Preclinical development of a humanized neutralizing antibody targeting HGF.

PubMed

Kim, Hyori; Hong, Sung Hee; Kim, Jung Yong; Kim, In-Chull; Park, Young-Whan; Lee, Song-Jae; Song, Seong-Won; Kim, Jung Ju; Park, Gunwoo; Kim, Tae Min; Kim, Yun-Hee; Park, Jong Bae; Chung, Junho; Kim, In-Hoo

2017-03-24

Hepatocyte growth factor (HGF) and its receptor, cMET, play critical roles in cell proliferation, angiogenesis and invasion in a wide variety of cancers. We therefore examined the anti-tumor activity of the humanized monoclonal anti-HGF antibody, YYB-101, in nude mice bearing human glioblastoma xenografts as a single agent or in combination with temozolomide. HGF neutralization, The extracellular signal-related kinases 1 and 2 (ERK1/2) phosphorylation, and HGF-induced scattering were assessed in HGF-expressing cell lines treated with YYB-101. To support clinical development, we also evaluated the preclinical pharmacokinetics and toxicokinetics in cynomolgus monkeys, and human and cynomolgus monkey tissue was stained with YYB-101 to test tissue cross-reactivity. We found that YYB-101 inhibited cMET activation in vitro and suppressed tumor growth in the orthotopic mouse model of human glioblastoma. Combination treatment with YYB-101 and temozolomide decreased tumor growth and increased overall survival compared with the effects of either agent alone. Five cancer-related genes (TMEM119, FST, RSPO3, ROS1 and NBL1) were overexpressed in YYB-101-treated mice that showed tumor regrowth. In the tissue cross-reactivity assay, critical cross-reactivity was not observed. The terminal elimination half-life was 21.7 days. Taken together, the in vitro and in vivo data demonstrated the anti-tumor efficacy of YYB-101, which appeared to be mediated by blocking the HGF/cMET interaction. The preclinical pharmacokinetics, toxicokinetics and tissue cross-reactivity data support the clinical development of YYB-101 for advanced cancer.

The Hepatocyte Growth Factor (HGF)/Met Axis: A Neglected Target in the Treatment of Chronic Myeloproliferative Neoplasms?

PubMed Central

Boissinot, Marjorie; Vilaine, Mathias; Hermouet, Sylvie

2014-01-01

Met is the receptor of hepatocyte growth factor (HGF), a cytoprotective cytokine. Disturbing the equilibrium between Met and its ligand may lead to inappropriate cell survival, accumulation of genetic abnormalities and eventually, malignancy. Abnormal activation of the HGF/Met axis is established in solid tumours and in chronic haematological malignancies, including myeloma, acute myeloid leukaemia, chronic myelogenous leukaemia (CML), and myeloproliferative neoplasms (MPNs). The molecular mechanisms potentially responsible for the abnormal activation of HGF/Met pathways are described and discussed. Importantly, inCML and in MPNs, the production of HGF is independent of Bcr-Abl and JAK2V617F, the main molecular markers of these diseases. In vitro studies showed that blocking HGF/Met function with neutralizing antibodies or Met inhibitors significantly impairs the growth of JAK2V617F-mutated cells. With personalised medicine and curative treatment in view, blocking activation of HGF/Met could be a useful addition in the treatment of CML and MPNs for those patients with high HGF/MET expression not controlled by current treatments (Bcr-Abl inhibitors in CML; phlebotomy, hydroxurea, JAK inhibitors in MPNs). PMID:25119536

Hepatocyte growth factor fusion protein having collagen-binding activity (CBD-HGF) accelerates re-endothelialization and intimal hyperplasia in balloon-injured rat carotid artery.

PubMed

Ohkawara, Nana; Ueda, Hiroki; Shinozaki, Shohei; Kitajima, Takashi; Ito, Yoshihiro; Asaoka, Hiroshi; Kawakami, Akio; Kaneko, Eiji; Shimokado, Kentaro

2007-08-01

Hepatocyte growth factor (HGF) is known to stimulate endothelial cell proliferation. However, re-endothelialization is not enhanced when the native protein is administered to the injured artery, probably due to the short half-life of HGF at the site of injury. Therefore, the effects of an HGF fusion protein having collagen-binding activity (CBD-HGF) on re-endothelialization and neointimal formation was studied in the balloon-injured rat carotid artery. The left common carotid artery of male Sprague-Dawley rats was injured with an inflated balloon catheter, and then treated with CBD-HGF 10 microg/mL), HGF (10 micro g/mL) or saline (control) for 15 min. After 14 days, the rats were injected with Evans blue and sacrificed. The re-endothelialized area was significantly greater in the CBD-HGF- treated rats than in the control or HGF -treated rats. Neointimal formation was significantly more pronounced in the CBD-HGF treated rats than in other rat groups. Both HGF and CBD-HGF stimulated proliferation of vascular smooth muscle cells as well as endothelial cells in vitro. Consistent with this, cultured smooth muscle cells were shown to express the HGF receptor (c-Met). CBD-HGF accelerates re-endothelialization and neointimal formation in vivo. CBD fusion protein is a useful vehicle to deliver vascular growth factors to injured arteries.

Hepatocyte growth factor: a regulator of extracellular matrix genes in mouse mesangial cells.

PubMed

Laping, N J; Olson, B A; Ho, T; Ziyadeh, F N; Albrightson, C R

2000-04-01

The potential role of hepatocyte growth factor (HGF) in regulating extracellular matrix in mouse mesangial cells (MMC) was evaluated. Functional HGF receptors were deed in MMC by HGF-induced extracellular acidification, a response that was inhibited by the HGF inhibitor HGF/NK2, a splice variant expressing the N-terminal domain through the second kringle domain HGF also increased fibronectin and collagen alpha1 (IV) mRNA levels in these cells; the increases were associated with a concentration-dependent increase in transcriptional activity of the collagen alpha1 (IV) gene. HGF also stimulated fibronectin and collagen alpha1 (IV) mRNA levels in primary rabbit proximal tubule epithelial cells To evaluate the potential consequences of chronic elevation of HGF on renal fuction, HGF was administered continuously for 18 days to normal and diabetic C57BLKS/J lepr(db) mice. In the diabetic mice, HGF reduced creatinine clearance and increased microalbuminuria, indicating that chronic exposure to HGF impairs renal function. Thus, chronically elevated HGF may contribute to the progression of chronic renal disease in diabetes by decreasing the glomerular filtration rate and possibly promoting the accumulation of extracellular matrix.

Expression of p53/HGF/c-met/STAT3 signal in fetuses with neural tube defects.

PubMed

Trovato, Maria; D'Armiento, Maria; Lavra, Luca; Ulivieri, Alessandra; Dominici, Roberto; Vitarelli, Enrica; Grosso, Maddalena; Vecchione, Raffaella; Barresi, Gaetano; Sciacchitano, Salvatore

2007-02-01

Neural tube defects (NTD) are morphogenetic alterations due to a defective closure of neural tube. Hepatocyte growth factor (HGF)/c-met system plays a role in morphogenesis of nervous system, lung, and kidney. HGF/c-met morphogenetic effects are mediated by signal transducers and activators of transcription (STAT)3 and both HGF and c-met genes are regulated from p53. The aim of our study was to analyze mRNA and protein expressions of p53, HGF, c-met, and STAT3 in fetuses with NTD. By reverse transcriptase-polymerase chain reaction and immunohistochemistry, we analyzed neural tissues from four NTD fetuses and the corresponding non-malformed lungs, kidneys and placentas. We found a reduced mRNA expression of HGF/c-met/STAT3 pathway, in the malformed nervous systems and placentas. The reduced expression of this pathway correlated with the absence of p53 in all these samples. On the contrary, detectable expression levels of p53, HGF, c-met, and STAT3 were observed in non-malformed lungs and kidneys obtained from the same fetuses. Comparable results were obtained by immunohistochemistry, with the exception of p53, which was undetected in all fetal tissues. In conclusion, in NTD fetuses, both the defective neural tube tissue and the placenta have a reduction in all components of the p53/HGF/c-met/STAT3 cascade. This raises the possibility of using the suppression of these genes for early diagnosis of NTD especially on chorionic villus sampling.

Triple Inhibition of EGFR, Met, and VEGF Suppresses Regrowth of HGF-Triggered, Erlotinib-Resistant Lung Cancer Harboring an EGFR Mutation

PubMed Central

Nakade, Junya; Takeuchi, Shinji; Nakagawa, Takayuki; Ishikawa, Daisuke; Sano, Takako; Nanjo, Shigeki; Yamada, Tadaaki; Ebi, Hiromichi; Zhao, Lu; Yasumoto, Kazuo; Matsumoto, Kunio; Yonekura, Kazuhiko

2014-01-01

Introduction: Met activation by gene amplification and its ligand, hepatocyte growth factor (HGF), imparts resistance to epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (TKIs) in EGFR-mutant lung cancer. We recently reported that Met activation by HGF stimulates the production of vascular endothelial growth factor (VEGF) and facilitates angiogenesis, which indicates that HGF induces EGFR-TKI resistance and angiogenesis. This study aimed to determine the effect of triple inhibition of EGFR, Met, and angiogenesis on HGF-triggered EGFR-TKI resistance in EGFR-mutant lung cancer. Methods: Three clinically approved drugs, erlotinib (an EGFR inhibitor), crizotinib (an inhibitor of anaplastic lymphoma kinase and Met), and bevacizumab (anti-VEGF antibody), and TAS-115, a novel dual TKI for Met and VEGF receptor 2, were used in this study. EGFR-mutant lung cancer cell lines PC-9, HCC827, and HGF-gene–transfected PC-9 (PC-9/HGF) cells were examined. Results: Crizotinib and TAS-115 inhibited Met phosphorylation and reversed erlotinib resistance and VEGF production triggered by HGF in PC-9 and HCC827 cells in vitro. Bevacizumab and TAS-115 inhibited angiogenesis in PC-9/HGF tumors in vivo. Moreover, the triplet erlotinib, crizotinib, and bevacizumab, or the doublet erlotinib and TAS-115 successfully inhibited PC-9/HGF tumor growth and delayed tumor regrowth associated with sustained tumor vasculature inhibition even after cessation of the treatment. Conclusion: These results suggest that triple inhibition of EGFR, HGF/Met, and VEGF/VEGF receptor 2, by either a triplet of clinical drugs or TAS-115 combined with erlotinib, may be useful for controlling progression of EGFR-mutant lung cancer by reversing EGFR-TKI resistance and for inhibiting angiogenesis. PMID:24828661

Cellular and molecular mechanisms of HGF/Met in the cardiovascular system.

PubMed

Gallo, Simona; Sala, Valentina; Gatti, Stefano; Crepaldi, Tiziana

2015-12-01

Met tyrosine kinase receptor, also known as c-Met, is the HGF (hepatocyte growth factor) receptor. The HGF/Met pathway has a prominent role in cardiovascular remodelling after tissue injury. The present review provides a synopsis of the cellular and molecular mechanisms underlying the effects of HGF/Met in the heart and blood vessels. In vivo, HGF/Met function is particularly important for the protection of the heart in response to both acute and chronic insults, including ischaemic injury and doxorubicin-induced cardiotoxicity. Accordingly, conditional deletion of Met in cardiomyocytes results in impaired organ defence against oxidative stress. After ischaemic injury, activation of Met provides strong anti-apoptotic stimuli for cardiomyocytes through PI3K (phosphoinositide 3-kinase)/Akt and MAPK (mitogen-activated protein kinase) cascades. Recently, we found that HGF/Met is also important for autophagy regulation in cardiomyocytes via the mTOR (mammalian target of rapamycin) pathway. HGF/Met induces proliferation and migration of endothelial cells through Rac1 (Ras-related C3 botulinum toxin substrate 1) activation. In fibroblasts, HGF/Met antagonizes the actions of TGFβ1 (transforming growth factor β1) and AngII (angiotensin II), thus preventing fibrosis. Moreover, HGF/Met influences the inflammatory response of macrophages and the immune response of dendritic cells, indicating its protective function against atherosclerotic and autoimmune diseases. The HGF/Met axis also plays an important role in regulating self-renewal and myocardial regeneration through the enhancement of cardiac progenitor cells. HGF/Met has beneficial effects against myocardial infarction and endothelial dysfunction: the cellular and molecular mechanisms underlying repair function in the heart and blood vessels are common and include pro-angiogenic, anti-inflammatory and anti-fibrotic actions. Thus administration of HGF or HGF mimetics may represent a promising therapeutic agent for the

[PLA-O-CMC nanoparticles: HGF loading and delivery behaviors in vitro].

PubMed

Li, Zhifeng; Chen, Zhong; Chang, Ren'an

2011-04-01

This paper is aimed to observe the hepatocyte growth factor (HGF) loading and delivery ability of polylactic acid and oxygen carboxymethylated chitosan copolyer nanoparticles (PLA-O-CMC NPs). We prepared PLA-O-CMC NPs loaded with HGF by ultrasound in combination with magnetic stirring method. The NPs were characterized by transmission electron microscopy, embedding ratio; drug loading and drug delivery behaviors were observed by ELISA. The characteristics of PLA-O-CMC NPs loaded with HGF showed that the mean size was 139. 82 nm, polydispersity was 0.108, maximal HGF-embedding ratio was 76. 32%. The cumulative HGF release gradually increased in the first 24 hours in vitro, with sharp increasing in the first 7 hours, and moderate and steady increasing in the following 17 hours. The HGF had a burst release in the first 24 hours, and in this process the released HGF took up 36.7% of the whole release. From the second day,the HGF release decreased obviously, while it kept on releasing steadily (45-55 ng/d) for quite long time up to 30 days. The experiment proved that PLA-O-CMC NPs is a favourable carrier of HGF. PLA-O-CMC NPs loaded with HGF could rapidly release HGF in vitro. The released HGF reached the effective drug concentration and maintained the certain effective drug concentration for a long time.

Hepatocyte growth factor induces resistance to anti-epidermal growth factor receptor antibody in lung cancer.

PubMed

Yamada, Tadaaki; Takeuchi, Shinji; Kita, Kenji; Bando, Hideaki; Nakamura, Takahiro; Matsumoto, Kunio; Yano, Seiji

2012-02-01

Epidermal growth factor receptor (EGFR) is an attractive drug target in lung cancer, with several anti-EGFR antibodies and small-molecule inhibitors showing efficacy in lung cancer patients. Patients, however, may develop resistance to EGFR inhibitors. We demonstrated previously that hepatocyte growth factor (HGF) induced resistance to EGFR tyrosine kinase inhibitors in lung cancers harboring EGFR mutations. We therefore determined whether HGF could induce resistance to the anti-EGFR antibody (EGFR Ab) cetuximab in lung cancer cells, regardless of EGFR gene status. Cetuximab sensitivity and signal transduction in lung cancer cells were examined in the presence or absence of HGF, HGF-producing fibroblasts, and cells tranfected with the HGF gene in vitro and in vivo. HGF induced resistance to cetuximab in H292 (EGFR wild) and Ma-1(EGFR mutant) cells. Western blotting showed that HGF-induced resistance was mediated by the Met/Gab1/Akt signaling pathway. Resistance of H292 and Ma-1 cells to cetuximab was also induced by coculture with lung fibroblasts producing high levels of HGF and by cells stably transfected with the HGF gene. This resistance was abrogated by treatment with anti-HGF neutralizing antibody. HGF-mediated resistance is a novel mechanism of resistance to EGFR Ab in lung cancers, with fibroblast-derived HGF inducing cetuximab resistance in H292 tumors in vivo. The involvement of HGF-Met-mediated signaling should be assessed in acquired resistance to EGFR Ab in lung cancer, regardless of EGFR gene status.

HGF and c-Met Interaction Promotes Migration in Human Chondrosarcoma Cells

PubMed Central

Tsou, Hsi-Kai; Chen, Hsien-Te; Hung, Ya-Huey; Chang, Chia-Hao; Li, Te-Mao; Fong, Yi-Chin; Tang, Chih-Hsin

2013-01-01

Chondrosarcoma is a type of highly malignant tumor with a potent capacity for local invasion and causing distant metastasis. Chondrosarcoma shows a predilection for metastasis to the lungs. Hepatocyte growth factor (HGF) has been demonstrated to stimulate cancer proliferation, migration, and metastasis. However, the effect of HGF on migration activity of human chondrosarcoma cells is not well known. Here, we found that human chondrosarcoma tissues demonstrated significant expression of HGF, which was higher than that in normal cartilage. We also found that HGF increased the migration and expression of matrix metalloproteinase (MMP)-2 in human chondrosarcoma cells. c-Met inhibitor and siRNA reduced HGF-increased cell migration and MMP-2 expression. HGF treatment resulted in activation of the phosphatidylinositol 3′-kinase (PI3K)/Akt/PKCδ/NF-κB pathway, and HGF-induced expression of MMP-2 and cell migration was inhibited by specific inhibitors or siRNA-knockdown of PI3K, Akt, PKCδ, and NF-κB cascades. Taken together, our results indicated that HGF enhances migration of chondrosarcoma cells by increasing MMP-2 expression through the c-Met receptor/PI3K/Akt/PKCδ/NF-κB signal transduction pathway. PMID:23320110

Role of HGF in epithelial–stromal cell interactions during progression from benign breast disease to ductal carcinoma in situ

PubMed Central

2013-01-01

Introduction Basal-like and luminal breast cancers have distinct stromal–epithelial interactions, which play a role in progression to invasive cancer. However, little is known about how stromal–epithelial interactions evolve in benign and pre-invasive lesions. Methods To study epithelial–stromal interactions in basal-like breast cancer progression, we cocultured reduction mammoplasty fibroblasts with the isogenic MCF10 series of cell lines (representing benign/normal, atypical hyperplasia, and ductal carcinoma in situ). We used gene expression microarrays to identify pathways induced by coculture in premalignant cells (MCF10DCIS) compared with normal and benign cells (MCF10A and MCF10AT1). Relevant pathways were then evaluated in vivo for associations with basal-like subtype and were targeted in vitro to evaluate effects on morphogenesis. Results Our results show that premalignant MCF10DCIS cells express characteristic gene expression patterns of invasive basal-like microenvironments. Furthermore, while hepatocyte growth factor (HGF) secretion is upregulated (relative to normal, MCF10A levels) when fibroblasts are cocultured with either atypical (MCF10AT1) or premalignant (MCF10DCIS) cells, only MCF10DCIS cells upregulated the HGF receptor MET. In three-dimensional cultures, upregulation of HGF/MET in MCF10DCIS cells induced morphological changes suggestive of invasive potential, and these changes were reversed by antibody-based blocking of HGF signaling. These results are relevant to in vivo progression because high expression of a novel MCF10DCIS-derived HGF signature was correlated with the basal-like subtype, with approximately 86% of basal-like cancers highly expressing the HGF signature, and because high expression of HGF signature was associated with poor survival. Conclusions Coordinated and complementary changes in HGF/MET expression occur in epithelium and stroma during progression of pre-invasive basal-like lesions. These results suggest that

Regulation of HGF and c-MET Interaction in Normal Ovary and Ovarian Cancer.

PubMed

Kwon, Youngjoo; Godwin, Andrew K

2017-04-01

Binding of hepatocyte growth factor (HGF) to the c-MET receptor has mitogenic, motogenic, and morphogenic effects on cells. The versatile biological effects of HGF and c-MET interactions make them important contributors to the development of malignant tumors. We and others have demonstrated a therapeutic value in targeting the interaction of c-MET and HGF in epithelial ovarian cancer (EOC). However, both HGF and c-MET are expressed in the normal ovary as well. Therefore, it is important to understand the differences in mechanisms that control HGF signaling activation and its functional role in the normal ovary and EOC. In the normal ovary, HGF signaling may be under hormonal regulation. During ovulation, HGF-converting proteases are secreted and the subsequent activation of HGF signaling enhances the proliferation of ovarian surface epithelium in order to replenish the area damaged due to expulsion of the ovum. In contrast, EOC cells that exhibit epithelial characteristics constitutively express both c-MET and HGF-converting proteases such as urokinase-type plasminogen activator. In EOC, mechanisms to control the activation of HGF signaling are absent since HGF is provided locally from the tissue microenvironment as well as remotely throughout the body. Potential incessant HGF signaling in EOC may lead to an increase in proliferation, invasion through the stroma, and migration to other tissues of cancer cells. Therefore, targeting the interaction of c-MET and HGF would be beneficial in treating EOC.

Regulation of HGF and c-MET Interaction in Normal Ovary and Ovarian Cancer

PubMed Central

Kwon, Youngjoo; Godwin, Andrew K.

2016-01-01

Binding of hepatocyte growth factor (HGF) to the c-MET receptor has mitogenic, motogenic, and morphogenic effects on cells. The versatile biological effects of HGF and c-MET interactions make them important contributors to the development of malignant tumors. We and others have demonstrated a therapeutic value in targeting the interaction of c-MET and HGF in epithelial ovarian cancer (EOC). However, both HGF and c-MET are expressed in the normal ovary as well. Therefore, it is important to understand the differences in mechanisms that control HGF signaling activation and its functional role in the normal ovary and EOC. In the normal ovary, HGF signaling may be under hormonal regulation. During ovulation, HGF-converting proteases are secreted and the subsequent activation of HGF signaling enhances the proliferation of ovarian surface epithelium in order to replenish the area damaged due to expulsion of the ovum. In contrast, EOC cells that exhibit epithelial characteristics constitutively express both c-MET and HGF-converting proteases such as urokinase-type plasminogen activator. In EOC, mechanisms to control the activation of HGF signaling are absent since HGF is provided locally from the tissue microenvironment as well as remotely throughout the body. Potential incessant HGF signaling in EOC may lead to an increase in proliferation, invasion through the stroma, and migration to other tissues of cancer cells. Therefore, targeting the interaction of c-MET and HGF would be beneficial in treating EOC. PMID:27170665

Recent Progress and Advances in HGF/MET-Targeted Therapeutic Agents for Cancer Treatment

PubMed Central

Zhang, Yilong; Jain, Rajul K.; Zhu, Min

2015-01-01

The hepatocyte growth factor (HGF): MET axis is a ligand-mediated receptor tyrosine kinase pathway that is involved in multiple cellular functions, including proliferation, survival, motility, and morphogenesis. Aberrancy in the HGF/MET pathway has been reported in multiple tumor types and is associated with tumor stage and prognosis. Thus, targeting the HGF/MET pathway has become a potential therapeutic strategy in oncology development in the last two decades. A number of novel therapeutic agents—either as therapeutic proteins or small molecules that target the HGF/MET pathway—have been tested in patients with different tumor types in clinical studies. In this review, recent progress in HGF/MET pathway-targeted therapy for cancer treatment, the therapeutic potential of HGF/MET-targeted agents, and challenges in the development of such agents will be discussed. PMID:28536405

Hepatocyte growth factor and transforming growth factor beta regulate 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase gene expression in rat hepatocyte primary cultures.

PubMed Central

Joaquin, M; Rosa, J L; Salvadó, C; López, S; Nakamura, T; Bartrons, R; Gil, J; Tauler, A

1996-01-01

Hepatocyte growth factor (HGF) and transforming growth factor beta (TGF-beta) are believed to be of major importance for hepatic regeneration after liver damage. We have studied the effect of these growth factors on fructose 2,6-bisphosphate (Fru-2,6-P2) levels and the expression of 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase (6PF2K/Fru-2,6-BPase) in rat hepatocyte primary cultures. Our results demonstrate that HGF activates the expression of the 6PF2K/Fru-2,6-BPase gene by increasing the levels of its mRNA. As a consequence of this activation, the amount of 6PF2K/Fru-2,6-BPase protein and 6-phosphofructo-2-kinase activity increased, which was reflected by a rise in Fru-2,6-P2 levels. In contrast, TGF-beta decreased the levels of 6PF2K/Fru-2,6-BPase mRNA, which led to a decrease in the amount of 6PF2K/Fru-2,6-BPase protein and Fru-2,6-P2. The different actions of HGF and TGF-beta on 6PF2K/Fru-2,6-BPase gene expression are concomitant with their effect on cell proliferation. Here we show that, in the absence of hormones, primary cultures of hepatocytes express the F-type isoenzyme. In addition, HGF increases the expression of this isoenzyme, and dexamethasone activates the L-type isoform. HGF and TGF-beta were able to inhibit this activation. PMID:8660288

Combined administration of mesenchymal stem cells overexpressing IGF-1 and HGF enhances neovascularization but moderately improves cardiac regeneration in a porcine model.

PubMed

Gómez-Mauricio, Guadalupe; Moscoso, Isabel; Martín-Cancho, María-Fernanda; Crisóstomo, Verónica; Prat-Vidal, Cristina; Báez-Díaz, Claudia; Sánchez-Margallo, Francisco M; Bernad, Antonio

2016-07-16

Insulin-like growth factor 1 (IGF-1) and hepatocyte growth factor (HGF) are among the most promising growth factors for promoting cardiorepair. Here, we evaluated the combination of cell- and gene-based therapy using mesenchymal stem cells (MSC) genetically modified to overexpress IGF-1 or HGF to treat acute myocardial infarction (AMI) in a porcine model. Pig MSC from adipose tissue (paMSC) were genetically modified for evaluation of different therapeutic strategies to improve AMI treatment. Three groups of infarcted Large White pigs were compared (I, control, non-transplanted; II, transplanted with paMSC-GFP (green fluorescent protein); III, transplanted with paMSC-IGF-1/HGF). Cardiac function was evaluated non-invasively using magnetic resonance imaging (MRI) for 1 month. After euthanasia and sampling of the animal, infarcted areas were studied by histology and immunohistochemistry. Intramyocardial transplant in a porcine infarct model demonstrated the safety of paMSC in short-term treatments. Treatment with paMSC-IGF-1/HGF (1:1) compared with the other groups showed a clear reduction in inflammation in some sections analyzed and promoted angiogenic processes in ischemic tissue. Although cardiac function parameters were not significantly improved, cell retention and IGF-1 overexpression was confirmed within the myocardium. The simultaneous administration of IGF-1- and HGF-overexpressing paMSC appears not to promote a synergistic effect or effective repair. The combined enhancement of neovascularization and fibrosis in paMSC-IGF-1/HGF-treated animals nonetheless suggests that sustained exposure to high IGF-1 + HGF levels promotes beneficial as well as deleterious effects that do not improve overall cardiac regeneration.

TEC protein tyrosine kinase is involved in the Erk signaling pathway induced by HGF

DOE Office of Scientific and Technical Information (OSTI.GOV)

Li, Feifei; Jiang, Yinan; Zheng, Qiping

Research highlights: {yields} TEC is rapidly tyrosine-phosphorylated and activated by HGF-stimulation in vivo or after partial hepatectomy in mice. {yields} TEC enhances the activity of Elk and serum response element (SRE) in HGF signaling pathway in hepatocyte. {yields} TEC promotes hepatocyte proliferation through the Erk-MAPK pathway. -- Abstract: Background/aims: TEC, a member of the TEC family of non-receptor type protein tyrosine kinases, has recently been suggested to play a role in hepatocyte proliferation and liver regeneration. This study aims to investigate the putative mechanisms of TEC kinase regulation of hepatocyte differentiation, i.e. to explore which signaling pathway TEC is involvedmore » in, and how TEC is activated in hepatocyte after hepatectomy and hepatocyte growth factor (HGF) stimulation. Methods: We performed immunoprecipitation (IP) and immunoblotting (IB) to examine TEC tyrosine phosphorylation after partial hepatectomy in mice and HGF stimulation in WB F-344 hepatic cells. The TEC kinase activity was determined by in vitro kinase assay. Reporter gene assay, antisense oligonucleotide and TEC dominant negative mutant (TEC{sup KM}) were used to examine the possible signaling pathways in which TEC is involved. The cell proliferation rate was evaluated by {sup 3}H-TdR incorporation. Results: TEC phosphorylation and kinase activity were increased in 1 h after hepatectomy or HGF treatment. TEC enhanced the activity of Elk and serum response element (SRE). Inhibition of MEK1 suppressed TEC phosphorylation. Blocking TEC activity dramatically decreased the activation of Erk. Reduced TEC kinase activity also suppressed the proliferation of WB F-344 cells. These results suggest TEC is involved in the Ras-MAPK pathway and acts between MEK1 and Erk. Conclusions: TEC promotes hepatocyte proliferation and regeneration and is involved in HGF-induced Erk signaling pathway.« less

Neural differentiation of mesenchymal stem cells influences chemotactic responses to HGF.

PubMed

Zheng, Bing; Wang, Chunyan; He, Lihong; Xu, Xiaojing; Qu, Jing; Hu, Jun; Zhang, Huanxiang

2013-01-01

Recently, mesenchymal stem cells (MSCs) have been extensively used for cell-based therapies in neuronal degenerative disease. Although much effort has been devoted to the delineation of factors involved in the migration of MSCs, the relationship between the chemotactic responses and the differentiation status of these cells remains elusive. Here, we report that MSCs in varying neural differentiation states display different chemotactic responses to hepatocyte growth factor (HGF): first, the number of chemotaxing MSCs and the optimal concentrations of HGF that induced the peak migration varied greatly; second, time-lapse video analysis showed that MSCs in certain differentiation state migrated more efficiently toward HGF; third, the phosphorylation levels of Akt, ERK1/2, SAPK/JNK, and p38MAPK were closely related to the differentiation levels of MSCs subjected to HGF; and finally, although inhibition of ERK1/2 signaling significantly attenuated HGF-stimulated transfilter migration of both undifferentiated and differentiating MSCs, abolishment of PI3K/Akt, p38MAPK, or SAPK/JNK signaling only decreased the number of migrated cells in certain differentiation state(s). Blocking of PI3K/Akt or MAPK signaling impaired the migration efficiency and/or speed, the extent of which depends on the cell differentiation states. Meanwhile, F-actin rearrangement, which is essential for MSCs chemotaxis, was induced by HGF, and the time points of cytoskeletal reorganization were different among these cells. Collectively, these results demonstrate that neural differentiation of MSCs influences their chemotactic responses to HGF: MSCs in varying differentiation states possess different migratory capacities, thereby shedding light on optimization of the therapeutic potential of MSCs to be employed for neural regeneration after injury. Copyright © 2012 Wiley Periodicals, Inc.

The loss of local HGF, an endogenous gastrotrophic factor, leads to mucosal injuries in the stomach of mice

DOE Office of Scientific and Technical Information (OSTI.GOV)

Nakahira, Rie; Department of Neurosurgery, Osaka University Graduate School of Medicine, Yamadaoka 2-2-E6, Suita 565-0871; Mizuno, Shinya

2006-03-24

The stomach is constantly exposed to mechanical and chemical stresses. Under persistent damages, epithelial cell proliferation is required to maintain mucosal integrity. Nevertheless, which ligand system(s) is physiologically involved in gastric defense remains unclear. Herein, we provide evidence that HGF is a key 'natural ligand' to reverse gastric injury. The injection of cisplatin in mice led to the loss of HGF in the gastric interstitium, associated with the decrease in proliferating epithelium and the progression of mucotitis. When c-Met tyrosine phosphorylation was abolished by anti-HGF IgG, mucosal cell proliferation became faint, leading to delayed recovery from mucotitis, and vice versamore » in cases of HGF supplementation. Our findings indicate that: (1) HGF/c-Met signal on mucosa is needed to restore gastric injuries; and (2) the loss of local HGF leads to manifestation of gastric lesions. This study provides a rationale that explains why HGF supplement is useful for reversing gastric diseases.« less

Pyridine-pyrimidine amides that prevent HGF-induced epithelial scattering by two distinct mechanisms.

PubMed

Siddiqui-Jain, Adam; Hoj, Jacob P; Hargiss, J Blade; Hoj, Taylor H; Payne, Carter J; Ritchie, Collin A; Herron, Steven R; Quinn, Colette; Schuler, Jeffrey T; Hansen, Marc D H

2017-09-01

Stimulation of cultured epithelial cells with scatter factor/hepatocyte growth factor (HGF) results in individual cells detaching and assuming a migratory and invasive phenotype. Epithelial scattering recapitulates cancer progression and studies have implicated HGF signaling as a driver of cancer metastasis. Inhibitors of HGF signaling have been proposed to act as anti-cancer agents. We previously screened a small molecule library for compounds that block HGF-induced epithelial scattering. Most hits identified in this screen exhibit anti-mitotic properties. Here we assess the biological mechanism of a compound that blocks HGF-induced scattering with limited anti-mitotic activity. Analogs of this compound have one of two distinct activities: inhibiting either cell migration or cell proliferation with cell cycle arrest in G2/M. Each activity bears unique structure-activity relationships. The mechanism of action of anti-mitotic compounds is by inhibition of microtubule polymerization; these compounds entropically and enthalpically bind tubulin in the colchicine binding site, generating a conformational change in the tubulin dimer. Copyright © 2017 Elsevier Ltd. All rights reserved.

BetaPIX and GIT1 regulate HGF-induced lamellipodia formation and WAVE2 transport.

PubMed

Morimura, Shigeru; Suzuki, Katsuo; Takahashi, Kazuhide

2009-05-08

Formation of lamellipodia is the first step during cell migration, and involves actin reassembly at the leading edge of migrating cells through the membrane transport of WAVE2. However, the factors that regulate WAVE2 transport to the cell periphery for initiating lamellipodia formation have not been elucidated. We report here that in human breast cancer MDA-MB-231 cells, the hepatocyte growth factor (HGF) induced the association between the constitutive complex of betaPIX and GIT1 with WAVE2, which was concomitant with the induction of lamellipodia formation and WAVE2 transport. Although depletion of betaPIX by RNA interference abrogated the HGF-induced WAVE2 transport and lamellipodia formation, GIT1 depletion caused HGF-independent WAVE2 transport and lamellipodia formation. Collectively, we suggest that betaPIX releases cells from the GIT1-mediated suppression of HGF-independent responses and recruits GIT1 to WAVE2, thereby facilitating HGF-induced WAVE2 transport and lamellipodia formation.

Targeted delivery of HGF to the skeletal muscle improves glucose homeostasis in diet-induced obese mice.

PubMed

Sanchez-Encinales, Viviana; Cozar-Castellano, Irene; Garcia-Ocaña, Adolfo; Perdomo, Germán

2015-12-01

Hepatocyte growth factor (HGF) is a cytokine that increases glucose transport ex vivo in skeletal muscle. The aim of this work was to decipher the impact of whether conditional overexpression of HGF in vivo could improve glucose homeostasis and insulin sensitivity in mouse skeletal muscle. Following tetracyclin administration, muscle HGF levels were augmented threefold in transgenic mice (SK-HGF) compared to control mice without altering plasma HGF levels. In conditions of normal diet, SK-HGF mice showed no differences in body weight, plasma triglycerides, blood glucose, plasma insulin and glucose tolerance compared to control mice. Importantly, obese SK-HGF mice exhibited improved whole-body glucose tolerance independently of changes in body weight or plasma triglyceride levels compared to control mice. This effect on glucose homeostasis was associated with significantly higher (∼80%) levels of phosphorylated protein kinase B in muscles from SK-HGF mice compared to control mice. In conclusion, muscle expression of HGF counteracts obesity-mediated muscle insulin resistance and improves glucose tolerance in mice.

Epigenetic Upregulation of HGF and c-Met Drives Metastasis in Hepatocellular Carcinoma

PubMed Central

Ogunwobi, Olorunseun O.; Puszyk, William; Dong, Hui-Jia; Liu, Chen

2013-01-01

Hepatocyte growth factor (HGF) and its receptor, c-Met, are important regulators of growth and differentiation of healthy hepatocytes. However, upregulation of HGF and c-Met have been associated with tumor progression and metastasis in hepatocellular carcinoma (HCC). Hematogenous dissemination is the most common route for cancer metastasis, but the role of HGF and c-Met in circulating tumor cells (CTCs) is unknown. We have isolated and established a circulating tumor cell line from the peripheral blood of a mouse HCC model. Our studies show that these CTCs have increased expression of HGF and c-Met in comparison to the primary tumor cells. The CTCs display phenotypic evidence of epithelial-mesenchymal transition (EMT) and the EMT appears to be inducible by HGF. Epigenetic analysis of the c-Met promoter identified significant loss of DNA methylation in CTCs which correlated with overexpression of c-Met and increased expression of HGF. Six specific CpG sites of c-Met promoter demethylation were identified. CTCs show significantly increased tumorigenicity and metastatic potential in a novel orthotopic syngeneic model of metastatic HCC. We conclude that during hematogenous dissemination in HCC, CTCs undergo EMT under the influence of increased HGF. This process also involves up regulation of c-Met via promoter demethylation at 6 CpG sites. Consequently, targeting HGF and c-Met expression by CTCs may be a novel non-invasive approach with potential clinical applications in HCC management. PMID:23723997

Dual inhibition of Met kinase and angiogenesis to overcome HGF-induced EGFR-TKI resistance in EGFR mutant lung cancer.

PubMed

Takeuchi, Shinji; Wang, Wei; Li, Qi; Yamada, Tadaaki; Kita, Kenji; Donev, Ivan S; Nakamura, Takahiro; Matsumoto, Kunio; Shimizu, Eiji; Nishioka, Yasuhiko; Sone, Saburo; Nakagawa, Takayuki; Uenaka, Toshimitsu; Yano, Seiji

2012-09-01

Acquired resistance to epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (TKIs) is a serious problem in the management of EGFR mutant lung cancer. We recently reported that hepatocyte growth factor (HGF) induces resistance to EGFR-TKIs by activating the Met/PI3K pathway. HGF is also known to induce angiogenesis in cooperation with vascular endothelial growth factor (VEGF), which is an important therapeutic target in lung cancer. Therefore, we hypothesized that dual inhibition of HGF and VEGF may be therapeutically useful for controlling HGF-induced EGFR-TKI-resistant lung cancer. We found that a dual Met/VEGF receptor 2 kinase inhibitor, E7050, circumvented HGF-induced EGFR-TKI resistance in EGFR mutant lung cancer cell lines by inhibiting the Met/Gab1/PI3K/Akt pathway in vitro. HGF stimulated VEGF production by activation of the Met/Gab1 signaling pathway in EGFR mutant lung cancer cell lines, and E7050 showed an inhibitory effect. In a xenograft model, tumors produced by HGF-transfected Ma-1 (Ma-1/HGF) cells were more angiogenic than vector control tumors and showed resistance to gefitinib. E7050 alone inhibited angiogenesis and retarded growth of Ma-1/HGF tumors. E7050 combined with gefitinib induced marked regression of tumor growth. Moreover, dual inhibition of HGF and VEGF by neutralizing antibodies combined with gefitinib also markedly regressed tumor growth. These results indicate the therapeutic rationale of dual targeting of HGF-Met and VEGF-VEGF receptor 2 for overcoming HGF-induced EGFR-TKI resistance in EGFR mutant lung cancer. Copyright © 2012 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.

Activated HGF-c-Met Axis in Head and Neck Cancer

PubMed Central

Arnold, Levi; Enders, Jonathan; Thomas, Sufi Mary

2017-01-01

Head and neck squamous cell carcinoma (HNSCC) is a highly morbid disease. Recent developments including Food and Drug Administration (FDA) approved molecular targeted agent’s pembrolizumab and cetuximab show promise but did not improve the five-year survival which is currently less than 40%. The hepatocyte growth factor receptor; also known as mesenchymal–epithelial transition factor (c-Met) and its ligand hepatocyte growth factor (HGF) are overexpressed in head and neck squamous cell carcinoma (HNSCC); and regulates tumor progression and response to therapy. The c-Met pathway has been shown to regulate many cellular processes such as cell proliferation, invasion, and angiogenesis. The c-Met pathway is involved in cross-talk, activation, and perpetuation of other signaling pathways, curbing the cogency of a blockade molecule on a single pathway. The receptor and its ligand act on several downstream effectors including phospholipase C gamma (PLCγ), cellular Src kinase (c-Src), phosphotidylinsitol-3-OH kinase (PI3K) alpha serine/threonine-protein kinase (Akt), mitogen activate protein kinase (MAPK), and wingless-related integration site (Wnt) pathways. They are also known to cross-talk with other receptors; namely epidermal growth factor receptor (EGFR) and vascular endothelial growth factor receptor (VEGFR) and specifically contribute to treatment resistance. Clinical trials targeting the c-Met axis in HNSCC have been undertaken because of significant preclinical work demonstrating a relationship between HGF/c-Met signaling and cancer cell survival. Here we focus on HGF/c-Met impact on cellular signaling in HNSCC to potentiate tumor growth and disrupt therapeutic efficacy. Herein we summarize the current understanding of HGF/c-Met signaling and its effects on HNSCC. The intertwining of c-Met signaling with other signaling pathways provides opportunities for more robust and specific therapies, leading to better clinical outcomes. PMID:29231907

TGF-beta and HGF transmit the signals through JNK-dependent Smad2/3 phosphorylation at the linker regions.

PubMed

Mori, Shigeo; Matsuzaki, Koichi; Yoshida, Katsunori; Furukawa, Fukiko; Tahashi, Yoshiya; Yamagata, Hideo; Sekimoto, Go; Seki, Toshihito; Matsui, Hirofumi; Nishizawa, Mikio; Fujisawa, Jun-ichi; Okazaki, Kazuichi

2004-09-23

Although hepatocyte growth factor (HGF) can act synergistically or antagonistically with transforming growth factor-beta (TGF-beta) signaling, molecular mechanism of their crosstalk remains unknown. Using antibodies which selectively distinguished receptor-regulated Smads (R-Smads) phosphorylated at linker regions from those at C-terminal regions, we herein showed that either HGF or TGF-beta treatment of normal stomach-origin cells activated the JNK pathway, thereafter inducing endogenous R-Smads phosphorylation at linker regions. However, the phosphorylation at their C-terminal regions was not induced by HGF treatment. The activated JNK could directly phosphorylate R-Smads in vitro at the same sites that were phosphorylated in response to TGF-beta or HGF in vivo. Thus, the linker regions of R-Smads were the common phosphorylation sites for HGF and TGF-beta signaling pathways. The phosphorylation induced by simultaneous treatment with HGF and TGF-beta allowed R-Smads to associate with Smad4 and to translocate into the nucleus. JNK pathway involved HGF and TGF-beta-mediated infiltration potency since a JNK inhibitor SP600125 caused the reduction of invasive capacity induced by HGF and TGF-beta signals. Moreover, a combined treatment with HGF and TGF-beta led to a potent increase in plasminogen activator inhibitor type 1 transcriptional activity through Smad3 phosphorylation at the linker region. In contrast, HGF treatment reduced TGF-beta-dependent activation of p15INK4B promoter, in which Smad3 phosphorylation at the C-terminal region was involved. In conclusion, HGF and TGF-beta transmit the signals through JNK-mediated R-Smads phosphorylation at linker regions.

HGF and IGF-1 promote protective effects of allogeneic BMSC transplantation in rabbit model of acute myocardial infarction.

PubMed

Zhang, Guang-Wei; Gu, Tian-Xiang; Guan, Xiao-Yu; Sun, Xue-Jun; Qi, Xun; Li, Xue-Yuan; Wang, Xiao-Bing; Lv, Feng; Yu, Lei; Jiang, Da-Qing; Tang, Rui

2015-12-01

To explore effects of hepatocyte growth factor (HGF) combined with insulin-like growth factor 1 (IGF-1) on transplanted bone marrow mesenchymal stem cells (BMSCs), for treatment of acute myocardial ischaemia. After ligation of the left anterior descending artery, rabbits were divided into a Control group, a Factors group (HGF+IGF-1), a BMSC group and a Factors+BMSCs group. Allogenous BMSCs (1 × 10(7)) and/or control-released microspheres of 2 μg HGF+2 μg IGF-1 were intramyocardially injected into infarcted regions. Apoptosis and differentiation of implanted BMSCs, histological and morphological results, and cardiac remodelling and function were evaluated at different time points. In vitro, BMSCs were exposed to HGF, IGF-1 and both (50 ng/ml) and subsequently proliferation, migration, myocardial differentiation and apoptosis induced by hypoxia, were analysed. Four weeks post-operatively, the above indices were significantly improved in Factors+BMSCs group compared to the others (P < 0.01), although Factors and BMSCs group also showed better results than Control group (P < 0.05). In vitro, HGF promoted BMSC migration and differentiation into cardiomyocytes, but inhibited proliferation (P < 0.05), while IGF-1 increased proliferation and migration, and inhibited apoptosis induced by hypoxia (P < 0.05), but did not induce myocardial differentiation. Combination of HGF and IGF-1 significantly promoted BMSCs capacity for migration, differentiation and lack of apoptosis (P < 0.05). Combination of HGF and IGF-1 activated BMSCs complementarily, and controlled release of the two factors promoted protective potential of transplanted BMSCs to repair infarcted myocardium. This suggests a new strategy for cell therapies to overcome acute ischemic myocardial injury. © 2015 John Wiley & Sons Ltd.

Hepatocyte Growth Factor Gene-Modified Mesenchymal Stem Cells Augment Sinonasal Wound Healing

PubMed Central

Li, Jing; Li, Yong; Yang, Chen; Lin, Hai; Duan, Hong-Gang

2015-01-01

This study was designed to investigate the effects of hepatocyte growth factor (HGF) transgenic mesenchymal stem cells (HGF-MSCs) on wound healing in the sinonasal mucosa and nasal epithelial cells (NECs). We also sought to determine whether HGF-MSCs and MSCs can migrate into the injured mucosa and differentiate into ciliated cells. Human HGF-overexpressing umbilical cord MSCs (hHGF-UCMSCs) were established, and upregulation of hHGF expression was confirmed by real-time PCR (RT-PCR) and enzyme-linked immunosorbant assay (ELISA). To investigate the paracrine effect of human MSCs (hMSCs) on nasal epithelial repair, hMSC- and HGF-MSC-conditioned media (CM) were used in NEC proliferation assays and in an in vitro scratch-wound repair model. The in vivo sinonasal wound-healing model was established, and all enrolled rabbits were randomly assigned to four groups: the GFP-MSC group, the HGF-MSC group, the Ad-HGF group, and the surgery control group. The average decreased diameter was recorded, and the medial wall of the maxillary sinus was removed for histological analysis and scanning electron microscopy. Collagen deposition in the wound tissue was detected via Masson trichrome (M&T) staining. The distribution of MSCs and HGF-MSCs was observed by immunofluorescence. MSCs improved nasal wound healing both in vivo and in vitro. HGF overexpression in MSCs augmented the curative effects. Reduced collagen deposition and transforming growth factor beta1 (TGF-β1) expression were detected in the HGF-MSC group compared with the MSC-, Ad-HGF-, and phosphate-buffered saline-treated groups based on M&T staining and ELISA. The enhanced therapeutic effects of HGF-MSCs were accompanied by decreased level of the fibrogenic cytokine TGF-β1. In addition, both HGF-MSCs and MSCs can migrate to the injured mucosa and epithelial layer. PMID:25835956

Hepatocyte Growth Factor Gene-Modified Mesenchymal Stem Cells Augment Sinonasal Wound Healing.

