the EGF receptor. EGF receptor hyperstimulation (TGF alpha overexpression) in the context of the whole animal leads to epithelial proliferations whereas hypostimulation (EGF receptor knock out) leads to epithelial immaturities. In the minipigs, the epithelia of the oesophagus, ducts of the pancreas, and the urothelium were hyperplastic, the latter two epithelia with accumulation of glycoconjugates. In the rats, the epithelial hyperplasias in these tissues and in the small and large intestines were without glycoconjugate accumulations. In rats, the mucosal proliferations in the intestines resulted in increased mucosal surface area. Mesenchymal growth effects were also noted. In the ureters of the minipigs, smooth muscle cell hyperplasia and hypertrophia were found. The heart of the minipigs was also enlarged, an interesting finding regarding interactions between the different parts of the EGF system, as knock out mice of the receptors erbB2 and erbB4 die due to maldevelopments in the heart. Measurements in blood and serum also revealed consistent changes. (ABSTRACT TRUNCATED) PMID:10424202

  13. Inhibition of histamine receptor 3 suppresses glioblastoma tumor growth, invasion, and epithelial-to-mesenchymal transition.