PubMed

Li, Jing; Zheng, Chun-Quan; Li, Yong; Yang, Chen; Lin, Hai; Duan, Hong-Gang

2015-08-01

This study was designed to investigate the effects of hepatocyte growth factor (HGF) transgenic mesenchymal stem cells (HGF-MSCs) on wound healing in the sinonasal mucosa and nasal epithelial cells (NECs). We also sought to determine whether HGF-MSCs and MSCs can migrate into the injured mucosa and differentiate into ciliated cells. Human HGF-overexpressing umbilical cord MSCs (hHGF-UCMSCs) were established, and upregulation of hHGF expression was confirmed by real-time PCR (RT-PCR) and enzyme-linked immunosorbant assay (ELISA). To investigate the paracrine effect of human MSCs (hMSCs) on nasal epithelial repair, hMSC- and HGF-MSC-conditioned media (CM) were used in NEC proliferation assays and in an in vitro scratch-wound repair model. The in vivo sinonasal wound-healing model was established, and all enrolled rabbits were randomly assigned to four groups: the GFP-MSC group, the HGF-MSC group, the Ad-HGF group, and the surgery control group. The average decreased diameter was recorded, and the medial wall of the maxillary sinus was removed for histological analysis and scanning electron microscopy. Collagen deposition in the wound tissue was detected via Masson trichrome (M&T) staining. The distribution of MSCs and HGF-MSCs was observed by immunofluorescence. MSCs improved nasal wound healing both in vivo and in vitro. HGF overexpression in MSCs augmented the curative effects. Reduced collagen deposition and transforming growth factor beta1 (TGF-β1) expression were detected in the HGF-MSC group compared with the MSC-, Ad-HGF-, and phosphate-buffered saline-treated groups based on M&T staining and ELISA. The enhanced therapeutic effects of HGF-MSCs were accompanied by decreased level of the fibrogenic cytokine TGF-β1. In addition, both HGF-MSCs and MSCs can migrate to the injured mucosa and epithelial layer.

HGF potentiates extracellular matrix-driven migration of human myoblasts: involvement of matrix metalloproteinases and MAPK/ERK pathway.

PubMed

González, Mariela Natacha; de Mello, Wallace; Butler-Browne, Gillian S; Silva-Barbosa, Suse Dayse; Mouly, Vincent; Savino, Wilson; Riederer, Ingo

2017-10-10

The hepatocyte growth factor (HGF) is required for the activation of muscle progenitor cells called satellite cells (SC), plays a role in the migration of proliferating SC (myoblasts), and is present as a soluble factor during muscle regeneration, along with extracellular matrix (ECM) molecules. In this study, we aimed at determining whether HGF is able to interact with ECM proteins, particularly laminin 111 and fibronectin, and to modulate human myoblast migration. We evaluated the expression of the HGF-receptor c-Met, laminin, and fibronectin receptors by immunoblotting, flow cytometry, or immunofluorescence and used Transwell assays to analyze myoblast migration on laminin 111 and fibronectin in the absence or presence of HGF. Zymography was used to check whether HGF could modulate the production of matrix metalloproteinases by human myoblasts, and the activation of MAPK/ERK pathways was evaluated by immunoblotting. We demonstrated that human myoblasts express c-Met, together with laminin and fibronectin receptors. We observed that human laminin 111 and fibronectin have a chemotactic effect on myoblast migration, and this was synergistically increased when low doses of HGF were added. We detected an increase in MMP-2 activity in myoblasts treated with HGF. Conversely, MMP-2 inhibition decreased the HGF-associated stimulation of cell migration triggered by laminin or fibronectin. HGF treatment also induced in human myoblasts activation of MAPK/ERK pathways, whose specific inhibition decreased the HGF-associated stimulus of cell migration triggered by laminin 111 or fibronectin. We demonstrate that HGF induces ERK phosphorylation and MMP production, thus stimulating human myoblast migration on ECM molecules. Conceptually, these data state that the mechanisms involved in the migration of human myoblasts comprise both soluble and insoluble moieties. This should be taken into account to optimize the design of therapeutic cell transplantation strategies by improving

Regulation of HGF and SDF-1 expression by oral fibroblasts--implications for invasion of oral cancer.

PubMed

Daly, Aisling J; McIlreavey, Leanne; Irwin, Chris R

2008-07-01

Invasion and metastasis of oral squamous cell carcinoma (OSCC) is dependent on signals received from stromal fibroblasts present in the surrounding connective tissue. The aim of this study was to investigate the regulation of expression of two important signaling molecules--HGF and SDF-1--by both stromal fibroblasts and their 'activated' form, myofibroblasts, and to determine the role of these two factors in stimulating OSCC cell invasion in vitro. Fibroblasts and myofibroblasts produced similar levels of HGF and SDF-1. IL-1alpha and OSCC cell conditioned medium both stimulated HGF and SDF-1 expression, while TGF-beta(1) inhibited production of each factor. Myofibroblast-derived conditioned medium stimulated OSCC cell invasion through matrigel. Blocking antibodies to both HGF and SDF-1 reduced the level of invasion. In fibroblast-free organotypic raft cultures, addition of HGF and SDF-1 stimulated OSCC cell invasion into the underlying collagen gel, although the pattern of invasion differed from that induced by fibroblasts. Fibroblast-derived HGF and SDF-1 appear to play central roles in the reciprocal interactions between OSCC cells and underlying stromal fibroblasts leading to the local invasion of oral cancer.

Predictive values of FAP and HGF for tumor angiogenesis and metastasis in colorectal cancer.

PubMed

Ma, T H; Gao, C C; Xie, R; Yang, X Z; Dai, W J; Zhang, J L; Yan, W; Wu, S N

2017-01-01

This study aims to explore the correlation of hepatocyte growth factor (HGF) and fibroblast activation protein (FAP) expressions with the angiogenesis and metastasis in colorectal cancer (CRC). The immunohistochemical SABC method was used to detect HGF and FAP expressions in 127 CRC tissues, 51 colorectal polyp tissues and 28 normal tissues. HGF and FAP expressions in liver metastasis were detected using western blot to analyze the correlation of their expressions with lymph node metastasis and liver metastasis. Micro-vessel density (MVD) and clinic-pathologic information of CRC patients were recorded and analyzed. In CRC group, HGF and FAP expressions were greatly higher than those in normal group and colorectal polyps group (P < 0.05). Moreover, the positive rates of HGF and FAP expressions in lymph node metastasis were evidently higher than those in non-lymph node metastasis (P < 0.05). In liver metastasis group, HGF and FAP expressions were obviously higher than non-liver metastasis group (P < 0.05). CRC group had much more MVD in comparison with normal group and colorectal polyps group (P < 0.05).When compared with negative group, MVD was significantly higher than that in CRC tissue with positive HGF and FAP (P < 0.05). Spearman rank correlation analysis showed that HGF and FAP were in positive correlation with MVD (r = 0.542, P < 0.001; r = 0.753, P < 0.001). These results indicate that FAP and HGF play an important role in CRC angiogenesis, and their expression levels are valuable to predict CRC liver metastasis and lymph node metastasis.

Therapeutic potential of fibroblast growth factor-2 for hypertrophic scars: upregulation of MMP-1 and HGF expression.

PubMed

Eto, Hitomi; Suga, Hirotaka; Aoi, Noriyuki; Kato, Harunosuke; Doi, Kentaro; Kuno, Shinichiro; Tabata, Yasuhiko; Yoshimura, Kotaro

2012-02-01

Although hypertrophic scars (HTSs) and keloids are challenging problems, their pathogenesis is not well understood, making therapy difficult. We showed that matrix metalloproteinase (MMP)-1 expression was downregulated in HTS compared with normal skin from the same patients, whereas type 1 and 3 collagen and transforming growth factor-β (TGF-β) were upregulated. These differences, however, were not seen in cultured fibroblasts, suggesting the involvement of microenvironmental factors in the pathogenesis of HTS. Fibroblast growth factor-2 (FGF-2) highly upregulated the expression of MMP-1 and hepatocyte growth factor (HGF) in both HTS-derived and control fibroblasts; the upregulation was reversed by extracellular signal-regulated kinase (ERK) and c-Jun N-terminal kinase (JNK) inhibitors. An animal study using human HTS tissue implanted into nude mice indicated that controlled-release FGF-2 resulted in significantly less weight and decreased hydroxyproline content in HTS. Degradation of collagen fibers in FGF-2-treated HTS was also confirmed histologically. Western blotting showed that FGF-2-treated HTS expressed significantly higher MMP-1 protein than control. Decreased MMP-1 expression may be an important transcriptional change in HTS, and its reversal as well as upregulation of HGF by FGF-2 could be a new therapeutic approach for HTS.

Signal transduction and downregulation of C-MET in HGF stimulated low and highly metastatic human osteosarcoma cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Husmann, Knut, E-mail: khusmann@research.balgrist.ch; Ducommun, Pascal; Division of Plastic Surgery and Hand Surgery, Department of Surgery, University Hospital Zurich, Zurich

2015-09-04

The poor outcome of osteosarcoma (OS), particularly in patients with metastatic disease and a five-year survival rate of only 20%, asks for more effective therapeutic strategies targeting malignancy-promoting mechanisms. Dysregulation of C-MET, its ligand hepatocyte growth factor (HGF) and the fusion oncogene product TPR-MET, first identified in human MNNG-HOS OS cells, have been described as cancer-causing factors in human cancers. Here, the expression of these molecules at the mRNA and the protein level and of HGF-stimulated signaling and downregulation of C-MET was compared in the parental low metastatic HOS and MG63 cell lines and the respective highly metastatic MNNG-HOS andmore » 143B and the MG63-M6 and MG63-M8 sublines. Interestingly, expression of TPR-MET was only observed in MNNG-HOS cells. HGF stimulated the phosphorylation of Akt and Erk1/2 in all cell lines investigated, but phospho-Stat3 remained at basal levels. Downregulation of HGF-stimulated Akt and Erk1/2 phosphorylation was much faster in the HGF expressing MG63-M8 cells than in HOS cells. Degradation of HGF-activated C-MET occurred predominantly through the proteasomal and to a lesser extent the lysosomal pathway in the cell lines investigated. Thus, HGF-stimulated Akt and Erk1/2 signaling as well as proteasomal degradation of HGF activated C-MET are potential therapeutic targets in OS. - Highlights: • Expression of TPR-MET was only observed in MNNG-HOS cells. • HGF stimulated the phosphorylation of Akt and Erk1/2 but not of Stat3 in osteosarcoma cell lines. • Degradation of HGF-activated C-MET occurred predominantly through the proteasomal pathway.« less

Role and Therapeutic Targeting of the HGF/MET Pathway in Glioblastoma

PubMed Central

Cruickshanks, Nichola; Zhang, Ying; Yuan, Fang; Pahuski, Mary; Gibert, Myron; Abounader, Roger

2017-01-01

Glioblastoma (GBM) is a lethal brain tumor with dismal prognosis. Current therapeutic options, consisting of surgery, chemotherapy and radiation, have only served to marginally increase patient survival. Receptor tyrosine kinases (RTKs) are dysregulated in approximately 90% of GBM; attributed to this, research has focused on inhibiting RTKs as a novel and effective therapy for GBM. Overexpression of RTK mesenchymal epithelial transition (MET), and its ligand, hepatocyte growth factor (HGF), in GBM highlights a promising new therapeutic target. This review will discuss the role of MET in cell cycle regulation, cell proliferation, evasion of apoptosis, cell migration and invasion, angiogenesis and therapeutic resistance in GBM. It will also discuss the modes of deregulation of HGF/MET and their regulation by microRNAs. As the HGF/MET pathway is a vital regulator of multiple pro-survival pathways, efforts and strategies for its exploitation for GBM therapy are also described. PMID:28696366

Hepatocyte growth factor (HGF) upregulates heparanase expression via the PI3K/Akt/NF-κB signaling pathway for gastric cancer metastasis.

PubMed

Hao, Ning-Bo; Tang, Bo; Wang, Guo-Zheng; Xie, Rui; Hu, Chang-Jiang; Wang, Su-Min; Wu, Yu-Yun; Liu, En; Xie, Xia; Yang, Shi-Ming

2015-05-28

Heparanase (HPA) is an endoglucuronidase that can promote the shedding of associated cytokines in several types of tumors. However, little is known about what controls the expression of HPA or its role in gastric cancer. In this study, we report for the first time that HGF regulates HPA expression to promote gastric cancer metastasis. In this study, HGF and HPA were found to be significantly expressed in 58 gastric cancer patients. High expression of both HGF and HPA was positively associated with TNM stage, invasion depth and poor prognosis. In MKN74 cells, exogenous HGF significantly increased HPA expression at both the mRNA and protein levels. Further study revealed that HGF first activated PI3K/Akt signaling. NF-κB signaling was activated downstream of PI3K/Akt and promoted HPA expression. However, when c-met, PI3K/Akt or NF-κB signal inhibitors were used, HPA expression was significantly decreased. All of these results indicate that HGF regulates HPA expression by PI3K/Akt and downstream NF-κB signaling. Using bioinformatics and the ChIP assay, p65 was observed to bind to the HPA promoter. Furthermore, HGF significantly induced tumor cell migration, whereas treatment with an NF-κB inhibitor decreased migration. Moreover, when HPA was overexpressed in MKN74 cells, migration was significantly enhanced, and the HGF concentration was increased. However, when HPA was down-regulated in MKN45 cells, migration and HGF levels decreased. Together, these results demonstrate that HGF/c-met can activate PI3K/Akt and downstream NF-κB signaling to promote HPA expression and subsequent tumor metastasis. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

Cooperative interaction of MUC1 with the HGF/c-Met pathway during hepatocarcinogenesis

PubMed Central

2012-01-01

Background Hepatocyte growth factor (HGF) induced c-Met activation is known as the main stimulus for hepatocyte proliferation and is essential for liver development and regeneration. Activation of HGF/c-Met signaling has been correlated with aggressive phenotype and poor prognosis in hepatocellular carcinoma (HCC). MUC1 is a transmembrane mucin, whose over-expression is reported in most cancers. Many of the oncogenic effects of MUC1 are believed to occur through the interaction of MUC1 with signaling molecules. To clarify the role of MUC1 in HGF/c-Met signaling, we determined whether MUC1 and c-Met interact cooperatively and what their role(s) is in hepatocarcinogenesis. Results MUC1 and c-Met over-expression levels were determined in highly motile and invasive, mesenchymal-like HCC cell lines, and in serial sections of cirrhotic and HCC tissues, and these levels were compared to those in normal liver tissues. Co-expression of both c-Met and MUC1 was found to be associated with the differentiation status of HCC. We further demonstrated an interaction between c-Met and MUC1 in HCC cells. HGF-induced c-Met phosphorylation decreased this interaction, and down-regulated MUC1 expression. Inhibition of c-Met activation restored HGF-mediated MUC1 down-regulation, and decreased the migratory and invasive abilities of HCC cells via inhibition of β-catenin activation and c-Myc expression. In contrast, siRNA silencing of MUC1 increased HGF-induced c-Met activation and HGF-induced cell motility and invasion. Conclusions These findings indicate that the crosstalk between MUC1 and c-Met in HCC could provide an advantage for invasion to HCC cells through the β-catenin/c-Myc pathway. Thus, MUC1 and c-Met could serve as potential therapeutic targets in HCC. PMID:22962849

Cooperative interaction of MUC1 with the HGF/c-Met pathway during hepatocarcinogenesis.

PubMed

Bozkaya, Giray; Korhan, Peyda; Cokaklı, Murat; Erdal, Esra; Sağol, Ozgül; Karademir, Sedat; Korch, Christopher; Atabey, Neşe

2012-09-11

Hepatocyte growth factor (HGF) induced c-Met activation is known as the main stimulus for hepatocyte proliferation and is essential for liver development and regeneration. Activation of HGF/c-Met signaling has been correlated with aggressive phenotype and poor prognosis in hepatocellular carcinoma (HCC). MUC1 is a transmembrane mucin, whose over-expression is reported in most cancers. Many of the oncogenic effects of MUC1 are believed to occur through the interaction of MUC1 with signaling molecules. To clarify the role of MUC1 in HGF/c-Met signaling, we determined whether MUC1 and c-Met interact cooperatively and what their role(s) is in hepatocarcinogenesis. MUC1 and c-Met over-expression levels were determined in highly motile and invasive, mesenchymal-like HCC cell lines, and in serial sections of cirrhotic and HCC tissues, and these levels were compared to those in normal liver tissues. Co-expression of both c-Met and MUC1 was found to be associated with the differentiation status of HCC. We further demonstrated an interaction between c-Met and MUC1 in HCC cells. HGF-induced c-Met phosphorylation decreased this interaction, and down-regulated MUC1 expression. Inhibition of c-Met activation restored HGF-mediated MUC1 down-regulation, and decreased the migratory and invasive abilities of HCC cells via inhibition of β-catenin activation and c-Myc expression. In contrast, siRNA silencing of MUC1 increased HGF-induced c-Met activation and HGF-induced cell motility and invasion. These findings indicate that the crosstalk between MUC1 and c-Met in HCC could provide an advantage for invasion to HCC cells through the β-catenin/c-Myc pathway. Thus, MUC1 and c-Met could serve as potential therapeutic targets in HCC.

Hepatocyte growth factor regulates cyclooxygenase-2 expression via β-catenin, Akt, and p42/p44 MAPK in human bronchial epithelial cells

PubMed Central

Lee, Young H.; Suzuki, Yuichiro J.; Griffin, Autumn J.; Day, Regina M.

2008-01-01

Hepatocyte growth factor (HGF) is upregulated in response to lung injury and has been implicated in tissue repair through its antiapoptotic and proliferative activities. Cyclooxygenase-2 (COX-2) is an inducible enzyme in the biosynthetic pathway of prostaglandins, and its activation has been shown to play a role in cell growth. Here, we report that HGF induces gene transcription of COX-2 in human bronchial epithelial cells (HBEpC). Treatment of HBEpC with HGF resulted in phosphorylation of the HGF receptor (c-Met), activation of Akt, and upregulation of COX-2 mRNA. Adenovirus-mediated gene transfer of a dominant negative (DN) Akt mutant revealed that HGF increased COX-2 mRNA in an Akt-dependent manner. COX-2 promoter analysis in luciferase reporter constructs showed that HGF regulation required the β-catenin-responsive T cell factor-4 binding element (TBE). The HGF activation of the COX-2 gene transcription was blocked by DN mutant of β-catenin or by inhibitors that blocked activation of Akt. Inhibition of p42/p44 MAPK pathway blocked HGF-mediated activation of β-catenin gene transcription but not Akt activation, suggesting that p42/p44 MAPK acts in a parallel mechanism for β-catenin activation. We also found that inhibition of COX-2 with NS-398 blocked HGF-induced growth in HBEpC. Together, the results show that the HGF increases COX-2 gene expression via an Akt-, MAPK-, and β-catenin-dependent pathway in HBEpC. PMID:18245266

Met receptor signaling is required for sensory nerve development and HGF promotes axonal growth and survival of sensory neurons

PubMed Central

Maina, Flavio; Hilton, Mark C.; Ponzetto, Carola; Davies, Alun M.; Klein, Rüdiger

1997-01-01

The development of the nervous system is a dynamic process during which factors act in an instructive fashion to direct the differentiation and survival of neurons, and to induce axonal outgrowth, guidance to, and terminal branching within the target tissue. Here we report that mice expressing signaling mutants of the hepatocyte growth factor (HGF) receptor, the Met tyrosine kinase, show a striking reduction of sensory nerves innervating the skin of the limbs and thorax, implicating the HGF/Met system in sensory neuron development. Using in vitro assays, we find that HGF cooperates with nerve growth factor (NGF) to enhance axonal outgrowth from cultured dorsal root ganglion (DRG) neurons. HGF also enhances the neurotrophic activities of NGF in vitro, and Met receptor signaling is required for the survival of a proportion of DRG neurons in vivo. This synergism is specific for NGF but not for the related neurotrophins BDNF and NT3. By using a mild signaling mutant of Met, we have demonstrated previously that Met requires signaling via the adapter molecule Grb2 to induce proliferation of myoblasts. In contrast, the actions of HGF on sensory neurons are mediated by Met effectors distinct from Grb2. Our findings demonstrate a requirement for Met signaling in neurons during development. PMID:9407027

HGF Secreted by Activated Kupffer Cells Induces Apoptosis of Plasmodium-Infected Hepatocytes

PubMed Central

Gonçalves, Lígia Antunes; Rodo, Joana; Rodrigues-Duarte, Lurdes; de Moraes, Luciana Vieira; Penha-Gonçalves, Carlos

2017-01-01

Malaria liver stage infection is an obligatory parasite development step and represents a population bottleneck in Plasmodium infections, providing an advantageous target for blocking parasite cycle progression. Parasite development inside hepatocytes implies a gross cellular insult evoking innate host responses to counteract intra-hepatocytic infection. Using primary hepatocyte cultures, we investigated the role of Kupffer cell-derived hepatocyte growth factor (HGF) in malaria liver stage infection. We found that Kupffer cells from Plasmodium-infected livers produced high levels of HGF, which trigger apoptosis of infected hepatocytes through a mitochondrial-independent apoptosis pathway. HGF action in infected hepatocyte primary cultures results in a potent reduction of parasite yield by specifically sensitizing hepatocytes carrying established parasite exo-erythrocytic forms to undergo apoptosis. This apoptosis mechanism is distinct from cell death that is spontaneously induced in infected cultures and is governed by Fas signaling modulation through a mitochondrial-dependent apoptosis pathway. This work indicates that HGF and Fas signaling pathways are part of an orchestrated host apoptosis response that occurs during malaria liver stage infection, decreasing the success of infection of individual hepatocytes. Our results raise the hypothesis that paracrine signals derived from Kupffer cell activation are implicated in directing death of hepatocytes infected with the malaria parasite. PMID:28220125

HGF Secreted by Activated Kupffer Cells Induces Apoptosis of Plasmodium-Infected Hepatocytes.

PubMed

Gonçalves, Lígia Antunes; Rodo, Joana; Rodrigues-Duarte, Lurdes; de Moraes, Luciana Vieira; Penha-Gonçalves, Carlos

2017-01-01

Malaria liver stage infection is an obligatory parasite development step and represents a population bottleneck in Plasmodium infections, providing an advantageous target for blocking parasite cycle progression. Parasite development inside hepatocytes implies a gross cellular insult evoking innate host responses to counteract intra-hepatocytic infection. Using primary hepatocyte cultures, we investigated the role of Kupffer cell-derived hepatocyte growth factor (HGF) in malaria liver stage infection. We found that Kupffer cells from Plasmodium -infected livers produced high levels of HGF, which trigger apoptosis of infected hepatocytes through a mitochondrial-independent apoptosis pathway. HGF action in infected hepatocyte primary cultures results in a potent reduction of parasite yield by specifically sensitizing hepatocytes carrying established parasite exo-erythrocytic forms to undergo apoptosis. This apoptosis mechanism is distinct from cell death that is spontaneously induced in infected cultures and is governed by Fas signaling modulation through a mitochondrial-dependent apoptosis pathway. This work indicates that HGF and Fas signaling pathways are part of an orchestrated host apoptosis response that occurs during malaria liver stage infection, decreasing the success of infection of individual hepatocytes. Our results raise the hypothesis that paracrine signals derived from Kupffer cell activation are implicated in directing death of hepatocytes infected with the malaria parasite.

Polyphyllin I inhibits gastric cancer cell proliferation by downregulating the expression of fibroblast activation protein alpha (FAP) and hepatocyte growth factor (HGF) in cancer-associated fibroblasts.

PubMed

Dong, Ruizeng; Guo, Jianmin; Zhang, Zewei; Zhou, Yimin; Hua, Yonghong

2018-03-18

The aim of this study was to identify the anti-cancer mechanism of Polyphyllin I (PPI) on gastric cancer cells via its activity on cancer-associated fibroblasts (CAFs). We cultured purified gastric CAFs obtained from fresh human gastric cancer tissue and examined the effect of Polyphyllin I on CAF proliferation using a colorimetric viability assay. In addition, we established a nude mouse xenograft model to examine the effect of Polyphyllin I administration on tumorigenesis. Using Western analysis, we quantified protein expression of the CAF-derived cytokines fibroblast activation protein alpha (FAP), secreted protein acidic and cysteine rich (SPARC), stromal cell-derived factor 1 (SDF-1), hepatocyte growth factor tenascin-C (TNC), and hepatocyte growth factor (HGF) in both in vitro and in vivo models. We found that Polyphyllin I inhibits the proliferation of CAFs in a concentration-dependent manner. Following treatment with 2 μg/ml PPI for 24 h in vitro, the expression of FAP, SDF-1 and HGF protein in CAFs was significantly lower than that in the control group, but there was no significant difference in SPARC and TNC protein expression between the two groups. In the nude mouse xenograft model, the tumor inhibition rate was 45.5% when PPI was administered early and 29.4% with administration in the third week. The expression of FAP and HGF in the xenografts was significantly decreased, while the expression of SPARC, SDF-1, and TNC was largely unaltered. Altogether, these data suggest that Polyphyllin I can inhibit the proliferation of gastric cancer cells by downregulating the expression of FAP and HGF in CAFs in vivo. Copyright © 2018 Elsevier Inc. All rights reserved.

Epigenetic regulation of HGF/Met receptor axis is critical for the outgrowth of bone metastasis from breast carcinoma.

PubMed

Bendinelli, Paola; Maroni, Paola; Matteucci, Emanuela; Desiderio, Maria Alfonsina

2017-02-02

Our translational research deals with the influence of microenvironment on the phenotype and colonization of bone metastases from breast carcinoma, and on pre-metastatic niche formation. The aim of the present study was to clarify the origin of hepatocyte growth factor (HGF), ligand of Met receptor, the control of the axis HGF/Met by DNA methylation, and its importance for the nexus supportive cells-metastatic cells and for metastasis outgrowth. In bone metastasis of the 1833-xenograft model, DNA methyltransferase blockade using the chemotherapic drug 5-aza-2'-deoxycytidine (decitabine) strongly reduced the expression of HGF/Met receptor axis and of E-cadherin, with decrease of metastasis wideness and osteolysis, prolonging mice survival. Thus, DNA methylation events acted as commanders of breast carcinoma cells metastatizing to bone influencing the epithelial phenotype. HGF emerged as a bone-marrow stimulus, and the exosomes seemed to furnish HGF to metastatic cells. In fact, decitabine treatment similarly affected some markers of these microvesicles and HGF, indicating that its supply to recipient cells was prevented. Notably, in bone metastasis the hypomethylation of HGF, Met and E-cadherin promoters did not appear responsible for their elevated expression, but we suggest the involvement of hypermethylated regulators and of Wwox oncosuppressor, the latter being affected by decitabine. Wwox expression increased under decitabine strongly localizing in nuclei of bone metastases. We hypothesize a role of Wwox in Met activity since in vitro Wwox overexpression downregulated the level of nuclear-Met protein fragment and Met stability, also under long exposure of 1833 cells to decitabine. HGF enhanced phosphoMet and the activity in nuclei, an effect partially prevented by decitabine. Altogether, the data indicated the importance to target the tumor microenvironment by blocking epigenetic mechanisms, which control critical events for colonization such as HGF/Met axis

Adenovirus-mediated transfer of hepatocyte growth factor gene to human dental pulp stem cells under good manufacturing practice improves their potential for periodontal regeneration in swine.

PubMed

Cao, Yu; Liu, Zhenhai; Xie, Yilin; Hu, Jingchao; Wang, Hua; Fan, Zhipeng; Zhang, Chunmei; Wang, Jingsong; Wu, Chu-Tse; Wang, Songlin

2015-12-15

Periodontitis is one of the most widespread infectious diseases in humans. We previously promoted significant periodontal tissue regeneration in swine models with the transplantation of autologous periodontal ligament stem cells (PDLSCs) and PDLSC sheet. We also promoted periodontal tissue regeneration in a rat model with a local injection of allogeneic bone marrow mesenchymal stem cells. The purpose of the present study is to investigate the roles of the hepatocyte growth factor (HGF) and human dental pulp stem cells (DPSCs) in periodontal tissue regeneration in swine. In the present study, we transferred an adenovirus that carried HGF gene into human DPSCs (HGF-hDPSCs) under good manufacturing practice (GMP) conditions. These cells were then transplanted into a swine model for periodontal regeneration. Twenty miniature pigs were used to generate periodontitis with bone defect of 5 mm in width, 7 mm in length, and 3 mm in depth. After 12 weeks, clinical, radiological, quantitative and histological assessment of regenerated periodontal tissues was performed to compare periodontal regeneration in swine treated with cell implantation. Our study showed that injecting HGF-hDPSCs into this large animal model could significantly improve periodontal bone regeneration and soft tissue healing. A hDPSC or HGF-hDPSC sheet showed superior periodontal tissue regeneration compared to the injection of dissociated cells. However, the sheets required surgical placement; thus, they were suitable for surgically-managed periodontitis treatments. The adenovirus-mediated transfer of the HGF gene markedly decreased hDPSC apoptosis in a hypoxic environment or in serum-free medium, and it increased blood vessel regeneration. This study indicated that HGF-hDPSCs produced under GMP conditions significantly improved periodontal bone regeneration in swine; thus, this method represents a potential clinical application for periodontal regeneration.

Hepatocyte growth factor limits autoimmune neuroinflammation via glucocorticoid-induced leucine zipper expression in dendritic cells.

PubMed

Benkhoucha, Mahdia; Molnarfi, Nicolas; Dunand-Sauthier, Isabelle; Merkler, Doron; Schneiter, Gregory; Bruscoli, Stefano; Riccardi, Carlo; Tabata, Yasuhiko; Funakoshi, Hiroshi; Nakamura, Toshikazu; Reith, Walter; Santiago-Raber, Marie-Laure; Lalive, Patrice H

2014-09-15

Autoimmune neuroinflammation, including multiple sclerosis and its animal model, experimental autoimmune encephalomyelitis (EAE), a prototype for T cell-mediated autoimmunity, is believed to result from immune tolerance dysfunction leading to demyelination and substantial neurodegeneration. We previously showed that CNS-restricted expression of hepatocyte growth factor (HGF), a potent neuroprotective factor, reduced CNS inflammation and clinical deficits associated with EAE. In this study, we demonstrate that systemic HGF treatment ameliorates EAE through the development of tolerogenic dendritic cells (DCs) with high expression levels of glucocorticoid-induced leucine zipper (GILZ), a transcriptional repressor of gene expression and a key endogenous regulator of the inflammatory response. RNA interference-directed neutralization of GILZ expression by DCs suppressed the induction of tolerance caused by HGF. Finally, adoptive transfer of HGF-treated DCs from wild-type but not GILZ gene-deficient mice potently mediated functional recovery in recipient mice with established EAE through effective modulation of autoaggressive T cell responses. Altogether, these results show that by inducing GILZ in DCs, HGF reproduces the mechanism of immune regulation induced by potent immunomodulatory factors such as IL-10, TGF-β1, and glucocorticoids and therefore that HGF therapy may have potential in the treatment of autoimmune dysfunctions. Copyright © 2014 by The American Association of Immunologists, Inc.

Involvement of activator protein 1 complexes in the epithelium-specific activation of the laminin gamma2-chain gene promoter by hepatocyte growth factor (scatter factor).

PubMed Central

Olsen, J; Lefebvre, O; Fritsch, C; Troelsen, J T; Orian-Rousseau, V; Kedinger, M; Simon-Assmann, P

2000-01-01

Laminin-5 is a trimer of laminin alpha3, beta3 and gamma2 chains that is found in the intestinal basement membrane. Deposition of the laminin gamma2 chain at the basement membrane is of great interest because it undergoes a developmental shift in its cellular expression. Here we study the regulatory elements that control basal and cytokine-activated transcriptional expression of the LAMC2 gene, which encodes the laminin gamma2 chain. By using transient transfection experiments we demonstrated the presence of constitutive and cytokine-responsive cis-elements. Comparison of the transcriptional activity of the LAMC2 promoter in the epithelial HT29mtx cells with that in small-intestinal fibroblastic cells (C20 cells) led us to conclude that two regions with constitutive epithelium-specific activity are present between positions -1.2 and -0.12 kb. This was further validated by transfections of primary foetal intestinal endoderm and mesenchyme. A 2.5 kb portion of the LAMC2 5' flanking region was equally responsive to PMA and hepatocyte growth factor (HGF), whereas it was less responsive to transforming growth factor beta1. A minimal promoter limited to the initial 120 bp upstream of the transcriptional start site maintained inducibility by PMA and HGF. This short promoter fragment contains two activator protein 1 (AP-1) elements and the 5'-most of these is a composite AP-1/Sp1 element. The 5'AP-1 element is crucial to the HGF-mediated activity of the promoter; analysis of interacting nuclear proteins demonstrated that AP-1 proteins containing JunD mediate the response to HGF. PMID:10749670

Trans-Ethnic Meta-Analysis Identifies Common and Rare Variants Associated with Hepatocyte Growth Factor Levels in the Multi-Ethnic Study of Atherosclerosis (MESA)

PubMed Central

Larson, Nicholas B.; Berardi, Cecilia; Decker, Paul A.; Wassel, Christina L.; Kirsch, Phillip S.; Pankow, James S.; Sale, Michele M.; de Andrade, Mariza; Sicotte, Hugues; Tang, Weihong; Hanson, Naomi Q.; Tsai, Michael Y.; Taylor, Kent D.; Bielinski, Suzette J.

2015-01-01

Summary Hepatocyte growth factor (HGF) is a mesenchyme-derived pleiotropic factor that regulates cell growth, motility, mitogenesis, and morphogenesis in a variety of cells, and increased serum levels of HGF have been linked to a number of clinical and subclinical cardiovascular disease phenotypes. However, little is currently known regarding what genetic factors influence HGF levels, despite evidence of substantial genetic contributions to HGF variation. Based upon ethnicity-stratified single-variant association analysis and trans-ethnic meta-analysis of 6201 participants of the Multi-Ethnic Study of Atherosclerosis (MESA), we discovered five statistically significant common and low-frequency variants: HGF missense polymorphism rs5745687 (p.E299K) as well as four variants (rs16844364, rs4690098, rs114303452, rs3748034) within or in proximity to HGFAC. We also identified two significant ethnicity-specific gene-level associations (A1BG in African Americans; FASN in Chinese Americans) based upon low-frequency/rare variants, while meta-analysis of gene-level results identified a significant association for HGFAC. However, identified single-variant associations explained modest proportions of the total trait variation and were not significantly associated with coronary artery calcium or coronary heart disease. Our findings indicate genetic factors influencing circulating HGF levels may be complex and ethnically diverse. PMID:25998175

HGF/c-Met related activation of β-catenin in hepatoblastoma

PubMed Central

2011-01-01

Background Activation of beta-catenin is a hallmark of hepatoblastoma (HB) and appears to play a crucial role in its pathogenesis. While aberrant accumulation of the beta-catenin is a common event in HB, mutations or deletions in CTNNB1 (beta-catenin gene) do not always account for the high frequency of protein expression. In this study we have investigated alternative activation of beta-catenin by HGF/c-Met signaling in a large cohort of 98 HB patients enrolled in the SIOPEL-3 clinical trial. Methods We performed immunohistochemistry, using antibodies to total beta-catenin and tyrosine654-phosphorylated beta-catenin, which is a good surrogate marker of HGF/c-Met activation. CTNNB1 mutation analysis was also carried out on all samples. We also investigated beta-catenin pathway activation in two liver cancer cell lines, HuH-6 and HuH-7. Results Aberrant beta-catenin expression was seen in the cytoplasm and/or nucleus of 87% of tumour samples. Our results also revealed a large subset of HB, 83%, with cytoplasmic expression of tyrosine654-phosphorylated beta-catenin and 30% showing additional nuclear accumulation. Sequence analysis revealed mutations in 15% of our cohort. Statistical analysis showed an association between nuclear expression of c-Met-activated beta-catenin and wild type CTNNB1 (P-value = 0.015). Analysis of total beta-catenin and Y654-beta-catenin in response to HGF activation in the cell lines, mirrors that observed in our HB tumour cohort. Results We identified a significant subset of hepatoblastoma patients for whom targeting of the c-Met pathway may be a treatment option and also demonstrate distinct mechanisms of beta-catenin activation in HB. PMID:21992464

The TAM-family receptor Mer mediates production of HGF through the RhoA-dependent pathway in response to apoptotic cells.

PubMed

Park, Hyun-Jung; Baen, Ji-Yeon; Lee, Ye-Ji; Choi, Youn-Hee; Kang, Jihee Lee

2012-08-01

The TAM receptor protein tyrosine kinases Tyro3, Axl, and Mer play important roles in macrophage function. We investigated the roles of the TAM receptors in mediating the induction of hepatocyte growth factor (HGF) during the interaction of macrophages with apoptotic cells. Mer-specific neutralizing antibody, small interfering RNA (siRNA), and a recombinant Mer protein (Mer/Fc) inhibited HGF mRNA and protein expression, as well as activation of RhoA, Akt, and specific mitogen-activated protein (MAP) kinases in response to apoptotic cells. Inhibition of Axl or Tyro3 with specific antibodies, siRNA, or Fc-fusion proteins did not prevent apoptotic cell-induced HGF mRNA and protein expression and did not inhibit activation of the postreceptor signaling molecules RhoA and certain MAP kinases, including extracellular signal-regulated protein kinase and c-Jun NH(2)-terminal kinase. However, Axl- and Tyro3-specific blockers did inhibit the activation of Akt and p38 MAP kinase in response to apoptotic cells. In addition, none of the TAM receptors mediated the effects of apoptotic cells on transforming growth factor-β or epidermal growth factor mRNA expression. However, they were involved in the induction of vascular endothelial growth factor mRNA expression. Our data provide evidence that when macrophages interact with apoptotic cells, only Mer of the TAM-family receptors is responsible for mediating transcriptional HGF production through a RhoA-dependent pathway.

Association of extracellular cleavage of E-cadherin mediated by MMP-7 with HGF-induced in vitro invasion in human stomach cancer cells.

PubMed

Lee, K H; Choi, E Y; Hyun, M S; Jang, B I; Kim, T N; Kim, S W; Song, S K; Kim, J H; Kim, J-R

2007-01-01

Proteolytic shedding of the ectodomain of a variety of transmembrane proteins, including cell-to-cell adhesion molecules, has been observed in solid cancers. We have investigated whether extracellular cleavage of E-cadherin mediated by matrix metalloproteinase-7 (MMP-7) is involved in hepatocyte growth factor (HGF) induced in vitro invasion in stomach cancer cells. The effects of HGF on the expression of E-cadherin/beta-catenin and MMP-7 at both the protein and mRNA levels were assessed in stomach cancer cells, NUGC-3 and MKN-28, and in cells in which the expression of MMP-7 was downregulated by transfection with a MMP-7 short hairpin RNA plasmid. Treatment with HGF increased the extracellular cleavage of E-cadherin and the release of MMP-7 and reduced the level of E-cadherin in a dose- and time-dependent manner. HGF treatment repressed the phosphorylation of beta-catenin in a Triton-soluble fraction, but enhanced this phosphorylation in a Triton-insoluble fraction. The association of E-cadherin with beta-catenin was decreased by HGF treatment in the Triton-soluble fraction. In addition, treatment of MMP-7 short hairpin RNA transfected NUGC-3 cells with HGF resulted in no extracellular cleavage of E-cadherin and also decreased the in vitro cell invasion. These results suggest that incubation with HGF mediated the release of MMP-7, resulting in extracellular cleavage of E-cadherin from stomach cancer cells. This might be a key mechanism in HGF-induced in vitro invasion and metastasis. Copyright 2007 S. Karger AG, Basel.

Supplementation with CTGF, SDF1, NGF, and HGF promotes ovine in vitro oocyte maturation and early embryo development.

PubMed

Wang, D H; Ren, J; Zhou, C J; Han, Z; Wang, L; Liang, C G

2018-05-17

The strategies for improving the in vitro maturation (IVM) of domestic animal oocytes focus on promoting nuclear and cytoplasmic maturation. The identification of paracrine factors and their supplementation in the culture medium represent effective approaches for oocyte maturation and embryo development. This study investigated the effects of paracrine factor supplementation including connective tissue growth factor (CTGF), nerve growth factor (NGF), hepatocyte growth factor (HGF), and stromal derived factor 1 (SDF1) on ovine oocytes and early parthenogenetic embryos using an in vitro culture system. First, we identified the optimal concentrations of CTGF (30 ng/mL), SDF1 (10 ng/mL), NGF (3 ng/mL), and HGF (100 ng/mL) for promoting oocyte maturation, which combined, induced nuclear maturation in 94.19% of oocytes. This combination also promoted cumulus cell expansion and inhibited oocyte/cumulus apoptosis, while enabling a larger proportion (33.04%) of embryos to develop into blastocysts than in the controls and prevented embryo apoptosis. These novel findings demonstrate that the paracrine factors CTGF, SDF1, NGF, and HGF facilitate ovine oocyte and early parthenogenetic embryo development in vitro. Thus, supplementation with these factors may help optimize the IVM of ovine oocytes and early parthenogenetic embryo development strategies. Copyright © 2018 Elsevier Inc. All rights reserved.

Clinical and prognostic value of the C-Met/HGF signaling pathway in cervical cancer.

PubMed

Boromand, Nadia; Hasanzadeh, Malihe; ShahidSales, Soodabeh; Farazestanian, Marjaneh; Gharib, Masoumeh; Fiuji, Hamid; Behboodi, Negin; Ghobadi, Niloofar; Hassanian, Seyed Mahdi; Ferns, Gordon A; Avan, Amir

2018-06-01

Aberrant activation of the HGF/c-Met signalling pathway is reported to be associated with cell proliferation, progression, and metastasis features of several tumor types, including cervical cancer, suggesting that it may be of potential value as a novel therapeutic target. Furthermore, HPV-positive patients had a higher serum level of HGF or c-Met protein, compared with HPV-negative patients. c-Met or HGF overexpression in lesions of cervical cancer is reported to be related to a poorer prognosis, and hence this may be of value as a prognostic and predictive biomarker. Several approaches have been developed for targeting HGF and/or c-Met. One of these is crizotinib (a dual c-Met/ALK inhibitor). This has been approved by FDA for the treatment of lung-cancer. Further investigations are required to evaluate and optimize the use of c-Met inhibitors in cervical cancer or parallel targeting signalling pathway associated/activated via MET/HGF pathway. The main aim of current review was to give an overview of the potential of the c-Met/HGF pathway as a prognostic, or predictive biomarker in cervical cancer. © 2017 Wiley Periodicals, Inc.

Population expansion, clonal growth, and specific differentiation patterns in primary cultures of hepatocytes induced by HGF/SF, EGF and TGF alpha in a chemically defined (HGM) medium

PubMed Central

1996-01-01

Mature adult parenchymal hepatocytes, typically of restricted capacity to proliferate in culture, can now enter into clonal growth under the influence of hepatocyte growth factor (scatter factor) (HGF/SF), epidermal growth factor (EGF), and transforming growth factor alpha (TGFalpha) in the presence of a new chemically defined medium (HGM). The expanding populations of hepatocytes lose expression of hepatocyte specific genes (albumin, cytochrome P450 IIB1), acquire expression of markers expressed by bile duct epithelium (cytokeratin 19), produce TGFalpha and acidic FGF and assume a very simplified morphologic phenotype by electron microscopy. A major change associated with this transition is the decrease in ratio between transcription factors C/EBPalpha and C/EBPbeta, as well as the emergence in the proliferating hepatocytes of transcription factors AP1, NFkappaB. The liver associated transcription factors HNFI, HNF3, and HNF4 are preserved throughout this process. After population expansion and clonal growth, the proliferating hepatocytes can return to mature hepatocyte phenotype in the presence of EHS gel (Matrigel). This includes complete restoration of electron microscopic structure and albumin expression. The hepatocyte cultures however can instead be induced to form acinar/ductular structures akin to bile ductules (in the presence of HGF/SF and type I collagen). These transformations affect the entire population of the hepatocytes and occur even when DNA synthesis is inhibited. Similar acinar/ductular structures are seen in embryonic liver when HGF/SF and its receptor are expressed at high levels. These findings strongly support the hypothesis that mature hepatocytes can function as or be a source of bipotential facultative hepatic stem cells (hepatoblasts). These studies also provide evidence for the growth factor and matrix signals that govern these complex phenotypic transitions of facultative stem cells which are crucial for recovery from acute and

Hepatocyte growth factor enhances the inflammation-alleviating effect of umbilical cord-derived mesenchymal stromal cells in a bronchiolitis obliterans model.

PubMed

Cao, Xiao-Pei; Han, Dong-Mei; Zhao, Li; Guo, Zi-Kuan; Xiao, Feng-Jun; Zhang, Yi-Kun; Zhang, Xiao-Yan; Wang, Li-Sheng; Wang, Heng-Xiang; Wang, Hua

2016-03-01

Specific and effective therapy for prevention or reversal of bronchiolitis obliterans (BO) is lacking. In this study, we evaluated the therapeutic effect of hepatocyte growth factor (HGF) gene modified mesenchymal stromal cells (MSCs) on BO. A mouse model of experimental BO was established by subcutaneously transplanting the tracheas from C57BL/6 mice into Balb/C recipients, which were then administered saline, Ad-HGF-modified human umbilical cord-MSCs (MSCs-HGF) or Ad-Null-modified MSCs (MSCs-Null). The therapeutic effects of MSCs-Null and MSCs-HGF were evaluated by using fluorescence-activated cell sorting (FACS) for lymphocyte immunophenotype of spleen, enzyme-linked immunosorbent assay (ELISA) and real-time polymerase chain reaction (rt-PCR) for cytokine expression, and histopathological analysis for the transplanted trachea. The histopathologic recovery of allograft tracheas was improved significantly after MSCs-Null and MSCs-HGF treatment and the beneficial effects were particularly observed in MSCs-HGF-treated mice. Furthermore, the allo-transplantation-induced immunophenotype disorders of the spleen, including regulatory T (Treg), T helper (Th)1, Th2 and Th17, were attenuated in both cell-treated groups. MSCs-HGF treatment reduced expression and secretion of inflammation cytokines interferon-gamma (IFN-γ), and increased expression of anti-inflammatory cytokine interleukin (IL)-4 and IL-10. It also decreased the expression level of the profibrosis factor transforming growth factor (TGF)-β. Treatment of BO with HGF gene modified MSCs results in reduction of local inflammation and promotion in recovery of allograft trachea histopathology. These findings might provide an effective therapeutic strategy for BO. Copyright © 2015 International Society for Cellular Therapy. Published by Elsevier Inc. All rights reserved.

Umbilical Cord-derived Mesenchymal Stem Cells Instruct Monocytes Towards an IL10-producing Phenotype by Secreting IL6 and HGF.

PubMed

Deng, Yinan; Zhang, Yingcai; Ye, Linsen; Zhang, Tong; Cheng, Jintao; Chen, Guihua; Zhang, Qi; Yang, Yang

2016-12-05

Human UC-MSCs are regarded as an attractive alternative to BM-MSCs for clinical applications due to their easy preparation, higher proliferation and lower immunogenicity. However, the mechanisms underlying immune suppression by UC-MSCs are still unclear. We studied the mechanism of inhibition by UC-MSCs during the differentiation of monocytes into DCs and focused on the specific source and the role of the involved cytokines. We found that UC-MSCs suppressed monocyte differentiation into DCs and instructed monocytes towards other cell types, with clear decreases in the expression of co-stimulatory molecules, in the secretion of inflammatory factors and in allostimulatory capacity. IL6, HGF and IL10 might be involved in this process because they were detected at higher levels in a coculture system. UC-MSCs produce IL-6 and HGF, and neutralization of IL-6 and HGF reversed the suppressive effect of UC-MSCs. IL10 was not produced by UC-MSCs but was exclusively produced by monocytes after exposure to UC-MSCs, IL-6 or HGF. In summary, we found that the UC-MSC-mediated inhibitory effect was dependent on IL6 and HGF secreted by UC-MSCs and that this effect induced monocyte-derived cells to produce IL10, which might indirectly strengthen the suppressive effect of UC-MSCs.

HGF/MET-directed therapeutics in gastroesophageal cancer: a review of clinical and biomarker development.

PubMed

Hack, Stephen P; Bruey, Jean-Marie; Koeppen, Hartmut

2014-05-30

Aberrant activation of the HGF/MET signaling axis has been strongly implicated in the malignant transformation and progression of gastroesophageal cancer (GEC). MET receptor overexpression in tumor samples from GEC patients has been consistently correlated with an aggressive metastatic phenotype and poor prognosis. In preclinical GEC models, abrogation of HGF/MET signaling has been shown to induce tumor regression as well as inhibition of metastatic dissemination. Promising clinical results in patient subsets in which MET is overexpressed have spurned several randomized studies of HGF/MET-directed agents, including two pivotal global Phase III trials. Available data highlight the need for predictive biomarkers in order to select patients most likely to benefit from HGF/MET inhibition. In this review, we discuss the current knowledge of mechanisms of MET activation in GEC, the current status of the clinical evaluation of MET-targeted therapies in GEC, characteristics of ongoing randomized GEC trials and the associated efforts to identify and validate biomarkers. We also discuss the considerations and challenges for HGF/MET inhibitor drug development in the GEC setting.

HGF/MET-directed therapeutics in gastroesophageal cancer: a review of clinical and biomarker development

PubMed Central

Hack, Stephen P.; Bruey, Jean-Marie; Koeppen, Hartmut

2014-01-01

Aberrant activation of the HGF/MET signaling axis has been strongly implicated in the malignant transformation and progression of gastroesophageal cancer (GEC). MET receptor overexpression in tumor samples from GEC patients has been consistently correlated with an aggressive metastatic phenotype and poor prognosis. In preclinical GEC models, abrogation of HGF/MET signaling has been shown to induce tumor regression as well as inhibition of metastatic dissemination. Promising clinical results in patient subsets in which MET is overexpressed have spurned several randomized studies of HGF/MET-directed agents, including two pivotal global Phase III trials. Available data highlight the need for predictive biomarkers in order to select patients most likely to benefit from HGF/MET inhibition. In this review, we discuss the current knowledge of mechanisms of MET activation in GEC, the current status of the clinical evaluation of MET-targeted therapies in GEC, characteristics of ongoing randomized GEC trials and the associated efforts to identify and validate biomarkers. We also discuss the considerations and challenges for HGF/MET inhibitor drug development in the GEC setting. PMID:24930887

Nonclinical evaluation of the serum pharmacodynamic biomarkers HGF and shed MET following dosing with the anti-MET monovalent monoclonal antibody onartuzumab.

PubMed

Mai, Elaine; Zheng, Zhong; Chen, Youjun; Peng, Jing; Severin, Christophe; Filvaroff, Ellen; Romero, Mally; Mallet, William; Kaur, Surinder; Gelzleichter, Thomas; Nijem, Ihsan; Merchant, Mark; Young, Judy C

2014-02-01

Onartuzumab, a humanized, monovalent monoclonal anti-MET antibody, antagonizes MET signaling by inhibiting binding of its ligand, hepatocyte growth factor (HGF). We investigated the effects of onartuzumab on cell-associated and circulating (shed) MET (sMET) and circulating HGF in vitro and nonclinically to determine their utility as pharmacodynamic biomarkers for onartuzumab. Effects of onartuzumab on cell-associated MET were assessed by flow cytometry and immunofluorescence. sMET and HGF were measured in cell supernatants and in serum or plasma from multiple species (mouse, cynomolgus monkey, and human) using plate-based immunoassays. Unlike bivalent anti-MET antibodies, onartuzumab stably associates with MET on the surface of cells without inducing MET internalization or shedding. Onartuzumab delayed the clearance of human xenograft tumor-produced sMET from the circulation of mice, and endogenous sMET in cynomolgus monkeys. In mice harboring MET-expressing xenograft tumors, in the absence of onartuzumab, levels of human sMET correlated with tumor size, and may be predictive of MET-expressing tumor burden. Because binding of sMET to onartuzumab in circulation resulted in increasing sMET serum concentrations due to reduced clearance, this likely renders sMET unsuitable as a pharmacodynamic biomarker for onartuzumab. There was no observed effect of onartuzumab on circulating HGF levels in xenograft tumor-bearing mice or endogenous HGF in cynomolgus monkeys. Although sMET and HGF may serve as predictive biomarkers for MET therapeutics, these data do not support their use as pharmacodynamic biomarkers for onartuzumab.

Apigenin inhibits HGF-promoted invasive growth and metastasis involving blocking PI3K/Akt pathway and {beta}4 integrin function in MDA-MB-231 breast cancer cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lee, W.-J.; Chen, W.-K.; Wang, C.-J.

2008-01-15

Hepatocyte growth factor (HGF) and its receptor, Met, known to control invasive growth program have recently been shown to play crucial roles in the survival of breast cancer patients. The diet-derived flavonoids have been reported to possess anti-invasion properties; however, knowledge on the pharmacological and molecular mechanisms in suppressing HGF/Met-mediated tumor invasion and metastasis is poorly understood. In our preliminary study, we use HGF as an invasive inducer to investigate the effect of flavonoids including apigenin, naringenin, genistein and kaempferol on HGF-dependent invasive growth of MDA-MB-231 human breast cancer cells. Results show that apigenin presents the most potent anti-migration andmore » anti-invasion properties by Boyden chamber assay. Furthermore, apigenin represses the HGF-induced cell motility and scattering and inhibits the HGF-promoted cell migration and invasion in a dose-dependent manner. The effect of apigenin on HGF-induced signaling activation involving invasive growth was evaluated by immunoblotting analysis, it shows that apigenin blocks the HGF-induced Akt phosphorylation but not Met, ERK, and JNK phosphorylation. In addition to MDA-MB-231 cells, apigenin exhibits inhibitory effect on HGF-induced Akt phosphorylation in hepatoma SK-Hep1 cells and lung carcinoma A549 cells. By indirect immunofluorescence microscopy assay, apigenin inhibits the HGF-induced clustering of {beta}4 integrin at actin-rich adhesive site and lamellipodia through PI3K-dependent manner. Treatment of apigenin inhibited HGF-stimulated integrin {beta}4 function including cell-matrix adhesion and cell-endothelial cells adhesion in MDA-MB-231 cells. By Akt-siRNA transfection analysis, it confirmed that apigenin inhibited HGF-promoted invasive growth involving blocking PI3K/Akt pathway. Finally, we evaluated the effect of apigenin on HGF-promoted metastasis by lung colonization of tumor cells in nude mice and organ metastasis of tumor cells in chick

Mesenchymal stem cells and myoblast differentiation under HGF and IGF-1 stimulation for 3D skeletal muscle tissue engineering.

PubMed

Witt, R; Weigand, A; Boos, A M; Cai, A; Dippold, D; Boccaccini, A R; Schubert, D W; Hardt, M; Lange, C; Arkudas, A; Horch, R E; Beier, J P

2017-02-28

Volumetric muscle loss caused by trauma or after tumour surgery exceeds the natural regeneration capacity of skeletal muscle. Hence, the future goal of tissue engineering (TE) is the replacement and repair of lost muscle tissue by newly generating skeletal muscle combining different cell sources, such as myoblasts and mesenchymal stem cells (MSCs), within a three-dimensional matrix. Latest research showed that seeding skeletal muscle cells on aligned constructs enhance the formation of myotubes as well as cell alignment and may provide a further step towards the clinical application of engineered skeletal muscle. In this study the myogenic differentiation potential of MSCs upon co-cultivation with myoblasts and under stimulation with hepatocyte growth factor (HGF) and insulin-like growth factor-1 (IGF-1) was evaluated. We further analysed the behaviour of MSC-myoblast co-cultures in different 3D matrices. Primary rat myoblasts and rat MSCs were mono- and co-cultivated for 2, 7 or 14 days. The effect of different concentrations of HGF and IGF-1 alone, as well as in combination, on myogenic differentiation was analysed using microscopy, multicolour flow cytometry and real-time PCR. Furthermore, the influence of different three-dimensional culture models, such as fibrin, fibrin-collagen-I gels and parallel aligned electrospun poly-ε-caprolacton collagen-I nanofibers, on myogenic differentiation was analysed. MSCs could be successfully differentiated into the myogenic lineage both in mono- and in co-cultures independent of HGF and IGF-1 stimulation by expressing desmin, myocyte enhancer factor 2, myosin heavy chain 2 and alpha-sarcomeric actinin. An increased expression of different myogenic key markers could be observed under HGF and IGF-1 stimulation. Even though, stimulation with HGF/IGF-1 does not seem essential for sufficient myogenic differentiation. Three-dimensional cultivation in fibrin-collagen-I gels induced higher levels of myogenic differentiation

Umbilical Cord-derived Mesenchymal Stem Cells Instruct Monocytes Towards an IL10-producing Phenotype by Secreting IL6 and HGF

PubMed Central

Deng, Yinan; Zhang, Yingcai; Ye, Linsen; Zhang, Tong; Cheng, Jintao; Chen, Guihua; Zhang, Qi; Yang, Yang

2016-01-01

Human UC-MSCs are regarded as an attractive alternative to BM-MSCs for clinical applications due to their easy preparation, higher proliferation and lower immunogenicity. However, the mechanisms underlying immune suppression by UC-MSCs are still unclear. We studied the mechanism of inhibition by UC-MSCs during the differentiation of monocytes into DCs and focused on the specific source and the role of the involved cytokines. We found that UC-MSCs suppressed monocyte differentiation into DCs and instructed monocytes towards other cell types, with clear decreases in the expression of co-stimulatory molecules, in the secretion of inflammatory factors and in allostimulatory capacity. IL6, HGF and IL10 might be involved in this process because they were detected at higher levels in a coculture system. UC-MSCs produce IL-6 and HGF, and neutralization of IL-6 and HGF reversed the suppressive effect of UC-MSCs. IL10 was not produced by UC-MSCs but was exclusively produced by monocytes after exposure to UC-MSCs, IL-6 or HGF. In summary, we found that the UC-MSC-mediated inhibitory effect was dependent on IL6 and HGF secreted by UC-MSCs and that this effect induced monocyte-derived cells to produce IL10, which might indirectly strengthen the suppressive effect of UC-MSCs. PMID:27917866

Human hepatocyte growth factor promotes functional recovery in primates after spinal cord injury.

PubMed

Kitamura, Kazuya; Fujiyoshi, Kanehiro; Yamane, Jun-Ichi; Toyota, Fumika; Hikishima, Keigo; Nomura, Tatsuji; Funakoshi, Hiroshi; Nakamura, Toshikazu; Aoki, Masashi; Toyama, Yoshiaki; Okano, Hideyuki; Nakamura, Masaya

2011-01-01

Many therapeutic interventions for spinal cord injury (SCI) using neurotrophic factors have focused on reducing the area damaged by secondary, post-injury degeneration, to promote functional recovery. Hepatocyte growth factor (HGF), which is a potent mitogen for mature hepatocytes and a mediator of the inflammatory responses to tissue injury, was recently highlighted as a potent neurotrophic factor in the central nervous system. We previously reported that introducing exogenous HGF into the injured rodent spinal cord using a herpes simplex virus-1 vector significantly reduces the area of damaged tissue and promotes functional recovery. However, that study did not examine the therapeutic effects of administering HGF after injury, which is the most critical issue for clinical application. To translate this strategy to human treatment, we induced a contusive cervical SCI in the common marmoset, a primate, and then administered recombinant human HGF (rhHGF) intrathecally. Motor function was assessed using an original open field scoring system focusing on manual function, including reach-and-grasp performance and hand placement in walking. The intrathecal rhHGF preserved the corticospinal fibers and myelinated areas, thereby promoting functional recovery. In vivo magnetic resonance imaging showed significant preservation of the intact spinal cord parenchyma. rhHGF-treatment did not give rise to an abnormal outgrowth of calcitonin gene related peptide positive fibers compared to the control group, indicating that this treatment did not induce or exacerbate allodynia. This is the first study to report the efficacy of rhHGF for treating SCI in non-human primates. In addition, this is the first presentation of a novel scale for assessing neurological motor performance in non-human primates after contusive cervical SCI.

Human Hepatocyte Growth Factor Promotes Functional Recovery in Primates after Spinal Cord Injury

PubMed Central

Kitamura, Kazuya; Fujiyoshi, Kanehiro; Yamane, Jun-ichi; Toyota, Fumika; Hikishima, Keigo; Nomura, Tatsuji; Funakoshi, Hiroshi; Nakamura, Toshikazu; Aoki, Masashi; Toyama, Yoshiaki; Okano, Hideyuki; Nakamura, Masaya

2011-01-01

Many therapeutic interventions for spinal cord injury (SCI) using neurotrophic factors have focused on reducing the area damaged by secondary, post-injury degeneration, to promote functional recovery. Hepatocyte growth factor (HGF), which is a potent mitogen for mature hepatocytes and a mediator of the inflammatory responses to tissue injury, was recently highlighted as a potent neurotrophic factor in the central nervous system. We previously reported that introducing exogenous HGF into the injured rodent spinal cord using a herpes simplex virus-1 vector significantly reduces the area of damaged tissue and promotes functional recovery. However, that study did not examine the therapeutic effects of administering HGF after injury, which is the most critical issue for clinical application. To translate this strategy to human treatment, we induced a contusive cervical SCI in the common marmoset, a primate, and then administered recombinant human HGF (rhHGF) intrathecally. Motor function was assessed using an original open field scoring system focusing on manual function, including reach-and-grasp performance and hand placement in walking. The intrathecal rhHGF preserved the corticospinal fibers and myelinated areas, thereby promoting functional recovery. In vivo magnetic resonance imaging showed significant preservation of the intact spinal cord parenchyma. rhHGF-treatment did not give rise to an abnormal outgrowth of calcitonin gene related peptide positive fibers compared to the control group, indicating that this treatment did not induce or exacerbate allodynia. This is the first study to report the efficacy of rhHGF for treating SCI in non-human primates. In addition, this is the first presentation of a novel scale for assessing neurological motor performance in non-human primates after contusive cervical SCI. PMID:22140459

Emergence of HGF/SF-Induced Coordinated Cellular Motility

PubMed Central

Zaritsky, Assaf; Natan, Sari; Ben-Jacob, Eshel; Tsarfaty, Ilan

2012-01-01

Collective cell migration plays a major role in embryonic morphogenesis, tissue remodeling, wound repair and cancer invasion. Despite many decades of extensive investigations, only few analytical tools have been developed to enhance the biological understanding of this important phenomenon. Here we present a novel quantitative approach to analyze long term kinetics of bright field time-lapse wound healing. Fully-automated spatiotemporal measures and visualization of cells' motility and implicit morphology were proven to be sound, repetitive and highly informative compared to single-cell tracking analysis. We study cellular collective migration induced by tyrosine kinase-growth factor signaling (Met-Hepatocyte Growth Factor/Scatter Factor (HGF/SF)). Our quantitative approach is applied to demonstrate that collective migration of the adenocarcinoma cell lines is characterized by simple morpho-kinetics. HGF/SF induces complex morpho-kinetic coordinated collective migration: cells at the front move faster and are more spread than those further away from the wound edge. As the wound heals, distant cells gradually accelerate and enhance spread and elongation –resembling the epithelial to mesenchymal transition (EMT), and then the cells become more spread and maintain higher velocity than cells located closer to the wound. Finally, upon wound closure, front cells halt, shrink and round up (resembling mesenchymal to epithelial transition (MET) phenotype) while distant cells undergo the same process gradually. Met inhibition experiments further validate that Met signaling dramatically alters the morpho-kinetic dynamics of the healing wound. Machine-learning classification was applied to demonstrate the generalization of our findings, revealing even subtle changes in motility patterns induced by Met-inhibition. It is concluded that activation of Met-signaling induces an elaborated model in which cells lead a coordinated increased motility along with gradual differentiation

Transplantation of Hepatocyte Growth Factor-Modified Dental Pulp Stem Cells Prevents Bone Loss in the Early Phase of Ovariectomy-Induced Osteoporosis.

PubMed

Kong, Fanxuan; Shi, Xuefeng; Xiao, Fengjun; Yang, Yuefeng; Zhang, Xiaoyan; Wang, Li-Sheng; Wu, Chu-Tse; Wang, Hua

2018-02-01

Investigations based on mesenchymal stem cells (MSCs) for osteoporosis have attracted attention recently. MSCs can be derived from various tissues, such as bone marrow, adipose, umbilical cord, placenta, and dental pulp. Among these, dental pulp-derived MSCs (DPSCs) and hepatocyte growth factor (HGF)-modified DPSCs (DPSCs-HGF) highly express osteogenic-related genes and have stronger osteogenic differentiation capacities. DPSCs have more benefits in treating osteoporosis. The purpose of this study was to investigate the roles of HGF gene-modified DPSCs in bone regeneration using a mouse model of ovariectomy (OVX)-induced bone loss. The HGF and luciferase genes were transferred into human DPSCs using recombinant adenovirus. These transduced cells were assayed for distribution or bone regeneration assay by transplantation into an OVX-induced osteoporosis model. By using bioluminogenic imaging, it was determined that some DPSCs could survive for >1 month in vivo. The DPSCs were mainly distributed to the lung in the early stage and to the liver in the late stage of OVX osteoporosis after administration, but they were scarcely distributed to the bone. The homing efficiency of DPSCs is higher when administrated in the early stage of a mouse OVX model. Micro-computed tomography indicated that DPSCs-Null or DPSCs-HGF transplantation significantly reduces OVX-induced bone loss in the trabecular bone of the distal femur metaphysis, and DPSCs-HGF show a stronger capacity to reduce bone loss. The data suggest that systemic infusion of DPSCs-HGF is a potential therapeutic approach for OVX-induced bone loss, which might be mediated by paracrine mechanisms.

Hepatocyte growth factor/scatter factor-MET signaling in neural crest-derived melanocyte development.

PubMed

Kos, L; Aronzon, A; Takayama, H; Maina, F; Ponzetto, C; Merlino, G; Pavan, W

1999-02-01

The mechanisms governing development of neural crest-derived melanocytes, and how alterations in these pathways lead to hypopigmentation disorders, are not completely understood. Hepatocyte growth factor/scatter factor (HGF/SF) signaling through the tyrosine-kinase receptor, MET, is capable of promoting the proliferation, increasing the motility, and maintaining high tyrosinase activity and melanin synthesis of melanocytes in vitro. In addition, transgenic mice that ubiquitously overexpress HGF/SF demonstrate hyperpigmentation in the skin and leptomenigenes and develop melanomas. To investigate whether HGF/ SF-MET signaling is involved in the development of neural crest-derived melanocytes, transgenic embryos, ubiquitously overexpressing HGF/SF, were analyzed. In HGF/SF transgenic embryos, the distribution of melanoblasts along the characteristic migratory pathway was not affected. However, additional ectopically localized melanoblasts were also observed in the dorsal root ganglia and neural tube, as early as 11.5 days post coitus (p.c.). We utilized an in vitro neural crest culture assay to further explore the role of HGF/SF-MET signaling in neural crest development. HGF/SF added to neural crest cultures increased melanoblast number, permitted differentiation into pigmented melanocytes, promoted melanoblast survival, and could replace mast-cell growth factor/Steel factor (MGF) in explant cultures. To examine whether HGF/SF-MET signaling is required for the proper development of melanocytes, embryos with a targeted Met null mutation (Met-/-) were analysed. In Met-/- embryos, melanoblast number and location were not overtly affected up to 14 days p.c. These results demonstrate that HGF/SF-MET signaling influences, but is not required for, the initial development of neural crest-derived melanocytes in vivo and in vitro.

HGF-induced serine 897 phosphorylation of EphA2 regulates epithelial morphogenesis of MDCK cells in 3D culture.

PubMed

Harada, Kohei; Negishi, Manabu; Katoh, Hironori

2015-05-15

Expression of EphA2 is upregulated in various cancers that are derived from epithelial cells and correlates with the ability of a cancer cell to undergo migration and invasion. Here we have investigated the role of EphA2 in the epithelial morphogenesis of Madin-Darby canine kidney (MDCK) cells in three-dimensional culture. We show that EphA2 is phosphorylated on serine residue 897 through hepatocyte growth factor (HGF) stimulation using a phosphatidylinositol 3-kinase (PI3K)-Akt-dependent mechanism and that this phosphorylation is required for the formation of extensions, the first step of tubulogenesis, in MDCK cysts. By contrast, stimulation using the ligand ephrinA1 dephosphorylates EphA2 on serine residue 897 and suppresses the HGF-induced morphological change. Furthermore, activation of the small GTPase RhoG is involved in the HGF-induced formation of extensions downstream of EphA2. These observations suggest that a ligand-independent activity of EphA2 contributes to epithelial morphogenesis. © 2015. Published by The Company of Biologists Ltd.

The HGF Receptor c-Met Is Overexpressed in Esophageal Adenocarcinoma1

PubMed Central

Herrera, Luis J; El-Hefnawy, Talal; Queiroz de Oliveira, Pierre E; Raja, Siva; Finkelstein, Sydney; Gooding, William; Luketich, James D; Godfrey, Tony E; Hughes, Steven J

2005-01-01

Abstract The hepatocyte growth factor (HGF) receptor, Met, has established oncogenic properties; however, its expression and function in esophageal adenocarcinoma (EA) remain poorly understood. We aimed to determine the expression and potential alterations in Met expression in EA. Met expression was investigated in surgical specimens of EA, Barrett's esophagus (BE), and normal esophagus (NE) using immunohistochemistry (IHC) and quantitative reverse transcriptase polymerase chain reaction. Met expression, phosphorylation, and the effect of COX-2 inhibition on expression were examined in EA cell lines. IHC demonstrated intense Met immunoreactivity in all (100%) EA and dysplastic BE specimens. In contrast, minimal immunostaining was observed in BE without dysplasia or NE specimens. Met mRNA and protein levels were increased in three EA cell lines, and Met protein was phosphorylated in the absence of serum. Sequence analysis found the kinase domain of c-met to be wild type in all three EA cell lines. HGF mRNA expression was identified in two EA cell lines. In COX-2-overexpressing cells, COX-2 inhibition decreased Met expression. Met is consistently overexpressed in EA surgical specimens and in three EA cell lines. Met dysregulation occurs early in Barrett's dysplasia to adenocarcinoma sequence. Future study of Met inhibition as a potential biologic therapy for EA is warranted. PMID:15720819

Hepatocyte growth factor acts as a mitogen for equine satellite cells via protein kinase C δ directed signaling.

PubMed

Brandt, Amanda M; Kania, Joanna M; Gonzalez, Madison L; Johnson, Sally E

2018-06-16

Hepatocyte growth factor (HGF) signals mediate mouse skeletal muscle stem cell, or satellite cell (SC), reentry into the cell cycle and myoblast proliferation. Because the athletic horse experiences exercise-induced muscle damage, the objective of the experiment was to determine the effect of HGF on equine SC (eqSC) bioactivity. Fresh isolates of adult eqSC were incubated with increasing concentrations of HGF and the initial time to DNA synthesis was measured. Media supplementation with HGF did not shorten (P > 0.05) the duration of G0/G1 transition suggesting the growth factor does not affect activation. Treatment with 25 ng/mL HGF increased (P < 0.05) eqSC proliferation that was coincident with phosphorylation of extracellular signal-regulated kinase (ERK)1/2 and AKT serine/threonine kinase 1 (AKT1). Chemical inhibition of the upstream effectors of ERK1/2 or AKT1 elicited no effect (P > 0.05) on HGF-mediated EdU incorporation. By contrast, treatment of eqSC with 2 µm Gö6983, a pan-protein kinase C (PKC) inhibitor, blocked (P < 0.05) HGF-initiated mitotic activity. Gene expression analysis revealed that eqSC express PKCα, -δ and -ε isoforms. Knockdown of PKCδ with a small interfering RNA (siRNA) prevented (P > 0.05) HGF-mediated EdU incorporation. The siPKCδ was specific to the kinase and did not affect (P > 0.05) expression of either PKCα or PKCε. Treatment of confluent eqSCs with 25 ng/mL HGF suppressed (P < 0.05) nuclear myogenin expression during the early stages of differentiation. These results demonstrate that HGF may not affect activation but can act as a mitogen and modest suppressor of differentiation.

Ad-HGF improves the cardiac remodeling of rat following myocardial infarction by upregulating autophagy and necroptosis and inhibiting apoptosis.

PubMed

Liu, Jiabao; Wu, Peng; Wang, Yunle; Du, Yingqiang; A, Nan; Liu, Shuiyuan; Zhang, Yiming; Zhou, Ningtian; Xu, Zhihui; Yang, Zhijian

2016-01-01

Cell death in MI is the most critical determinant of subsequent left ventricular remodeling and heart failure. Besides apoptosis, autophagy and necroptosis have been recently found to be another two regulated cell death styles. HGF has been reported to have a protective role in MI, but its impact on the three death styles remains unclear. Thus, our study was performed to investigate the distribution of autophagy, apoptosis and necroptosis in cardiac tissues after MI and explore the role and mechanism of Ad-HGF on cardiac remodeling by regulating the three death styles. We firstly showed the distribution of autophagy, apoptosis and necroptosis differs in temporal and spatial context after MI using immunofluorescence. Notably, Ad-HGF treatment improves the cardiac remodeling of SD rats following MI by preserving the heart function, reducing the scar size and aggresomes. Further mechanism study reveals Ad-HGF promotes autophagy and necroptosis and inhibits apoptosis in vivo and in vitro . Co-immunoprecipitation assays showed Ad-HGF treatment significantly decreased the binding of Bcl-2 to Beclin1 but enhanced Bcl-2 binding to Bax in H9c2 cells under hypoxia. Moreover, HGF-induced sequestration of Bax by Bcl-2 allows Bax to become inactive, thereby inhibiting apoptosis. In addition, Ad-HGF markedly increased the formation of Beclin1-Vps34-Atg14L complex, which accounted for promoting autophagy. Both the western blot and activity assay showed Ad-HGF significantly decreased the caspase 8 protein and activity levels, which obligated the cell to undergo necroptosis under hypoxia and block apoptosis. Thus, our findings offer new evidence and strategies for the treatment of MI and post-MI cardiac remodeling.

Ad-HGF improves the cardiac remodeling of rat following myocardial infarction by upregulating autophagy and necroptosis and inhibiting apoptosis

PubMed Central

Liu, Jiabao; Wu, Peng; Wang, Yunle; Du, Yingqiang; A, Nan; Liu, Shuiyuan; Zhang, Yiming; Zhou, Ningtian; Xu, Zhihui; Yang, Zhijian

2016-01-01

Cell death in MI is the most critical determinant of subsequent left ventricular remodeling and heart failure. Besides apoptosis, autophagy and necroptosis have been recently found to be another two regulated cell death styles. HGF has been reported to have a protective role in MI, but its impact on the three death styles remains unclear. Thus, our study was performed to investigate the distribution of autophagy, apoptosis and necroptosis in cardiac tissues after MI and explore the role and mechanism of Ad-HGF on cardiac remodeling by regulating the three death styles. We firstly showed the distribution of autophagy, apoptosis and necroptosis differs in temporal and spatial context after MI using immunofluorescence. Notably, Ad-HGF treatment improves the cardiac remodeling of SD rats following MI by preserving the heart function, reducing the scar size and aggresomes. Further mechanism study reveals Ad-HGF promotes autophagy and necroptosis and inhibits apoptosis in vivo and in vitro. Co-immunoprecipitation assays showed Ad-HGF treatment significantly decreased the binding of Bcl-2 to Beclin1 but enhanced Bcl-2 binding to Bax in H9c2 cells under hypoxia. Moreover, HGF-induced sequestration of Bax by Bcl-2 allows Bax to become inactive, thereby inhibiting apoptosis. In addition, Ad-HGF markedly increased the formation of Beclin1-Vps34-Atg14L complex, which accounted for promoting autophagy. Both the western blot and activity assay showed Ad-HGF significantly decreased the caspase 8 protein and activity levels, which obligated the cell to undergo necroptosis under hypoxia and block apoptosis. Thus, our findings offer new evidence and strategies for the treatment of MI and post-MI cardiac remodeling. PMID:27904666

[Ischemic brain injury and hepatocyte growth factor].

PubMed

Takeo, Satoshi; Takagi, Norio; Takagi, Keiko

2007-11-01

Cerebral ischemia causes an irreversible and neurodegenerative disorder that may lead to progressive dementia and global cognitive deterioration. Since the overall process of ischemic brain injuries is extremely complex, treatment with endogenous multifunctional factors would be better choices for preventing complicated ischemic brain injuries. Hepatocyte growth factor, HGF, is a multifunctional cytokine originally identified and purified as a potent mitogen for hepatocyte. The activation of the c-Met/HGF receptor evokes diverse cellular responses, including mitogenic, morphogenic, angiogenic and anti-apoptotic activities in various types of cell. Previous studies showed that HGF and c-Met were expressed in various brain regions under normal conditions and that HGF enhanced the survival of hippocampal and cortical neurons during the aging of cells in culture. The protective effects of HGF on in vivo ischemic brain injuries and their mechanisms have not fully understood. To elucidate therapeutic potencies of HGF for ischemic brain injuries, we examined effects of HGF on ischemia-induced learning and memory dysfunction, neuronal cell death and endothelial cell damage by using the 4-vessel occlusion model and the microsphere embolism model in rats. Our findings suggested that treatment with HGF was capable of protecting hippocampal neurons against ischemia-induced cell death through the prevention of apoptosis-inducing factor translocation to the nucleus. Furthermore, we demonstrated that HGF had the ability to prevent tissue degeneration and improved learning and memory function after cerebral embolism, possibly through prevention of cerebral vessel injuries. As HGF has a potent cerebroprotective effect, it could be a prospective agent for the therapy against complicated ischemic brain diseases.

Acute tissue injury activates satellite cells and promotes sarcoma formation via the HGF/c-MET signaling pathway.

PubMed

Van Mater, David; Añó, Leonor; Blum, Jordan M; Webster, Micah T; Huang, WeiQiao; Williams, Nerissa; Ma, Yan; Cardona, Diana M; Fan, Chen-Ming; Kirsch, David G

2015-02-01

Some patients with soft-tissue sarcoma (STS) report a history of injury at the site of their tumor. Although this phenomenon is widely reported, there are relatively few experimental systems that have directly assessed the role of injury in sarcoma formation. We recently described a mouse model of STS whereby p53 is deleted and oncogenic Kras is activated in muscle satellite cells via a Pax7(CreER) driver following intraperitoneal injection with tamoxifen. Here, we report that after systemic injection of tamoxifen, the vast majority of Pax7-expressing cells remain quiescent despite mutation of p53 and Kras. The fate of these muscle progenitors is dramatically altered by tissue injury, which leads to faster kinetics of sarcoma formation. In adult muscle, quiescent satellite cells will transition into an active state in response to hepatocyte growth factor (HGF). We show that modulating satellite cell quiescence via intramuscular injection of HGF increases the penetrance of sarcoma formation at the site of injection, which is dependent on its cognate receptor c-MET. Unexpectedly, the tumor-promoting effect of tissue injury also requires c-Met. These results reveal a mechanism by which HGF/c-MET signaling promotes tumor formation after tissue injury in a mouse model of primary STS, and they may explain why some patients develop a STS at the site of injury. ©2014 American Association for Cancer Research.

Cloning of a human hepatocyte growth factor/scatter factor transcription variant from a gastric cancer cell line HSC-39.

PubMed

Yokozaki, H; Tahara, H; Oue, N; Tahara, E

2000-01-01

A new transcription variant of hepatocyte growth factor/scatter factor (HGF/SF) was cloned from human gastric cancer cell line HSC-39. Northern blot analysis of eight human gastric cancer cell lines (TMK-1, MKN-1, MKN-7, MKN-28, MKN-45, MKN-74, KATO-III and HSC-39) demonstrated that HSC-39 cells expressed a 1.3 kb abnormal HGF/SF transcript. Screening of 1 x 10(6) colonies of cDNA library from HSC-39 constructed in pAP3neo mammalian expression vector selected four positive clones containing HGF/SF transcript. Among them, two contained a 1.3 kbp insert detecting the identical transcript to that obtained with HGF/SF probe by Northern blotting. Deoxynucleotide sequencing of the 1.3 kbp insert revealed that it was composed of a part of HGF/SF cDNA from exon 14 to exon 18, corresponding to the whole sequence of HGF/SF light chain, with 5' 75 nucleotides unrelated to any sequence involved in HGF/SF.

Erythropoietin induces production of hepatocyte growth factor from bone marrow mesenchymal stem cells in vitro.

PubMed

Tari, Kaveh; Atashi, Amir; Kaviani, Saied; AkhavanRahnama, Mahshid; Anbarlou, Azadeh; Mossahebi-Mohammadi, Majid

2017-01-01

Hepatocyte Growth Factor (HGF) plays a pivotal role in hematopoiesis, motility, growth and mobilization of hematopoietic stem/progenitor cells (HSPCs). HGF mainly is produced by bone marrow mesenchymal stem cells (BM-MSCs). MSCs express erythropoietin (EPO) receptor. In this study, we aimed to assess the effect of EPO on HGF secretion in BM-MSCs. The BM-MSCs treated with EPO (4 IU/ml) for 6, 24 and 48 h. HGF gene expression and protein level were assessed using quantitative real time PCR (qRT-PCR) and Enzyme-linked immunosorbant Assay. In order to show the effect of secreted HGF on migration of HSPCs, hematopoietic stem cells (HSCs) were isolated from cord blood and evaluated using transwell migration assay. We observed a significant increase in level of HGF in cell supernatant after 48 h compared to control group (P < 0.05). Also, qRT-PCR results demonstrated a significant elevation in HGF expression level after 24 and 48 h treatment with EPO compared to control group (P < 0.05). Finally, migration assay results showed a significant increase in migration of HSCs in treated group after 48 h. Our data indicated that EPO may play an important role in stem cell mobilization through up regulating HGF in MSCs and inducing migration of HSCs. Copyright © 2016 International Alliance for Biological Standardization. Published by Elsevier Ltd. All rights reserved.

Hepatocyte growth factor, a biomarker of macroangiopathy in diabetes mellitus

PubMed Central

Konya, Hiroyuki; Miuchi, Masayuki; Satani, Kahori; Matsutani, Satoshi; Tsunoda, Taku; Yano, Yuzo; Katsuno, Tomoyuki; Hamaguchi, Tomoya; Miyagawa, Jun-Ichiro; Namba, Mitsuyoshi

2014-01-01

Atherosclerotic involvements are an essential causal element of prospect in diabetes mellitus (DM), with carotid atherosclerosis (CA) being a common risk-factor for prospective crisis of coronary artery diseases (CAD) and/or cerebral infarction (CI) in DM subjects. From another point of view, several reports have supplied augmenting proof that hepatocyte growth factor (HGF) has a physiopathological part in DM involvements. HGF has been a mesenchymal-derived polyphenic factor which modulates development, motion, and morphosis of diverse cells, and has been regarded as a humor intermediator of epithelial-mesenchymal interplays. The serum concentrations of HGF have been elevated in subjects with CAD and CI, especially during the acute phase of both disturbances. In our study with 89 type 2 DM patients, the association between serum concentrations of HGF and risk-factors for macrovascular complications inclusive of CA were examined. The average of serum HGF levels in the subjects was more elevated than the reference interval. The serum HGF concentrations associated positively with both intimal-media thickness (IMT) (r = 0.24, P = 0.0248) and plaque score (r = 0.27, P = 0.0126), indicating a relationship between the elevated HGF concentrations and advancement of CA involvements. Multivariate statistical analysis accentuated that serum concentrations of HGF would be associated independently with IMT (standardized = 0.28, P = 0.0499). The review indicates what is presently known regarding serum HGF might be a new and meaningful biomarker of macroangiopathy in DM subjects. PMID:25317245

Hepatocyte growth factor in renal failure: promise and reality.

PubMed

Vargas, G A; Hoeflich, A; Jehle, P M

2000-04-01

Can science discover some secrets of Greek mythology? In the case of Prometheus, we can now suppose that his amazing hepatic regeneration was caused by a peptide growth factor called hepatocyte growth factor (HGF). Increasing evidence indicates that HGF acts as a multifunctional cytokine on different cell types. This review addresses the molecular mechanisms that are responsible for the pleiotropic effects of HGF. HGF binds with high affinity to its specific tyrosine kinase receptor c-met, thereby stimulating not only cell proliferation and differentiation, but also cell migration and tumorigenesis. The three fundamental principles of medicine-prevention, diagnosis, and therapy-may be benefited by the rational use of HGF. In renal tubular cells, HGF induces mitogenic and morphogenetic responses. In animal models of toxic or ischemic acute renal failure, HGF acts in a renotropic and nephroprotective manner. HGF expression is rapidly up-regulated in the remnant kidney of nephrectomized rats, inducing compensatory growth. In a mouse model of chronic renal disease, HGF inhibits the progression of tubulointerstitial fibrosis and kidney dysfunction. Increased HGF mRNA transcripts were detected in mesenchymal and tubular epithelial cells of rejecting kidney. In transplanted patients, elevated HGF levels may indicate renal rejection. When HGF is considered as a therapeutic agent in human medicine, for example, to stimulate kidney regeneration after acute injury, strategies need to be developed to stimulate cell regeneration and differentiation without an induction of tumorigenesis.

Microvesicles derived from human umbilical cord mesenchymal stem cells facilitate tubular epithelial cell dedifferentiation and growth via hepatocyte growth factor induction.

PubMed

Ju, Guan-qun; Cheng, Jun; Zhong, Liang; 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.

Hepatocyte growth factor incorporated chitosan nanoparticles augment the differentiation of stem cell into hepatocytes for the recovery of liver cirrhosis in mice

PubMed Central

2011-01-01

Background Short half-life and low levels of growth factors in the niche of injured microenvironment necessitates the exogenous and sustainable delivery of growth factors along with stem cells to augment the regeneration of injured tissues. Methods Here, recombinant human hepatocyte growth factor (HGF) was incorporated into chitosan nanoparticles (CNP) by ionic gelation method and studied for its morphological and physiological characteristics. Cirrhotic mice received either hematopoietic stem cells (HSC) or mesenchymal stemcells (MSC) with or without HGF incorporated chitosan nanoparticles (HGF-CNP) and saline as control. Biochemical, histological, immunostaining and gene expression assays were carried out using serum and liver tissue samples. One way analysis of variance was used for statics application Results Serum levels of selected liver protein and enzymes were significantly increased in the combination of MSC and HGF-CNP (MSC+HGF-CNP) treated group. Immunopositive staining for albumin (Alb) and cytokeratin 18 (CK18), and reverse transcription-polymerase chain reaction (RT-PCR) for Alb, alpha fetoprotein (AFP), CK18, cytokeratin 19 (CK19) ascertained that MSC-HGF-CNP treatment could be an effective combination to repopulate liver parenchymal cells in the liver cirrhosis. Zymogram and western blotting for matrix metalloproteinases 2 and 9 (MMP2 and MMP9) revealed that MMP2 actively involved in the fibrolysis of cirrhotic tissue. Immunostaining for alpha smooth muscle actin (αSMA) and type I collagen showed decreased expression in the MSC+HGF-CNP treatment. These results indicated that HGF-CNP enhanced the differentiation of stem cells into hepatocytes and supported the reversal of fibrolysis of extracellular matrix (ECM). Conclusion Bone marrow stem cells were isolated, characterized and transplanted in mice model. Biodegradable biopolymeric nanoparticles were prepared with the pleotrophic protein molecule and it worked well for the differentiation of stem

Sequence analysis of the Ras-MAPK pathway genes SOS1, EGFR & GRB2 in silver foxes (Vulpes vulpes): candidate genes for hereditary hyperplastic gingivitis.

PubMed

Clark, Jo-Anna B J; Tully, Sara J; Dawn Marshall, H

2014-12-01

Hereditary hyperplastic gingivitis (HHG) is an autosomal recessive disease that presents with progressive gingival proliferation in farmed silver foxes. Hereditary gingival fibromatosis (HGF) is an analogous condition in humans that is genetically heterogeneous with several known autosomal dominant loci. For one locus the causative mutation is in the Son of sevenless homologue 1 (SOS1) gene. For the remaining loci, the molecular mechanisms are unknown but Ras pathway involvement is suspected. Here we compare sequences for the SOS1 gene, and two adjacent genes in the Ras pathway, growth receptor bound protein 2 (GRB2) and epidermal growth factor receptor (EGFR), between HHG-affected and unaffected foxes. We conclude that the known HGF causative mutation does not cause HHG in foxes, nor do the coding regions or intron-exon boundaries of these three genes contain any candidate mutations for fox gum disease. Patterns of molecular evolution among foxes and other mammals reflect high conservation and strong functional constraints for SOS1 and GRB2 but reveal a lineage-specific pattern of variability in EGFR consistent with mutational rate differences, relaxed functional constraints, and possibly positive selection.

IL-6 modulates hepatocyte proliferation via induction of HGF/p21{sup cip1}: Regulation by SOCS3

DOE Office of Scientific and Technical Information (OSTI.GOV)

Sun Rui; Jaruga, Barbara; Kulkarni, Shailin

2005-12-30

The precise role of IL-6 in liver regeneration and hepatocyte proliferation is controversial and the role of SOCS3 in liver regeneration remains unknown. Here we show that in vitro treatment with IL-6 inhibited primary mouse hepatocyte proliferation. IL-6 induced p21{sup cip1} protein expression in primary mouse hepatocytes. Disruption of the p21{sup cip1} gene abolished the inhibitory effect of IL-6 on cell proliferation. Co-culture with nonparenchymal liver cells diminished IL-6 inhibition of hepatocyte proliferation, which was likely due to IL-6 stimulation of nonparenchymal cells to produce HGF. Finally, IL-6 induced higher levels of p21{sup cip1} protein expression and a slightly strongermore » inhibition of cell proliferation in SOCS3{sup +/-} mouse hepatocytes compared to wild-type hepatocytes, while liver regeneration was enhanced and prolonged in SOCS3{sup +/-} mice. Our findings suggest that IL-6 directly inhibits hepatocyte proliferation via a p21{sup cip1}-dependent mechanism and indirectly enhances hepatocyte proliferation via stimulating nonparenchymal cells to produce HGF. SOCS3 negatively regulates liver regeneration.« less

Establishment of human hair follicle mesenchymal stem cells with overexpressed human hepatocyte growth factor.

PubMed

Zhou, Dan; Cheng, Hongjing; Liu, Jinyu; Zhang, Lei

2017-06-01

Chronic liver disease has become a major health problem that causes serious damage to human health. Since the existing treatment effect was not ideal, we need to seek new treatment methods. We utilized the gene recombination technology to obtain the human hair mesenchymal stem cells which overexpression of human hepatocyte growth factor (hHGF). Furthermore, we verified the property of transfected cells through detecting surface marker by flow cytometry. We show here establishment of the hHGF-overexpressing lentivirus vector, and successfully transfection to human hair follicle mesenchymal stem cells. The verified experiments could demonstrate the human hair follicle mesenchymal stem cells which have been transfected still have the properties of stem cells. We successfully constructed human hair follicle mesenchymal stem cells which overexpression hHGF, and maintain the same properties compared with pro-transfected cells.

Hepatocyte growth factor is crucial for development of the carapace in turtles

PubMed Central

Kawashima-Ohya, Yoshie; Narita, Yuichi; Nagashima, Hiroshi; Usuda, Ryo; Kuratani, Shigeru

2011-01-01

Turtles are characterized by their shell, composed of a dorsal carapace and a ventral plastron. The carapace first appears as the turtle-specific carapacial ridge (CR) on the lateral aspect of the embryonic flank. Accompanying the acquisition of the shell, unlike in other amniotes, hypaxial muscles in turtle embryos appear as thin threads of fibrous tissue. To understand carapacial evolution from the perspective of muscle development, we compared the development of the muscle plate, the anlage of hypaxial muscles, between the Chinese soft-shelled turtle, Pelodiscus sinensis, and chicken embryos. We found that the ventrolateral lip (VLL) of the thoracic dermomyotome of P. sinensis delaminates early and produces sparse muscle plate in the lateral body wall. Expression patterns of the regulatory genes for myotome differentiation, such as Myf5, myogenin, Pax3, and Pax7 have been conserved among amniotes, including turtles. However, in P. sinensis embryos, the gene hepatocyte growth factor (HGF), encoding a regulatory factor for delamination of the dermomyotomal VLL, was uniquely expressed in sclerotome and the lateral body wall at the interlimb level. Implantation of COS-7 cells expressing a HGF antagonist into the turtle embryo inhibited CR formation. We conclude that the de novo expression of HGF in the turtle mesoderm would have played an innovative role resulting in the acquisition of the turtle-specific body plan. PMID:21535464

The Involvement of Hepatocyte Growth Factor-MET-Matrix Metalloproteinase 1 Signaling in Bladder Cancer Invasiveness and Proliferation. Effect of the MET Inhibitor, Cabozantinib (XL184), on Bladder Cancer Cells.

PubMed

Shintani, Terumichi; Kusuhara, Yoshito; Daizumoto, Kei; Dondoo, Tsogt-Ochir; Yamamoto, Hiroki; Mori, Hidehisa; Fukawa, Tomoya; Nakatsuji, Hiroyoshi; Fukumori, Tomoharu; Takahashi, Masayuki; Kanayama, Hiroomi

2017-03-01

To clarify the invasive mechanisms of muscle-invasive bladder cancer (BCa) would be useful for the determination of appropriate treatment strategies. We previously showed that hepatocyte growth factor (HGF)-MET signaling is correlated with invasiveness of BCa cells. Here, we investigated the effects of the MET inhibitor, cabozantinib (XL184), on BCa cells. We first conducted Western blot analysis to investigate MET expression in BCa cell lines. Next, we examined the effect of cabozantinib on their proliferation and invasive abilities using MTT and Matrigel invasion assays, respectively. Invasion assays were performed using the xCELLigence system. Additionally, to investigate the biological function of HGF-MET signaling, we analyzed gene expression profiles and performed real-time polymerase chain reaction analyses of 5637 cells that were cultivated with or without HGF stimulation, with or without cabozantinib. MET was highly expressed in 4 of 5 BCa cell lines, and 5637 and T24 cells showed especially high protein expression of MET. Cabozantinib suppressed cell proliferation and invasion (cell index; mock, 1.49 vs HGF, 2.26 vs HGF + XL184, 1.47, P < .05). Gene expression profile analysis indicated that matrix metalloproteinase 1 (MMP1) was significantly elevated at the mRNA level with addition of HGF. Moreover, cabozantinib suppressed HGF-induced MMP1 expression in 5637 T24 cells. These data indicate that cabozantinib suppressed MMP1 expression by blocking HGF-MET signaling and that HGF-MET-MMP1 signaling is involved in the invasiveness and proliferation of BCa cells. These results suggest that cabozantinib might prove useful for future treatment of muscle-invasive BCa. Copyright © 2016 Elsevier Inc. All rights reserved.

[Cloning and characterization of genes differentially expressed in human dental pulp cells and gingival fibroblasts].

PubMed

Wang, Zhong-dong; Wu, Ji-nan; Zhou, Lin; Ling, Jun-qi; Guo, Xi-min; Xiao, Ming-zhen; Zhu, Feng; Pu, Qin; Chai, Yu-bo; Zhao, Zhong-liang

2007-02-01

To study the biological properties of human dental pulp cells (HDPC) by cloning and analysis of genes differentially expressed in HDPC in comparison with human gingival fibroblasts (HGF). HDPC and HGF were cultured and identified by immunocytochemistry. HPDC and HGF subtractive cDNA library was established by PCR-based modified subtractive hybridization, genes differentially expressed by HPDC were cloned, sequenced and compared to find homogeneous sequence in GenBank by BLAST. Cloning and sequencing analysis indicate 12 genes differentially expressed were obtained, in which two were unknown genes. Among the 10 known genes, 4 were related to signal transduction, 2 were related to trans-membrane transportation (both cell membrane and nuclear membrane), and 2 were related to RNA splicing mechanisms. The biological properties of HPDC are determined by the differential expression of some genes and the growth and differentiation of HPDC are associated to the dynamic protein synthesis and secretion activities of the cell.

MiR-16 regulates the pro-tumorigenic potential of lung fibroblasts through the inhibition of HGF production in an FGFR-1- and MEK1-dependent manner.

PubMed

Andriani, Francesca; Majorini, Maria Teresa; Mano, Miguel; Landoni, Elena; Miceli, Rosalba; Facchinetti, Federica; Mensah, Mavis; Fontanella, Enrico; Dugo, Matteo; Giacca, Mauro; Pastorino, Ugo; Sozzi, Gabriella; Delia, Domenico; Roz, Luca; Lecis, Daniele

2018-03-20

Fibroblasts are crucial mediators of tumor-stroma cross-talk through synthesis and remodeling of the extracellular matrix and production of multiple soluble factors. Nonetheless, little is still known about specific determinants of fibroblast pro-tumorigenic activity in lung cancer. Here, we aimed at understanding the role of miRNAs, which are often altered in stromal cells, in reprogramming fibroblasts towards a tumor-supporting phenotype. We employed a co-culture-based high-throughput screening to identify specific miRNAs modulating the pro-tumorigenic potential of lung fibroblasts. Multiplex assays and ELISA were instrumental to study the effect of miRNAs on the secretome of both primary and immortalized lung fibroblasts from lung cancer patients and to evaluate plasmatic levels of HGF in heavy smokers. Direct mRNA targeting by miRNAs was investigated through dual-luciferase reporter assay and western blot. Finally, the pro-tumorigenic activity of fibroblasts and their conditioned media was tested by employing in vitro migration experiments and mouse xenografts. We identified miR-16 as a master regulator of fibroblast secretome and showed that its upregulation reduces HGF secretion by fibroblasts, impairing their capacity to promote cancer cell migration. This effect is due to a pleiotropic activity of miR-16 which prevents HGF expression through direct inhibition of FGFR-1 signaling and targeting of HGF mRNA. Mechanistically, miR-16 targets FGFR-1 downstream mediator MEK1, thus reducing ERK1/2 activation. Consistently, chemical or genetic inhibition of FGFR-1 mimics miR-16 activity and prevents HGF production. Of note, we report that primary fibroblast cell lines derived from lungs of heavy smokers express reduced miR-16 levels compared to those from lungs not exposed to smoke and that HGF concentration in heavy smokers' plasma correlates with levels of tobacco exposure. Finally, in vivo experiments confirmed that restoration of miR-16 expression in fibroblasts

Role of hepatocyte growth factor in the development of dendritic cells from CD34+ bone marrow cells.

PubMed

Ovali, E; Ratip, S; Kibaroglu, A; Tekelioglu, Y; Cetiner, M; Karti, S; Aydin, F; Bayik, M; Akoglu, T

2000-05-01

Hepatocyte growth factor (HGF) is known to augment the effects of stem cell factor, interleukin-3, granulocyte-macrophage colony-stimulating factor (GM-CSF), erythropoetin, and granulocyte colony-stimulating factor, all of which are involved in hematopoiesis. HGF is also known to have a role in immune responses. The aim of this study was to investigate whether HGF is involved in the development of dendritic cells (DC) from CD34+ bone marrow cells. CD34+ cells obtained from three healthy donors were incubated in various combinations of HGF, GM-CSF, and tumor necrosis factor (TNF) for 12 days. Developing cell populations were analyzed for surface markers, morphology and functional capacities by flow cytometry, light microscopy and mixed lymphocyte reaction, respectively. Incubation with HGF alone generated greater number of dendritic cells from CD34+ bone marrow cells than incubation with GM-CSF, or a combination of GM-CSF with TNF. HGF was also found to potentiate the effect of GM-CSF on DC and monocyte development. The effects of HGF were inhibited by the concurrent use of TNF. HGF appears to be a significant factor in the development of dendritic cells from CD34+ bone marrow cells.

Hepatocyte growth factor/scatter factor enhances the invasion of mesothelioma cell lines and the expression of matrix metalloproteinases

PubMed Central

Harvey, P; Clark, I M; Jaurand, M-C; Warn, R M; Edwards, D R

2000-01-01

Hepatocyte growth factor/scatter factor (HGF/SF) is a multifunctional factor involved both in development and tissue repair, as well as pathological processes such as cancer and metastasis. It has been identified in vivo in many types of tumours together with its tyrosine kinase receptor, Met. We show here that exogenous HGF/SF acts as a strong chemoattractant for human mesothelioma cell lines. The factor also enhanced cell adhesion to and invasion into Matrigel. The mesothelioma cell lines synthesized a panel of matrix metalloproteinases critical for tumour progression such as MMP-1, 2, 3, 9 and membrane-bound MT1-MMP. HGF/SF stimulated the expression of MMP-1, 9 and MT1-MMP and had a slight effect on expression of the MMP inhibitor TIMP-1 but not TIMP-2. However, there was no simple correlation between the levels of MMPs and TIMPs of the cell lines and their different invasion properties or between HGF/SF stimulatory effects on MMP expression and invasion. In addition, effects of protease inhibitors on invasion suggested that serine proteases were also expressed in human mesothelioma cell lines and were involved in HGF/SF-induced invasion. The results show a predominant role for HGF/SF in mesothelioma cell invasion, stimulating simultaneously adhesion, motility, invasion and regulation of MMP and TIMP levels. © 2000 Cancer Research Campaign PMID:11027427

Substrate-mediated delivery of gene complex nanoparticles via polydopamine coating for enhancing competitiveness of endothelial cells.

PubMed

Li, Bo-Chao; Chang, Hao; Ren, Ke-Feng; Ji, Jian

2016-11-01

Substrate-mediated delivery of functional plasmid DNA (pDNA) has been proven to be a promising strategy to promote competitiveness of endothelial cells (ECs) over smooth muscle cells (SMCs), which is beneficial to inducing fast endothelialization of implanted vascular devices. Thus, it is of great importance to develop universal approaches with simplicity and easiness to immobilize DNA complex nanoparticles on substrates. In this study, the bioinspired polydopamine (PDA) coating was employed in immobilization of DNA complex nanoparticles, which were composed of protamine (PrS) and plasmid DNA encoding with hepatocyte growth factor (HGF-pDNA) gene. We demonstrated that the DNA complex nanoparticles can be successfully immobilized onto the PDA surface. Consequently, the HGF expression of both ECs and SMCs were significantly improved when they cultured on the DNA complex nanoparticles-immobilized substrates. Furthermore, EC proliferation was specifically promoted due to bioactivity of HGF, leading to an enhancement of EC competitiveness over SMCs. Our findings demonstrated the substrate-mediated functional gene nanoparticle delivery through PDA coating as a simple and efficient approach. It may hold great potential in the field of interventional cardiovascular implants. Copyright © 2016 Elsevier B.V. All rights reserved.

Semi-synthesis of a HGF/SF kringle one (K1) domain scaffold generates a potent in vivo MET receptor agonist.

PubMed

Simonneau, Claire; Bérénice Leclercq; Mougel, Alexandra; Adriaenssens, Eric; Paquet, Charlotte; Raibaut, Laurent; Ollivier, Nathalie; Drobecq, Hervé; Marcoux, Julien; Cianférani, Sarah; Tulasne, David; de Jonge, Hugo; Melnyk, Oleg; Vicogne, Jérôme

2015-03-01

The development of MET receptor agonists is an important goal in regenerative medicine, but is limited by the complexity and incomplete understanding of its interaction with HGF/SF (Hepatocyte Growth Factor/Scatter Factor). NK1 is a natural occurring agonist comprising the N-terminal (N) and the first kringle (K1) domains of HGF/SF. In the presence of heparin, NK1 can self-associate into a "head to tail" dimer which is considered as the minimal structural module able to trigger MET dimerization and activation whereas isolated K1 and N domains showed a weak or a complete lack of agonistic activity respectively. Starting from these structural and biological observations, we investigated whether it was possible to recapitulate the biological properties of NK1 using a new molecular architecture of isolated N or K1 domains. Therefore, we engineered multivalent N or K1 scaffolds by combining synthetic and homogeneous site-specifically biotinylated N and K1 domains (NB and K1B) and streptavidin (S). NB alone or in complex failed to activate MET signaling and to trigger cellular phenotypes. Importantly and to the contrary of K1B alone, the semi-synthetic K1B/S complex mimicked NK1 MET agonist activity in cell scattering, morphogenesis and survival phenotypic assays. Impressively, K1B/S complex stimulated in vivo angiogenesis and, when injected in mice, protected the liver against fulminant hepatitis in a MET dependent manner whereas NK1 and HGF were substantially less potent. These data reveal that without N domain, proper multimerization of K1 domain is a promising strategy for the rational design of powerful MET agonists.

The autism susceptibility gene met regulates zebrafish cerebellar development and facial motor neuron migration

PubMed Central

Elsen, Gina E.; Choi, Louis Y.; Prince, Victoria E.; Ho, Robert K.

2009-01-01

During development, Met signaling regulates a range of cellular processes including growth, differentiation, survival and migration. The Met gene encodes a tyrosine kinase receptor, which is activated by Hgf (hepatocyte growth factor) ligand. Altered regulation of human MET expression has been implicated in autism. In mouse, Met signaling has been shown to regulate cerebellum development. Since abnormalities in cerebellar structure have been reported in some autistic patients, we have used the zebrafish to address the role of Met signaling during cerebellar development and thus further our understanding of the molecular basis of autism. We find that zebrafish met is expressed in the cerebellar primordium, later localizing to the ventricular zone (VZ), with the hgf1 and hgf2 ligand genes expressed in surrounding tissues. Morpholino knockdown of either Met or its Hgf ligands leads to a significant reduction in the size of the cerebellum, primarily as a consequence of reduced proliferation. Met signaling knockdown disrupts specification of VZ-derived cell types, and also reduces granule cell numbers, due to an early effect on cerebellar proliferation and/or as an indirect consequence of loss of signals from VZ-derived cells later in development. These patterning defects preclude analysis of cerebellar neuronal migration, but we have found that Met signaling is necessary for migration of hindbrain facial motor neurons. In summary, we have described roles for Met signaling in coordinating growth and cell type specification within the developing cerebellum, and in migration of hindbrain neurons. These functions may underlie the correlation between altered MET regulation and Autism Spectrum Disorders. PMID:19732764

An investigation into the stability of commercial versus MG63-derived hepatocyte growth factor under flow cultivation conditions.

PubMed

Meneghello, Giulia; Storm, Michael P; Chaudhuri, Julian B; De Bank, Paul A; Ellis, Marianne J

2015-03-01

The scale-up of tissue engineering cell culture must ensure that conditions are maintained while also being cost effective. Here we analyse the stability of hepatocyte growth factor (HGF) to investigate whether concentrations change under dynamic conditions, and compare commercial recombinant human HGF as an additive in 'standard medium', to HGF secreted by the osteosarcoma cell line MG63 as a 'preconditioned medium'. After 3 h under flow conditions, HGF in the standard medium degraded to 40% of its original concentration but HGF in the preconditioned medium remained at 100%. The concentration of secreted HGF was 10 times greater than the working concentration of commercially-available HGF. Thus HGF within this medium has increased stability; MG63-derived HGF should therefore be investigated as a cost-effective alternative to current lyophilised powders for use in in vitro models. Furthermore, we recommend that those intending to use HGF (or other growth factors) should consider similar stability testing before embarking on experiments with media flow.

HEPATOCYTE GROWTH FACTOR ACTS AS A MITOGEN AND CHEMOATTRACTANT FOR POSTNATAL SUBVENTRICULAR ZONE-OLFACTORY BULB NEUROGENESIS

PubMed Central

Wang, Tsu-Wei; Zhang, Huailin; Gyetko, Margaret R.; Parent, Jack M.

2011-01-01

Neural progenitor cells persist throughout life in the forebrain subventricular zone (SVZ). They generate neuroblasts that migrate to the olfactory bulb and differentiate into interneurons, but mechanisms underlying these processes are poorly understood. Hepatocyte growth factor/scatter factor (HGF/SF) is a pleiotropic factor that influences cell motility, proliferation and morphogenesis in neural and non-neural tissues. HGF and its receptor, c-Met, are present in the rodent SVZ-olfactory bulb pathway. Using in vitro neurogenesis assays and in vivo studies of partially HGF-deficient mice, we find that HGF promotes SVZ cell proliferation and progenitor cell maintenance, while slowing differentiation and possibly altering cell fate choices. HGF also acts as a chemoattractant for SVZ neuroblasts in co-culture assays. Decreased HGF signaling induces ectopic SVZ neuroblast migration and alters the timing of migration to the olfactory bulb. These results suggest that HGF influences multiple steps in postnatal forebrain neurogenesis. HGF is a mitogen for SVZ neural progenitors, and regulates their differentiation and olfactory bulb migration. PMID:21683144

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

In vitro modulation of microglia motility by glioma cells is mediated by hepatocyte growth factor/scatter factor.

PubMed

Badie, B; Schartner, J; Klaver, J; Vorpahl, J

1999-05-01

Considered as immune effector cells of the central nervous system, microglia represent a major component of the inflammatory cells found in malignant gliomas. Although their role in brain tumor biology is unclear, accumulation of microglia in malignant brain tumors may be mediated through active secretion of cytokines by glioma cells. Because hepatocyte growth factor/scatter factor (HGF/SF) has been shown to modulate glioma motility through an autocrine mechanism, and because microglia have been reported to express the HGF/SF receptor Met, we hypothesized that microglia recruitment by gliomas may also occur through the secretion of HGF/SF. The effect of glioma cells in augmenting BV-2 murine microglia motility was studied by using an in vitro Boyden chamber migration assay. To determine the chemokines involved in microglia migration, neutralizing monoclonal antibodies against monocyte chemotactic protein-1 and HGF/SF were tested. Immunoblotting was used to check for the expression of HGF/SF by glioma cells, and the expression of Met by BV-2 cells was examined by flow cytometry. BV-2 migration was noted within 7 hours of incubation with both human (U251 MG and U373 MG) and murine (GL261) glioma cell lines. This migration corresponded to HGF/SF secretion by glioma cells and was completely inhibited by neutralizing monoclonal antibody against HGF/SF, but not monocyte chemotactic protein-1. Exposure of BV-2 cells to recombinant HGF/SF, but not monocyte chemotactic protein-1, resulted in their migration and down-regulation of Met in a dose-dependent fashion. HGF/SF, which plays a role in glioma motility and mitogenesis, may also act as a chemokine for microglia and may be responsible for the microglia infiltration in malignant gliomas. This active recruitment of microglia may play an important role in glioma biology.

Multifaceted Therapeutic Benefits of Factors Derived From Dental Pulp Stem Cells for Mouse Liver Fibrosis.

PubMed

Hirata, Marina; Ishigami, Masatoshi; Matsushita, Yoshihiro; Ito, Takanori; Hattori, Hisashi; Hibi, Hideharu; Goto, Hidemi; Ueda, Minoru; Yamamoto, Akihito

2016-10-01

: Chronic liver injury from various causes often results in liver fibrosis (LF). Although the liver possesses endogenous tissue-repairing activities, these can be overcome by sustained inflammation and excessive fibrotic scar formation. Advanced LF leads to irreversible cirrhosis and subsequent liver failure and/or hepatic cancer. Here, using the mouse carbon tetrachloride (CCl 4 )-induced LF model, we showed that a single intravenous administration of stem cells derived from human exfoliated deciduous teeth (SHEDs) or of SHED-derived serum-free conditioned medium (SHED-CM) resulted in fibrotic scar resolution. SHED-CM suppressed the gene expression of proinflammatory mediators, such as TNF-α, IL-1β, and iNOS, and eliminated activated hepatic stellate cells by inducing their apoptosis, but protected parenchymal hepatocytes from undergoing apoptosis. In addition, SHED-CM induced tissue-repairing macrophages that expressed high levels of the profibrinolytic factor, matrix metalloproteinase 13. Furthermore, SHED-CM suppressed the CCl 4 -induced apoptosis of primary cultured hepatocytes. SHED-CM contained a high level of hepatocyte growth factor (HGF). Notably, HGF-depleted SHED-CM (dHGF-CM) did not suppress the proinflammatory response or resolve fibrotic scarring. Furthermore, SHED-CM, but not dHGF-CM, inhibited CCl 4 -induced hepatocyte apoptosis. These results suggest that HGF plays a central role in the SHED-CM-mediated resolution of LF. Taken together, our findings suggest that SHED-CM provides multifaceted therapeutic benefits for the treatment of LF. This study demonstrated that a single intravenous administration of stem cells from human exfoliated deciduous teeth (SHEDs) or of the serum-free conditioned medium (CM) derived from SHEDs markedly improved mouse liver fibrosis (LF). SHED-CM suppressed chronic inflammation, eliminated activated hepatic stellate cells by inducing their apoptosis, protected hepatocytes from undergoing apoptosis, and induced

Hepatocyte growth factor induces proliferation and differentiation of multipotent and erythroid hemopoietic progenitors.

PubMed

Galimi, F; Bagnara, G P; Bonsi, L; Cottone, E; Follenzi, A; Simeone, A; Comoglio, P M

1994-12-01

Hepatocyte growth factor (HGF) is a mesenchymal derived growth factor known to induce proliferation and "scattering" of epithelial and endothelial cells. Its receptor is the tyrosine kinase encoded by the c-MET protooncogene. Here we show that highly purified recombinant HGF stimulates hemopoietic progenitors to form colonies in vitro. In the presence of erythropoietin, picomolar concentrations of HGF induced the formation of erythroid burst-forming unit colonies from CD34-positive cells purified from human bone marrow, peripheral blood, or umbilical cord blood. The growth stimulatory activity was restricted to the erythroid lineage. HGF also stimulated the formation of multipotent CFU-GEMM colonies. This effect is synergized by stem cell factor, the ligand of the tyrosine kinase receptor encoded by the c-KIT protooncogene, which is active on early hemopoietic progenitors. By flow cytometry analysis, the receptor for HGF was found to be expressed on the cell surface in a fraction of CD34+ progenitors. Moreover, in situ hybridization experiments showed that HGF receptor mRNA is highly expressed in embryonic erythroid cells (megaloblasts). HGF mRNA was also found to be produced in the embryonal liver. These data show that HGF plays a direct role in the control of proliferation and differentiation of erythroid progenitors, and they suggest that it may be one of the long-sought mediators of paracrine interactions between stromal and hemopoietic cells within the hemopoietic microenvironment.

Blood vessel endothelium-directed tumor cell streaming in breast tumors requires the HGF/C-Met signaling pathway

PubMed Central

Leung, E; Xue, A; Wang, Y; Rougerie, P; Sharma, V P; Eddy, R; Cox, D; Condeelis, J

2017-01-01

During metastasis to distant sites, tumor cells migrate to blood vessels. In vivo, breast tumor cells utilize a specialized mode of migration known as streaming, where a linear assembly of tumor cells migrate directionally towards blood vessels on fibronectin-collagen I-containing extracellular matrix (ECM) fibers in response to chemotactic signals. We have successfully reconstructed tumor cell streaming in vitro by co-plating tumors cells, macrophages and endothelial cells on 2.5 μm thick ECM-coated micro-patterned substrates. We found that tumor cells and macrophages, when plated together on the micro-patterned substrates, do not demonstrate sustained directional migration in only one direction (sustained directionality) but show random bi-directional walking. Sustained directionality of tumor cells as seen in vivo was established in vitro when beads coated with human umbilical vein endothelial cells were placed at one end of the micro-patterned ‘ECM fibers' within the assay. We demonstrated that these endothelial cells supply the hepatocyte growth factor (HGF) required for the chemotactic gradient responsible for sustained directionality. Using this in vitro reconstituted streaming system, we found that directional streaming is dependent on, and most effectively blocked, by inhibiting the HGF/C-Met signaling pathway between endothelial cells and tumor cells. Key observations made with the in vitro reconstituted system implicating C-Met signaling were confirmed in vivo in mammary tumors using the in vivo invasion assay and intravital multiphoton imaging of tumor cell streaming. These results establish HGF/C-Met as a central organizing signal in blood vessel-directed tumor cell migration in vivo and highlight a promising role for C-Met inhibitors in blocking tumor cell streaming and metastasis in vivo, and for use in human trials. PMID:27893712

Hepatocyte Growth Factor Improves the Therapeutic Efficacy of Human Bone Marrow Mesenchymal Stem Cells via RAD51.

PubMed

Lee, Eun Ju; Hwang, Injoo; Lee, Ji Yeon; Park, Jong Nam; Kim, Keun Cheon; Kim, Gi-Hwan; Kang, Chang-Mo; Kim, Irene; Lee, Seo-Yeon; Kim, Hyo-Soo

2018-03-07

Human embryonic stem cell-derived mesenchymal stem cells (hE-MSCs) have greater proliferative capacity than other human mesenchymal stem cells (hMSCs), suggesting that they may have wider applications in regenerative cellular therapy. In this study, to uncover the anti-senescence mechanism in hE-MSCs, we compared hE-MSCs with adult bone marrow (hBM-MSCs) and found that hepatocyte growth factor (HGF) was more abundantly expressed in hE-MSCs than in hBM-MSCs and that it induced the transcription of RAD51 and facilitated its SUMOylation at K70. RAD51 induction/modification by HGF not only increased telomere length but also increased mtDNA replication, leading to increased ATP generation. Moreover, HGF-treated hBM-MSCs showed significantly better therapeutic efficacy than naive hBM-MSCs. Together, the data suggest that the RAD51-mediated effects of HGF prevent hMSC senescence by promoting telomere lengthening and inducing mtDNA replication and function, which opens the prospect of developing novel therapies for liver disease. Copyright © 2017 The American Society of Gene and Cell Therapy. Published by Elsevier Inc. All rights reserved.

Effect of novobiocin on the viability of human gingival fibroblasts (HGF-1)

PubMed Central

2014-01-01

Background Novobiocin is a coumarin antibiotic, which affects also eukaryotic cells inhibiting activity of Heat shock protein 90 (Hsp90). The Hsp90 represents a molecular chaperone critical for stabilization and activation of many proteins, particularly oncoproteins that drive cancer progression. Currently, Hsp90 inhibitors focus a significant attention since they form a potentially new class of drugs in therapy of cancer. However, in the process of tumorigenesis a significant role is played also by the microenvironment of the tumour, and, in particular, by cancer-associated fibroblasts (CAFs). This study aimed at examination of the effect played by novobiocin on viability of human gingival fibroblasts (HGF-1). Methods The studies were conducted using 24 h cultures of human gingival fibroblasts – HGF-1 (CRL-2014) in Chamber Slides, in presence of 0.1, 0.5, 1.0, 2.5 or 5.0 mM novobiocin. Cell viability was evaluated using fluorescence test, ATP assay and LDH release. Results Viability of HGF-1 was drastically reduced after 5 hour treatment with novobiocin in concentrations of 1 mM or higher. In turn, the percentage of LDH-releasing cells after 5 h did not differ from control value although it significantly increased after 10 h incubation with 1 mM and continued to increase till the 20th hour. Conclusions The obtained data indicate that novobiocin may induce death of human gingival fibroblasts. Therefore, application of the Hsp90 inhibitor in neoplastic therapy seems controversial: on one hand novobiocin reduces tumour-associated CAFs but, on the other, it may induce a significant destruction of periodontium. PMID:24887242

Hepatocyte growth factor improves bone regeneration via the bone morphogenetic protein‑2‑mediated NF‑κB signaling pathway.

PubMed

Zhen, Ruixin; Yang, Jianing; Wang, Yu; Li, Yubo; Chen, Bin; Song, Youxin; Ma, Guiyun; Yang, Bo

2018-04-01

Bone regeneration is an important process associated with the treatment of osteonecrosis, which is caused by various factors. Hepatocyte growth factor (HGF) is an active biological factor that has multifunctional roles in cell biology, life sciences and clinical medicine. It has previously been suggested that bone morphogenetic protein (BMP)‑2 exerts beneficial roles in bone formation, repair and angiogenesis in the femoral head. The present study aimed to investigate the benefits and molecular mechanisms of HGF in bone regeneration. The viability of osteoblasts and osteoclasts were studied in vitro. In addition, the expression levels of tumor necrosis factor (TNF)‑α, monocyte chemotactic protein (MCP)‑1, interleukin (IL)‑1 and IL‑6 were detected in a mouse fracture model following treatment with HGF. The expression and activity of nuclear factor (NF)‑κB were also analyzed in osteocytes post‑treatment with HGF. Histological analysis was used to determine the therapeutic effects of HGF on mice with fractures. The migration and differentiation of osteoblasts and osteoclasts were investigated in HGF‑incubated cells. Furthermore, angiogenesis and BMP‑2 expression were analyzed in the mouse fracture model post‑treatment with HGF. The results indicated that HGF regulates the cell viability of osteoblasts and osteoclasts, and also balanced the ratio between osteoblasts and osteoclasts. In addition, HGF decreased the serum expression levels of TNF‑α, MCP‑1, IL‑1 and IL‑6 in experimental mice. The results of a mechanistic analysis demonstrated that HGF upregulated p65, IκB kinase‑β and IκBα expression in osteoblasts from experimental mice. In addition, the expression levels of vascular endothelial growth factor, BMP‑2 receptor, receptor activator of NF‑κB ligand and macrophage colony‑stimulating factor were upregulated by HGF, which may effectively promote blood vessel regeneration, and contribute to the formation and

Mechanisms of Hepatocyte Growth Factor Activation in Cancer Tissues

PubMed Central

Kawaguchi, Makiko; Kataoka, Hiroaki

2014-01-01

Hepatocyte growth factor/scatter factor (HGF/SF) plays critical roles in cancer progression through its specific receptor, MET. HGF/SF is usually synthesized and secreted as an inactive proform (pro-HGF/SF) by stromal cells, such as fibroblasts. Several serine proteases are reported to convert pro-HGF/SF to mature HGF/SF and among these, HGF activator (HGFA) and matriptase are the most potent activators. Increased activities of both proteases have been observed in various cancers. HGFA is synthesized mainly by the liver and secreted as an inactive pro-form. In cancer tissues, pro-HGFA is likely activated by thrombin and/or human kallikrein 1-related peptidase (KLK)-4 and KLK-5. Matriptase is a type II transmembrane serine protease that is expressed by most epithelial cells and is also synthesized as an inactive zymogen. Matriptase activation is likely to be mediated by autoactivation or by other trypsin-like proteases. Recent studies revealed that matriptase autoactivation is promoted by an acidic environment. Given the mildly acidic extracellular environment of solid tumors, matriptase activation may, thus, be accelerated in the tumor microenvironment. HGFA and matriptase activities are regulated by HGFA inhibitor (HAI)-1 (HAI-1) and/or HAI-2 in the pericellular microenvironment. HAIs may have an important role in cancer cell biology by regulating HGF/SF-activating proteases. PMID:25268161

Gene expression profiles of putative biomarker candidates in Mycobacterium avium subsp. paratuberculosis-infected cattle.

PubMed

Park, Hyun-Eui; Shin, Min-Kyoung; Park, Hong-Tae; Jung, Myunghwan; Cho, Yong Il; Yoo, Han Sang

2016-06-01

This study was conducted to analyze the gene expression of prognostic potential biomarker candidates using the whole blood of cattle naturally infected with ITALIC! Mycobacterium aviumsubsp. ITALIC! paratuberculosis(MAP). We conducted real-time PCR to evaluate 23 potential biomarker candidates. Experimental animals were divided into four groups based on fecal MAP PCR and serum ELISA. Seven ( ITALIC! KLRB1, ITALIC! HGF, ITALIC! MPO, ITALIC! LTF, ITALIC! SERPINE1, ITALIC! S100A8and ITALIC! S100A9) genes were up-regulated in fecal MAP-positive cattle and three ( ITALIC! KLRB1, ITALIC! MPOand ITALIC! S100A9) were up-regulated in MAP-seropositive cattle relative to uninfected cattle. In subclinically infected animals, 17 genes ( ITALIC! TFRC, ITALIC! S100A8, ITALIC! S100A9, ITALIC! MPO, ITALIC! GBP6, ITALIC! LTF, ITALIC! KLRB1, ITALIC! SERPINE1, ITALIC! PIGR, ITALIC! IL-10, ITALIC! CXCR3, ITALIC! CD14, ITALIC! MMP9, ITALIC! ELANE, ITALIC! CHI3L1, ITALIC! HPand ITALIC! HGF) were up-regulated compared with the control group. Moreover, six genes ( ITALIC! CXCR3, ITALIC! HP, ITALIC! HGF, ITALIC! LTF, ITALIC! TFRCand ITALIC! GBP6) showed significant differences between experimental groups. Taken together, our data suggest that six genes ( ITALIC! LTF, ITALIC! HGF, ITALIC! HP, ITALIC! CXCR3, ITALIC! GBP6and ITALIC! TFRC) played essential roles in the immune response to MAP during the subclinical stage and therefore might be useful as prognostic biomarkers. © FEMS 2016. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Enhancement of Gastric Ulcer Healing and Angiogenesis by Hepatocyte Growth Factor Gene Mediated by Attenuated Salmonella in Rats.

PubMed

Ha, Xiaoqin; Peng, Junhua; Zhao, Hongbin; Deng, Zhiyun; Dong, Juzi; Fan, Hongyan; Zhao, Yong; Li, Bing; Feng, Qiangsheng; Yang, Zhihua

2017-02-01

The present study developed an oral hepatocyte growth factor (HGF) gene therapy strategy for gastric ulcers treatment. An attenuated Salmonella typhimurium that stably expressed high HGF (named as TPH) was constructed, and the antiulcerogenic effect of TPH was evaluated in a rat model of gastric ulcers that created by acetic acid subserosal injection. From day 5 after injection, TPH (1 × 10⁹ cfu), vehicle (TP, 1 × 10⁹ cfu), or sodium bicarbonate (model control) was administered orally every alternate day for three times. Then ulcer size was measured at day 21 after ulcer induction. The ulcer area in TPH-treated group was 10.56 ± 3.30 mm², which was smaller when compared with those in the TP-treated and model control groups (43.47 ± 4.18 and 56.25 ± 6.38 mm², respectively). A higher level of reepithelialization was found in TPH-treated group and the crawling length of gastric epithelial cells was significantly longer than in the other two groups (P < 0.05). The microvessel density in the ulcer granulation tissues of the TPH-treated rats was 39.9 vessels/mm², which was greater than in the TP-treated and model control rats, with a significant statistical difference. These results suggest that TPH treatment significantly accelerates the healing of gastric ulcers via stimulating proliferation of gastric epithelial cells and enhancing angiogenesis on gastric ulcer site.

Enhancement of Gastric Ulcer Healing and Angiogenesis by Hepatocyte Growth Factor Gene Mediated by Attenuated Salmonella in Rats

PubMed Central

2017-01-01

The present study developed an oral hepatocyte growth factor (HGF) gene therapy strategy for gastric ulcers treatment. An attenuated Salmonella typhimurium that stably expressed high HGF (named as TPH) was constructed, and the antiulcerogenic effect of TPH was evaluated in a rat model of gastric ulcers that created by acetic acid subserosal injection. From day 5 after injection, TPH (1 × 109 cfu), vehicle (TP, 1 × 109 cfu), or sodium bicarbonate (model control) was administered orally every alternate day for three times. Then ulcer size was measured at day 21 after ulcer induction. The ulcer area in TPH-treated group was 10.56 ± 3.30 mm2, which was smaller when compared with those in the TP-treated and model control groups (43.47 ± 4.18 and 56.25 ± 6.38 mm2, respectively). A higher level of reepithelialization was found in TPH-treated group and the crawling length of gastric epithelial cells was significantly longer than in the other two groups (P < 0.05). The microvessel density in the ulcer granulation tissues of the TPH-treated rats was 39.9 vessels/mm2, which was greater than in the TP-treated and model control rats, with a significant statistical difference. These results suggest that TPH treatment significantly accelerates the healing of gastric ulcers via stimulating proliferation of gastric epithelial cells and enhancing angiogenesis on gastric ulcer site. PMID:28049228

The HGF/c-MET Axis as a Critical Driver of Resistance to Androgen Suppression in Metastatic Castrate-Resistant Prostate Cancer

DTIC Science & Technology

2015-10-01

HGF and/or various treatment doses of cabozantinib ( Cabo ) in MET high/AR-negative prostate cancer cells for 24 hours. Subtask 4: Assess whether c...multityrosine kinase inhibitor cabozantinib ( Cabo ) as indicated. Left panel is representative pictures of indicated treatment results by fluorescent...phospho-MET and total MET protein levels. (C) Invasion assays were performed in the presence of HGF and/or various treatment doses of cabozantinib ( Cabo

The effect of hepatocyte growth factor on secretory functions in human eosinophils.

PubMed

Yamauchi, Yumiko; Ueki, Shigeharu; Konno, Yasunori; Ito, Wataru; Takeda, Masahide; Nakamura, Yuka; Nishikawa, Junko; Moritoki, Yuki; Omokawa, Ayumi; Saga, Tomoo; Hirokawa, Makoto

2016-12-01

Hepatocyte growth factor (HGF), originally identified as a potent mitogen for mature hepatocytes, is now recognized as a humoral mediator in inflammatory and immune responses. Previous studies indicated that HGF negatively regulated allergic airway inflammation. In view of eosinophils playing a role in the pathogenesis of asthma, especially in airway remodeling as a rich source of pro-fibrogenic mediators, the effects of HGF on the different types of eosinophil secretory functions were examined in this study. We found that HGF significantly inhibited IL-5-induced secretion of TGF-β and VEGF from human eosinophils. The inhibitory effect is not associated with TGF-β transcription; rather, it is associated with ultrastructural granule emptying and loss of intracellular TGF-β contents, indicating HGF inhibits the process of piecemeal degranulation. The effect of HGF on extracellular trap cell death (ETosis) that mediates cytolytic degranulation was also investigated; however, immobilized IgG- or phorbol myristate acetate-induced ETosis was only minimally attenuated by HGF. These results reveal the effect of HGF on the distinct pathways of eosinophil secretory functions and also provide novel insights into the role of HGF in the pathogenesis of allergic inflammation. Copyright © 2016 Elsevier Ltd. All rights reserved.

Peritumoral stromal remodeling, pattern of invasion and expression of c-met/HGF in advanced squamous cell carcinoma of the cervix uteri, FIGO stages III and IV.

PubMed

Horn, L-C; Hommel, N; Roschlau, U; Bilek, K; Hentschel, B; Einenkel, J

2012-07-01

Different patterns of invasion (PIs) have prognostic impact in several types of cancer and are associated with different grades of peritumoral stromal remodeling, characterized by the desmoplastic stromal response (DSR). One key regulator influencing cellular motility and peritumoral stromal response is c-met/HGF. This study evaluates the association between different PI, peritumoral DSR and its correlation to the expression of c-met/HGF in squamous cell carcinomas of the uterine cervix (CX). 131 advanced stage CX (FIGO III/IV) were re-evaluated histologically regarding PI, using a two-level scoring system. The tumor grows in solid cords/trabeculae in finger-like PI and in very small groups or single cells in spray-like PI. DSR was categorized as none/weak and moderate/strong. The tumors were stained with antibodies against c-met and HGF. The staining of >30% of tumor cells was defined as overexpression. The PI was correlated to the prognostic outcome, different categories of DSR and expression status of c-met and HGF. 66.4% of the tumors showed a finger-like, and 33.6% a spray-like PI. The spray-like PI showed a reduced two-year overall survival when compared to the finger-like PI (14.0% vs. 29.1%, respectively; p=0.012), and was associated with moderate/strong DSR. The majority of the tumors showed overexpression of c-met (85.4%) and HGF (74.8%). There was no correlation between the expression status of c-met/HGF and the FIGO stage, peritumoral DSR or the prognostic outcome. Spray-like PI is of prognostic impact in cervical carcinoma FIGO III/IV and is associated with strong peritumoral stromal remodeling. There is no prognostic impact of the immunohistochemical expression of c-met/HGF in advanced stage cervical carcinomas. Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ding, Ying; Adachi, Hiroaki, E-mail: hadachi-ns@umin.org; Department of Neurology, University of Occupational and Environmental Health School of Medicine, 1-1 Iseigaoka, Yahata-nishi-ku, Kitakyushu 807-8555

Spinal and bulbar muscular atrophy (SBMA) is an inherited motor neuron disease caused by the expansion of a polyglutamine (polyQ)-encoding tract within the androgen receptor (AR) gene. The pathologic features of SBMA are motor neuron loss in the spinal cord and brainstem and diffuse nuclear accumulation and nuclear inclusions of mutant AR in residual motor neurons and certain visceral organs. Hepatocyte growth factor (HGF) is a polypeptide growth factor which has neuroprotective properties. To investigate whether HGF overexpression can affect disease progression in a mouse model of SBMA, we crossed SBMA transgenic model mice expressing an AR gene with anmore » expanded CAG repeat with mice overexpressing HGF. Here, we report that high expression of HGF induces Akt phosphorylation and modestly ameliorated motor symptoms in an SBMA transgenic mouse model treated with or without castration. These findings suggest that HGF overexpression can provide a potential therapeutic avenue as a combination therapy with disease-modifying therapies in SBMA. - Highlights: • HGF overexpression ameliorates the motor phenotypes of the SBMA mouse model. • HGF overexpression induces Akt phosphorylation in the SBMA mouse model. • This is the first report of combination therapy in a mouse model of polyQ diseases.« less

c-Met and its ligand hepatocyte growth factor/scatter factor regulate mature B cell survival in a pathway induced by CD74.

PubMed

Gordin, Maya; Tesio, Melania; Cohen, Sivan; Gore, Yael; Lantner, Frida; Leng, Lin; Bucala, Richard; Shachar, Idit

2010-08-15

The signals regulating the survival of mature splenic B cells have become a major focus in recent studies of B cell immunology. Durable B cell persistence in the periphery is dependent on survival signals that are transduced by cell surface receptors. In this study, we describe a novel biological mechanism involved in mature B cell homeostasis, the hepatocyte growth factor/scatter factor (HGF)/c-Met pathway. We demonstrate that c-Met activation by HGF leads to a survival cascade, whereas its blockade results in induction of mature B cell death. Our results emphasize a unique and critical function for c-Met signaling in the previously described macrophage migration inhibitory factor/CD74-induced survival pathway. Macrophage migration inhibitory factor recruits c-Met to the CD74/CD44 complex and thereby enables the induction of a signaling cascade within the cell. This signal results in HGF secretion, which stimulates the survival of the mature B cell population in an autocrine manner. Thus, the CD74-HGF/c-Met axis defines a novel physiologic survival pathway in mature B cells, resulting in the control of the humoral immune response.

Renoprotective effects of hepatocyte growth factor in the stenotic kidney

PubMed Central

Stewart, Nicholas

2013-01-01

Renal microvascular (MV) damage and loss contribute to the progression of renal injury in renal artery stenosis (RAS). Hepatocyte growth factor (HGF) is a powerful angiogenic and antifibrotic cytokine that we showed to be decreased in the stenotic kidney. We hypothesized that renal HGF therapy will improve renal function mainly by protecting the renal microcirculation. Unilateral RAS was induced in 15 pigs. Six weeks later, single-kidney RBF and GFR were quantified in vivo using multidetector computed tomography (CT). Then, intrarenal rh-HGF or vehicle was randomly administered into the stenotic kidney (RAS, n = 8; RAS+HGF, n = 7). Pigs were observed for 4 additional weeks before CT studies were repeated. Renal MV density was quantified by 3D micro-CT ex vivo and histology, and expression of angiogenic and inflammatory factors, apoptosis, and fibrosis was determined. HGF therapy improved RBF and GFR compared with vehicle-treated pigs. This was accompanied by improved renal expression of angiogenic cytokines (VEGF, p-Akt) and tissue-healing promoters (SDF-1, CXCR4, MMP-9), reduced MV remodeling, apoptosis, and fibrosis, and attenuated renal inflammation. However, HGF therapy did not improve renal MV density, which was similarly reduced in RAS and RAS+HGF compared with controls. Using a clinically relevant animal model of RAS, we showed novel therapeutic effects of a targeted renal intervention. Our results show distinct actions on the existing renal microcirculation and promising renoprotective effects of HGF therapy in RAS. Furthermore, these effects imply plasticity of the stenotic kidney to recuperate its function and underscore the importance of MV integrity in the progression of renal injury in RAS. PMID:23269649

Correlation between increasing tissue ischemia and circulating levels of angiogenic growth factors in peripheral artery disease.

PubMed

Jalkanen, Juho; Hautero, Olli; Maksimow, Mikael; Jalkanen, Sirpa; Hakovirta, Harri

2018-04-21

The aim of the present study was to assess the circulating levels of vascular endothelial growth factor (VEGF) and other suggested therapeutic growth factors with the degree of ischemia in patients with different clinical manifestations of peripheral arterial disease (PAD) according to the Rutherford grades. The study cohort consists of 226 consecutive patients admitted to a Department of Vascular Surgery for elective invasive procedures. PAD patients were grouped according to the Rutherford grades after a clinical assessment. Ankle-brachial pressure indices (ABI) and absolute toe pressure (TP) values were measured. Serum levels of circulating VEGF, hepatocyte growth factor (HGF), basic fibroblast growth factor (bFGF), and platelet derived growth factor (PDGF) were measured from serum and analysed against Rutherford grades and peripheral hemodynamic measurements. The levels of VEGF (P = 0.009) and HGF (P < 0.001) increased significantly as the ischaemic burden became more severe according to the Rutherford grades. PDGF behaved in opposite manner and declined along increasing Rutherford grades (P = 0.004). A significant, inverse correlations between Rutherford grades was detected as follows; VEGF (Pearson's correlation = 0.183, P = 0.004), HGF (Pearson's correlation = 0.253, P < 0.001), bFGF (Pearson's correlation = 0.169, P = 0.008) and PDGF (Pearson's correlation = 0.296, P < 0.001). In addition, VEGF had a clear direct negative correlation with ABI (Pearson's correlation -0.19, P = 0.009) and TP (Pearson's correlation -0.20, P = 0.005) measurements. Our present observations show that the circulating levels of VEGF and other suggested therapeutic growth factors are significantly increased along with increasing ischemia. These findings present a new perspective to anticipated positive effects of gene therapies utilizing VEGF, HGF, and bFGF, because the levels of these growth factors are endogenously high in end

Decreased expression of hepatocyte growth factor in the nitrofen model of congenital diaphragmatic hernia.

PubMed

Takahashi, Toshiaki; Friedmacher, Florian; Zimmer, Julia; Puri, Prem

2016-10-01

Pleuroperitoneal folds (PPFs) are essential for normal diaphragmatic development, representing the only source of the diaphragm's muscle connective tissue. Hepatocyte growth factor (Hgf), which is secreted in PPFs, plays a crucial role in the formation of the muscular diaphragmatic components by regulating the migration of myogenic progenitor cells into the primordial diaphragm. Hgf is also a known downstream target of Gata4 and it has been demonstrated that the expression of Hgf was significantly downregulated in PPF cells of Gata4 knockouts with congenital diaphragmatic hernia (CDH). Furthermore, mutations in PPF-derived cells have been shown to result in CDH. We hypothesized that Hgf expression is decreased in developing diaphragms of fetal rats with nitrofen-induced CDH. Timed-pregnant rats were exposed to either nitrofen or vehicle on gestational day 9 (D9). Fetuses were harvested on selected time-points D13, D15 and D18. Dissected diaphragms (n = 72) were divided into control and nitrofen-exposed specimens (n = 12 per time-point and experimental group, respectively). Diaphragmatic gene expression of Hgf was analyzed by qRT-PCR. Immunofluorescence double staining for Hgf and the mesenchymal marker Gata4 or muscular progenitor marker Myogenin was performed to evaluate protein expression and localization in fetal diaphragms. Relative mRNA expression of Hgf was significantly downregulated in PPFs of nitrofen-exposed fetuses on D13 (3.08 ± 1.46 vs. 5.24 ± 1.93; p < 0.05), developing diaphragms of nitrofen-exposed fetuses on D15 (2.01 ± 0.79 vs. 4.10 ± 1.50; p < 0.05) and fully muscularized diaphragms of nitrofen-exposed fetuses on D18 (1.60 ± 0.78 vs. 3.21 ± 1.89; p < 0.05) compared to controls. Confocal laser scanning microscopy revealed markedly diminished diaphragmatic immunofluorescence of Hgf in nitrofen-exposed fetuses on D13, D15 and D18 compared to controls, which was associated with disruptions in muscle connective tissue

Lipopolysaccharide potentiates the effect of hepatocyte growth factor on hepatocyte replication in rats by augmenting AP-1 activity.

PubMed

Gao, C; Jokerst, R; Gondipalli, P; Cai, S R; Kennedy, S; Flye, M W; Ponder, K P

1999-12-01

The liver regenerates by replication of differentiated hepatocytes after damage or removal of part of the liver. Although several growth factors and signaling pathways are activated during regeneration, it is unclear as to which of these are essential for hepatocyte replication. We show here that low- (1 mg/kg) and high- (10 mg/kg) dose hepatocyte growth factor (HGF) induced replication of 2.1% and 11.1% of hepatocytes in rats, respectively. Lipopolysaccharide (LPS), an inducer of the acute phase response, augmented hepatocyte replication in response to low- and high-dose HGF by 4- and 2-fold, respectively. HGF alone induced moderate levels of c-Jun-N-terminal kinase (JNK) and p44/p42 mitogen-activated protein kinase (MAPK), resulting in moderate levels of AP-1-DNA binding activity. The combination of LPS + HGF increased JNK and AP-1-DNA binding activity more than levels seen with LPS or HGF alone. The activation of Stat3 that was observed after administration of LPS + HGF, but not HGF alone, could contribute to increased transcription of AP-1 components. Because phosphorylation of the c-Jun component of AP-1 by JNK increases its ability to activate transcription, the AP-1 in hepatocytes from animals treated with LPS + HGF may be more active than in rats treated with LPS or HGF alone. LPS may contribute to hepatocyte replication by potentiating the effect of HGF on the activation of both AP-1-DNA binding and transcriptional activity.

L-ascorbic acid 2-phosphate and fibroblast growth factor-2 treatment maintains differentiation potential in bone marrow-derived mesenchymal stem cells through expression of hepatocyte growth factor.

PubMed

Bae, Sung Hae; Ryu, Hoon; Rhee, Ki-Jong; Oh, Ji-Eun; Baik, Soon Koo; Shim, Kwang Yong; Kong, Jee Hyun; Hyun, Shin Young; Pack, Hyun Sung; Im, Changjo; Shin, Ha Cheol; Kim, Yong Man; Kim, Hyun Soo; Eom, Young Woo; Lee, Jong In

2015-04-01

l-ascorbic acid 2-phosphate (Asc-2P) acts as an antioxidant and a stimulator of hepatocyte growth factor (HGF) production. Previously, we reported that depletion of growth factors such as fibroblast growth factor (FGF)-2, epidermal growth factor (EGF), FGF-4 and HGF during serial passage could induce autophagy, senescence and down-regulation of stemness (proliferation via FGF-2/-4 and differentiation via HGF). In this study, we investigated the proliferation and differentiation potential of BMSCs by FGF-2 and Asc-2P. Co-treatment with FGF-2 and Asc-2P induced optimal proliferation of BMSCs and increased the accumulation rate of BMSC numbers during a 2-month culture period. Moreover, differentiation potential was maintained by co-treatment with FGF-2 and Asc-2P via HGF expression. Adipogenic differentiation potential by FGF-2 and Asc-2P was dramatically suppressed by c-Met inhibitors (SU11274). These data suggest that co-treatment with FGF-2 and Asc-2P would be beneficial in obtaining BMSCs that possess "stemness" during long-term culture.

Combination strategy targeting VEGF and HGF/c-met in human renal cell carcinoma models.

PubMed

Ciamporcero, Eric; Miles, Kiersten Marie; Adelaiye, Remi; Ramakrishnan, Swathi; Shen, Li; Ku, ShengYu; Pizzimenti, Stefania; Sennino, Barbara; Barrera, Giuseppina; Pili, Roberto

2015-01-01

Alternative pathways to the VEGF, such as hepatocyte growth factor or HGF/c-met, are emerging as key players in tumor angiogenesis and resistance to anti-VEGF therapies. The aim of this study was to assess the effects of a combination strategy targeting the VEGF and c-met pathways in clear cell renal cell carcinoma (ccRCC) models. Male SCID mice (8/group) were implanted with 786-O tumor pieces and treated with either a selective VEGF receptor tyrosine kinase inhibitor, axitinib (36 mg/kg, 2×/day); a c-met inhibitor, crizotinib (25 mg/kg, 1×/day); or combination. We further tested this drug combination in a human ccRCC patient-derived xenograft, RP-R-01, in both VEGF-targeted therapy-sensitive and -resistant models. To evaluate the resistant phenotype, we established an RP-R-01 sunitinib-resistant model by continuous sunitinib treatment (60 mg/kg, 1×/day) of RP-R-01-bearing mice. Treatment with single-agent crizotinib reduced tumor vascularization but failed to inhibit tumor growth in either model, despite also a significant increase of c-met expression and phosphorylation in the sunitinib-resistant tumors. In contrast, axitinib treatment was effective in inhibiting angiogenesis and tumor growth in both models, with its antitumor effect significantly increased by the combined treatment with crizotinib, independently from c-met expression. Combination treatment also induced prolonged survival and significant tumor growth inhibition in the 786-O human RCC model. Overall, our results support the rationale for the clinical testing of combined VEGF and HGF/c-met pathway blockade in the treatment of ccRCC, both in first- and second-line setting. ©2014 American Association for Cancer Research.

Imbalance in the pro-hepatocyte growth factor activation system in bleomycin-induced lung fibrosis in mice.

PubMed

Phin, Sophie; Marchand-Adam, Sylvain; Fabre, Aurélie; Marchal-Somme, Joëlle; Bantsimba-Malanda, Claudie; Kataoka, Hiroaki; Soler, Paul; Crestani, Bruno

2010-03-01

Hepatocyte growth factor (HGF) is a growth factor for alveolar epithelial cells. Activation of pro-HGF to HGF is regulated by the HGF activator (HGFA), a serine protease, and a specific inhibitor (HGFA inhibitor-1, HAI-1). An imbalance in the HGFA/HAI-1 system might contribute to lung fibrosis. Pro-HGF activation capacity from bronchoalveolar lavage (BAL) fluid was evaluated 3, 7, and 14 days after the intratracheal bleomycin injection (Bleo) in mice with or without thrombin. BAL fluid from naïve mice was used as control. HGFA and HAI-1 mRNA were evaluated by QPCR in the whole lung or by Western blot in BAL fluid. BAL fluid from control mice and Bleo mice activated pro-HGF in vitro at a similar degree. Thrombin accelerated proHGF activation by Bleo BAL on Day 3 and Day 7, but not on Day 14, or in control BAL. Incubation of pro-HGF with BAL from Bleo Day 3 and Day 7 mice increased phosphorylation of HGFR on A549 cells. Thrombin-induced pro-HGF activation was inhibited by an anti-HGFA antibody and accelerated by an anti-HAI-1 antibody. Active HGFA was not detected in control BAL and was strongly induced in Bleo BAL. HGFA concentrations were higher on Day 3 and Day 7 than on Day 14. HAI-1 was detected at low levels in control BAL and increased strongly by Day 3 with stable concentrations until Day 14. By demonstrating an imbalance between HGFA and HAI-1 expression in BAL fluid, our results highlight a defective thrombin-dependent proHGF activation system at the fibrotic phase of bleomycin-induced pulmonary fibrosis.

A novel mechanism by which hepatocyte growth factor blocks tubular epithelial to mesenchymal transition.

PubMed

Yang, Junwei; Dai, Chunsun; Liu, Youhua

2005-01-01

Hepatocyte growth factor (HGF) is a potent antifibrotic cytokine that blocks tubular epithelial to mesenchymal transition (EMT) induced by TGF-beta1. However, the underlying mechanism remains largely unknown. This study investigated the signaling events that lead to HGF blockade of the TGF-beta1-initiated EMT. Incubation of human kidney epithelial cells HKC with HGF only marginally affected the expression of TGF-beta1 and its type I and type II receptors, suggesting that disruption of TGF-beta1 signaling likely plays a critical role in mediating HGF inhibition of TGF-beta1 action. However, HGF neither affected TGF-beta1-induced Smad-2 phosphorylation and its subsequent nuclear translocation nor influenced the expression of inhibitory Smad-6 and -7 in tubular epithelial cells. HGF specifically induced the expression of Smad transcriptional co-repressor SnoN but not Ski and TG-interacting factor at both mRNA and protein levels in HKC cells. SnoN physically interacted with activated Smad-2 by forming transcriptionally inactive complex and overrode the profibrotic action of TGF-beta1. In vivo, HGF did not affect Smad-2 activation and its nuclear accumulation in tubular epithelium, but it restored SnoN protein abundance in the fibrotic kidney in obstructive nephropathy. Hence, HGF blocks EMT by antagonizing TGF-beta1's action via upregulating Smad transcriptional co-repressor SnoN expression. These findings not only identify a novel mode of interaction between the signals activated by HGF receptor tyrosine kinase and TGF-beta receptor serine/threonine kinases but also illustrate the feasibility of confining Smad activity as an effective strategy for blocking renal fibrosis.

Cloning of a cancer cell-producing hepatocyte growth factor, vascular endothelial growth factor, and interleukin-8 from gastric cancer cells.

PubMed

Iwai, Mineko; Matsuda, Masahiko; Iwai, Yoshiaki

2003-01-01

A cell colony (IM95m) that produces hepatocyte growth factor (HGF), vascular endothelial growth factor (VEGF), and interleukin-8 (IL-8) was cloned from gastric cancer cells (IM95 cell line). In culture medium, the highest levels of HGF, VEGF, and IL-8 were about 1.1, 0.9, and 0.17 ng/ml culture medium at 3 d from 10(5) cells. IM95m may be useful in elucidating the role of tumor cells in angiogenesis.

A novel rabbit anti-hepatocyte growth factor monoclonal neutralizing antibody inhibits tumor growth in prostate cancer cells and mouse xenografts

DOE Office of Scientific and Technical Information (OSTI.GOV)

Yu, Yanlan; Chen, Yicheng; Ding, Guoqing

The hepatocyte growth factor and its receptor c-Met are correlated with castration-resistance in prostate cancer. Although HGF has been considered as an attractive target for therapeutic antibodies, the lack of cross-reactivity of monoclonal antibodies with human/mouse HGFs is a major obstacle in preclinical developments. We generated a panel of anti-HGF RabMAbs either blocking HGF/c-Met interaction or inhibiting c-Met phosphorylation. We selected one RabMAb with mouse cross-reactivity and demonstrated that it blocked HGF-stimulated downstream activation in PC-3 and DU145 cells. Anti-HGF RabMAb inhibited not only the growth of PC-3 cells but also HGF-dependent proliferation in HUVECs. We further demonstrated the efficacymore » and potency of the anti-HGF RabMAb in tumor xenograft mice models. Through these in vitro and in vivo experiments, we explored a novel therapeutic antibody for advanced prostate cancer. - Highlights: • HGF is an attractive target for castration-refractory prostate cancer. • We generated and characterized a panel of anti-HGF rabbit monoclonal antibodies. • More than half of these anti-HGF RabMAbs was cross-reactive with mouse HGF. • Anti-HGF RabMAb blocks HGF-stimulated phosphorylation and cell growth in vitro. • Anti-HGF RabMAb inhibits tumor growth and angiogenesis in xenograft mice.« less

Effects of hepatocyte growth factor in myocarditis rats induced by immunization with porcine cardiac myosin

PubMed Central

Nakano, Jota; Marui, Akira; Muranaka, Hiroyuki; Masumoto, Hidetoshi; Noma, Hisashi; Tabata, Yasuhiko; Ido, Akio; Tsubouchi, Hirohito; Ikeda, Tadashi; Sakata, Ryuzo

2014-01-01

OBJECTIVES Myocarditis is considered one of the major causes of dilated cardiomyopathy. Hepatocyte growth factor (HGF) has pleiotropic activities that promote tissue regeneration and facilitate functional improvement of injured tissue. We investigated whether the epicardial sustained-release of HGF, using gelatin hydrogel sheets, improves cardiac function in a chronic myocarditis rat model. METHODS Six weeks after Lewis rats were immunized with porcine cardiac myosin to establish autoimmune myocarditis, HGF- or normal saline (NS)-incorporated gelatin hydrogel sheets were applied to the epicardium (G-HGF and G-NS, respectively). At either 2 or 4 weeks after treatment, these were compared with the Control myocarditis group. Cardiac function was evaluated by echocardiography and cardiac catheterization. Development of fibrosis was determined by histological study and expression of transforming growth factor-β1 (TGF-β1). Bax and Bcl-2 levels were measured to evaluate apoptotic activity. RESULTS At both points, fractional shortening and end-systolic elastance were higher in the G-HGF group than in the Control and G-NS groups (P < 0.01). Fractional shortening at 2 weeks of each group were as follows: 31.0 ± 0.9%, 24.8 ± 2.7% and 48.6 ± 2.6% (Control, G-NS and G-HGF, respectively). The ratio of the fibrotic area of the myocardium was lower in the G-HGF group than in the Control and G-NS groups at 2 weeks (G-HGF, 8.8 ± 0.9%; Control, 17.5 ± 0.2%; G-NS, 15.6 ± 0.7%; P < 0.01). The ratio at 4 weeks was lower in the G-HGF group than in the G-NS group (10.9 ± 1.4% vs 18.5 ± 1.3%; P < 0.01). The mRNA expression of TGF-β1 in the G-HGF group was lower than in the Control group at 2 weeks (0.6 ± 0.1 vs 1.1 ± 0.2) and lower than that in the G-NS group at 4 weeks (0.7 ± 0.1 vs 1.3 ± 0.2). The Bax-to-Bcl-2 ratios at both points were lower in the G-HGF group than in the Control group. CONCLUSIONS Sustained-released HGF markedly improves cardiac function in chronic

Pro-osteogenic effects of fibrin glue in treatment of avascular necrosis of the femoral head in vivo by hepatocyte growth factor-transgenic mesenchymal stem cells

PubMed Central

2014-01-01

Background Autologous transplantation of modified mesenchymal stem cells (MSCs) is a promising candidate for the treatment of the refractory clinical disease, avascular necrosis of the femoral head (ANFH). Our previous attempts by compounding MSCs with medical fibrin glue to treat ANFH in animal model have achieved excellent effects. However, the underlying molecular mechanism is unclear, especially on the transgenic gene expression. Methods Rabbit MSCs were isolated and compounded with fibrin glue. Following degrading of fibrin glue, proliferation, viability, expression of transgenic hepatocyte growth factor gene as well as osteogenic differentiation of MSCs were evaluated together with that of uncompounded MSCs. Fibrin glue-compounded MSCs were transplanted into the lesion of ANFH model, and the therapeutic efficacy was compared with uncompounded MSCs. One-Way ANOVA was used to determine the statistical significance among treatment groups. Results Fibrin glue compounding will not affect molecular activities of MSCs, including hepatocyte growth factor (HGF) secretion, cell proliferation and viability, and osteogenic differentiation in vitro. When applying fibrin glue-compounded MSCs for the therapy of ANFH in vivo, fibrin glue functioned as a drug delivery system and provided a sustaining microenvironment for MSCs which helped the relatively long-term secretion of HGF in the femoral head lesion and resulted in improved therapeutic efficacy when compared with uncompounded MSCs as indicated by hematoxylin-eosin staining and immunohistochemistry of osteocalcin, CD105 and HGF. Conclusion Transplantation of fibrin glue-compounding MSCs is a promising novel method for ANFH therapy. PMID:24885252

Cancer-associated fibroblasts promote epithelial-mesenchymal transition and EGFR-TKI resistance of non-small cell lung cancers via HGF/IGF-1/ANXA2 signaling.

PubMed

Yi, Yanmei; Zeng, Shanshan; Wang, Zhaotong; Wu, Minhua; Ma, Yuanhuan; Ye, Xiaoxia; Zhang, Biao; Liu, Hao

2018-03-01

The involvement of the tumor stromal cells in acquired resistance of non-small cell lung cancers (NSCLCs) to tyrosine kinase inhibitors (TKIs) has previously been reported, but the precise mechanism remains unclear. In the present study, we investigated the role and mechanism underlying Cancer-associated fibroblasts (CAFs) in TKI resistance of NSCLCs. In vitro and in vivo experiments showed that HCC827 and PC9 cells, non-small cell lung cancer cells with EGFR-activating mutations, became resistant to the EGFR-TKI gefitinib when cultured with CAFs isolated from NSCLC tissues. Moreover, we showed that CAFs could induce epithelial-mesenchymal transition (EMT) phenotype of HCC827 and PC9 cells, with an associated change in the expression of epithelial to mesenchymal transition markers. Using proteomics-based method, we identified that CAFs significantly increased the expression of the Annexin A2 (ANXA2). More importantly, knockdown of ANXA2 completely reversed EMT phenotype and gefitinib resistance induced by CAFs. Furthermore, we found that CAFs increased the expression and phosphorylation of ANXA2 by secretion of growth factors HGF and IGF-1 and by activation of the corresponding receptors c-met and IGF-1R. Dual inhibition of HGF/c-met and IGF-1/IGF-1R pathways could significantly suppress ANXA2, and markedly reduced CAFs-induced EMT and gefitinib resistance. Taken together, these findings indicate that CAFs promote EGFR-TKIs resistance through HGF/IGF-1/ANXA2/EMT signaling and may be an ideal therapeutic target in NSCLCs with EGFR-activating mutations. Copyright © 2017 Elsevier B.V. All rights reserved.

Does Platelet-Rich Plasma Freeze-Thawing Influence Growth Factor Release and Their Effects on Chondrocytes and Synoviocytes?

PubMed Central

Cavallo, Carola; Cenacchi, Annarita; Facchini, Andrea; Grigolo, Brunella; Kon, Elizaveta; Mariani, Erminia; Pratelli, Loredana; Marcacci, Maurilio

2014-01-01

PRP cryopreservation remains a controversial point. Our purpose was to investigate the effect of freezing/thawing on PRP molecule release, and its effects on the metabolism of chondrocytes and synoviocytes. PRP was prepared from 10 volunteers, and a half volume underwent one freezing/thawing cycle. IL-1β, HGF, PDGF AB/BB, TGF-β1, and VEGF were assayed 1 hour and 7 days after activation. Culture media of chondrocytes and synoviocytes were supplemented with fresh or frozen PRP, and, at 7 days, proliferation, gene expression, and secreted proteins levels were evaluated. Results showed that in the freeze-thawed PRP the immediate and delayed molecule releases were similar or slightly lower than those in fresh PRP. TGF-β1 and PDGF AB/BB concentrations were significantly reduced after freezing both at 1 hour and at 7 days, whereas HGF concentration was significantly lower in frozen PRP at 7 days. In fresh PRP IL-1β and HGF concentrations underwent a significant further increase after 7 days. Similar gene expression was found in chondrocytes cultured with both PRPs, whereas in synoviocytes HGF gene expression was higher in frozen PRP. PRP cryopreservation is a safe procedure, which sufficiently preserves PRP quality and its ability to induce proliferation and the production of ECM components in chondrocytes and synoviocytes. PMID:25136613

Therapeutic Angiogenesis by Gene Therapy for Critical Limb Ischemia: Choice of Biological Agent.

PubMed

Sanada, Fumihiro; Taniyama, Yoshiaki; Azuma, Junya; Yuka, Ikeda-Iwabe; Kanbara, Yasuhiro; Iwabayashi, Masaaki; Rakugi, Hiromi; Morishita, Ryuichi

2014-04-01

Peripheral artery disease (PAD) is caused by atherosclerosis, hardening and narrowing arteries over time due to buildup of fatty deposit in vascular bed called plaque. Severe blockage of an artery of the lower extremity markedly reduce blood flow, resulting in critical limb ischemia (CLI) manifested by a variety of clinical syndromes including rest pain in the feet or toes, ulcer and gangrene with infection. Despite significant advances in clinical care and interventions for revascularization, patients with CLI remain at high risk for amputation and cardiovascular death. To overcome this unmet need, therapeutic angiogenesis using angiogenic growth factors has evolved in an attempt to increase blood flow in ischemic limb. Initial animal studies and phase I clinical trials with vascular endothelial growth factor (VEGF) or fibroblast growth factor (FGF) demonstrated promising results, inspiring scientists to progress forward. However, more rigorous phase II and III clinical trials have failed to demonstrate beneficial effects of these angiogenic growth factors to date. Recently, two multicenter, double-blind, placebo-controlled clinical trials in Japan (phase III) and US (phase II) demonstrated that hepatocyte growth factor (HGF) gene therapy for CLI significant improved primary end points and tissue oxygenation up to two years in comparison to placebo. These clinical results implicate a distinct action of HGF on cellular processes involved in vascular remodeling under pathological condition. This review presents data from phase I-III clinical trials of therapeutic angiogenesis by gene therapy in patients with PAD. Further, we discuss the potential explanation for the success or failure of clinical trials in the context of the biological mechanisms underlying angiogenesis and vascular remodeling, including cellular senescence, inflammation, and tissue fibrosis.

Progesterone dose-dependently modulates hepatocyte growth factor production in 3T3-L1 mouse preadipocytes.

PubMed

Ito, Tomoki; Yamaji, Daisuke; Kamikawa, Akihiro; Abd Eldaim, Mabrouk Attia; Okamatsu-Ogura, Yuko; Terao, Akira; Saito, Masayuki; Kimura, Kazuhiro

2017-08-30

It is well documented that estrogen is predominant inducer of hepatocyte growth factor (HGF) in a variety of cell types. However, the effect of progesterone (P) remains to be elusive. Thus, in the present study, we examined the effect of P and combined effect of P and 17β-estradiol (E2) on HGF expression and production in 3T3-L1 fibroblastic preadipocytes and mature adipocytes, as a model of stromal cells. Northern blot analysis showed that hgf mRNA expressed in preadipocytes was notably higher than that of mature adipocytes, and increased by treatment of preadipocytes with E2 or 10 nM P, but not with 1,000 nM P. The E2-induced hgf mRNA expression was enhanced by 10 nM P, but suppressed by 1,000 nM P. Western blot analysis revealed that biological active forms of HGF protein was found in the preadipocyte culture medium, while the lesser amount of HGF precursor protein was detected in the mature adipocyte culture medium. The amounts of HGF were changed dependently on the hgf mRNA expression levels. These results indicate that HGF production is intricately regulated by E2 and P at the transcriptional levels in 3T3-L1 cells, and may explain the changes in the HGF production during the mammary gland development, especially decrease in HGF expression during pregnancy when P concentration is high.

Phosphorylation of hepatocyte growth factor receptor and epidermal growth factor receptor of human hepatocytes can be maintained in a (3D) collagen sandwich culture system.

PubMed

Engl, Tobias; Boost, Kim A; Leckel, Kerstin; Beecken, Wolf-Dietrich; Jonas, Dietger; Oppermann, Elsie; Auth, Marcus K H; Schaudt, André; Bechstein, Wolf-Otto; Blaheta, Roman A

2004-08-01

In vitro culture models that employ human liver cells could be potent tools for predictive studies on drug toxicity and metabolism in the pharmaceutical industry. However, an adequate receptor responsiveness is necessary to allow intracellular signalling and metabolic activity. We tested the ability of three-dimensionally arranged human hepatocytes to respond to the growth factors hepatocyte growth factor (HGF) or epidermal growth factor (EGF). Isolated adult human hepatocytes were cultivated within a three-dimensional collagen gel (sandwich) or on a two-dimensional collagen matrix. Cells were treated with HGF or EGF and expression and phosphorylative activity of HGF receptors (HGFr, c-met) or EGF receptors (EGFr) were measured by flow cytometry and Western blot. Increasing HGFr and EGFr levels were detected in hepatocytes growing two-dimensionally. However, both receptors were not activated in presence of growth factors. In contrast, when hepatocytes were plated within a three-dimensional matrix, HGFr and EGFr levels remained constantly low. However, both receptors became strongly phosphorylated by soluble HGF or EGF. We conclude that cultivation of human hepatocytes in a three-dimensionally arranged in vitro system allows the maintenance of specific functional activities. The necessity of cell dimensionality for HGFr and EGFr function should be considered when an adequate in vitro system has to be introduced for drug testing.

Changes in serum growth factors in stroke rehabilitation patients and their relation to hemiparesis improvement.

PubMed

Okazaki, Hideto; Beppu, Hidehiko; Mizutani, Kenmei; Okamoto, Sayaka; Sonoda, Shigeru

2014-07-01

Predicting recovery from hemiparesis after stroke is important for rehabilitation. A few recent studies reported that the levels of some growth factors shortly after stroke were positively correlated with the clinical outcomes during the chronic phase. The aim of this study was to examine the relationships between the serum levels of growth factors (vascular endothelial growth factor [VEGF], insulin-like growth factor-I [IGF-I], and hepatocyte growth factor [HGF]) and improvement in hemiparesis in stroke patients who received rehabilitation in a postacute rehabilitation hospital. Subjects were 32 stroke patients (cerebral infarction: 21 and intracerebral hemorrhage [ICH]: 11). We measured serum levels of VEGF, IGF-I, and HGF and 5 items of the Stroke Impairment Assessment Set (SIAS) for hemiparesis on admission and at discharge. Age-matched healthy subjects (n=15) served as controls. Serum levels of VEGF and HGF in cerebral infarct patients on admission were higher than those in control subjects, and the serum levels of IGF-I in stroke patients were lower than those in controls. The level of HGF in ICH patients on admission was negatively correlated with gains in SIAS, and higher outliers in HGF concentration were correlated with lower gains in SIAS. Focusing on the extremely high levels of these factors may be a predictor of the low recovery from hemiparesis after stroke. Copyright © 2014 National Stroke Association. Published by Elsevier Inc. All rights reserved.

Serum Hepatocyte Growth Factor Is Probably Associated With 3-Month Prognosis of Acute Ischemic Stroke.

PubMed

Zhu, Zhengbao; Xu, Tan; Guo, Daoxia; Huangfu, Xinfeng; Zhong, Chongke; Yang, Jingyuan; Wang, Aili; Chen, Chung-Shiuan; Peng, Yanbo; Xu, Tian; Wang, Jinchao; Sun, Yingxian; Peng, Hao; Li, Qunwei; Ju, Zhong; Geng, Deqin; Chen, Jing; Zhang, Yonghong; He, Jiang

2018-02-01

Serum hepatocyte growth factor (HGF) is positively associated with poor prognosis of heart failure and myocardial infarction, and it can also predict the risk of ischemic stroke in population. The goal of this study was to investigate the association between serum HGF and prognosis of ischemic stroke. A total of 3027 acute ischemic stroke patients were included in this post hoc analysis of the CATIS (China Antihypertensive Trial in Acute Ischemic Stroke). The primary outcome was composite outcome of death or major disability (modified Rankin Scale score ≥3) within 3 months. After multivariate adjustment, elevated HGF levels were associated with an increased risk of primary outcome (odds ratio, 1.50; 95% confidence interval, 1.10-2.03; P trend =0.015) when 2 extreme quartiles were compared. Each SD increase of log-transformed HGF was associated with 14% (95% confidence interval, 2%-27%) increased risk of primary outcome. Adding HGF quartiles to a model containing conventional risk factors improved the predictive power for primary outcome (net reclassification improvement: 17.50%, P <0.001; integrated discrimination index: 0.23%, P =0.022). The association between serum HGF and primary outcome could be modified by heparin pre-treatment ( P interaction =0.001), and a positive linear dose-response relationship between HGF and primary outcome was observed in patients without heparin pre-treatment ( P linearity <0.001) but not in those with heparin pre-treatment. Serum HGF levels were higher in the more severe stroke at baseline, and elevated HGF levels were probably associated with 3-month poor prognosis independently of stroke severity among ischemic stroke patients, especially in those without heparin pre-treatment. Further studies from other samples of ischemic stroke patients are needed to validate our findings. © 2018 American Heart Association, Inc.

Hepatocyte growth factor/c-MET axis-mediated tropism of cord blood-derived unrestricted somatic stem cells for neuronal injury.

PubMed

Trapp, Thorsten; Kögler, Gesine; El-Khattouti, Abdelouahid; Sorg, Rüdiger V; Besselmann, Michael; Föcking, Melanie; Bührle, Christian P; Trompeter, Ingo; Fischer, Johannes C; Wernet, Peter

2008-11-21

An under-agarose chemotaxis assay was used to investigate whether unrestricted somatic stem cells (USSC) that were recently characterized in human cord blood are attracted by neuronal injury in vitro. USSC migrated toward extracts of post-ischemic brain tissue of mice in which stroke had been induced. Moreover, apoptotic neurons secrete factors that strongly attracted USSC, whereas necrotic and healthy neurons did not. Investigating the expression of growth factors and chemokines in lesioned brain tissue and neurons and of their respective receptors in USSC revealed expression of hepatocyte growth factor (HGF) in post-ischemic brain and in apoptotic but not in necrotic neurons and of the HGF receptor c-MET in USSC. Neuronal lesion-triggered migration was observed in vitro and in vivo only when c-MET was expressed at a high level in USSC. Neutralization of the bioactivity of HGF with an antibody inhibited migration of USSC toward neuronal injury. This, together with the finding that human recombinant HGF attracts USSC, document that HGF signaling is necessary for the tropism of USSC for neuronal injury. Our data demonstrate that USSC have the capacity to migrate toward apoptotic neurons and injured brain. Together with their neural differentiation potential, this suggests a neuroregenerative potential of USSC. Moreover, we provide evidence for a hitherto unrecognized pivotal role of the HGF/c-MET axis in guiding stem cells toward brain injury, which may partly account for the capability of HGF to improve function in the diseased central nervous system.

Hepatocyte growth factor demonstrates racial heterogeneity as a biomarker for coronary heart disease.

PubMed

Bielinski, Suzette J; Berardi, Cecilia; Decker, Paul A; Larson, Nicholas B; Bell, Elizabeth J; Pankow, James S; Sale, Michele M; Tang, Weihong; Hanson, Naomi Q; Wassel, Christina L; de Andrade, Mariza; Budoff, Matthew J; Polak, Joseph F; Sicotte, Hugues; Tsai, Michael Y

2017-08-01

To determine if hepatocyte growth factor (HGF), a promising biomarker of coronary heart disease (CHD) given its release into circulation in response to endothelial damage, is associated with subclinical and clinical CHD in a racial/ethnic diverse population. HGF was measured in 6738 participants of the Multi-Ethnic Study of Atherosclerosis (MESA). Highest mean HGF values (pg/mL) were observed in Hispanic, followed by African, non-Hispanic white, then Chinese Americans. In all races/ethnicities, HGF levels were associated with older age, higher systolic blood pressure (SBP) and body mass index, lower high-density lipoprotein, diabetes and current smoking. In fully adjusted models, each SD higher HGF was associated with an average increase in coronary artery calcium (CAC) of 55 Agatston units for non-Hispanic whites (p<0.001) and 51 Agatston units for African-Americans (p=0.007) but was not in the other race/ethnic groups (interaction p=0.02). There were 529 incident CHD events, and CHD risk was 41% higher in African (p<0.001), 17% in non-Hispanic white (p=0.026) and Chinese (p=0.36), and 6% in Hispanic Americans (p=0.56) per SD increase in HGF. In a large and diverse population-based cohort, we report that HGF is associated with subclinical and incident CHD. We demonstrate evidence of racial/ethnic heterogeneity within these associations, as the results are most compelling in African-Americans and non-Hispanic white Americans. We provide evidence that HGF is a biomarker of atherosclerotic disease that is independent of traditional risk factors. © Article author(s) (or their employer(s) unless otherwise stated in the text of the article) 2017. All rights reserved. No commercial use is permitted unless otherwise expressly granted.

MEK, p38, and PI-3K mediate cross talk between EGFR and TNFR in enhancing hepatocyte growth factor production from human mesenchymal stem cells

PubMed Central

Wang, Yue; Weil, Brent R.; Herrmann, Jeremy L.; Abarbanell, Aaron M.; Tan, Jiangning; Markel, Troy A.; Kelly, Megan L.

2009-01-01

Human bone marrow mesenchymal stem cells (MSCs) are a potent source of growth factors, which are partly responsible for their beneficial paracrine effects. We reported previously that transforming growth factor-α (TGF-α), a putative mediator of wound healing and the injury response, increases the release of vascular endothelial growth factor (VEGF), augments tumor necrosis factor-α (TNF-α)-stimulated VEGF production, and activates mitogen-activated protein kinases and phosphatidylinositol 3-kinase (PI-3K) pathway in human MSCs. The experiments described in this report indicate that TGF-α increases MSC-derived hepatocyte growth factor (HGF) production. TGF-α-stimulated HGF production was abolished by inhibition of MEK, p38, PI-3K, or by small interfering RNA (siRNA) targeting TNF receptor 2 (TNFR2), but was not attenuated by siRNA targeting TNF receptor 1 (TNFR1). Ablation of TNFR1 significantly increased basal and stimulated HGF. A potent synergy between TGF-α and TNF-α was noted in MSC HGF production. This synergistic effect was abolished by MEK, P38, PI-3K inhibition, or by ablation of both TNF receptors using siRNA. We conclude that 1) novel cross talk occurs between tumor necrosis factor receptor and TGF-α/epidermal growth factor receptor in stimulating MSC HGF production; 2) this cross talk is mediated, at least partially, via activation of MEK, p38, and PI-3K; 3) TGF-α stimulates MSCs to produce HGF by MEK, p38, PI-3K, and TNFR2-dependent mechanisms; and 4) TNFR1 acts to decrease basal TGF-α and TNF-α-stimulated HGF. PMID:19692652

Analysis of gene expression profiles in tympanic membrane following perforation using PCR Array in rats--preliminary investigation.

PubMed

Hassmann-Poznańska, Elżbieta; Taranta, Andrzej; Bialuk, Izabela; Poznańska, Maria; Zajączkiewicz, Hanna; Winnicka, Maria Małgorzata

2013-10-01

The goal of this work was to identify genes, known to be involved in the skin wound healing, that express differentially in the healthy and injured tympanic membrane (TM), and designate the molecules potentially beneficial for treatment of TM perforation. The molecular mechanisms controlling the course of TM regeneration are far from being elucidated. Twenty rats had their tympanic membranes perforated, while four served as a control. Animals were sacrificed on either days 1, 2, 3, 5 and 10 post injury, and TMs were immediately dissected and frozen in liquid nitrogen. Total TM RNA was isolated and reversely transcribed. qPCR was performed using Rat Wound Healing RT(2) Profiler PCR Array (QIAGEN) containing primers for 84 genes. Statistically significant changes in the expression of 42 genes were found in various stages of TM healing. The increased expression of genes taking part in the inflammatory reaction (interleukin 6, granulocyte and macrophage chemotactic proteins) was observed from day 2. The expression of several genes of extracellular matrix components and their remodeling enzymes was also changed. Among growth factor genes: Vegfa, Igf1 and Hbegf showed increased expression at the beginning of the healing process, while Hgf expression was highest on day 3. Several changes in the expression of genes involved in remodeling of extracellular matrix point to important role of connective tissue in TM healing. The molecules accelerating this process, like HbEGF and HGF, seem to be good candidates for further evaluation of their possible use in clinical treatment. Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.

Modulation of Myostatin/Hepatocyte Growth Factor Balance by Different Hemodialysis Modalities.

PubMed

Esposito, Pasquale; La Porta, Edoardo; Calatroni, Marta; Grignano, Maria Antonietta; Milanesi, Samantha; Verzola, Daniela; Battaglia, Yuri; Gregorini, Marilena; Libetta, Carmelo; Garibotto, Giacomo; Rampino, Teresa

2017-01-01

Background. In this study we investigated the relevance of myostatin and Hepatocyte Growth Factor (HGF) in patients undergoing hemodialysis HD and the influence of different HD modalities on their levels. Methods. We performed a prospective crossover study in which HD patients were randomized to undergo 3-month treatment periods with bicarbonate hemodialysis (BHD) followed by online hemodiafiltration (HDF). Clinical data, laboratory parameters, and myostatin and HGF serum levels were collected and compared. Results. Ten patients and six controls (C) were evaluated. In any experimental condition myostatin and HGF levels were higher in HD than in C. At enrollment and after BHD there were not significant correlations, whereas at the end of the HDF treatment period myostatin and HGF were inversely correlated ( r   -0.65, p < 0.05), myostatin serum levels inversely correlated with transferrin ( r   -0.73, p < 0.05), and HGF levels that resulted positively correlated with BMI ( r 0.67, p < 0.05). Moving from BHD to HDF, clinical and laboratory parameters were unchanged, as well as serum HGF, whereas myostatin levels significantly decreased (6.3 ± 4.1 versus 4.3 ± 3.1 ng/ml, p < 0.05). Conclusions. Modulation of myostatin levels and myostatin/HGF balance by the use of different HD modalities might represent a novel approach to the prevention and treatment of HD-related muscle wasting syndrome.

Anti-apoptotic Role of Caspase-cleaved GAB1 Adaptor Protein in Hepatocyte Growth Factor/Scatter Factor-MET Receptor Protein Signaling*

PubMed Central

Le Goff, Arnaud; Ji, Zongling; Leclercq, Bérénice; Bourette, Roland P.; Mougel, Alexandra; Guerardel, Cateline; de Launoit, Yvan; Vicogne, Jérôme; Goormachtigh, Gautier; Fafeur, Véronique

2012-01-01

The GRB2-associated binder 1 (GAB1) docking/scaffold protein is a key mediator of the MET-tyrosine kinase receptor activated by hepatocyte growth factor/scatter factor (HGF/SF). Activated MET promotes recruitment and tyrosine phosphorylation of GAB1, which in turn recruits multiple proteins and mediates MET signaling leading to cell survival, motility, and morphogenesis. We previously reported that, without its ligand, MET is a functional caspase target during apoptosis, allowing the generation of a p40-MET fragment that amplifies apoptosis. In this study we established that GAB1 is also a functional caspase target by evidencing a caspase-cleaved p35-GAB1 fragment that contains the MET binding domain. GAB1 is cleaved by caspases before MET, and the resulting p35-GAB1 fragment is phosphorylated by MET upon HGF/SF binding and can interact with a subset of GAB1 partners, PI3K, and GRB2 but not with SHP2. This p35-GAB1 fragment favors cell survival by maintaining HGF/SF-induced MET activation of AKT and by hindering p40-MET pro-apoptotic function. These data demonstrate an anti-apoptotic role of caspase-cleaved GAB1 in HGF/SF-MET signaling. PMID:22915589

Modulation of hepatocyte growth factor secretion in human female reproductive tract stromal fibroblasts by poly (I:C) and estradiol.

PubMed

Coleman, Kimberly D; Ghosh, Mimi; Crist, Sarah G; Wright, Jacqueline A; Rossoll, Richard M; Wira, Charles R; Fahey, John V

2012-01-01

Hepatocyte Growth Factor (HGF) secretion facilitates epithelial cell growth and development in the female reproductive tract (FRT) and may contribute to pathological conditions such as cancer and endometriosis. We hypothesized that estradiol and poly (I:C), a synthetic RNA mimic, may have a regulatory effect on HGF secretion by stromal fibroblasts from FRT tissues. Following hysterectomies, normal tissue from the uterus, endocervix, and ectocervix were dispersed into stromal cell fractions by enzymatic digestion and differential filtering. Stromal fibroblasts were cultured and treated with estradiol and/or poly (I:C), and conditioned media were analyzed for HGF via enzyme-linked immunosorbent assay. Treating uterine fibroblasts with estradiol or poly (I:C) significantly increased HGF secretion. When uterine fibroblasts were co-treated with estradiol and poly (I:C), the effect on HGF secretion was additive. In contrast, stromal fibroblasts from endo- and ecto-cervix were unresponsive to estradiol, but were stimulated to secrete HGF by poly (I:C). HGF secretion is uniquely regulated in the uterus, but not in ecto- and endo-cervix, by estradiol. Moreover, potential viral pathogens further induce HGF. These findings have potential applications in understanding both hormonal regulation of normal tissue as well as the role of HGF in tumorogenesis, endometriosis, and human immunodeficiency virus infection. © 2011 John Wiley & Sons A/S.

Circulating level of hepatocyte growth factor predicts incidence of type 2 diabetes mellitus: The Multi-Ethnic Study of Atherosclerosis (MESA)

PubMed Central

Bancks, Michael P.; Bielinski, Suzette J.; Decker, Paul A.; Hanson, Naomi Q.; Larson, Nicholas B.; Sicotte, Hugues; Wassel, Christina L.; Pankow, James S.

2016-01-01

Background Hepatocyte growth factor (HGF) is a pleotropic factor posited to have metabolic homeostatic properties. The purpose of this study is to examine whether level of HGF is associated with the development of type 2 diabetes. Methods Data from the Multi-Ethnic Study of Atherosclerosis (MESA) were used to examine the prospective association between serum level of HGF and incident diabetes. Fasting HGF was measured at Exam 1 (2000–2002) in 5395 participants free from diabetes (61.5 ± 10.2 years old) and incidence of diabetes was determined at four subsequent follow-up exams over 12 years. Hazard ratios (HR) for incident diabetes were estimated according to 1 standard deviation (SD) unit increment of HGF (1 SD =26 μg/l), before and after adjustment for age, sex, race/ethnicity, education, study center, smoking status, alcohol consumption, body mass index, waist circumference, fasting glucose and insulin, C-reactive protein, and interleukin-6 levels. Results A 1 SD increment of baseline HGF was associated with a 46% (95% CI =1.37, 1.56) increased risk of diabetes before adjustment. After adjustment, diabetes risk per 1 SD increment of HGF was attenuated but remained significantly increased (HR=1.21; 95% CI=1.12, 1.32). Men had a significantly greater HR compared to women per equivalent increase of HGF (p-value for sex interaction=0.04). There was no evidence of effect modification by race/ethnicity. Conclusions This study advances understanding from cross-sectional studies and investigation of incident insulin resistance, demonstrating higher level of HGF is associated with incident diabetes and may reflect a unique type of impaired metabolism. PMID:26892517

Circulating level of hepatocyte growth factor predicts incidence of type 2 diabetes mellitus: The Multi-Ethnic Study of Atherosclerosis (MESA).

PubMed

Bancks, Michael P; Bielinski, Suzette J; Decker, Paul A; Hanson, Naomi Q; Larson, Nicholas B; Sicotte, Hugues; Wassel, Christina L; Pankow, James S

2016-03-01

Hepatocyte growth factor (HGF) is a pleotropic factor posited to have metabolic homeostatic properties. The purpose of this study is to examine whether level of HGF is associated with the development of type 2 diabetes. Data from the Multi-Ethnic Study of Atherosclerosis (MESA) were used to examine the prospective association between serum level of HGF and incident diabetes. Fasting HGF was measured at Exam 1 (2000-2002) in 5395 participants free from diabetes (61.5±10.2 years old) and incidence of diabetes was determined at four subsequent follow-up exams over 12 years. Hazard ratios (HR) for incident diabetes were estimated according to 1 standard deviation (SD) unit increment of HGF (1 SD=26 μg/l), before and after adjustment for age, sex, race/ethnicity, education, study center, smoking status, alcohol consumption, body mass index, waist circumference, fasting glucose and insulin, C-reactive protein, and interleukin-6 levels. A 1 SD increment of baseline HGF was associated with a 46% (95% CI=1.37, 1.56) increased risk of diabetes before adjustment. After adjustment, diabetes risk per 1 SD increment of HGF was attenuated but remained significantly increased (HR=1.21; 95% CI=1.12, 1.32). Men had a significantly greater HR compared to women per equivalent increase of HGF (p-value for sex interaction=0.04). There was no evidence of effect modification by race/ethnicity. This study advances understanding from cross-sectional studies and investigation of incident insulin resistance, demonstrating higher level of HGF is associated with incident diabetes and may reflect a unique type of impaired metabolism. Copyright © 2015 Elsevier Inc. All rights reserved.

Effect of mitomycin C on IL-1R expression, IL-1-related hepatocyte growth factor secretion and corneal epithelial cell migration.

PubMed

Chen, Tsan-Chi; Chang, Shu-Wen

2010-03-01

To investigate how mitomycin C (MMC) modulates hepatocyte growth factor (HGF) and keratinocyte growth factor (KGF) secretions in human corneal fibroblasts and regulates human corneal epithelial (HCE) cell migration. Primary human corneal fibroblasts were treated with MMC (0.05, 0.1, or 0.2 mg/mL for 5 minutes) and were cultivated with or without interleukin (IL)-1beta. Transcript and secretion of HGF and KGF were determined by quantitative real-time RT-PCR and Western blot analysis, respectively. The effect of MMC-treated fibroblasts on HCE cell migration was evaluated using a transwell migration assay. The influence of MMC on HGF expression/secretion and HCE cell migration was further confirmed by RNA interference. The number of IL-1 receptors (IL-1R) on the fibroblast surface was analyzed by flow cytometry. MMC alone did not affect endogenous HGF expression, whereas IL-1beta alone significantly upregulated HGF transcripts and secretion. By modifying IL-1R numbers, MMC further upregulated IL-1beta-related HGF expression at a concentration of 0.05 mg/mL but to a lesser extent at 0.1 and 0.2 mg/mL. KGF transcripts and intracellular expression were suppressed by MMC dose dependently in the presence or absence of IL-1beta, whereas KGF secretion was not affected. Conditioned medium from MMC-treated fibroblasts exerted a similar concentration-dependent effect on HCE cell migration, enhancing migration most significantly at 0.05 mg/mL MMC in the presence of IL-1beta. The MMC dose-dependent modulation of HCE cell migration was abolished in HGF-silenced fibroblasts. MMC differentially modulated IL-1R expression at various concentrations and regulated HGF and KGF differently. MMC alone did not alter HGF expression. In the presence of IL-1beta, MMC-treated corneal fibroblasts modified HCE cell migration through IL-1beta-induced HGF secretion.

Increase in the circulating level of hepatocyte growth factor in pancreatic cancer patients.

PubMed

Kemik, Ozgur; Purisa, Sevim; Kemik, Ahu Sarbay; Tuzun, Sefa

2009-01-01

Hepatocyte growth factor (HGF) has been reported the cause of many biological events, including cell proliferation, invasiveness, morphogenesis, and angiogenesis. Elevated HGF content in tumor tissue was reported to predict a more aggressive biology in breast and gastric cancer patients. Eighty patients with invasive pancreatic cancer investigated. Venous blood samples were collected before the surgery. Sera were obtained by centrifugation and stored at -70 degrees C until assayed. The control group created from healthy individuals. Serum concentrations of soluble HGF were measured by the quantitative sandwich enzyme immunoassay technique. The mean value of serum soluble HGF in patients with invasive pancreatic cancer was 497.2 +/- 53.8 pg/ml and that of control group was 53.6 +/- 7.5 pg/ml and the difference was significant (p < 0.001). The serum levels of soluble HGF might reflect the severity of invasive pancreatic cancer and deserve further evaluation (Tab. 2, Ref. 19). Full Text (Free, PDF) www.bmj.sk.

Preventive effects of the deleted form of hepatocyte growth factor against various liver injuries.

PubMed

Masunaga, H; Fujise, N; Shiota, A; Ogawa, H; Sato, Y; Imai, E; Yasuda, H; Higashio, K

1998-01-26

The effects of a naturally occurring deleted form of hepatocyte growth factor (HGF) on hepatic disorder were studied in various models of hepatic failure. The pretreatment of rats and mice with the deleted form of HGF prevented the liver injuries and coagulopathy induced by endotoxin, dimethylnitrosamine and acetaminophen and reduced the mortality due to hepatic dysfunction induced by these hepatotoxins. The concurrent administration of the deleted form of HGF also prevented the liver injury and hepatic fibrosis in mice treated with alpha-naphthylisothiocyanate and in rats treated with dimethylnitrosamine. Moreover, the deleted form of HGF normalized the results of the bromosulphalein-clearance test and ameliorated jaundice in rats with periportal cholangiolitic hepatopathy induced by alpha-naphthylisothiocyanate. The deleted form of HGF also reversed the coagulopathy in rats with hepatic disorder induced by dimethylnitrosamine or by 70% resection of cirrhotic liver (induced by carbon tetrachloride). In Long Evans cinnamon rats receiving vehicle, 20 out of 21 animals died within 4 days after the onset of jaundice. After infusion of the deleted form of HGF for 4 days, 7 out of 20 Long-Evans cinnamon rats survived. These results indicate that the deleted form of HGF could have therapeutic potency in patients with severe hepatic failure.

Enhanced cell survival and paracrine effects of mesenchymal stem cells overexpressing hepatocyte growth factor promote cardioprotection in myocardial infarction

DOE Office of Scientific and Technical Information (OSTI.GOV)

Zhao, Liyan; Liu, Xiaolin; Zhang, Yuelin

Poor cell survival post transplantation compromises the therapeutic benefits of mesenchymal stem cells (MSCs) in myocardial infarction (MI). Hepatocyte growth factor (HGF) is an important cytokine for angiogenesis, anti-inflammation and anti-apoptosis. This study aimed to evaluate the cardioprotective effects of MSCs overexpressing HGF in a mouse model of MI. The apoptosis of umbilical cord-derived MSCs (UC-MSCs) and HGF-UC-MSCs under normoxic and hypoxic conditions was detected. The conditioned medium (CdM) of UC-MSCs and HGF-UC-MSCs under a hypoxic condition was harvested and its protective effect on neonatal cardiomyocytes (NCMs) exposed to a hypoxic challenge was examined. UC-MSCs and HGF-UC-MSCs were transplanted intomore » the peri-infarct region in mice following MI and heart function assessed 4 weeks post transplantation. The apoptosis of HGF-UC-MSCs under hypoxic conditions was markedly decreased compared with that of UC-MSCs. NCMs treated with HGF-UC-MSC hypoxic CdM (HGF-UC-MSCs-hy-CdM) exhibited less cell apoptosis in response to hypoxic challenge than those treated with UC-MSC hypoxic CdM (UC-MSCs-hy-CdM). HGF-UC-MSCs-hy-CdM released the inhibited p-Akt and lowered the enhanced ratio of Bax/Bcl-2 induced by hypoxia in the NCMs. HGF-UC-MSCs-hy-CdM expressed higher levels of HGF, EGF, bFGF and VEGF than UC-MSCs-hy-CdM. Transplantation of HGF-UC-MSCs or UC-MSCs greatly improved heart function in the mouse model of MI. Compared with UC-MSCs, transplantation of HGF-UC-MSCs was associated with less cardiomyocyte apoptosis, enhanced angiogenesis and increased proliferation of cardiomyocytes. This study may provide a novel therapeutic strategy for MSC-based therapy in cardiovascular disease.« less

Effect of hepatocyte growth factor on endogenous hepatocarcinogenesis in rats fed a choline-deficient L-amino acid-defined diet.

PubMed

Nakanishi, Chihiro; Moriuchi, Akihiro; Ido, Akio; Numata, Masatsugu; Kim, Il-Deok; Kusumoto, Kazunori; Hasuike, Satoru; Abe, Hiroo; Nagata, Kenji; Akiyama, Yutaka; Uto, Hirofumi; Kataoka, Hiroaki; Tsubouchi, Hirohito

2006-07-01

Hepatocyte growth factor (HGF) is a promising agent for the treatment of intractable liver disease, due to its mitogenic, anti-apoptotic, and anti-fibrotic effects. We investigated the effect of recombinant human HGF (rh-HGF) on the development of both hepatocellular carcinoma (HCC) and preneoplastic nodules in rats fed a choline-deficient L-amino acid-defined (CDAA) diet, an animal model of hepatocarcinogenesis resembling human development of HCC with cirrhosis. From weeks 13 to 48 of the CDAA diet, rh-HGF (0.1 or 0.5 mg/kg/day) was administered intravenously to rats in four-week cycles, with treatment for five consecutive days of each week for two weeks, followed by a two-week washout period. Treatment with rh-HGF significantly inhibited the development of preneoplastic nodules in a dose-dependent manner at 24 weeks. Although the numbers and areas of the preneoplastic nodules in rats treated with rh-HGF were equivalent to those in mock-treated rats by 60 weeks, the incidence of HCC was reduced by HGF treatment. Although one rat treated with low-dose rh-HGF exhibited a massive HCC, which occupied almost the whole liver, and lung metastases, HGF treatment did not increase the overall frequency of HCC. Administration of high-dose rh-HGF, however, induced an increase in the urinary excretion of albumin, leading to decreased serum albumin at 60 weeks. These results indicate that long-term administration of rh-HGF does not accelerate hepatocarcinogenesis in rats fed a CDAA diet. However, these findings do not completely exclude the potential of HGF-induced hepatocarcinogenesis; this issue must be resolved before rh-HGF can be used for patients with intractable liver diseases, especially those with cirrhosis.

Hepatitis A complicated with acute renal failure and high hepatocyte growth factor: A case report.

PubMed

Oe, Shinji; Shibata, Michihiko; Miyagawa, Koichiro; Honma, Yuichi; Hiura, Masaaki; Abe, Shintaro; Harada, Masaru

2015-08-28

A 58-year-old man was admitted to our hospital. Laboratory data showed severe liver injury and that the patient was positive for immunoglobulin M anti-hepatitis A virus (HAV) antibodies. He was also complicated with severe renal dysfunction and had an extremely high level of serum hepatocyte growth factor (HGF). Therefore, he was diagnosed with severe acute liver failure with acute renal failure (ARF) caused by HAV infection. Prognosis was expected to be poor because of complications by ARF and high serum HGF. However, liver and renal functions both improved rapidly without intensive treatment, and he was subsequently discharged from our hospital on the 21(st) hospital day. Although complication with ARF and high levels of serum HGF are both important factors predicting poor prognosis in acute liver failure patients, the present case achieved a favorable outcome. Endogenous HGF might play an important role as a regenerative effector in injured livers and kidneys.

Phenotypic differences between oral and skin fibroblasts in wound contraction and growth factor expression.

PubMed

Shannon, Diane B; McKeown, Scott T W; Lundy, Fionnuala T; Irwin, Chris R

2006-01-01

Wounds of the oral mucosa heal in an accelerated fashion with reduced scarring compared with cutaneous wounds. The differences in healing outcome between oral mucosa and skin could be because of phenotypic differences between the respective fibroblast populations. This study compared paired mucosal and dermal fibroblasts in terms of collagen gel contraction, alpha-smooth muscle actin expression (alpha-SMA), and production of the epithelial growth factors: keratinocyte growth factor (KGF) and hepatocyte growth factor/scatter factor (HGF). The effects of transforming growth factor -beta1 and -beta3 on each parameter were also determined. Gel contraction in floating collagen lattices was determined over a 7-day period. alpha-SMA expression by fibroblasts was determined by Western blotting. KGF and HGF expression were determined by an enzyme-linked immunosorbent assay. Oral fibroblasts induced accelerated collagen gel contraction, yet surprisingly expressed lower levels of alpha-SMA. Oral cells also produced significantly greater levels of both KGF and HGF than their dermal counterparts. Transforming growth factor-beta1 and -beta3, over the concentration range of 0.1-10 ng/mL, had similar effects on cell function, stimulating both gel contraction and alpha-SMA production, but inhibiting KGF and HGF production by both cell types. These data indicate phenotypic differences between oral and dermal fibroblasts that may well contribute to the differences in healing outcome between these two tissues.

Hepatocyte growth factor secreted by ovarian cancer cells stimulates peritoneal implantation via the mesothelial-mesenchymal transition of the peritoneum.

PubMed

Nakamura, Michihiko; Ono, Yoshihiro J; Kanemura, Masanori; Tanaka, Tomohito; Hayashi, Masami; Terai, Yoshito; Ohmichi, Masahide

2015-11-01

A current working model for the metastatic process of ovarian carcinoma suggests that cancer cells are shed from the ovarian tumor into the peritoneal cavity and attach to the layer of mesothelial cells that line the inner surface of the peritoneum, and several studies suggest that hepatocyte growth factor (HGF) plays an important role in the dissemination of ovarian cancer. Our objectives were to evaluate the HGF expression of ovarian cancer using clinical data and assess the effect of HGF secreted from human ovarian cancer cells to human mesothelial cells. HGF expression was immunohistochemically evaluated in 165 epithelial ovarian cancer patients arranged as tissue microarrays. HGF expression in four ovarian cancer cell lines was evaluated by using semi-quantitative polymerase chain reaction, Western blotting and enzyme-linked immunosorbent assay. The effect of ovarian cancer cell derived HGF to the human mesothelial cells was assessed by using morphologic analysis, Western blotting and cell invasion assay. The effect of HGF on ovarian cancer metastasis was assessed by using in vivo experimental model. The clinical data showed a significantly high correlation between the HGF expression and the cancer stage. The in vivo and in vitro experimental models revealed that HGF secreted by ovarian cancer cells induces the mesothelial-to-mesenchymal transition and stimulates the invasion of mesothelial cells. Furthermore, manipulating the HGF activity affected the degree of dissemination and ascite formation. We demonstrated that HGF secreted by ovarian cancer cells plays an important role in cancer peritoneal implantation. Copyright © 2015 Elsevier Inc. All rights reserved.

Hepatocyte growth factor is elevated in amniotic fluid from obese women and regulates placental glucose and fatty acid metabolism.

PubMed

Visiedo, F; Bugatto, F; Carrasco-Fernández, C; Sáez-Benito, A; Mateos, R M; Cózar-Castellano, I; Bartha, J L; Perdomo, G

2015-04-01

To evaluate the impact of the pro-inflammatory cytokine hepatocyte growth factor (HGF) on the regulation of glucose and lipid placental metabolism. HGF levels were quantified in amniotic fluid and placenta from control and obese women. 2-deoxy-glucose (2-DOG) uptake, glycolysis, fatty acid oxidation (FAO), fatty acid esterification, de novo fatty acid synthesis, triglyceride levels and carnitine palmitoyltransferase activities (CPT) were measured in placental explants upon addition of pathophysiological HGF levels. In obese women, total- and -activated-HGF levels in amniotic fluid were elevated ∼24%, and placental HGF levels were ∼3-fold higher than in control women. At a similar dose to that present in amniotic fluid of obese women, HGF (30 ng/mL) increased Glut-1 levels and 2-DOG uptake by ∼25-30% in placental explants. HGF-mediated effect on 2-DOG uptake was dependent on the activation of phosphatidylinositol 3-kinase signaling pathway. In addition, HGF decreased ∼20% FAO, whereas esterification and de novo fatty acid synthesis increased ∼15% and ∼25% respectively, leading to 2-fold triglyceride accumulation in placental explants. In parallel, HGF reduced CPT-I activity ∼70%. HGF is a cytokine elevated in amniotic fluid and placental tissue of obese women, which through its ability to stimulate 2-DOG uptake and metabolism impairs FAO and enhances esterification and de novo fatty acid synthesis, leading to accumulation of placental triglycerides. Copyright © 2015 Elsevier Ltd. All rights reserved.

Priming Dental Pulp Stem Cells With Fibroblast Growth Factor-2 Increases Angiogenesis of Implanted Tissue-Engineered Constructs Through Hepatocyte Growth Factor and Vascular Endothelial Growth Factor Secretion.

PubMed

Gorin, Caroline; Rochefort, Gael Y; Bascetin, Rumeyza; Ying, Hanru; Lesieur, Julie; Sadoine, Jérémy; Beckouche, Nathan; Berndt, Sarah; Novais, Anita; Lesage, Matthieu; Hosten, Benoit; Vercellino, Laetitia; Merlet, Pascal; Le-Denmat, Dominique; Marchiol, Carmen; Letourneur, Didier; Nicoletti, Antonino; Vital, Sibylle Opsahl; Poliard, Anne; Salmon, Benjamin; Muller, Laurent; Chaussain, Catherine; Germain, Stéphane

2016-03-01

Tissue engineering strategies based on implanting cellularized biomaterials are promising therapeutic approaches for the reconstruction of large tissue defects. A major hurdle for the reliable establishment of such therapeutic approaches is the lack of rapid blood perfusion of the tissue construct to provide oxygen and nutrients. Numerous sources of mesenchymal stem cells (MSCs) displaying angiogenic potential have been characterized in the past years, including the adult dental pulp. Establishment of efficient strategies for improving angiogenesis in tissue constructs is nevertheless still an important challenge. Hypoxia was proposed as a priming treatment owing to its capacity to enhance the angiogenic potential of stem cells through vascular endothelial growth factor (VEGF) release. The present study aimed to characterize additional key factors regulating the angiogenic capacity of such MSCs, namely, dental pulp stem cells derived from deciduous teeth (SHED). We identified fibroblast growth factor-2 (FGF-2) as a potent inducer of the release of VEGF and hepatocyte growth factor (HGF) by SHED. We found that FGF-2 limited hypoxia-induced downregulation of HGF release. Using three-dimensional culture models of angiogenesis, we demonstrated that VEGF and HGF were both responsible for the high angiogenic potential of SHED through direct targeting of endothelial cells. In addition, FGF-2 treatment increased the fraction of Stro-1+/CD146+ progenitor cells. We then applied in vitro FGF-2 priming to SHED before encapsulation in hydrogels and in vivo subcutaneous implantation. Our results showed that FGF-2 priming is more efficient than hypoxia at increasing SHED-induced vascularization compared with nonprimed controls. Altogether, these data demonstrate that FGF-2 priming enhances the angiogenic potential of SHED through the secretion of both HGF and VEGF. ©AlphaMed Press.

Exosomes serve as nanoparticles to suppress tumor growth and angiogenesis in gastric cancer by delivering hepatocyte growth factor siRNA.

PubMed

Zhang, Haiyang; Wang, Yi; Bai, Ming; Wang, Junyi; Zhu, Kegan; Liu, Rui; Ge, Shaohua; Li, JiaLu; Ning, Tao; Deng, Ting; Fan, Qian; Li, Hongli; Sun, Wu; Ying, Guoguang; Ba, Yi

2018-03-01

Exosomes derived from cells have been found to mediate signal transduction between cells and to act as efficient carriers to deliver drugs and small RNA. Hepatocyte growth factor (HGF) is known to promote the growth of both cancer cells and vascular cells, and the HGF-cMET pathway is a potential clinical target. Here, we characterized the inhibitory effect of HGF siRNA on tumor growth and angiogenesis in gastric cancer. In addition, we showed that HGF siRNA packed in exosomes can be transported into cancer cells, where it dramatically downregulates HGF expression. A cell co-culture model was used to show that exosomes loaded with HGF siRNA suppress proliferation and migration of both cancer cells and vascular cells. Moreover, exosomes were able to transfer HGF siRNA in vivo, decreasing the growth rates of tumors and blood vessels. The results of our study demonstrate that exosomes have potential for use in targeted cancer therapy by delivering siRNA. © 2018 The Authors. Cancer Science published by John Wiley & Sons Australia, Ltd on behalf of Japanese Cancer Association.

Genomic profiling of a Hepatocyte growth factor-dependent signature for MET-targeted therapy in glioblastoma.

PubMed

Johnson, Jennifer; Ascierto, Maria Libera; Mittal, Sandeep; Newsome, David; Kang, Liang; Briggs, Michael; Tanner, Kirk; Marincola, Francesco M; Berens, Michael E; Vande Woude, George F; Xie, Qian

2015-09-17

Constitutive MET signaling promotes invasiveness in most primary and recurrent GBM. However, deployment of available MET-targeting agents is confounded by lack of effective biomarkers for selecting suitable patients for treatment. Because endogenous HGF overexpression often causes autocrine MET activation, and also indicates sensitivity to MET inhibitors, we investigated whether it drives the expression of distinct genes which could serve as a signature indicating vulnerability to MET-targeted therapy in GBM. Interrogation of genomic data from TCGA GBM (Student's t test, GBM patients with high and low HGF expression, p ≤ 0.00001) referenced against patient-derived xenograft (PDX) models (Student's t test, sensitive vs. insensitive models, p ≤ 0.005) was used to identify the HGF-dependent signature. Genomic analysis of GBM xenograft models using both human and mouse gene expression microarrays (Student's t test, treated vs. vehicle tumors, p ≤ 0.01) were performed to elucidate the tumor and microenvironment cross talk. A PDX model with EGFR(amp) was tested for MET activation as a mechanism of erlotinib resistance. We identified a group of 20 genes highly associated with HGF overexpression in GBM and were up- or down-regulated only in tumors sensitive to MET inhibitor. The MET inhibitors regulate tumor (human) and host (mouse) cells within the tumor via distinct molecular processes, but overall impede tumor growth by inhibiting cell cycle progression. EGFR (amp) tumors undergo erlotinib resistance responded to a combination of MET and EGFR inhibitors. Combining TCGA primary tumor datasets (human) and xenograft tumor model datasets (human tumor grown in mice) using therapeutic efficacy as an endpoint may serve as a useful approach to discover and develop molecular signatures as therapeutic biomarkers for targeted therapy. The HGF dependent signature may serve as a candidate predictive signature for patient enrollment in clinical trials using MET inhibitors

Assessment of Growth Factors Secreted by Human Breastmilk Mesenchymal Stem Cells.

PubMed

Kaingade, Pankaj Mahipatrao; Somasundaram, Indumathi; Nikam, Amar Babaso; Sarang, Shabari Amit; Patel, Jagdish Shantilal

2016-01-01

Human breastmilk is a dynamic, multifaceted biological fluid containing nutrients, bioactive substances, and growth factors. It is effective in supporting growth and development of an infant. As breastmilk has been found to possess mesenchymal stem cells, the importance of the components of breastmilk and their physiological roles is increasing day by day. The present study was intended to identify the secretions of growth factors, mainly vascular endothelial growth factor (VEGF) and hepatocyte growth factor (HGF), from human breastmilk mesenchymal stem cells under basal conditions of in vitro cell culture using synthetic media and human cord serum. The growth factors were analyzed with the enzyme-linked immunosorbent assay technique. The cultured mesenchymal stem cells of breastmilk without serum revealed significant differences in secretions of the VEGF and HGF growth factors (8.55 ± 2.26402 pg/mL and 230.8 ± 45.9861 pg/mL, respectively) compared with mesenchymal stem cells of breastmilk with serum (21.31 ± 4.69 pg/mL and 2,404.42 ± 481.593 pg/mL, respectively). Results obtained from our study demonstrate that both VEGF and HGF are secreted in vitro by human breastmilk mesenchymal stem cells. The roles of VEGF and HGF in surfactant secretion, pulmonary maturation, and neonatal maturity have been well established. Thus, we emphasize that breastmilk-derived MSCs could be a potent therapeutic source in treating neonatal diseases. Besides, due to its immense potency, the study also emphasizes the importance of breastfeeding, which is promoted by organizations like the World Heatlh Organization and UNICEF.

Hepatocyte Growth Factor Is Required for Mesenchymal Stromal Cell Protection Against Bleomycin-Induced Pulmonary Fibrosis

PubMed Central

Cahill, Emer F.; Kennelly, Helen; Carty, Fiona; Mahon, Bernard P.

2016-01-01

The incidence of idiopathic pulmonary fibrosis is on the rise and existing treatments have failed to halt or reverse disease progression. Mesenchymal stromal cells (MSCs) have potent cytoprotective effects, can promote tissue repair, and have demonstrated efficacy in a range of fibrotic lung diseases; however, the exact mechanisms of action remain to be elucidated. Chemical antagonists and short hairpin RNA knockdown were used to identify the mechanisms of action used by MSCs in promoting wound healing, proliferation, and inhibiting apoptosis. Using the bleomycin induced fibrosis model, the protective effects of early or late MSC administration were examined. The role for hepatocyte growth factor (HGF) in MSC protection against bleomycin lung injury was examined using HGF knockdown MSC. Terminal deoxynucleotidyl transferase (TdT) dUTP nick-end labeling assay was performed on ex vivo lung sections to examine the effects of MSC on apoptosis. MSC conditioned media (CM) enhanced wound closure and inhibited apoptosis of pulmonary cells in vitro. HGF was required for MSC CM enhancement of epithelial cell proliferation and inhibition of apoptosis. In contrast, MSC required COX-2 for CM to inhibit fibroblast proliferation. In a murine model, early administration of MSC protected against bleomycin induced lung fibrosis and correlated with reduced levels of the proinflammatory cytokine interleukin-1β, reduced levels of apoptosis, and significantly increased levels of HGF. These protective effects were in part mediated by MSC derived HGF as HGF knockdown MSC were unable to protect against fibrosis in vivo. These findings delineate the mechanisms of MSC protection in a preclinical model of fibrotic lung disease. Significance The mechanisms used by mesenchymal stromal cells (MSCs) in mediating protective effects in chronic models of lung disease are not understood and remain to be elucidated. These findings from in vitro studies highlight an important role for the MSC

Placental insufficiency decreases pancreatic vascularity and disrupts hepatocyte growth factor signaling in the pancreatic islet endothelial cell in fetal sheep.

PubMed

Rozance, Paul J; Anderson, Miranda; Martinez, Marina; Fahy, Anna; Macko, Antoni R; Kailey, Jenai; Seedorf, Gregory J; Abman, Steven H; Hay, William W; Limesand, Sean W

2015-02-01

Hepatocyte growth factor (HGF) and vascular endothelial growth factor A (VEGFA) are paracrine hormones that mediate communication between pancreatic islet endothelial cells (ECs) and β-cells. Our objective was to determine the impact of intrauterine growth restriction (IUGR) on pancreatic vascularity and paracrine signaling between the EC and β-cell. Vessel density was less in IUGR pancreata than in controls. HGF concentrations were also lower in islet EC-conditioned media (ECCM) from IUGR, and islets incubated with control islet ECCM responded by increasing insulin content, which was absent with IUGR ECCM. The effect of ECCM on islet insulin content was blocked with an inhibitory anti-HGF antibody. The HGF receptor was not different between control and IUGR islets, but VEGFA was lower and the high-affinity VEGF receptor was higher in IUGR islets and ECs, respectively. These findings show that paracrine actions from ECs increase islet insulin content, and in IUGR ECs, secretion of HGF was diminished. Given the potential feed-forward regulation of β-cell VEGFA and islet EC HGF, these two growth factors are highly integrated in normal pancreatic islet development, and this regulation is decreased in IUGR fetuses, resulting in lower pancreatic islet insulin concentrations and insulin secretion. © 2015 by the American Diabetes Association. Readers may use this article as long as the work is properly cited, the use is educational and not for profit, and the work is not altered.

c-Met in esophageal squamous cell carcinoma: an independent prognostic factor and potential therapeutic target.

PubMed

Ozawa, Yohei; Nakamura, Yasuhiro; Fujishima, Fumiyoshi; Felizola, Saulo J A; Takeda, Kenichiro; Okamoto, Hiroshi; Ito, Ken; Ishida, Hirotaka; Konno, Takuro; Kamei, Takashi; Miyata, Go; Ohuchi, Noriaki; Sasano, Hironobu

2015-06-03

c-Met is widely known as a poor prognostic factor in various human malignancies. Previous studies have suggested the involvement of c-Met and/or its ligand, hepatocyte growth factor (HGF), in esophageal squamous cell carcinoma (ESCC), but the correlation between c-Met status and clinical outcome remains unclear. Furthermore, the identification of a novel molecular therapeutic target might potentially help improve the clinical outcome of ESCC patients. The expression of c-Met and HGF was immunohistochemically assessed in 104 surgically obtained tissue specimens. The correlation between c-Met/HGF expression and patients' clinicopathological features, including survival, was evaluated. We also investigated changes in cell functions and protein expression of c-Met and its downstream signaling pathway components under treatments with HGF and/or c-Met inhibitor in ESCC cell lines. Elevated expression of c-Met was significantly correlated with tumor depth and pathological stage. Patients with high c-Met expression had significantly worse survival. In addition, multivariate analysis identified the high expression of c-Met as an independent prognostic factor. Treatment with c-Met inhibitor under HGF stimulation significantly inhibited the invasive capacity of an ESCC cell line with elevated c-Met mRNA expression. Moreover, c-Met and its downstream signaling inactivation was also detected after treatment with c-Met inhibitor. The results of our study identified c-Met expression as an independent prognostic factor in ESCC patients and demonstrated that c-Met could be a potential molecular therapeutic target for the treatment of ESCC with elevated c-Met expression.

Hepatocyte growth factor is associated with progression of atherosclerosis: The Multi-Ethnic Study of Atherosclerosis (MESA).

PubMed

Bell, Elizabeth J; Decker, Paul A; Tsai, Michael Y; Pankow, James S; Hanson, Naomi Q; Wassel, Christina L; Larson, Nicholas B; Cohoon, Kevin P; Budoff, Matthew J; Polak, Joseph F; Stein, James H; Bielinski, Suzette J

2018-05-01

Hepatocyte growth factor (HGF) has previously been associated with risk of stroke, coronary heart disease, and atherosclerosis. We hypothesized that higher circulating HGF is associated with greater progression of measures of atherosclerosis: coronary artery calcium (CAC) and carotid plaque. Participants aged 45-84 years from the prospective cohort study Multi-Ethnic Study of Atherosclerosis had HGF measured at baseline (between 2000 and 2002) and were followed for progression of atherosclerosis for up to 12 years. CAC was measured at all five exams using the Agatston method. Mixed-effects models were used to examine the association of HGF and CAC progression among 6695 participants with available data. Relative risk regression was used to assess the association between HGF and new or additional carotid plaque between exams 1 and 5 in 3400 participants with available data. All point estimates were adjusted for potential confounding variables. Each standard deviation higher HGF at baseline was associated with 2.9 Agatston units/year greater CAC progression (95% CI: 1.6-4.2, p < 0.0001), and the magnitude of this association differed by race/ethnicity (p value for interaction by race = 0.003). Each standard deviation higher HGF at baseline was associated with a 4% higher risk of new or additional carotid plaque (95% CI: 1.01-1.08, p = 0.005). Higher levels of HGF were significantly associated with greater progression of atherosclerosis in this large and diverse population. Circulating HGF continues to show promise as a potential clinical biomarker for cardiovascular disease. Copyright © 2018 Elsevier B.V. All rights reserved.

The Multisubstrate Adapter Gab1 Regulates Hepatocyte Growth Factor (Scatter Factor)–c-Met Signaling for Cell Survival and DNA Repair

PubMed Central

Fan, Saijun; Ma, Yong Xian; Gao, Min; Yuan, Ren-Qi; Meng, Qinghui; Goldberg, Itzhak D.; Rosen, Eliot M.

2001-01-01

Hepatocyte growth factor (scatter factor) (HGF/SF) is a pleiotrophic mediator of epithelial cell motility, morphogenesis, angiogenesis, and tumorigenesis. HGF/SF protects cells against DNA damage by a pathway from its receptor c-Met to phosphatidylinositol 3-kinase (PI3K) to c-Akt, resulting in enhanced DNA repair and decreased apoptosis. We now show that protection against the DNA-damaging agent adriamycin (ADR; topoisomerase IIα inhibitor) requires the Grb2-binding site of c-Met, and overexpression of the Grb2-associated binder Gab1 (a multisubstrate adapter required for epithelial morphogenesis) inhibits the ability of HGF/SF to protect MDCK epithelial cells against ADR. In contrast to Gab1 and its homolog Gab2, overexpression of c-Cb1, another multisubstrate adapter that associates with c-Met, did not affect protection. Gab1 blocked the ability of HGF/SF to cause the sustained activation of c-Akt and c-Akt signaling (FKHR phosphorylation). The Gab1 inhibition of sustained c-Akt activation and of cell protection did not require the Gab1 pleckstrin homology or SHP2 phosphatase-binding domain but did require the PI3K-binding domain. HGF/SF protection of parental MDCK cells was blocked by wortmannin, expression of PTEN, and dominant negative mutants of p85 (regulatory subunit of PI3K), Akt, and Pak1; the protection of cells overexpressing Gab1 was restored by wild-type or activated mutants of p85, Akt, and Pak1. These findings suggest that the adapter Gab1 may redirect c-Met signaling through PI3K away from a c-Akt/Pak1 cell survival pathway. PMID:11438654

The association between thyroid volume, L-thyroxine therapy and hepatocyte growth factor levels among patients with euthyroid and hypothyroid goitrous and non-goitrous Hashimoto's thyroiditis versus healthy subjects.

PubMed

Kilic, Mustafa Kemal; Yesilkaya, Yakup; Tezcan, Kadriye; Cinar, Nese; Akin, Safak; Karakaya, Jale; Akata, Deniz; Usman, Aydan; Gurlek, Alper

2016-05-01

Hashimoto's thyroiditis (HT) is the most common etiology of hypothyroidism in regions where iodine deficiency is not a concern. To date, many clinical investigations have been conducted to elucidate its pathogenesis. Several growth factors have been shown to have a role in its development. Hepatocyte growth factor (HGF) is one of the aforementioned molecules. We aimed to demonstrate whether HGF is responsible for HT and goiter development. Also, we aimed to test the hypothesis that levo-thyroxine sodium therapy will suppress HGF levels. Sixty-one premenopausal women who were admitted to our outpatient clinic between November 2010 and September 2011 were enrolled. Three groups were determined according to their thyroid function tests (TFTs) as euthyroid Hashimoto's, control and subclinical hypothyroid Hashimoto's groups. Basal TFTs, anti-thyroid peroxidase (anti-TPO), anti-thyroglobulin (anti-tg), thyroid ultrasonography (USG) and HGF were studied and recorded. Subclinical hypothyroid HT patients received levo-thyroxine sodium replacement therapy, and were re-assessed for the same laboratory and radiologic features after a median 3.5 month follow-up. Basal HGF levels were not different between groups. In the subclinical hypothyroidism group, HGF levels (752.75 ± 144.91 pg/ml vs. 719.37 ± 128.05 pg/ml; p = 0.496) and thyroid volumes (12.51 ± 3.67 cc vs. 12.18 ± 4.26 cc; p = 0.7) before and after treatment did not change significantly. No correlations were found between HGF and other parameters. HGF levels were similar between subjects with nodular goiter and normal thyroid structure. HGF was not shown to be associated with HT and goiter development. In addition, levo-thyroxine sodium replacement therapy did not alter serum HGF levels significantly.

Matriptase is required for the active form of hepatocyte growth factor induced Met, focal adhesion kinase and protein kinase B activation on neural stem/progenitor cell motility.

PubMed

Fang, Jung-Da; Lee, Sheau-Ling

2014-07-01

Hepatocyte growth factor (HGF) is a chemoattractant and inducer for neural stem/progenitor (NS/P) cell migration. Although the type II transmembrane serine protease, matriptase (MTP) is an activator of the latent HGF, MTP is indispensable on NS/P cell motility induced by the active form of HGF. This suggests that MTP's action on NS/P cell motility involves mechanisms other than proteolytic activation of HGF. In the present study, we investigate the role of MTP in HGF-stimulated signaling events. Using specific inhibitors of phosphatidylinositol-3-kinase (PI3K), protein kinase B (Akt) or focal adhesion kinase (FAK), we demonstrated that in NS/P cells HGF-activated c-Met induces PI3k-Akt signaling which then leads to FAK activation. This signaling pathway ultimately induces MMP2 expression and NS/P cell motility. Knocking down of MTP in NS/P cells with specific siRNA impaired HGF-stimulation of c-Met, Akt and FAK activation, blocked HGF-induced production of MMP2 and inhibited HGF-stimulated NS/P cell motility. MTP-knockdown NS/P cells cultured in the presence of recombinant protein of MTP protease domain or transfected with the full-length wild-type but not the protease-defected MTP restored HGF-responsive events in NS/P cells. In addition to functioning as HGF activator, our data revealed novel function of MTP on HGF-stimulated c-Met signaling activation. Copyright © 2014. Published by Elsevier B.V.

Hepatocyte growth factor upregulation promotes carcinogenesis and epithelial-mesenchymal transition in hepatocellular carcinoma via Akt and COX-2 pathways

PubMed Central

Ogunwobi, Olorunseun O.

2013-01-01

Advanced hepatocellular carcinoma (HCC) is an important cause of cancer mortality. Epithelial-mesenchymal transition (EMT) has been shown to be an important biological process in cancer progression and metastasis. We have focused on elucidating factors that induce EMT to promote carcinogenesis and subsequent metastasis in HCC using the BNL CL.2 (BNL) and BNL 1ME A. 7R.1 (1MEA) cell lines. BNL cells are normal hepatocytes whereas the 1MEA cells are HCC cells derived from chemical transformation of the BNL cells. Their morphological characteristics were examined. Expression levels of hepatocyte growth factor (HGF), markers of EMT and mediators of HGF signaling were determined and functional characteristics were compared. BNL cells were treated with HGF and effects on EMT-marker and mediators of HGF signaling were analyzed. BNL cells display characteristic epithelial morphology whereas 1MEA cells display mesenchymal characteristics. 1MEA cells express and secrete more HGF than BNL cells. There was significantly decreased expression of E-cadherin, albumin, AAT and increased expression of fibronectin, collagen-1, vimentin, snail and slug in 1MEA cells. There was also increased expression of cyclooxygenase-2 (COX-2), Akt and phosphorylated Akt (pAkt) in 1MEA cells. Moreover, 1MEA cells had increased migratory capacity inhibited by inhibition of COX-2 and Akt but not extracellular signal regulated kinase (ERK). Molecular mesenchymal characteristics of 1MEA cells were reversed by inhibition of COX-2, Akt and ERK. Treatment of BNL cells with HGF led to decreased expression of E-cadherin and increased expression of fibronectin, vimentin, snail, slug, COX-2, Akt, pAkt and increased migration, invasiveness and clonogenicity. We conclude that development of HCC is associated with upregulation of HGF which promotes EMT and carcinogenesis via upregulation of COX-2 and Akt. Consequently, HGF signaling may be targeted for therapy in advanced and metastatic HCC. PMID:21744257

Hepatocyte growth factor upregulation promotes carcinogenesis and epithelial-mesenchymal transition in hepatocellular carcinoma via Akt and COX-2 pathways.

PubMed

Ogunwobi, Olorunseun O; Liu, Chen

2011-12-01

Advanced hepatocellular carcinoma (HCC) is an important cause of cancer mortality. Epithelial-mesenchymal transition (EMT) has been shown to be an important biological process in cancer progression and metastasis. We have focused on elucidating factors that induce EMT to promote carcinogenesis and subsequent metastasis in HCC using the BNL CL.2 (BNL) and BNL 1ME A. 7R.1 (1MEA) cell lines. BNL cells are normal hepatocytes whereas the 1MEA cells are HCC cells derived from chemical transformation of the BNL cells. Their morphological characteristics were examined. Expression levels of hepatocyte growth factor (HGF), markers of EMT and mediators of HGF signaling were determined and functional characteristics were compared. BNL cells were treated with HGF and effects on EMT-marker and mediators of HGF signaling were analyzed. BNL cells display characteristic epithelial morphology whereas 1MEA cells display mesenchymal characteristics. 1MEA cells express and secrete more HGF than BNL cells. There was significantly decreased expression of E-cadherin, albumin, AAT and increased expression of fibronectin, collagen-1, vimentin, snail and slug in 1MEA cells. There was also increased expression of cyclooxygenase-2 (COX-2), Akt and phosphorylated Akt (pAkt) in 1MEA cells. Moreover, 1MEA cells had increased migratory capacity inhibited by inhibition of COX-2 and Akt but not extracellular signal regulated kinase (ERK). Molecular mesenchymal characteristics of 1MEA cells were reversed by inhibition of COX-2, Akt and ERK. Treatment of BNL cells with HGF led to decreased expression of E-cadherin and increased expression of fibronectin, vimentin, snail, slug, COX-2, Akt, pAkt and increased migration, invasiveness and clonogenicity. We conclude that development of HCC is associated with upregulation of HGF which promotes EMT and carcinogenesis via upregulation of COX-2 and Akt. Consequently, HGF signaling may be targeted for therapy in advanced and metastatic HCC.

Up-Regulation of Bcl-xl by Hepatocyte Growth Factor in Human Mesothelioma Cells Involves ETS Transcription Factors

PubMed Central

Cao, Xiaobo; Littlejohn, James; Rodarte, Charles; Zhang, Lidong; Martino, Benjamin; Rascoe, Philip; Hamid, Kamran; Jupiter, Daniel; Smythe, W. Roy

2009-01-01

Bcl-xl and the hepatocyte growth factor (HGF) receptor c-Met are both highly expressed in mesotheliomas, where they protect cells from apoptosis and can confer resistance to conventional therapeutic agents. In our current study, we investigate a model for the transcriptional control of Bcl-xl that involves ETS transcription factors and the HGF/Met axis. In addition, the effects of activated c-Met on the phosphorylation of the ETS family transcriptional factors were examined. The transient expression of ETS-2 and PU.1 cDNAs in mesothelioma cell lines resulted in an increase in the promoter activity of Bcl-xl and consequently in its mRNA and protein expression levels, whereas the transcriptional repressor Tel suppressed Bcl-xl transcription. The activation of the HGF/Met axis led to rapid phosphorylation of ETS family transcription factors in mesothelioma cells through the mitogen-activated protein kinase pathway and via nuclear accumulation of ETS-2 and PU.1. A chromatin immunoprecipitation assay further demonstrated that the activation of c-Met enhanced the binding of ETS transcriptional factors to the Bcl-x promoter. Finally, we determined the Bcl-xl and phosphorylated c-Met expression levels in mesothelioma patient samples; these data suggest a strong correlation between Bcl-xl and phosphorylated c-Met levels. Taken together, these findings support a role for c-Met as an inhibitor of apoptosis and an activator of Bcl-xl. PMID:19834061

Analysis of hepatocyte growth factor immunostaining in the placenta of HIV-infected normotensive versus preeclamptic pregnant women.

PubMed

Cele, S B; Odun-Ayo, F; Onyangunga, O A; Moodley, J; Naicker, T

2018-08-01

Hepatocyte Growth Factor (HGF) plays a role in the migration and morphogenesis of different cell types and tissues. Preeclampsia (PE) is associated with deficient trophoblast invasion and placental insufficiency; hence HGF production is expected to be compromised. This study therefore aimed to immunolocalize and morphometrically analyse placental HGF in normotensive versus PE pregnancies stratified by HIV status and gestational age. Normotensive (N; n = 40) and preeclamptic (PE; n = 80) women were stratified by HIV status (HIV- and HIV+), and gestational age i.e. early onset of PE (EOPE; <34 weeks) and late onset of PE (LOPE; ≥34 weeks). Placental tissues were stained using conventional immunohistochemistry, performed using mouse anti-human HGF antibody. Morphometric image analysis was performed using Zeiss Axio-Vision software. HGF was immuno-localized within the syncytiotrophoblast, cytotrophoblast, endothelial and fibroblast-like cell populations of both conducting and exchange villi. Based on pregnancy type, HGF immunoexpression within the conducting villi was significantly different between Nvs EOPE (p = 0.0372) and EOPE vs LOPE (p = 0.0006). Within the exchange villi, no significant difference of HGF immunostaining was noted between N vs EOPE and N vs LOPE. A down-regulation of HGF immuno-expression was observed in LOPE compared to other groups within both villi types, albeit non-significant. Based on HIV status, no significant difference in HGF immuno-expression was demonstrated between HIV- vs HIV + within the exchange and conducting villi. However, the expression of HGF in HIV- group was elevated in both villi types. Across the groups, a significant difference was found between N+ vs EOPE- (p  = 0.0207), EOPE+ vs LOPE- (p = 0.0036) and EOPE- vs LOPE- (p = 0.0016) of the conducting villi while no significant difference was found within the exchange villi. This novel study demonstrates that HGF was two-fold higher in conducting compared to

Oral microbe-host interactions: influence of β-glucans on gene expression of inflammatory cytokines and metabolome profile.

PubMed

Silva, Viviam de Oliveira; Pereira, Luciano José; Murata, Ramiro Mendonça

2017-03-07

The aim of this study was to evaluate the effects of β-glucan on the expression of inflammatory mediators and metabolomic profile of oral cells [keratinocytes (OBA-9) and fibroblasts (HGF-1) in a dual-chamber model] infected by Aggregatibacter actinomycetemcomitans. The periodontopathogen was applied and allowed to cross the top layer of cells (OBA-9) to reach the bottom layer of cells (HGF-1) and induce the synthesis of immune factors and cytokines in the host cells. β-glucan (10 μg/mL or 20 μg/mL) were added, and the transcriptional factors and metabolites produced were quantified in the remaining cell layers and supernatant. The relative expression of interleukin (IL)-1-α and IL-18 genes in HGF-1 decreased with 10 μg/mL or 20 μg/mL of β-glucan, where as the expression of PTGS-2 decreased only with 10 μg/mL. The expression of IL-1-α increased with 20 μg/mL and that of IL-18 increased with 10 μg/mL in OBA-9; the expression of BCL 2, EP 300, and PTGS-2 decreased with the higher dose of β-glucan. The production of the metabolite 4-aminobutyric acid presented lower concentrations under 20 μg/mL, whereas the concentrations of 2-deoxytetronic acid NIST and oxalic acid decreased at both concentrations used. Acetophenone, benzoic acid, and pinitol presented reduced concentrations only when treated with 10 μg/mL of β-glucan. Treatment with β-glucans positively modulated the immune response and production of metabolites.

Soluble CD30 and Hepatocyte growth factor as predictive markers of antibody-mediated rejection of the kidney allograft.

PubMed

Pavlova, Yelena; Viklicky, Ondrej; Slatinska, Janka; Bürgelova, Marcela; Süsal, Caner; Skibova, Jelena; Honsová, Eva; Striz, Ilja; Kolesar, Libor; Slavcev, Antonij

2011-07-01

Our retrospective study was aimed to assess the relevance of pre- and post-transplant measurements of serum concentrations of the soluble CD30 molecule (soluble CD30, sCD30) and the cytokine Hepatocyte growth factor (HGF) for prediction of the risk for development of antibody-mediated rejection (AMR) in kidney transplant patients. Evaluation of sCD30, HGF levels and the presence of HLA-specific antibodies in a cohort of 205 patients was performed before, 2weeks and 6months after transplantation. Patients were followed up for kidney graft function and survival for two years. We found a tendency of higher incidence of AMR in retransplanted patients with elevated pre-transplant sCD30 (≥100U/ml) (p=0.051), however no such correlation was observed in first-transplant patients. Kidney recipients with simultaneously high sCD30 and HLA-specific antibodies (sCD30+/Ab+) before transplantation had significantly lower AMR-free survival compared to the other patient groups (p<0.001). HGF concentrations were not associated with the incidence of AMR at any time point of measurement, nevertheless, the combined analysis HGF and sCD30 showed increased incidence of AMR in recipients with elevated pretransplant sCD30 and low HGF levels. the predictive value of pretransplant sCD30 for the development of antibody-mediated rejection after transplantation is significantly potentiated by the co-presence of HLA specific antibodies. The role of HGF as a rejection-protective factor in patients with high pretransplant HGF levels would need further investigation. Copyright © 2011 Elsevier B.V. All rights reserved.

89Zr-DFO-AMG102 Immuno-PET to Determine Local Hepatocyte Growth Factor Protein Levels in Tumors for Enhanced Patient Selection.

PubMed

Price, Eric W; Carnazza, Kathryn E; Carlin, Sean D; Cho, Andrew; Edwards, Kimberly J; Sevak, Kuntal K; Glaser, Jonathan M; de Stanchina, Elisa; Janjigian, Yelena Y; Lewis, Jason S

2017-09-01

The hepatocyte growth factor (HGF) binding antibody rilotumumab (AMG102) was modified for use as a 89 Zr-based immuno-PET imaging agent to noninvasively determine the local levels of HGF protein in tumors. Because recent clinical trials of HGF-targeting therapies have been largely unsuccessful in several different cancers (e.g., gastric, brain, lung), we have synthesized and validated 89 Zr-DFO-AMG102 as a companion diagnostic for improved identification and selection of patients having high local levels of HGF in tumors. To date, patient selection has not been performed using the local levels of HGF protein in tumors. Methods: The chelator p -SCN-Bn-DFO was conjugated to AMG102, radiolabeling with 89 Zr was performed in high radiochemical yields and purity (>99%), and binding affinity of the modified antibody was confirmed using an enzyme-linked immunosorbent assay (ELISA)-type binding assay. PET imaging, biodistribution, autoradiography and immunohistochemistry, and ex vivo HGF ELISA experiments were performed on murine xenografts of U87MG (HGF-positive, MET-positive) and MKN45 (HGF-negative, MET-positive) and 4 patient-derived xenografts (MET-positive, HGF unknown). Results: Tumor uptake of 89 Zr-DFO-AMG102 at 120 h after injection in U87MG xenografts (HGF-positive) was high (36.8 ± 7.8 percentage injected dose per gram [%ID/g]), whereas uptake in MKN45 xenografts (HGF-negative) was 5.0 ± 1.3 %ID/g and a control of nonspecific human IgG 89 Zr-DFO-IgG in U87MG tumors was 11.5 ± 3.3 %ID/g, demonstrating selective uptake in HGF-positive tumors. Similar experiments performed in 4 different gastric cancer patient-derived xenograft models showed low uptake of 89 Zr-DFO-AMG102 (∼4-7 %ID/g), which corresponded with low HGF levels in these tumors (ex vivo ELISA). Autoradiography, immunohistochemical staining, and HGF ELISA assays confirmed that elevated levels of HGF protein were present only in U87MG tumors and that 89 Zr-DFO-AMG102 uptake was closely correlated

Plasma hepatocyte growth factor is a novel marker of AL cardiac amyloidosis.

PubMed

Swiger, Kristopher J; Friedman, Eitan A; Brittain, Evan L; Tomasek, Kelsey A; Huang, Shi; Su, Yan R; Sawyer, Douglas B; Lenihan, Daniel J

2016-12-01

Cardiac amyloidosis is an infiltrative cardiomyopathy that is challenging to diagnose. We hypothesized that the novel biomarkers hepatocyte growth factor (HGF), galectin-3 (GAL-3), interleukin-6 (IL-6), and vascular endothelial growth factor (VEGF) would be elevated in cardiac amyloidosis and may be able to discriminate from non-cardiac systemic amyloidosis or other cardiomyopathies with similar clinical or morphologic characteristics. Patients were selected from the Vanderbilt Main Heart Registry according to the following groups: (1) amyloid light-chain (AL) cardiac amyloidosis (n = 26); (2) transthyretin (ATTR) cardiac amyloidosis (n = 7); (3) left ventricular hypertrophy (LVH) (n = 45); (4) systolic heart failure (n = 42); and (5) non-cardiac systemic amyloidosis (n = 7). Biomarkers were measured in stored plasma samples. Biomarkers' discrimination performance in predicting AL cardiac amyloidosis (i.e., Concordance index) was reported. A survival analysis was used to explore the relationship between HGF levels and mortality among AL cardiac amyloidosis patients. HGF levels were markedly elevated in patients with AL cardiac amyloidosis (median = 622, interquartile range (IQR): 299-1228 pg/mL) compared with the other groups, including those with non-cardiac systemic amyloidosis (median = 134, IQR: 94-163 pg/mL, p < 0.001). HGF was not a specific marker for ATTR amyloidosis. Gal-3 was elevated in all groups with amyloidosis but could not differentiate between those with and without cardiac involvement. There was no difference in IL-6 or VEGF between those with AL cardiac amyloidosis compared to other groups (p = 0.13 and 0.057, respectively). HGF may be a specific marker that distinguishes AL cardiac amyloidosis from other cardiomyopathies with similar clinical or morphologic characteristics. Further studies are necessary to determine whether HGF levels predict the likelihood of survival.

Priming Dental Pulp Stem Cells With Fibroblast Growth Factor-2 Increases Angiogenesis of Implanted Tissue-Engineered Constructs Through Hepatocyte Growth Factor and Vascular Endothelial Growth Factor Secretion

PubMed Central

Gorin, Caroline; Rochefort, Gael Y.; Bascetin, Rumeyza; Ying, Hanru; Lesieur, Julie; Sadoine, Jérémy; Beckouche, Nathan; Berndt, Sarah; Novais, Anita; Lesage, Matthieu; Hosten, Benoit; Vercellino, Laetitia; Merlet, Pascal; Le-Denmat, Dominique; Marchiol, Carmen; Letourneur, Didier; Nicoletti, Antonino; Vital, Sibylle Opsahl; Poliard, Anne; Salmon, Benjamin; Germain, Stéphane

2016-01-01

Tissue engineering strategies based on implanting cellularized biomaterials are promising therapeutic approaches for the reconstruction of large tissue defects. A major hurdle for the reliable establishment of such therapeutic approaches is the lack of rapid blood perfusion of the tissue construct to provide oxygen and nutrients. Numerous sources of mesenchymal stem cells (MSCs) displaying angiogenic potential have been characterized in the past years, including the adult dental pulp. Establishment of efficient strategies for improving angiogenesis in tissue constructs is nevertheless still an important challenge. Hypoxia was proposed as a priming treatment owing to its capacity to enhance the angiogenic potential of stem cells through vascular endothelial growth factor (VEGF) release. The present study aimed to characterize additional key factors regulating the angiogenic capacity of such MSCs, namely, dental pulp stem cells derived from deciduous teeth (SHED). We identified fibroblast growth factor-2 (FGF-2) as a potent inducer of the release of VEGF and hepatocyte growth factor (HGF) by SHED. We found that FGF-2 limited hypoxia-induced downregulation of HGF release. Using three-dimensional culture models of angiogenesis, we demonstrated that VEGF and HGF were both responsible for the high angiogenic potential of SHED through direct targeting of endothelial cells. In addition, FGF-2 treatment increased the fraction of Stro-1+/CD146+ progenitor cells. We then applied in vitro FGF-2 priming to SHED before encapsulation in hydrogels and in vivo subcutaneous implantation. Our results showed that FGF-2 priming is more efficient than hypoxia at increasing SHED-induced vascularization compared with nonprimed controls. Altogether, these data demonstrate that FGF-2 priming enhances the angiogenic potential of SHED through the secretion of both HGF and VEGF. Significance The results from the present study show that fibroblast growth factor-2 (FGF-2) priming is more

Intra-myocardial injection of both growth factors and heart derived Sca-1+/CD31- cells attenuates post-MI LV remodeling more than does cell transplantation alone: neither intervention enhances functionally significant cardiomyocyte regeneration.

PubMed

Wang, Xiaohong; Li, Qinglu; Hu, Qingsong; Suntharalingam, Piradeep; From, Arthur H L; Zhang, Jianyi

2014-01-01

Insulin-like growth factor 1 (IGF-1) and hepatocyte growth factor (HGF) are two potent cell survival and regenerative factors in response to myocardial injury (MI). We hypothesized that simultaneous delivery of IGF+HGF combined with Sca-1+/CD31- cells would improve the outcome of transplantation therapy in response to the altered hostile microenvironment post MI. One million adenovirus nuclear LacZ-labeled Sca-1+/CD31- cells were injected into the peri-infarction area after left anterior descending coronary artery (LAD) ligation in mice. Recombinant mouse IGF-1+HGF was added to the cell suspension prior to the injection. The left ventricular (LV) function was assessed by echocardiography 4 weeks after the transplantation. The cell engraftment, differentiation and cardiomyocyte regeneration were evaluated by histological analysis. Sca-1+/CD31- cells formed viable grafts and improved LV ejection fraction (EF) (Control, 54.5+/-2.4; MI, 17.6+/-3.1; Cell, 28.2+/-4.2, n = 9, P<0.01). IGF+HGF significantly enhanced the benefits of cell transplantation as evidenced by increased EF (38.8+/-2.2; n = 9, P<0.01) and attenuated adverse structural remodeling. Furthermore, IGF+HGF supplementation increased the cell engraftment rate, promoted the transplanted cell survival, enhanced angiogenesis, and minimally stimulated endogenous cardiomyocyte regeneration in vivo. The in vitro experiments showed that IGF+HGF treatment stimulated Sca-1+/CD31- cell proliferation and inhibited serum free medium induced apoptosis. Supperarray profiling of Sca-1+/CD31- cells revealed that Sca-1+/CD31- cells highly expressed various trophic factor mRNAs and IGF+HGF treatment altered the mRNAs expression patterns of these cells. These data indicate that IGF-1+HGF could serve as an adjuvant to cell transplantation for myocardial repair by stimulating donor cell and endogenous cardiac stem cell survival, regeneration and promoting angiogenesis.

[Intramuscular injection of lentivirus-mediated EPAS1 gene improves hind limb ischemia and its mechanism in a rat model of peripheral artery vascular disease].

PubMed

Wang, Zhihong; Gu, Hongbin; Yang, Fan; Xie, Huajie; Sheng, Lei; Li, Mingfei

2017-11-01

Objective To investigate the effect of over-expressed endothelial Per-Arnt-Sim domain protein 1 (EPAS1) on peripheral arterial disease (PAD) in a rat model. Methods PAD rat model was established by external iliac artery ligation followed by lentivirus-mediated EPAS1 gene injection into rat right adductor magnus. The models were evaluated by quantitative analysis of gait disturbance. The changes of blood flow in the posterior extremity of the rats were detected using laser Doppler. The expressions of EPAS1, hepatocyte growth factor (HGF), basic fibroblast growth factor (bFGF), and vascular endothelial growth factor (VEGF) mRNAs were tested by real-time quantitative PCR. The expression of α-smooth muscle actin (αSMA) was detected by immunohistochemical staining. Results Compared with lenti-EGFP group, rat hind limb function and circulation got recovered obviously 7 days after lenti-EPAS1 injection. The mRNA expressions of EPAS1, HGF, bFGF, and VEGF were up-regulated in the lenti-EPAS1-treated sites.The expression of αSMA showed an obvious increase in the lenti-EPAS1-treated muscles. Conclusion Over-expressed lenti-EPAS1 can promote angiogenesis via the up-regulation of EPAS1-related angiogenic factors in the muscles of the affected hind limb and reduce gait disturbance.

Prognostic implication of serum hepatocyte growth factor in stage II/III breast cancer patients who received neoadjuvant chemotherapy.

PubMed

Kim, Hyori; Youk, Jeonghwan; Yang, Yaewon; Kim, Tae-Yong; Min, Ahrum; Ham, Hye-Seon; Cho, Seongcheol; Lee, Kyung-Hun; Keam, Bhumsuk; Han, Sae-Won; Oh, Do-Youn; Ryu, Han Suk; Han, Wonshik; Park, In Ae; Kim, Tae-You; Noh, Dong-Young; Im, Seock-Ah

2016-03-01

In stage II/III breast cancer, neoadjuvant chemotherapy (NAC) is a standard treatment. Although several biomarkers are used to predict prognosis in breast cancer, there is no reliable predictive biomarker for NAC success. Recently, the hepatocyte growth factor (HGF) and cMet signaling pathway demonstrated to be involved in breast cancer tumor progression, and its potential as a biomarker is under active investigation. In this study, we assessed the potential of serum HGF as a prognostic biomarker for NAC efficacy. Venous blood samples were drawn from patients diagnosed with stage II/III breast cancer and treated with NAC in Seoul National University Hospital from August 2004 to November 2009. Serum HGF level was determined using an ELISA system. We reviewed the medical records of the patients and investigated the association of HGF level with patients' clinicopathologic characteristics. A total of 121 female patients (median age = 45 years old) were included. Median level of HGF was 934 pg/ml (lower quartile: 772, upper quartile: 1145 pg/ml). Patients with higher HGF level than median value were significantly more likely to have clinically detectable regional node metastasis (p = 0.017, Fisher's exact test). Patients with complete and partial response according to the American Joint Committee on Cancer 7th Edition criteria tended to have higher HGF level (p = 0.105 by t test). Patients with an HGF level higher than the upper quartile value had longer relapse-free survival than the other patients (106 vs. 85 months, p = 0.008). High serum HGF levels in breast cancer patients are associated with clinically detectable regional node metastasis and, paradoxically, with longer relapse-free survival in stage II/III breast cancer.

Obesity-Mediated Regulation of HGF/c-Met Is Associated with Reduced Basal-Like Breast Cancer Latency in Parous Mice

PubMed Central

Sundaram, Sneha; Freemerman, Alex J.; Galanko, Joseph A.; McNaughton, Kirk K.; Bendt, Katharine M.; Darr, David B.; Troester, Melissa A.; Makowski, Liza

2014-01-01

It is widely thought that pregnancy reduces breast cancer risk, but this lacks consideration of breast cancer subtypes. While a full term pregnancy reduces risk for estrogen receptor positive (ER+) and luminal breast cancers, parity is associated with increased risk of basal-like breast cancer (BBC) subtype. Basal-like subtypes represent less than 10% of breast cancers and are highly aggressive, affecting primarily young, African American women. Our previous work demonstrated that high fat diet-induced obesity in nulliparous mice significantly blunted latency in C3(1)-TAg mice, a model of BBC, potentially through the hepatocyte growth factor (HGF)/c-Met oncogenic pathway. Experimental studies have examined parity and obesity individually, but to date, the joint effects of parity and obesity have not been studied. We investigated the role of obesity in parous mice on BBC. Parity alone dramatically blunted tumor latency compared to nulliparous controls with no effects on tumor number or growth, while obesity had only a minor role in further reducing latency. Obesity-associated metabolic mediators and hormones such as insulin, estrogen, and progesterone were not significantly regulated by obesity. Plasma IL-6 was also significantly elevated by obesity in parous mice. We have previously reported a potential role for stromal-derived hepatocyte growth factor (HGF) via its cognate receptor c-Met in the etiology of obesity-induced BBC tumor onset and in both human and murine primary coculture models of BBC-aggressiveness. Obesity-associated c-Met concentrations were 2.5-fold greater in normal mammary glands of parous mice. Taken together, our studies demonstrate that, parity in C3(1)-TAg mice dramatically reduced BBC latency compared to nulliparous mice. In parous mice, c-Met is regulated by obesity in unaffected mammary gland and is associated with tumor onset. C3(1)-TAg mice recapitulate epidemiologic findings such that parity drives increased BBC risk and potential

Hepatocyte growth factor modulates interleukin-6 production in bone marrow derived macrophages: implications for inflammatory mediated diseases.

PubMed

Coudriet, Gina M; He, Jing; Trucco, Massimo; Mars, Wendy M; Piganelli, Jon D

2010-11-02

The generation of the pro-inflammatory cytokines IL-6, TNF-α, and IL-1β fuel the acute phase response (APR). To maintain body homeostasis, the increase of inflammatory proteins is resolved by acute phase proteins via presently unknown mechanisms. Hepatocyte growth factor (HGF) is transcribed in response to IL-6. Since IL-6 production promotes the generation of HGF and induces the APR, we posited that accumulating HGF might be a likely candidate for quelling excess inflammation under non-pathological conditions. We sought to assess the role of HGF and how it influences the regulation of inflammation utilizing a well-defined model of inflammatory activation, lipopolysaccharide (LPS)-stimulation of bone marrow derived macrophages (BMM). BMM were isolated from C57BL6 mice and were stimulated with LPS in the presence or absence of HGF. When HGF was present, there was a decrease in production of the pro-inflammatory cytokine IL-6, along with an increase in the anti-inflammatory cytokine IL-10. Altered cytokine production correlated with an increase in phosphorylated GSK3β, increased retention of the phosphorylated NFκB p65 subunit in the cytoplasm, and an enhanced interaction between CBP and phospho-CREB. These changes were a direct result of signaling through the HGF receptor, MET, as effects were reversed in the presence of a selective inhibitor of MET (SU11274) or when using BMM from macrophage-specific conditional MET knockout mice. Combined, these data provide compelling evidence that under normal circumstances, HGF acts to suppress the inflammatory response.

STK/RON receptor tyrosine kinase mediates both apoptotic and growth signals via the multifunctional docking site conserved among the HGF receptor family.

PubMed Central

Iwama, A; Yamaguchi, N; Suda, T

1996-01-01

STK/RON tyrosine kinase, a member of the hepatocyte growth factor (HGF) receptor family, is a receptor for macrophage-stimulating protein (MSP). To examine the STK/RON signalling pathway, we generated STK/ RON transfectants showing opposite features in growth. STK/RON-expressing Ba/F3 pro-B cells (BaF/STK) exhibited MSP-dependent growth, whereas STK/ RON-expressing mouse erythroleukaemia cells (MEL/ STK) displayed MSP-induced apoptosis. This apoptosis was accompanied by the prolonged activation of c-Jun N-terminal kinase (JNK), which has recently been implicated in the initiation of apoptosis. Co-immunoprecipitation analyses showed that autophosphorylated STK/RON associated with PLC-gamma, P13-kinase, Shc and Grb2 in both transfectants. However, major tyrosine-phosphorylated proteins, p61 and p65, specifically associated with STK/RON in MEL/STK cells. Mutations at two C-terminal tyrosine residues, Y1330 and Y1337, in the counterpart of the multifunctional docking site of the HGF receptor abolished both MSP-induced growth and apoptosis. Analyses of these mutants and in vitro association revealed that signalling proteins including p61 and p65 directly bound to the phosphotyrosines in the multifunctional docking site. These results demonstrate that positive or negative signals toward cell growth are generated through the multifunctional docking site and suggest the involvement of p61 and p65 as well as JNK in apoptosis. Our findings provide the first evidence for apoptosis via a receptor tyrosine kinase. Images PMID:8918464

The role of growth factors in maintenance of stemness in bone marrow-derived mesenchymal stem cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Eom, Young Woo; Oh, Ji-Eun; Lee, Jong In

2014-02-28

Highlights: • Expression of FGF-2, FGF-4, EGF, and HGF decreased during long-term culture of BMSCs. • Loss of growth factors induced autophagy, senescence and decrease of stemness. • FGF-2 increased proliferation potential via AKT and ERK activation in BMSCs. • FGF-2 suppressed LC3-II expression and down-regulated senescence of BMSCs. • HGF was important in maintenance of the differentiation potential of BMSCs. - Abstract: Mesenchymal stem cells (MSCs) are an active topic of research in regenerative medicine due to their ability to secrete a variety of growth factors and cytokines that promote healing of damaged tissues and organs. In addition, thesemore » secreted growth factors and cytokines have been shown to exert an autocrine effect by regulating MSC proliferation and differentiation. We found that expression of EGF, FGF-4 and HGF were down-regulated during serial passage of bone marrow-derived mesenchymal stem cells (BMSCs). Proliferation and differentiation potentials of BMSCs treated with these growth factors for 2 months were evaluated and compared to BMSCs treated with FGF-2, which increased proliferation of BMSCs. FGF-2 and -4 increased proliferation potentials at high levels, about 76- and 26-fold, respectively, for 2 months, while EGF and HGF increased proliferation of BMSCs by less than 2.8-fold. Interestingly, differentiation potential, especially adipogenesis, was maintained only by HGF treatment. Treatment with FGF-2 rapidly induced activation of AKT and later induced ERK activation. The basal level of phosphorylated ERK increased during serial passage of BMSCs treated with FGF-2. The expression of LC3-II, an autophagy marker, was gradually increased and the population of senescent cells was increased dramatically at passage 7 in non-treated controls. But FGF-2 and FGF-4 suppressed LC3-II expression and down-regulated senescent cells during long-term (i.e. 2 month) cultures. Taken together, depletion of growth factors during serial

[Growth Factors and Interleukins in Amniotic Membrane Tissue Homogenate].

PubMed

Stachon, T; Bischoff, M; Seitz, B; Huber, M; Zawada, M; Langenbucher, A; Szentmáry, N

2015-07-01

Application of amniotic membrane homogenate eye drops may be a potential treatment alternative for therapy resistant corneal epithelial defects. The purpose of this study was to determine the concentrations of epidermal growth factor (EGF), fibroblast growth factor basic (bFGF), hepatocyte growth factor (HGF), keratinocyte growth factor (KGF), interleukin-6 (IL-6) and interleukin-8 (IL-8) in amniotic membrane homogenates. Amniotic membranes of 8 placentas were prepared and thereafter stored at - 80 °C using the standard methods of the LIONS Cornea Bank Saar-Lor-Lux, Trier/Westpfalz. Following defreezing, amniotic membranes were cut in two pieces and homogenized in liquid nitrogen. One part of the homogenate was prepared in cell-lysis buffer, the other part was prepared in PBS. The tissue homogenates were stored at - 20 °C until enzyme-linked immunosorbent assay (ELISA) analysis for EGF, bFGF, HGF, KGF, IL-6 and IL-8 concentrations. Concentrations of KGF, IL-6 and IL-8 were below the detection limit using both preparation techniques. The EGF concentration in tissue homogenates treated with cell-lysis buffer (2412 pg/g tissue) was not significantly different compared to that of tissue homogenates treated with PBS (1586 pg/g tissue, p = 0.72). bFGF release was also not significantly different using cell-lysis buffer (3606 pg/g tissue) or PBS treated tissue homogenates (4649 pg/g tissue, p = 0.35). HGF release was significantly lower using cell-lysis buffer (23,555 pg/g tissue), compared to PBS treated tissue (47,766 pg/g tissue, p = 0.007). Containing EGF, bFGF and HGF, and lacking IL-6 and IL-8, the application of amniotic membrane homogenate eye drops may be a potential treatment alternative for therapy-resistant corneal epithelial defects. Georg Thieme Verlag KG Stuttgart · New York.

HSP27 is required for invasion and metastasis triggered by hepatocyte growth factor.

PubMed

Pavan, Simona; Musiani, Daniele; Torchiaro, Erica; Migliardi, Giorgia; Gai, Marta; Di Cunto, Ferdinando; Erriquez, Jessica; Olivero, Martina; Di Renzo, Maria Flavia

2014-03-15

The hepatocyte growth factor (HGF) also known as scatter factor activates cancer cell invasion and metastasis. We show that in ovarian cancer cells HGF induced the phosphorylation of the small heat shock protein of 27 kDa (HSP27) by activating the p38MAPK. HSP27 is increased in many cancers at advanced stage including ovarian cancer and associated with cancer resistance to therapy and poor patients' survival. The phosphorylation of HSP27 regulates both its chaperone activity and its control of cytoskeletal stability. We show that HSP27 was necessary for the remodeling of actin filaments induced by HGF and that motility in vitro depended on the p38MAPK-MK2 axis. In vivo, HSP27 silencing impaired the ability of the highly metastatic, HGF-secreting ovarian cancer cells to give rise to spontaneous metastases. This was due to defective motility across the vessel wall and reduced growth. Indeed, HSP27 silencing impaired the ability of circulating ovarian cancer cells to home to the lungs and to form experimental hematogenous metastases and the capability of cancer cells to grow as subcutaneous xenografts. Moreover, HSP27 suppression resulted in the sensitization of xenografts to low doses of the chemotherapeutic paclitaxel, likely because HSP27 protected microtubules from bundling caused by the drug. Altogether, these data show that the HSP27 is required for the proinvasive and prometastatic activity of HGF and suggest that HSP27 might be not only a marker of progression of ovarian cancer, but also a suitable target for therapy. © 2013 UICC.

Macrophage depletion impairs skeletal muscle regeneration: The roles of regulatory factors for muscle regeneration.

PubMed

Liu, Xiaoguang; Liu, Yu; Zhao, Linlin; Zeng, Zhigang; Xiao, Weihua; Chen, Peijie

2017-03-01

Though macrophages are essential for skeletal muscle regeneration, which is a complex process, the roles and mechanisms of the macrophages in the process of muscle regeneration are still not fully understood. The objective of this study is to explore the roles of macrophages and the mechanisms involved in the regeneration of injured skeletal muscle. One hundred and twelve C57BL/6 mice were randomly divided into muscle contusion and macrophages depleted groups. Their gastrocnemius muscles were harvested at the time points of 12 h, 1, 3, 5, 7, 14 d post-injury. The changes in skeletal muscle morphology were assessed by hematoxylin and eosin (HE) stain. The gene expression was analyzed by real-time polymerase chain reaction. The data showed that CL-liposomes treatment did affect the expression of myogenic regulatory factors (MyoD, myogenin) after injury. In addition, CL-liposomes treatment decreased the expression of regulatory factors of muscle regeneration (HGF, uPA, COX-2, IGF-1, MGF, FGF6) and increased the expression of inflammatory cytokines (TGF-β1, TNF-α, IL-1β, RANTES) in the late stage of regeneration. Moreover, there were significant correlations between macrophages and some regulatory factors (such as HGF, uPA) for muscle regeneration. These results suggested that macrophages depletion impairs skeletal muscle regeneration and that the regulatory factors for muscle regeneration may play important roles in this process. © 2017 International Federation for Cell Biology.

[Combined assay of soluble CD30 and hepatocyte growth factor for diagnosis of acute renal allograft rejection].

PubMed

Li, Chuan-jiang; Yu, Li-xin; Xu, Jian; Fu, Shao-jie; Deng, Wen-feng; Du, Chuan-fu; Wang, Yi-bin

2008-02-01

To study the value of detection of both preoperative soluble CD30 (sCD30) and hepatocyte growth factor (HGF) level 5 days after transplantation in the diagnosis of acute rejection of renal allograft. Preoperative serum sCD30 levels and HGF level 5 days after transplantation were determined in 65 renal-transplant recipients using enzyme-linked immunosorbent assay. The recipients were divided according to the sCD30 levels positivity. Receiver operating characteristic (ROC) curves were used to assess the value of HGF level on day 5 posttransplantation for diagnosis of acute renal allograft rejection, and the value of combined assay of the sCD30 and HGF levels was also estimated. After transplantation, 26 recipients developed graft rejection and 39 had uneventful recovery without rejection. With the cut-off value of sCD30 of 120 U/ml, the positivity rate of sCD30 was significantly higher in recipients with graft rejection than in those without (61.5% vs 17.9%, P<0.05). Recipients with acute rejection showed also significantly higher HGF levels on day 5 posttransplantation than those without rejection (P<0.05). ROC curve analysis indicated that HGF levels on day 5 posttransplantation was a good marker for diagnosis of acute renal allograft rejection, and at the cut-off value of 90 ug/L, the diagnostic sensitivity was 84.6% and specificity 76.9%. Evaluation of both the sCD30 and HGF levels significantly enhanced the diagnostic accuracy of acute graft rejection. Combined assay of serum sCD30 and HGF levels offers a useful means for diagnosis of acute renal allograft rejection.

MET inhibitor PHA-665752 suppresses the hepatocyte growth factor-induced cell proliferation and radioresistance in nasopharyngeal carcinoma cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Liu, Tongxin; Li, Qi; Sun, Quanquan

2014-06-20

Highlights: • We demonstrated that irradiation induced MET overexpression and activation. • The aberrant MET signal mediated by HGF induced proliferation and radioresistance of NPC cells. • MET inhibitor PHA-665752 effectively suppressed HGF induced cell proliferation and radioresistance in NPC cells. • PHA-665752 suppressed the three downstream pathway of HGF/MET signal in a dose-dependent manner. - Abstract: Although ionizing radiation (IR) has provided considerable improvements in nasopharyngeal carcinoma (NPC), in subsets of patients, radioresistance is still a major problem in the treatment. In this study, we demonstrated that irradiation induced MET overexpression and activation, and the aberrant MET signal mediatedmore » by hepatocyte growth factor (HGF) induced radioresistance. We also found that MET inhibitor PHA-665752 effectively suppressed HGF induced cell proliferation and radioresistance in NPC cells. Further investigation indicated that PHA-665752 suppressed the phosphorylation of the Akt, ERK1/2, and STAT3 proteins in a dose-dependent manner. Our data indicated that the combination of IR with a MET inhibitor, such as PHA-665752, might be a promising therapeutic strategy for NPC.« less

Gab1 Mediates Hepatocyte Growth Factor-Stimulated Mitogenicity and Morphogenesis in Multipotent Myeloid Cells

PubMed Central

Felici, Angelina; Giubellino, Alessio; Bottaro, Donald P.

2012-01-01

Hepatocyte growth factor (HGF)-stimulated mitogenesis, motogenesis and morphogenesis in various cell types begins with activation of the Met receptor tyrosine kinase and the recruitment of intracellular adaptors and kinase substrates. The adapter protein Gab1 is a critical effector and substrate of activated Met, mediating morphogenesis, among other activities, in epithelial cells. To define its role downstream of Met in hematopoietic cells, Gab1 was expressed in the HGF-responsive, Gab1-negative murine myeloid cell line 32D. Interestingly, the adhesion and motility of Gab1-expressing cells were significantly greater than parental cells, independent of growth factor treatment. Downstream of activated Met, Gab1 expression was specifically associated with rapid Shp-2 recruitment and activation, increased mitogenic potency, suppression of GATA-1 expression and concomitant upregulation of GATA-2 transcription. In addition to enhanced proliferation, continuous culture of Gab1-expressing 32D cells in HGF resulted in cell attachment, filopodia extension and phenotypic changes suggestive of monocytic differentiation. Our results suggest that in myeloid cells, Gab1 is likely to enhance HGF mitogenicity by coupling Met to Shp-2 and GATA-2 expression, thereby potentially contributing to normal myeloid differentiation as well as oncogenic transformation. PMID:20506405

Met-Activating Genetically Improved Chimeric Factor-1 Promotes Angiogenesis and Hypertrophy in Adult Myogenesis.

PubMed

Ronzoni, Flavio; Ceccarelli, Gabriele; Perini, Ilaria; Benedetti, Laura; Galli, Daniela; Mulas, Francesca; Balli, Martina; Magenes, Giovanni; Bellazzi, Riccardo; De Angelis, Gabriella C; Sampaolesi, Maurilio

2017-01-01

Myogenic progenitor cells (activated satellite cells) are able to express both HGF and its receptor cMet. After muscle injury, HGF-Met stimulation promotes activation and primary division of satellite cells. MAGIC-F1 (Met-Activating Genetically Improved Chimeric Factor-1) is an engineered protein that contains two human Met-binding domains that promotes muscle hypertrophy. MAGIC-F1 protects myogenic precursors against apoptosis and increases their fusion ability enhancing muscle differentiation. Hemizygous and homozygous Magic-F1 transgenic mice displayed constitutive muscle hypertrophy. Here we describe microarray analysis on Magic-F1 myogenic progenitor cells showing an altered gene signatures on muscular hypertrophy and angiogenesis compared to wild-type cells. In addition, we performed a functional analysis on Magic-F1+/+ transgenic mice versus controls using treadmill test. We demonstrated that Magic-F1+/+ mice display an increase in muscle mass and cross-sectional area leading to an improvement in running performance. Moreover, the presence of MAGIC-F1 affected positively the vascular network, increasing the vessel number in fast twitch fibers. Finally, the gene expression profile analysis of Magic-F1+/+ satellite cells evidenced transcriptomic changes in genes involved in the control of muscle growth, development and vascularisation. We showed that MAGIC-F1-induced muscle hypertrophy affects positively vascular network, increasing vessel number in fast twitch fibers. This was due to unique features of mammalian skeletal muscle and its remarkable ability to adapt promptly to different physiological demands by modulating the gene expression profile in myogenic progenitors. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

Inhibitory effects of hepatocyte growth factor and interleukin-6 on transforming growth factor-beta1 mediated vocal fold fibroblast-myofibroblast differentiation.

PubMed

Vyas, Bimal; Ishikawa, Keiko; Duflo, Suzy; Chen, Xia; Thibeault, Susan L

2010-05-01

The role of myofibroblasts in vocal fold scarring has not been extensively studied, partly because of the lack of a robust in vitro model. The objective of this investigation was to develop and characterize a myofibroblast in vitro model that could be utilized to investigate the molecular mechanism of myofibroblast differentiation and function in injured vocal fold tissue. Differentiation of human primary vocal fold fibroblasts (hVFFs) to myofibroblasts was stimulated with 5, 10, or 20 ng/mL of recombinant transforming growth factor-beta1 (TGF-beta1). Cultures were analyzed by immunofluorescence and Western blotting, with an alpha-smooth muscle actin (alpha-SMA) antibody used as a myofibroblast marker. Normal rabbit vocal folds were treated with 10 ng/mL of TGF-beta1 for 7 days for in vivo corroboration. The effects of interleukin-6 (IL-6) and hepatocyte growth factor (HGF) on myofibroblast differentiation were studied with Western blots. The hVFFs demonstrated positive alpha-SMA labeling in cells stimulated by 10 and 20 ng/mL TGF-beta1, indicating that hVFFs were capable of differentiation to myofibroblasts. Transforming growth factor-beta1 induced the largest increase in alpha-SMA at 10 ng/mL on day 5 of treatment. Both HGF and IL-6 suppressed the expression of TGF-beta1-induced alpha-SMA. Our work characterizes a useful in vitro model of TGF-beta1-mediated vocal fold fibroblast-myofibroblast differentiation. The extent of differentiation appears to be attenuated by HGF, suggesting a potential mechanism to support prior work indicating that HGF plays a protective role in reducing scar formation in vocal fold injuries. Paradoxically, IL-6, which has been shown to play a profibrotic role in dermal studies, also attenuated the TGF-beta1 response.

E2F1 transcription factor and its impact on growth factor and cytokine signaling.

PubMed

Ertosun, Mustafa Gokhan; Hapil, Fatma Zehra; Osman Nidai, Ozes

2016-10-01

E2F1 is a transcription factor involved in cell cycle regulation and apoptosis. The transactivation capacity of E2F1 is regulated by pRb. In its hypophosphorylated form, pRb binds and inactivates DNA binding and transactivating functions of E2F1. The growth factor stimulation of cells leads to activation of CDKs (cyclin dependent kinases), which in turn phosphorylate Rb and hyperphosphorylated Rb is released from E2F1 or E2F1/DP complex, and free E2F1 can induce transcription of several genes involved in cell cycle entry, induction or inhibition of apoptosis. Thus, growth factors and cytokines generally utilize E2F1 to direct cells to either fate. Furthermore, E2F1 regulates expressions of various cytokines and growth factor receptors, establishing positive or negative feedback mechanisms. This review focuses on the relationship between E2F1 transcription factor and cytokines (IL-1, IL-2, IL-3, IL-6, TGF-beta, G-CSF, LIF), growth factors (EGF, KGF, VEGF, IGF, FGF, PDGF, HGF, NGF), and interferons (IFN-α, IFN-β and IFN-γ). Copyright © 2016 Elsevier Ltd. All rights reserved.

The role of FGF2 in migration and tubulogenesis of endothelial progenitor cells in relation to pro-angiogenic growth factor production.

PubMed

Litwin, Monika; Radwańska, Agata; Paprocka, Maria; Kieda, Claudine; Dobosz, Tadeusz; Witkiewicz, Wojciech; Baczyńska, Dagmara

2015-12-01

In recent years, special attention has been paid to finding new pro-angiogenic factors which could be used in gene therapy of vascular diseases such as critical limb ischaemia (CLI). Angiogenesis, the formation of new blood vessels, is a complex process dependent on different cytokines, matrix proteins, growth factors and other pro- or anti-angiogenic stimuli. Numerous lines of evidence suggest that key mediators of angiogenesis, vascular endothelial growth factor (VEGF) and hepatocyte growth factor (HGF) together with fibroblast growth factor2 (FGF2) are involved in regulation of the normal and pathological process of angiogenesis. However, less information is available on the complex interactions between these and other angiogenic factors. The aim of this study was to characterise the effect of fibroblast growth factor2 on biological properties of human endothelial progenitor cells with respect to the expression level of other regulatory cytokines. Ectopic expression of FGF2 in EP cells stimulates their pro-angiogenic behaviour, leading to increased proliferation, migration and tube formation abilities. Moreover, we show that the expression profile of VEGF and other pro-angiogenic cytokines, such as HGF, MCP2, and interleukins, is affected differently by FGF2 in EPC. In conclusion, we provide evidence that FGF2 directly affects not only the biological properties of EP cells but also the expression pattern and secretion of numerous chemocytokines. Our results suggest that FGF2 could be applied in therapeutic approaches for CLI and other ischaemic diseases of the vascular system in vivo.

Sustained release of hepatocyte growth factor by cationic self-assembling peptide/heparin hybrid hydrogel improves β-cell survival and function through modulating inflammatory response

PubMed Central

Liu, Shuyun; Zhang, Lanlan; Cheng, Jingqiu; Lu, Yanrong; Liu, Jingping

2016-01-01

Inflammatory response is a major cause of grafts dysfunction in islet transplantation. Hepatocyte growth factor (HGF) had shown anti-inflammatory activity in multiple diseases. In this study, we aim to deliver HGF by self-assembling peptide/heparin (SAP/Hep) hybrid gel to protect β-cell from inflammatory injury. The morphological and slow release properties of SAPs were analyzed. Rat INS-1 β-cell line was treated with tumor necrosis factor α in vitro and transplanted into rat kidney capsule in vivo, and the viability, apoptosis, function, and inflammation of β-cells were evaluated. Cationic KLD1R and KLD2R self-assembled to nanofiber hydrogel, which showed higher binding affinity for Hep and HGF because of electrostatic interaction. Slow release of HGF from cationic SAP/Hep gel is a two-step mechanism involving binding affinity with Hep and molecular diffusion. In vitro and in vivo results showed that HGF-loaded KLD2R/Hep gel promoted β-cell survival and insulin secretion, and inhibited cell apoptosis, cytokine release, T-cell infiltration, and activation of NFκB/p38 MAPK pathways in β-cells. This study suggested that SAP/Hep gel is a promising carrier for local delivery of bioactive proteins in islet transplantation. PMID:27729786

PHA665752, a small-molecule inhibitor of c-Met, inhibits hepatocyte growth factor-stimulated migration and proliferation of c-Met-positive neuroblastoma cells.

PubMed

Crosswell, Hal E; Dasgupta, Anindya; Alvarado, Carlos S; Watt, Tanya; Christensen, James G; De, Pradip; Durden, Donald L; Findley, Harry W

2009-11-25

c-Met is a tyrosine kinase receptor for hepatocyte growth factor/scatter factor (HGF/SF), and both c-Met and its ligand are expressed in a variety of tissues. C-Met/HGF/SF signaling is essential for normal embryogenesis, organogenesis, and tissue regeneration. Abnormal c-Met/HGF/SF signaling has been demonstrated in different tumors and linked to aggressive and metastatic tumor phenotypes. In vitro and in vivo studies have demonstrated inhibition of c-Met/HGF/SF signaling by the small-molecule inhibitor PHA665752. This study investigated c-Met and HGF expression in two neuroblastoma (NBL) cell lines and tumor tissue from patients with NBL, as well as the effects of PHA665752 on growth and motility of NBL cell lines. The effect of the tumor suppressor protein PTEN on migration and proliferation of tumor cells treated with PHA665752 was also evaluated. Expression of c-Met and HGF in NBL cell lines SH-EP and SH-SY5Y and primary tumor tissue was assessed by immunohistochemistry and quantitative RT-PCR. The effect of PHA665752 on c-Met/HGF signaling involved in NBL cell proliferation and migration was evaluated in c-Met-positive cells and c-Met-transfected cells. The transwell chemotaxis assay and the MTT assay were used to measure migration and proliferation/cell-survival of tumor cells, respectively. The PPAR-gamma agonist rosiglitazone was used to assess the effect of PTEN on PHA665752-induced inhibition of NBL cell proliferation/cell-survival and migration High c-Met expression was detected in SH-EP cells and primary tumors from patients with advanced-stage disease. C-Met/HGF signaling induced both migration and proliferation of SH-EP cells. Migration and proliferation/cell-survival were inhibited by PHA665752 in a dose-dependent manner. We also found that induced overexpression of PTEN following treatment with rosiglitazone significantly enhanced the inhibitory effect of PHA665752 on NBL-cell migration and proliferation. c-Met is highly expressed in most tumors

Potent blockade of hepatocyte growth factor-stimulated cell motility, matrix invasion and branching morphogenesis by antagonists of Grb2 Src homology 2 domain interactions.

PubMed

Atabey, N; Gao, Y; Yao, Z J; Breckenridge, D; Soon, L; Soriano, J V; Burke, T R; Bottaro, D P

2001-04-27

Hepatocyte growth factor (HGF) stimulates mitogenesis, motogenesis, and morphogenesis in a wide range of cellular targets during development, homeostasis and tissue regeneration. Inappropriate HGF signaling occurs in several human cancers, and the ability of HGF to initiate a program of protease production, cell dissociation, and motility has been shown to promote cellular invasion and is strongly linked to tumor metastasis. Upon HGF binding, several tyrosines within the intracellular domain of its receptor, c-Met, become phosphorylated and mediate the binding of effector proteins, such as Grb2. Grb2 binding through its SH2 domain is thought to link c-Met with downstream mediators of cell proliferation, shape change, and motility. We analyzed the effects of Grb2 SH2 domain antagonists on HGF signaling and observed potent blockade of cell motility, matrix invasion, and branching morphogenesis, with ED(50) values of 30 nm or less, but only modest inhibition of mitogenesis. These compounds are 1000-10,000-fold more potent anti-motility agents than any previously characterized Grb2 SH2 domain antagonists. Our results suggest that SH2 domain-mediated c-Met-Grb2 interaction contributes primarily to the motogenic and morphogenic responses to HGF, and that these compounds may have therapeutic application as anti-metastatic agents for tumors where the HGF signaling pathway is active.

Analysis of soluble factors in conditioned media derived from primary cultures of cirrhotic liver of biliary atresia.

PubMed

Yamazaki, Taisuke; Wakai, Mariko; Enosawa, Shin; Tokiwa, Takayoshi

2017-06-01

Biliary atresia (BA) is a rare and serious liver disease in newborn infants. Previously, we reported that non-parenchymal cell (NPC) fractions from cirrhotic liver of BA may contain hepatic stem/progenitor cells in primary culture of NPC fractions. In this study, NPC fractions were subjected to primary or passage culture and found that clusters of hepatocyte-like cells appear even without adding hepatocyte growth factor (HGF) to the culture medium, but not in their passage culture used as a control. Based on these findings, conditioned media (CMs) were collected and soluble factors in the CMs were analyzed in order to elucidate the mechanism of the appearance of hepatocyte-like cells or their clusters. A large amount of active HGF consisting of α and β chains was detected in CMs derived from primary culture, but not in CMs from passage culture, as determined by western blot analysis, bone morphogenetic protein (BMP)-4, oncostatin M (OSM), and transforming growth factor (TGF)-β1 were not detected in any of the CMs. The number of hepatocyte-like cells in primary culture tended to decrease following treatment with the HGF receptor c-Met inhibitor, SU11274 in a dose-dependent manner. Furthermore, the clusters of hepatocyte-like cells tended to increase in size and number when freshly isolated NPC fractions were cultured in the presence of 10% of CMs collected after 3-4 wk of primary culture. In conclusion, these findings indicate that CMs derived from primary culture of NPC fractions of BA liver contain a large amount of active HGF, which may activate hepatic stem/progenitor cells and promote the appearance of hepatocyte-like cells or their clusters through HGF/c-Met signaling. The present study would lead to cell therapy using the patient's own cells for the treatment of BA.

N-WASP and WAVE2 acting downstream of phosphatidylinositol 3-kinase are required for myogenic cell migration induced by hepatocyte growth factor.

PubMed

Kawamura, Kazuhiro; Takano, Kazunori; Suetsugu, Shiro; Kurisu, Shusaku; Yamazaki, Daisuke; Miki, Hiroaki; Takenawa, Tadaomi; Endo, Takeshi

2004-12-24

During skeletal muscle regeneration caused by injury, muscle satellite cells proliferate and migrate toward the site of muscle injury. This migration is mainly induced by hepatocyte growth factor (HGF) secreted by intact myofibers and also released from injured muscle. However, the intracellular machinery for the satellite cell migration has not been elucidated. To examine the mechanisms of satellite cell migration, we utilized satellite cell-derived mouse C2C12 skeletal muscle cells. HGF induced reorganization of actin cytoskeleton to form lamellipodia in C2C12 myoblasts. HGF treatment facilitated both nondirectional migration of the myoblasts in phagokinetic track assay and directional chemotactic migration toward HGF in a three-dimensional migration chamber assay. Endogenous N-WASP and WAVE2 were concentrated in the lamellipodia at the leading edge of the migrating cells. Moreover, exogenous expression of wild-type N-WASP or WAVE2 promoted lamellipodial formation and migration. By contrast, expression of the dominant-negative mutant of N-WASP or WAVE2 and knockdown of N-WASP or WAVE2 expression by the RNA interference prevented the HGF-induced lamellipodial formation and migration. When the cells were treated with LY294002, an inhibitor of phosphatidylinositol 3-kinase, the HGF-induced lamellipodial formation and migration were abrogated. These results imply that both N-WASP and WAVE2, which are activated downstream of phosphati-dylinositol 3-kinase, are required for the migration through the lamellipodial formation of C2C12 cells induced by HGF.

Independent association of elevated serum hepatocyte growth factor levels with development of insulin resistance in a 10-year prospective study.

PubMed

Tsukagawa, Eri; Adachi, Hisashi; Hirai, Yuji; Enomoto, Mika; Fukami, Ako; Ogata, Kinuka; Kasahara, Akiko; Yokoi, Kanako; Imaizumi, Tsutomu

2013-07-01

Hepatocyte growth factor (HGF) receptors form a hybrid complex with insulin receptors in the liver of mice, which lead to robust signalling to regulate glucose metabolism. Serum HGF levels are high in subjects with metabolic syndrome and/or obesity. Accordingly, we prospectively investigated the relationship between HGF and the development of insulin resistance (IR) in a general population without IR at baseline. A total of 1492 subjects received health examinations. After excluding subjects with diabetes and/or IR (n = 402) at baseline, the remaining subjects (n = 1090) were followed-up 10 years later. Complete data sets were available from 716 subjects for prospective analysis. Logistic regression was performed to determine factors associated with the development of IR after 10 years. In subjects without diabetes at baseline, serum HGF levels were higher (0·26 ± 0·10 ng/ml, n = 259) in subjects with IR than without it (0·22 ± 0·09 ng/ml, n = 1090). After deleting subjects who developed liver disease during follow-up, 188 were found to have developed IR at 10 years after the original screening. HGF (P < 0·05), age (P < 0·001), homoeostasis model assessment index (P < 0·001), HDL-c (P < 0·05; inversely) and hypertensive medication (P < 0·05) were significantly associated with the development of IR by multivariate stepwise logistic regression analysis. A significant (P < 0·05) relative risk [1·75 (95%CI: 1·01-3·12)] for the development of IR was observed in the highest (≥0·30 ng/ml) vs the lowest categories (<0·15 ng/ml) of HGF after adjustments for confounders. Our 10-year prospective study suggests that elevated serum HGF levels were significantly associated with the development of IR. © 2012 John Wiley & Sons Ltd.

Quantitative Analysis of Cancer Cell Migration in Gradients Of EGF, HGF, and SDF-alpha Using a Microfluidic Chemotaxis Device

DTIC Science & Technology

2005-01-01

Quantitative Analysis of Cancer Cell Migration in Gradients of EGF, HGF, and SDF-alpha Using a Microfluidic Chemotaxis Device The University of California...allowing for parallel analysis . Additionally, simple methods of localizing gels into microdevices are demonstrated. The device was characterized by...To overcome some of these drawbacks, several approaches have utilized free diffusion to produce gradients in static environ - ments.5-9 However

Markers of hepatic regeneration associated with surgical attenuation of congenital portosystemic shunts in dogs.

PubMed

Tivers, Michael S; Lipscomb, Victoria J; Smith, Kenneth C; Wheeler-Jones, Caroline P D; House, Arthur K

2014-05-01

Dogs with congenital portosystemic shunts (CPSS) have liver hypoplasia and hepatic insufficiency. Surgical CPSS attenuation results in liver growth associated with clinical improvement. The mechanism of this hepatic response is unknown, although liver regeneration is suspected. This study investigated whether markers of liver regeneration were associated with CPSS attenuation. Dogs treated with CPSS attenuation were prospectively recruited. Residual liver tissue was collected for gene expression analysis (seven genes) from 24 CPSS dogs that tolerated complete attenuation, 25 dogs that tolerated partial attenuation and seven control dogs. Relative gene expression was measured using quantitative polymerase chain reaction (qPCR). Blood samples were collected before, 24 h and 48 h post-surgery from 36 CPSS dogs and from 10 control dogs. Serum hepatocyte growth factor (HGF) concentration was measured using a canine specific enzyme-linked immunosorbent assay (ELISA). HGF mRNA expression was significantly decreased in CPSS compared with control dogs (P = 0.046). There were significant increases in HGF (P = 0.050) and methionine adenosyltransferase 2 A (MAT2A; P = 0.002) mRNA expression following partial CPSS attenuation. Dogs with complete attenuation had significantly greater MAT2A (P = 0.024) mRNA expression compared with dogs with partial attenuation. Serum HGF concentration significantly increased 24 h following CPSS attenuation (P < 0.001). Hepatic mRNA expression of two markers of hepatocyte proliferation (HGF and MAT2A) was associated with the response to surgery in dogs with CPSS, and serum HGF significantly increased following surgery, suggesting hepatocyte proliferation. These findings support the concept that hepatic regeneration is important in the hepatic response to CPSS surgery. Copyright © 2014 Elsevier Ltd. All rights reserved.

Clinical Implications of Hepatocyte Growth Factor, Interleukin-20, and Interleukin-22 in Serum and Bronchoalveolar Fluid of Patients with Non-Small Cell Lung Cancer.

PubMed

Naumnik, W; Naumnik, B; Niklińska, W; Ossolińska, M; Chyczewska, E

2016-01-01

Hepatocyte growth factor (HGF) is involved in tumorigenesis, interleukin-20 (IL-20) is an inhibitor of angiogenesis, and interleukin-22 (IL-22) stimulates tumor growth. The aim of this study was to determine the level of HGF, IL-20, and IL-22 in both serum and bronchoalveolar lavage fluid (BALF) of non-small cell lung cancer (NSCLC) patients before onset of chemotherapy, the nature of the interrelationships between these markers, and their prognostic significance regarding post-chemotherapy survival time. We studied 46 NSCLC patients and 15 healthy subjects as a control group. We found significantly higher serum levels of HGF and IL-22 in the NSCLC patients than those in controls [pg/ml: HGF - 1911 (693-6510) vs. 1333 (838-3667), p = 0.0004; IL-22 - 10.66 (1.44-70.34) vs. 4.69 (0.35-12.29), p = 0.0007]. In contrast, concentrations of HGF and IL-22 in BALF were lower in NSCLC patients than those in controls [pg/ml: HGF - 72 (6-561) vs. 488 (14-2003), p = 0.0002; IL-22 - 2.28 (0.70-6.52) vs. 3.72 (2.76-5.64), p = 0.002]. In the NSCLC patients, there was a negative correlation between the serum level of IL-20 and time to tumor progression (r = -0.405, p = 0.04) and between the serum level of HGF and survival time (r = -0.41, p = 0.005). In addition, a higher serum level of HGF and a higher BALF level of IL-22 in patients were linked with a shorter overall survival. We conclude that HGF, IL-20, and IL-22 in the serum and BALF of NSCLC patients before chemotherapy may be a prognostic of cancer progression.

Preclinical Evaluation of MET Inhibitor INC-280 With or Without the Epidermal Growth Factor Receptor Inhibitor Erlotinib in Non–Small-Cell Lung Cancer

PubMed Central

Lara, Matthew S.; Holland, William S.; Chinn, Danielle; Burich, Rebekah A.; Lara, Primo N.; Gandara, David R.; Kelly, Karen; Mack, Philip C.

2018-01-01

The MET inhibitor INC-280 restored sensitivity to erlotinib and promoted apoptosis in non–small-cell lung cancer models rendered resistant to erlotinib by hepatocyte growth factor. Background Although the epidermal growth factor receptor (EGFR) inhibitor erlotinib is initially effective in non–small-cell lung cancer (NSCLC) patients with tumors harboring activating mutations of EGFR, most subsequently develop acquired resistance. One recognized resistance mechanism occurs through activation of bypass signaling via the hepatocyte growth factor (HGF)-MET pathway. INC-280 is a small molecule kinase inhibitor of MET. We sought to demonstrate the activity of INC-280 on select NSCLC cell lines both as a single agent and in combination with erlotinib using exogenous HGF to simulate MET up-regulation. Methods Four NSCLC cell lines (HCC827, PC9, H1666, and H358) were treated with either single-agent INC-280 or in combination with erlotinib with or without HGF. The activity of the drug treatments was measured by cell viability assays. Immunoblotting was used to monitor expression of EGFR/pEGFR, MET/pMET, GAB1/pGAB1, AKT/pAKT, and ERK/pERK as well as markers of apoptosis (PARP and capase-3 cleavage) in H1666, HCC827, and PC9. Results As a single agent, INC-280 showed minimal cytotoxicity despite potent inhibition of MET kinase activity at concentrations as low as 10 nM. Addition of HGF prevented erlotinib-induced cell death. The addition of INC280 to HGF-mediated erlotinib-resistant models restored erlotinib sensitivity for all cell lines tested, associated with cleavage of both PARP and caspase-3. In these models, INC-280 treatment was sufficient to restore erlotinib-induced inhibition of MET, GAB1, AKT, and ERK in the presence of HGF. Conclusion Although the MET inhibitor INC-280 alone had no discernible effect on cell growth, it was able to restore sensitivity to erlotinib and promote apoptosis in NSCLC models rendered erlotinib resistant by HGF. These data provide a

[The expression of serum hepatocyte growth factor in OSAHS].

PubMed

Zhou, S L; Meng, B; Ding, J H

2017-05-05

Objective: To investigate the clinical significance of detecting peripheral blood hepatocyte growth factor（HGF） in OSAHS patients. Method: Ninety-six cases of OSAHS patients in our hospital were selected as OSAHS group,and were divided into 3 subgroups according to the PSG results：mild,medium and severe. Each group included 32 cases,Thirty-five cases of healthy persons were selected as control group. ELISA method was utilized to detect the HGF level of peripheral blood. HGF concentration was measured in 32 patients with severe OSAHS after 3 months of comprehensive treatment. The relationship between serum HGF and sleep respiration events was further analyzed. Result: The HGF concentration of peripheral blood increased with the severity of OSAHS.The serum levels of HGF in the control,mild,medium and severe group were（487.75±46.74）pg/ml，（519.44±50.77）pg/ml，（753.52±58.91） pg/ml and（829.49±61.74）pg/ml,respectively. There were significant differences among groups（ F =117．733, P <0.01). HGF concentration in peripheral blood of OSAHS patients was unrelated to sex,age,and BMI（ P >0.05）,and positively correlated with AHI,negatively correlated with LSaO₂（ P <0.01）. After comprehensive treatment,the serum HGF concentration and AHI in severe OSAHS group were significantly decreased,while LSaO₂ was significantly increased. Conclusion: The level of HGF was increased in OSAHS patients and was positively correlated with the severity of OSAHS. Determining the level of HGF in peripheral blood is important for evaluating the severity of OSAHS and the degree of vascular endothelial dysfunction,and assessing the risk of cardiovascular disease. Copyright© by the Editorial Department of Journal of Clinical Otorhinolaryngology Head and Neck Surgery.

[Influence of hepatocyte growth factor on iNOS, NO and IL-1β in the cerebrum during cerebral ischemia/reperfusion in rats].

PubMed

He, Fang; Ye, Bei; Chen, Jianzhen; Sun, Xiaoyan; Li, Chang

2014-01-01

To explore the effect of hepatocyte growth factor (HGF) on inducible nitric oxide synthase (iNOS), NO and interleukin-1β (IL-1β) in the cerebrum of rats subjected to cerebral ischemia/reperfusion (I/R). Sprague-Dawley rats were randomly divided into 5 groups: a sham group, an I/R group,an HGF1 group, an HGF2 group, and an HGF3 group. The latter 3 groups were respectively injected 15, 30 and 60 μg/kg HGF. The focal cerebral I/R model was established by sutureoccluded method. After 1.5 h ischemia followed by 24 h reperfusion, the iNOS activity and NO content in the ischemic cerebral tissue were assessed. The expression of iNOS mRNA and IL-1β mRNA was detected. The level of iNOS protein and IL-1β content were determined. In addition, cultured cerebral cortical neurons in vitro were exposed to I/R. Then the expression of iNOS and IL-1β protein in the neurons was detected, and NO content was assessed. The iNOS activity and NO content in the ischemic cerebral tissue were increased. The expression of iNOS mRNA and IL-1β mRNA was upregulated. The level of iNOS protein and IL- 1β content were increased. Administration of HGF decreased the iNOS activity and NO content, and downregulated the expression of iNOS mRNA, IL-1β mRNA, iNOS protein and IL-1β content in the ischemic cerebral tissue. HGF decreased the expression of IL-1β, iNOS protein and NO content in the cortical neurons exposed to I/R in vitro. HGF can inhibit the expression of IL-1β and decrease the expression of iNOS and content of NO, which is probably one of the mechanisms mediating the protection of HGF against cerebral ischemia injury.

Chromatin remodeling agent trichostatin A: a key-factor in the hepatic differentiation of human mesenchymal stem cells derived of adult bone marrow

PubMed Central

Snykers, Sarah; Vanhaecke, Tamara; De Becker, Ann; Papeleu, Peggy; Vinken, Mathieu; Van Riet, Ivan; Rogiers, Vera

2007-01-01

Background The capability of human mesenchymal stem cells (hMSC) derived of adult bone marrow to undergo in vitro hepatic differentiation was investigated. Results Exposure of hMSC to a cocktail of hepatogenic factors [(fibroblast growth factor-4 (FGF-4), hepatocyte growth factor (HGF), insulin-transferrin-sodium-selenite (ITS) and dexamethasone)] failed to induce hepatic differentiation. Sequential exposure to these factors (FGF-4, followed by HGF, followed by HGF+ITS+dexamethasone), however, resembling the order of secretion during liver embryogenesis, induced both glycogen-storage and cytokeratin (CK)18 expression. Additional exposure of the cells to trichostatin A (TSA) considerably improved endodermal differentiation, as evidenced by acquisition of an epithelial morphology, chronological expression of hepatic proteins, including hepatocyte-nuclear factor (HNF)-3β, alpha-fetoprotein (AFP), CK18, albumin (ALB), HNF1α, multidrug resistance-associated protein (MRP)2 and CCAAT-enhancer binding protein (C/EBP)α, and functional maturation, i.e. upregulated ALB secretion, urea production and inducible cytochrome P450 (CYP)-dependent activity. Conclusion hMSC are able to undergo mesenchymal-to-epithelial transition. TSA is hereby essential to promote differentiation of hMSC towards functional hepatocyte-like cells. PMID:17407549

Hepatocyte Growth Factor-c-MET Signaling Mediates the Development of Nonsensory Structures of the Mammalian Cochlea and Hearing.

PubMed

Shibata, Shumei; Miwa, Toru; Wu, Hsiao-Huei; Levitt, Pat; Ohyama, Takahiro

2016-08-03

The stria vascularis is a nonsensory structure that is essential for auditory hair cell function by maintaining potassium concentration of the scala media. During mouse embryonic development, a subpopulation of neural crest cell-derived melanocytes migrates and incorporates into a subregion of the cochlear epithelium, forming the intermediate cell layer of the stria vascularis. The relation of this developmental process to stria vascularis function is currently unknown. In characterizing the molecular differentiation of developing peripheral auditory structures, we discovered that hepatocyte growth factor (Hgf) is expressed in the future stria vascularis of the cochlear epithelium. Its receptor tyrosine kinase, c-Met, is expressed in the cochlear epithelium and melanocyte-derived intermediate cells in the stria vascularis. Genetic dissection of HGF signaling via c-MET reveals that the incorporation of the melanocytes into the future stria vascularis of the cochlear duct requires c-MET signaling. In addition, inactivation of either the ligand or receptor developmentally resulted in a profound hearing loss at young adult stages. These results suggest a novel connection between HGF signaling and deafness via melanocyte deficiencies. We found the roles of hepatocyte growth factor (HGF) signaling in stria vascularis development for the first time and that lack of HGF signaling in the inner ear leads to profound hearing loss in the mouse. Our findings reveal a novel mechanism that may underlie human deafness DFNB39 and DFNB97. Our findings reveal an additional example of context-dependent c-MET signaling diversity, required here for proper cellular invasion developmentally that is essential for specific aspects of auditory-related organogenesis. Copyright © 2016 the authors 0270-6474/16/368200-10$15.00/0.

Plasma vemurafenib exposure and pre-treatment hepatocyte growth factor level are two factors contributing to the early peripheral lymphocytes depletion in BRAF-mutated melanoma patients.

PubMed

Puszkiel, Alicja; White-Koning, Mélanie; Dupin, Nicolas; Kramkimel, Nora; Thomas-Schoemann, Audrey; Noé, Gaëlle; Chapuis, Nicolas; Vidal, Michel; Goldwasser, François; Chatelut, Etienne; Blanchet, Benoit

2016-11-01

The therapeutic response to vemurafenib, a BRAF serine-threonine kinase inhibitor, exhibits large variations between patients. Evaluation of factors predicting the clinical efficacy of vemurafenib may help to identify patients at high risk of non-response in the early phase of treatment. The aim of this study was to analyze the pharmacokinetics of vemurafenib by a population approach and to evaluate the relationship between plasma drug exposure and pre-treatment plasma hepatocyte growth factor (HGF) levels with clinical effects (progression-free survival (PFS), peripheral lymphocytes depletion) in patients with metastatic BRAF V600 mutated melanoma treated with single agent vemurafenib. Concentration-time data (n=332) obtained in 44 patients were analyzed using the NONMEM program. Pre-treatment plasma levels of HGF (n=36) were assayed by ELISA method. A Cox model was used to identify prognostic factors associated with progression-free survival (PFS), and a linear regression to identify factors contributing to the depletion of peripheral lymphocytes at day 15. Steady-state pharmacokinetics of vemurafenib was described by a one compartment model with first order absorption and first order elimination. None of the tested covariates explained the inter-patient variability in CL/F. A significant decrease in total lymphocytes count was observed within the first 15days (median ratio Day15/Day0=0.66, p<0.0001). Patients with Day15/Day0 ratio below 0.66 had longer PFS (14 vs 4 months, HR=0.41, CI95%=[0.15-0.77], p=0.0095). In the multivariate Cox model analysis, ECOG PS was the only parameter independently associated with PFS (grade 1 vs 0, HR=3.26, CI95%=[1.29-8.22], p=0.01 and grade ≥2 vs 0, HR=4.77, CI95%=[1.52-14.95], p=0.007). Plasma vemurafenib exposure (p=0.046) and pre-treatment HGF levels (p=0.003) were independently associated with the total lymphocyte ratio Day15/Day0. These findings show that plasma vemurafenib exposure and pre-treatment HGF levels are two

Characterization and profiling of immunomodulatory genes of equine mesenchymal stromal cells from non-invasive sources

PubMed Central

2014-01-01

only chondro- and adipogenic differentiation could be confirmed. Regardless of the source, equine MSCs expressed the immunomodulatory genes CD40, CD80, HGF, and transforming growth factor-beta (TGFβ). In contrast, no mRNA expression was found for CD86, indoleamine 2,3-dioxygenase (IDO), and tumor necrosis factor-alpha (TNFα). Conclusions Whereas UCM seems less feasible because of the high contamination risks and low isolation success rates, UCB seems a promising alternative MSC source, especially when considering allogeneic MSC use. PMID:24418262

Potent HGF/c-Met axis inhibitors from Eucalyptus globulus: the coupling of phloroglucinol and sesquiterpenoid is essential for the activity.

PubMed

Yang, Sheng-Ping; Zhang, Xiao-Wei; Ai, Jing; Gan, Li-She; Xu, Jin-Biao; Wang, Ying; Su, Zu-Shang; Wang, Lu; Ding, Jian; Geng, Mei-Yu; Yue, Jian-Min

2012-09-27

Eucalyptin A (1), together with two known compounds 2 and 3 exhibiting potent inhibition on HGF/c-Met axis, was discovered from the fruits of Eucalyptus globulus. 1 possessed an unprecedented carbon framework of phloroglucinol-coupled sesquiterpenoid, and its structure was elucidated by spectroscopic method and ECD calculation. A brief structure-activity relationship discussion indicated that the coupling of a phloroglucinol and a sesquiterpenoid is essential for the activity.

Monocarboxylate transporter 1 contributes to growth factor-induced tumor cell migration independent of transporter activity

PubMed Central

Gray, Alana L.; Coleman, David T.; Shi, Runhua; Cardelli, James A.

2016-01-01

Tumor progression to metastatic disease contributes to the vast majority of incurable cancer. Understanding the processes leading to advanced stage cancer is important for the development of future therapeutic strategies. Here, we establish a connection between tumor cell migration, a prerequisite to metastasis, and monocarboxylate transporter 1 (MCT1). MCT1 transporter activity is known to regulate aspects of tumor progression and, as such, is a clinically relevant target for treating cancer. Knockdown of MCT1 expression caused decreased hepatocyte growth factor (HGF)-induced as well as epidermal growth factor (EGF)-induced tumor cell scattering and wound healing. Western blot analysis suggested that MCT1 knockdown (KD) hinders signaling through the HGF receptor (c-Met) but not the EGF receptor. Exogenous, membrane-permeable MCT1 substrates were not able to rescue motility in MCT1 KD cells, nor was pharmacologic inhibition of MCT1 able to recapitulate decreased cell motility as seen with MCT1 KD cells, indicating transporter activity of MCT1 was dispensable for EGF- and HGF-induced motility. These results indicate MCT1 expression, independent of transporter activity, is required for growth factor-induced tumor cell motility. The findings presented herein suggest a novel function for MCT1 in tumor progression independent of its role as a monocarboxylate transporter. PMID:27127175

Physiological factors that regulate skin pigmentation

PubMed Central

Yamaguchi, Yuji; Hearing, Vincent J.

2009-01-01

More than 150 genes have been identified that affect skin color either directly or indirectly, and we review current understanding of physiological factors that regulate skin pigmentation. We focus on melanosome biogenesis, transport and transfer, melanogenic regulators in melanocytes and factors derived from keratinocytes, fibroblasts, endothelial cells, hormones, inflammatory cells and nerves. Enzymatic components of melanosomes include tyrosinase, tyrosinase-related protein 1 and dopachrome tautomerase, which depend on the functions of OA1, P, MATP, ATP7A and BLOC-1 to synthesize eumelanins and pheomelanins. The main structural component of melanosomes is Pmel17/gp100/Silv, whose sorting involves adaptor protein 1A (AP1A), AP1B, AP2 and spectrin, as well as a chaperone-like component, MART-1. During their maturation, melanosomes move from the perinuclear area toward the plasma membrane. Microtubules, dynein, kinesin, actin filaments, Rab27a, melanophilin, myosin Va and Slp2-a are involved in melanosome transport. Foxn1 and p53 up-regulate skin pigmentation via bFGF and POMC derivatives including α-MSH and ACTH, respectively. Other critical factors that affect skin pigmentation include MC1R, CREB, ASP, MITF, PAX3, SOX9/10, LEF-1/TCF, PAR-2, DKK1, SCF, HGF, GM-CSF, endothelin-1, prostaglandins, leukotrienes, thromboxanes, neurotrophins and neuropeptides. UV radiation up-regulates most factors that increase melanogenesis. Further studies will elucidate the currently unknown functions of many other pigment genes/proteins. PMID:19449448

Migration of bone marrow and cord blood mesenchymal stem cells in vitro is regulated by stromal-derived factor-1-CXCR4 and hepatocyte growth factor-c-met axes and involves matrix metalloproteinases.

PubMed

Son, Bo-Ra; Marquez-Curtis, Leah A; Kucia, Magda; Wysoczynski, Marcin; Turner, A Robert; Ratajczak, Janina; Ratajczak, Mariusz Z; Janowska-Wieczorek, Anna

2006-05-01

Human mesenchymal stem cells (MSCs) are increasingly being considered in cell-based therapeutic strategies for regeneration of various organs/tissues. However, the signals required for their homing and recruitment to injured sites are not yet fully understood. Because stromal-derived factor (SDF)-1 and hepatocyte growth factor (HGF) become up-regulated during tissue/organ damage, in this study we examined whether these factors chemoattract ex vivo-expanded MSCs derived from bone marrow (BM) and umbilical cord blood (CB). Specifically, we investigated the expression by MSCs of CXCR4 and c-met, the cognate receptors of SDF-1 and HGF, and their functionality after early and late passages of MSCs. We also determined whether MSCs express matrix metalloproteinases (MMPs), including membrane type 1 (MT1)-MMP, matrix-degrading enzymes that facilitate the trafficking of hematopoietic stem cells. We maintained expanded BM- or CB-derived MSCs for up to 15-18 passages with monitoring of the expression of 1) various tissue markers (cardiac and skeletal muscle, neural, liver, and endothelial cells), 2) functional CXCR4 and c-met, and 3) MMPs. We found that for up to 15-18 passages, both BM- and CB-derived MSCs 1) express mRNA for cardiac, muscle, neural, and liver markers, as well as the vascular endothelial (VE) marker VE-cadherin; 2) express CXCR4 and c-met receptors and are strongly attracted by SDF-1 and HGF gradients; 3) express MMP-2 and MT1-MMP transcripts and proteins; and 4) are chemo-invasive across the reconstituted basement membrane Matrigel. These in vitro results suggest that the SDF-1-CXCR4 and HGF-c-met axes, along with MMPs, may be involved in recruitment of expanded MSCs to damaged tissues.

Effect of two different preparations of platelet-rich plasma on synoviocytes.

PubMed

Assirelli, Elisa; Filardo, Giuseppe; Mariani, Erminia; Kon, Elizaveta; Roffi, Alice; Vaccaro, Franca; Marcacci, Maurilio; Facchini, Andrea; Pulsatelli, Lia

2015-09-01

To analyse the modifications induced by two different platelet-rich plasma (PRP) preparations on osteoarthritis (OA) synoviocytes, by documenting changes in gene expression of factors involved in joint physiopathology. OA synoviocytes were cultured for 7 days in medium with different concentrations of either P-PRP (a pure platelet concentrate without leucocytes but with a limited number of platelets), L-PRP (a higher platelet concentrate containing leucocytes) or platelet-poor plasma (PPP). Gene expression of interleukin (IL)-1beta, IL-6, IL-8/CXCL8, tumour necrosis factor alpha, IL-10, IL-4, IL-13, metalloproteinase-13, tissue inhibitor of metalloproteinase (TIMP)-1, (TIMP)-3, (TIMP)-4, vascular endothelial growth factor, transforming growth factor beta1, fibroblast growth factor (FGF)-2, hepatocyte growth factor (HGF), hyaluronic acid (HA) synthases (HAS)-1, (HAS)-2, and (HAS)-3 was analysed by RT-PCR. HA production was determined in culture supernatants by ELISA. IL-1β, IL-8 and FGF-2 were significantly induced by L-PRP compared to both P-PRP and PPP; HGF was down-modulated by L-PRP versus both P-PRP and PPP, and an inverse dose-response influence was shown for all preparations. Expression level of TIMP-4 was lower in the presence of L-PRP compared with P-PRP. HA production and HAS gene expression did not seem to be modulated by PRP. L-PRP is able to sustain the up-regulation of proinflammatory factors, (IL-1beta, IL-8 and FGF-2), together with a down-modulation of HGF and TIMP-4 expression, two factors that have been recognized as anti-catabolic mediators in cartilage, thus supporting the need to further optimize the PRP preparations to be applied in clinical practice.

Sustained functional improvement by hepatocyte growth factor-like small molecule BB3 after focal cerebral ischemia in rats and mice

PubMed Central

Chaparro, Rafael E; Izutsu, Miwa; Sasaki, Toshihiro; Sheng, Huaxin; Zheng, Yi; Sadeghian, Homa; Qin, Tao; von Bornstadt, Daniel; Herisson, Fanny; Duan, Bin; Li, Jing-Song; Jiang, Kai; Pearlstein, Molly; Pearlstein, Robert D; Smith, David E; Goldberg, Itzhak D; Ayata, Cenk; Warner, David S

2015-01-01

Hepatocyte growth factor (HGF), efficacious in preclinical models of acute central nervous system injury, is burdened by administration of full-length proteins. A multiinstitutional consortium investigated the efficacy of BB3, a small molecule with HGF-like activity that crosses the blood–brain barrier in rodent focal ischemic stroke using Stroke Therapy Academic Industry Roundtable (STAIR) and Good Laboratory Practice guidelines. In rats, BB3, begun 6 hours after temporary middle cerebral artery occlusion (tMCAO) reperfusion, or permanent middle cerebral artery occlusion (pMCAO) onset, and continued for 14 days consistently improved long-term neurologic function independent of sex, age, or laboratory. BB3 had little effect on cerebral infarct size and no effect on blood pressure. BB3 increased HGF receptor c-Met phosphorylation and synaptophysin expression in penumbral tissue consistent with a neurorestorative mechanism from HGF-like activity. In mouse tMCAO, BB3 starting 10 minutes after reperfusion and continued for 14 days improved neurologic function that persisted for 8 weeks in some, but not all measures. Study in animals with comorbidities and those exposed to common stroke drugs are the next steps to complete preclinical assessment. These data, generated in independent, masked, and rigorously controlled settings, are the first to suggest that the HGF pathway can potentially be harnessed by BB3 for neurologic benefit after ischemic stroke. PMID:25712497

The anti-fibrotic effects of mesenchymal stem cells on irradiated lungs via stimulating endogenous secretion of HGF and PGE2

PubMed Central

Dong, Li-Hua; Jiang, Yi-Yao; Liu, Yong-Jun; Cui, Shuang; Xia, Cheng-Cheng; Qu, Chao; Jiang, Xin; Qu, Ya-Qin; Chang, Peng-Yu; Liu, Feng

2015-01-01

Radiation-induced pulmonary fibrosis is a common disease and has a poor prognosis owing to the progressive breakdown of gas exchange regions in the lung. Recently, a novel strategy of administering mesenchymal stem cells for pulmonary fibrosis has achieved high therapeutic efficacy. In the present study, we attempted to use human adipose tissue-derived mesenchymal stem cells to prevent disease in Sprague-Dawley rats that received semi-thoracic irradiation (15 Gy). To investigate the specific roles of mesenchymal stem cells in ameliorating radiation-induced pulmonary fibrosis, we treated control groups of irradiated rats with human skin fibroblasts or phosphate-buffered saline. After mesenchymal stem cells were infused, host secretions of hepatocyte growth factor (HGF) and prostaglandin E2 (PGE2) were elevated compared with those of the controls. In contrast, tumour necrosis factor-alpha (TNF-α) and transforming growth factor-beta1 (TGF-β1) levels were decreased after infusion of mesenchymal stem cells. Consequently, the architecture of the irradiated lungs was preserved without marked activation of fibroblasts or collagen deposition within the injured sites. Moreover, mesenchymal stem cells were able to prevent the irradiated type II alveolar epithelial cells from undergoing epithelial-mesenchymal transition. Collectively, these data confirmed that mesenchymal stem cells have the potential to limit pulmonary fibrosis after exposure to ionising irradiation. PMID:25736907

Requirement of kinesin-mediated membrane transport of WAVE2 along microtubules for lamellipodia formation promoted by hepatocyte growth factor.

PubMed

Takahashi, Kazuhide; Suzuki, Katsuo

2008-07-01

Lamellipodia formation necessary for epithelial cell migration and invasion is accomplished by rearrangement of the actin cytoskeleton at the leading edge through membrane transport of WAVE2. However, how WAVE2 is transported to the cell periphery where lamellipodia are formed remains to be established. We report here that hepatocyte growth factor (HGF) promoted lamellipodia formation and intracellular transport of WAVE2 to the cell periphery, depending on Rac1 activity, in MDA-MB-231 human breast cancer cells. Immunoblot analyses indicating the coimmunoprecipitation of WAVE2 with kinesin heavy chain KIF5B, one of the motor proteins, and IQGAP1 suggest that KIF5B and IQGAP1 formed a complex with WAVE2 in serum-starved cells and increased in their amount after HGF stimulation. Both downregulation of KIF5B by the small interfering RNA and depolymerization of microtubules with nocodazole abrogated the HGF-induced lamellipodia formation and WAVE2 transport. Therefore, we propose here that the promotion of lamellipodia formation by HGF in MDA-MB-231 cells is Rac1-dependent and requires KIF5B-mediated transport of WAVE2 and IQGAP1 to the cell periphery along microtubules.

Cell Population Kinetics of Collagen Scaffolds in Ex Vivo Oral Wound Repair

PubMed Central

Agis, Hermann; Collins, Amy; Taut, Andrei D.; Jin, Qiming; Kruger, Laura; Görlach, Christoph; Giannobile, William V.

2014-01-01

Biodegradable collagen scaffolds are used clinically for oral soft tissue augmentation to support wound healing. This study sought to provide a novel ex vivo model for analyzing healing kinetics and gene expression of primary human gingival fibroblasts (hGF) within collagen scaffolds. Sponge type and gel type scaffolds with and without platelet-derived growth factor-BB (PDGF) were assessed in an hGF containing matrix. Morphology was evaluated with scanning electron microscopy, and hGF metabolic activity using MTT. We quantitated the population kinetics within the scaffolds based on cell density and distance from the scaffold border of DiI-labled hGFs over a two-week observation period. Gene expression was evaluated with gene array and qPCR. The sponge type scaffolds showed a porous morphology. Absolute cell number and distance was higher in sponge type scaffolds when compared to gel type scaffolds, in particular during the first week of observation. PDGF incorporated scaffolds increased cell numbers, distance, and formazan formation in the MTT assay. Gene expression dynamics revealed the induction of key genes associated with the generation of oral tissue. DKK1, CYR61, CTGF, TGFBR1 levels were increased and integrin ITGA2 levels were decreased in the sponge type scaffolds compared to the gel type scaffold. The results suggest that this novel model of oral wound healing provides insights into population kinetics and gene expression dynamics of biodegradable scaffolds. PMID:25397671

Hepatocyte growth factor activator inhibitors (HAI-1 and HAI-2): Emerging key players in epithelial integrity and cancer.

PubMed

Kataoka, Hiroaki; Kawaguchi, Makiko; Fukushima, Tsuyoshi; Shimomura, Takeshi

2018-03-01

The growth, survival, and metabolic activities of multicellular organisms at the cellular level are regulated by intracellular signaling, systemic homeostasis and the pericellular microenvironment. Pericellular proteolysis has a crucial role in processing bioactive molecules in the microenvironment and thereby has profound effects on cellular functions. Hepatocyte growth factor activator inhibitor type 1 (HAI-1) and HAI-2 are type I transmembrane serine protease inhibitors expressed by most epithelial cells. They regulate the pericellular activities of circulating hepatocyte growth factor activator and cellular type II transmembrane serine proteases (TTSPs), proteases required for the activation of hepatocyte growth factor (HGF)/scatter factor (SF). Activated HGF/SF transduces pleiotropic signals through its receptor tyrosine kinase, MET (coded by the proto-oncogene MET), which are necessary for cellular migration, survival, growth and triggering stem cells for accelerated healing. HAI-1 and HAI-2 are also required for normal epithelial functions through regulation of TTSP-mediated activation of other proteases and protease-activated receptor 2, and also through suppressing excess degradation of epithelial junctional proteins. This review summarizes current knowledge regarding the mechanism of pericellular HGF/SF activation and highlights emerging roles of HAIs in epithelial development and integrity, as well as tumorigenesis and progression of transformed epithelial cells. © 2018 Japanese Society of Pathology and John Wiley & Sons Australia, Ltd.

Physiological factors that regulate skin pigmentation.

PubMed

Yamaguchi, Yuji; Hearing, Vincent J

2009-01-01

More than 150 genes have been identified that affect skin color either directly or indirectly, and we review current understanding of physiological factors that regulate skin pigmentation. We focus on melanosome biogenesis, transport and transfer, melanogenic regulators in melanocytes, and factors derived from keratinocytes, fibroblasts, endothelial cells, hormones, inflammatory cells, and nerves. Enzymatic components of melanosomes include tyrosinase, tyrosinase-related protein 1, and dopachrome tautomerase, which depend on the functions of OA1, P, MATP, ATP7A, and BLOC-1 to synthesize eumelanins and pheomelanins. The main structural component of melanosomes is Pmel17/gp100/Silv, whose sorting involves adaptor protein 1A (AP1A), AP1B, AP2, and spectrin, as well as a chaperone-like component, MART-1. During their maturation, melanosomes move from the perinuclear area toward the plasma membrane. Microtubules, dynein, kinesin, actin filaments, Rab27a, melanophilin, myosin Va, and Slp2-a are involved in melanosome transport. Foxn1 and p53 up-regulate skin pigmentation via bFGF and POMC derivatives including alpha-MSH and ACTH, respectively. Other critical factors that affect skin pigmentation include MC1R, CREB, ASP, MITF, PAX3, SOX9/10, LEF-1/TCF, PAR-2, DKK1, SCF, HGF, GM-CSF, endothelin-1, prostaglandins, leukotrienes, thromboxanes, neurotrophins, and neuropeptides. UV radiation up-regulates most factors that increase melanogenesis. Further studies will elucidate the currently unknown functions of many other pigment genes/proteins. (c) 2009 International Union of Biochemistry and Molecular Biology, Inc.

Factor IX gene haplotypes in Amerindians.

PubMed

Franco, R F; Araújo, A G; Zago, M A; Guerreiro, J F; Figueiredo, M S

1997-02-01

We have determined the haplotypes of the factor IX gene for 95 Indians from 5 Brazilian Amazon tribes: Wayampí, Wayana-Apalaí, Kayapó, Arára, and Yanomámi. Eight polymorphisms linked to the factor IX gene were investigated: MseI (at 5', nt -698), BamHI (at 5', nt -561), DdeI (intron 1), BamHI (intron 2), XmnI (intron 3), TaqI (intron 4), MspI (intron 4), and HhaI (at 3', approximately 8 kb). The results of the haplotype distribution and the allele frequencies for each of the factor IX gene polymorphisms in Amerindians were similar to the results reported for Asian populations but differed from results for other ethnic groups. Only five haplotypes were identified within the entire Amerindian study population, and the haplotype distribution was significantly different among the five tribes, with one (Arára) to four (Wayampí) haplotypes being found per tribe. These findings indicate a significant heterogeneity among the Indian tribes and contrast with the homogeneous distribution of the beta-globin gene cluster haplotypes but agree with our recent findings on the distribution of alpha-globin gene cluster haplotypes and the allele frequencies for six VNTRs in the same Amerindian tribes. Our data represent the first study of factor IX-associated polymorphisms in Amerindian populations and emphasizes the applicability of these genetic markers for population and human evolution studies.

Major psychological factors affecting acceptance of gene-recombination technology.

PubMed

Tanaka, Yutaka

2004-12-01

The purpose of this study was to verify the validity of a causal model that was made to predict the acceptance of gene-recombination technology. A structural equation model was used as a causal model. First of all, based on preceding studies, the factors of perceived risk, perceived benefit, and trust were set up as important psychological factors determining acceptance of gene-recombination technology in the structural equation model. An additional factor, "sense of bioethics," which I consider to be important for acceptance of biotechnology, was added to the model. Based on previous studies, trust was set up to have an indirect influence on the acceptance of gene-recombination technology through perceived risk and perceived benefit in the model. Participants were 231 undergraduate students in Japan who answered a questionnaire with a 5-point bipolar scale. The results indicated that the proposed model fits the data well, and showed that acceptance of gene-recombination technology is explained largely by four factors, that is, perceived risk, perceived benefit, trust, and sense of bioethics, whether the technology is applied to plants, animals, or human beings. However, the relative importance of the four factors was found to vary depending on whether the gene-recombination technology was applied to plants, animals, or human beings. Specifically, the factor of sense of bioethics is the most important factor in acceptance of plant gene-recombination technology and animal gene-recombination technology, and the factors of trust and perceived risk are the most important factors in acceptance of human being gene-recombination technology.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Hu Shunying; Chen Yundai; Li Libing

Purpose: Irradiation to the heart may lead to late cardiovascular complications. The purpose of this study was to investigate whether adenovirus-mediated delivery of the human hepatocyte growth factor gene could reduce post-irradiation damage of the rat heart and improve heart function. Methods and Materials: Twenty rats received single-dose irradiation of 20 Gy gamma ray locally to the heart and were randomized into two groups. Two weeks after irradiation, these two groups of rats received Ad-HGF or mock adenovirus vector intramyocardial injection, respectively. Another 10 rats served as sham-irradiated controls. At post-irradiation Day 120, myocardial perfusion was tested by myocardial contrastmore » echocardiography with contrast agent injected intravenously. At post-irradiation Day 180, cardiac function was assessed using the Langendorff technique with an isolated working heart model, after which heart samples were collected for histological evaluation. Results: Myocardial blood flow was significantly improved in HGF-treated animals as measured by myocardial contrast echocardiography at post-irradiation Day 120 . At post-irradiation Day 180, cardiac function was significantly improved in the HGF group compared with mock vector group, as measured by left ventricular peak systolic pressure (58.80 +- 9.01 vs. 41.94 +- 6.65 mm Hg, p < 0.05), the maximum dP/dt (5634 +- 1303 vs. 1667 +- 304 mm Hg/s, p < 0.01), and the minimum dP/dt (3477 +- 1084 vs. 1566 +- 499 mm Hg/s, p < 0.05). Picrosirius red staining analysis also revealed a significant reduction of fibrosis in the HGF group. Conclusion: Based on the study findings, hepatocyte growth factor gene transfer can attenuate radiation-induced cardiac injury and can preserve cardiac function.« less

Tissue-specific epigenetics in gene neighborhoods: myogenic transcription factor genes

PubMed Central

Chandra, Sruti; Terragni, Jolyon; Zhang, Guoqiang; Pradhan, Sriharsa; Haushka, Stephen; Johnston, Douglas; Baribault, Carl; Lacey, Michelle; Ehrlich, Melanie

2015-01-01

Myogenic regulatory factor (MRF) genes, MYOD1, MYOG, MYF6 and MYF5, are critical for the skeletal muscle lineage. Here, we used various epigenome profiles from human myoblasts (Mb), myotubes (Mt), muscle and diverse non-muscle samples to elucidate the involvement of multigene neighborhoods in the regulation of MRF genes. We found more far-distal enhancer chromatin associated with MRF genes in Mb and Mt than previously reported from studies in mice. For the MYF5/MYF6 gene-pair, regions of Mb-associated enhancer chromatin were located throughout the adjacent 236-kb PTPRQ gene even though Mb expressed negligible amounts of PTPRQ mRNA. Some enhancer chromatin regions inside PTPRQ in Mb were also seen in PTPRQ mRNA-expressing non-myogenic cells. This suggests dual-purpose PTPRQ enhancers that upregulate expression of PTPRQ in non-myogenic cells and MYF5/MYF6 in myogenic cells. In contrast, the myogenic enhancer chromatin regions distal to MYOD1 were intergenic and up to 19 kb long. Two of them contain small, known MYOD1 enhancers, and one displayed an unusually high level of 5-hydroxymethylcytosine in a quantitative DNA hydroxymethylation assay. Unexpectedly, three regions of MYOD1-distal enhancer chromatin in Mb and Mt overlapped enhancer chromatin in umbilical vein endothelial cells, which might upregulate a distant gene (PIK3C2A). Lastly, genes surrounding MYOG were preferentially transcribed in Mt, like MYOG itself, and exhibited nearby myogenic enhancer chromatin. These neighboring chromatin regions may be enhancers acting in concert to regulate myogenic expression of multiple adjacent genes. Our findings reveal the very different and complex organization of gene neighborhoods containing closely related transcription factor genes. PMID:26041816

Hepatocyte growth factor sensitizes brain tumors to c-MET kinase inhibition

PubMed Central

Zhang, Ying; Farenholtz, Kaitlyn E.; Yang, Yanzhi; Guessous, Fadila; diPierro, Charles G.; Calvert, Valerie S.; Deng, Jianghong; Schiff, David; Xin, Wenjun; Lee, Jae K.; Purow, Benjamin; Christensen, James; Petricoin, Emanuel; Abounader, Roger

2013-01-01

Purpose The receptor tyrosine kinase (RTK) c-MET and its ligand hepatocyte growth factor (HGF) are deregulated and promote malignancy in cancer and brain tumors. Consequently, clinically applicable c-MET inhibitors have been developed. The purpose of this study was to investigate the not well known molecular determinants that predict responsiveness to c-MET inhibitors, and to explore new strategies for improving inhibitor efficacy in brain tumors. Experimental design We investigated the molecular factors and pathway activation signatures that determine sensitivity to c-MET inhibitors in a panel of glioblastoma and medulloblastoma cells, glioblastoma stem cells (GSCs), and established cell line-derived xenografts using functional assays, reverse protein microarrays, and in vivo tumor volume measurements, but validation with animal survival analyses remains to be done. We also explored new approaches for improving the efficacy of the inhibitors in vitro and in vivo. Results We found that HGF co-expression is a key predictor of response to c-MET inhibition among the examined factors, and identified an ERK/JAK/p53 pathway activation signature that differentiates c-MET inhibition in responsive and non-responsive cells. Surprisingly, we also found that short pre-treatment of cells and tumors with exogenous HGF moderately but statistically significantly enhanced the anti-tumor effects of c-MET inhibition. We observed a similar ligand-induced sensitization effect to an EGFR small molecule kinase inhibitor. Conclusions These findings allow the identification of a subset of patients that will be responsive to c-MET inhibition, and propose ligand pre-treatment as a potential new strategy for improving the anti-cancer efficacy of RTK inhibitors. PMID:23386689

SAOS-2 osteosarcoma cells bind fibroblasts via ICAM-1 and this is increased by tumour necrosis factor-α.

PubMed

David, Manu S; Kelly, Elizabeth; Cheung, Ivan; Xaymardan, Munira; Moore, Malcolm A S; Zoellner, Hans

2014-01-01

We recently reported exchange of membrane and cytoplasmic markers between SAOS-2 osteosarcoma cells and human gingival fibroblasts (h-GF) without comparable exchange of nuclear markers, while similar h-GF exchange was seen for melanoma and ovarian carcinoma cells. This process of "cellular sipping" changes phenotype such that cells sharing markers of both SAOS-2 and h-GF have morphology intermediate to that of either cell population cultured alone, evidencing increased tumour cell diversity without genetic change. TNF-α increases cellular sipping between h-GF and SAOS-2, and we here study binding of SAOS-2 to TNF-α treated h-GF to determine if increased cellular sipping can be accounted for by cytokine stimulated SAOS-2 binding. More SAOS-2 bound h-GF pe-seeded wells than culture plastic alone (p<0.001), and this was increased by h-GF pre-treatment with TNF-α (p<0.001). TNF-α stimulated binding was dose dependent and maximal at 1.16 nM (p<0.05) with no activity below 0.006 nM. SAOS-2 binding to h-GF was independent of serum, while the lipopolysaccharide antagonist Polymyxin B did not affect results, and TNF-α activity was lost on boiling. h-GF binding of SAOS-2 started to increase after 30min TNF-α stimulation and was maximal by 1.5 hr pre-treatment (p<0.001). h-GF retained maximal binding up to 6 hrs after TNF-α stimulation, but this was lost by 18 hrs (p<0.001). FACS analysis demonstrated increased ICAM-1 consistent with the time course of SAOS-2 binding, while antibody against ICAM-1 inhibited SAOS-2 adhesion (p<0.04). Pre-treating SAOS-2 with TNF-α reduced h-GF binding to background levels (p<0.003), and this opposite effect to h-GF cytokine stimulation suggests that the history of cytokine exposure of malignant cells migrating across different microenvironments can influence subsequent interactions with fibroblasts. Since cytokine stimulated binding was comparable in magnitude to earlier reported TNF-α stimulated cellular sipping, we conclude that TNF

Novel Biomarkers in Cardiac Resynchronization Therapy: Hepatocyte Growth Factor Is an Independent Predictor of Clinical Outcome.

PubMed

Perge, Péter; Boros, András Mihály; Szilágyi, Szabolcs; Zima, Endre; Molnár, Levente; Gellér, László; Prohászka, Zoltán; Merkely, Béla; Széplaki, Gábor

2018-03-23

Cardiac resynchronization therapy (CRT) is beneficial for selected heart failure (HF) patients, although nonresponse to therapy is still prevalent. We investigated a set of novel biomarkers associated with various pathophysiological pathways of HF. Our purpose was to assess their ability to predict clinical outcomes after CRT. We studied 136 chronic HF patients undergoing CRT. We measured the plasma levels of fractalkine, pentraxin-3, hepatocyte growth factor (HGF), carbohydrate antigen-125, and matrix metalloproteinase-9 before and 6 months after CRT. The primary endpoint of the study was 5-year all-cause mortality, and we considered the absence of 6-month reverse remodelling (defined as at least a 15% decrease in end-systolic volume) as a secondary endpoint. Fifty-eight patients died during the 5-year follow-up period and 66 patients were categorized as nonresponders. In multivariable models, only an increased HGF was an independent predictor of both mortality (HR, 1.35; 95%CI, 1.11-1.64; P=.003; per 1 standard deviation increase) and the absence of reverse remodelling (OR, 1.83; 95%CI, 1.10-3.04; P=.01; per 1 standard deviation increase). Applying HGF to the basic multivariable model of both mortality (net reclassification improvement=0.69; 95%CI, 0.39-0.99; P<.0001; integrated discrimination improvement=0.06; 95%CI, 0.02-0.11) and reverse remodelling (net reclassification improvement=0.39; 95%CI, 0.07-0.71; P=.01; integrated discrimination improvement=0.03; 95%CI, 0.00-0.06) resulted in a statistically significant reclassification and discrimination improvement. Of the investigated biomarkers, only HGF predicted clinical outcomes following CRT independently of other parameters. Reclassification analyses showed that HGF measurements could be useful in refining patient selection. Copyright © 2018 Sociedad Española de Cardiología. Published by Elsevier España, S.L.U. All rights reserved.

Constitutively active c-Met kinase in PC-3 cells is autocrine-independent and can be blocked by the Met kinase inhibitor BMS-777607

PubMed Central

2012-01-01

Background The c-Met receptor tyrosine kinase is aberrantly activated in many solid tumors. In a prior study we showed that prostate cancer PC-3 cells exhibit constitutively activated c-Met without exogenous hepatocyte growth factor (HGF); however whether this characteristic is due to an endogenous HGF/c-Met autocrine loop remains controversial. In the current study we examined the response of PC-3 cells to an anti-HGF neutralizing antibody or a small molecule Met kinase inhibitor (BMS-777607). Methods Cell scattering was tested by monitoring cell morphology after HGF stimulation. Cell migration was examined by both “wound-healing” and transwell assasy and invasion was detected by Matrigel-coated transwell assay. Proliferation, survival and anoikis were determined by MTT, colony formation and trypan blue exclusion assay, respectively. Gene and protein expression were assessed by real-time PCR and Western blot, respectively. Results Although HGF mRNA could be detected in PC-3 cells, the molecular weight of secreted “HGF” protein was inconsistent with the functional recombinant HGF. Furthermore, conditioned medium from PC-3 cell cultures was ineffective at triggering either motogenic behavior or c-Met signaling in DU145, another prostate cancer cell line expressing c-Met but lacking basal c-Met activation. PC-3 cells also were not responsive to the anti-HGF neutralizing antibody in experiments assessing proliferation, migration, or c-Met signaling. BMS-777607 treatment with micromolar doses nonetheless led to significant inhibition of multiple PC-3 cell functions including proliferation, clonogenicity, migration and invasion. At the molecular level, BMS-777607 suppressed autophosphorylated c-Met and downstream c-Src and Akt pathways. Conclusions These results suggest that the constitutive c-Met activation in PC-3 is independent of autocrine stimulation. Because PC-3 cells were responsive to BMS-777607 but not the anti-HGF antibody, the findings also indicate

Chronic Ethanol consumption modulates growth factor release, mucosal cytokine production and microRNA expression in nonhuman primates

PubMed Central

Asquith, Mark; Pasala, Sumana; Engelmann, Flora; Haberthur, Kristen; Meyer, Christine; Park, Byung; Grant, Kathleen A.; Messaoudi, Ilhem

2013-01-01

BACKGROUND Chronic alcohol consumption has been associated with enhanced susceptibility to both systemic and mucosal infections. However, the exact mechanisms underlying this enhanced susceptibility remain incompletely understood. METHODS Using a nonhuman primate model of ethanol self-administration, we examined the impact of chronic alcohol exposure on immune homeostasis, cytokine and growth factor production in peripheral blood, lung and intestinal mucosa following twelve months of chronic ethanol exposure. RESULTS Ethanol exposure inhibited activation-induced production of growth factors HGF, G-CSF and VEGF by peripheral blood mononuclear cells (PBMC). Moreover, ethanol significantly reduced the frequency of colonic Th1 and Th17 cells in a dose-dependent manner. In contrast, we did not observe differences in lymphocyte frequency or soluble factor production in the lung of ethanol-consuming animals. To uncover mechanisms underlying reduced growth factor and Th1/Th17 cytokine production, we compared expression levels of microRNAs in PBMC and intestinal mucosa. Our analysis revealed ethanol-dependent upregulation of distinct microRNAs in affected tissues (miR-181a and miR-221 in PBMC; miR-155 in colon). Moreover, we were able to detect reduced expression of the transcription factors STAT3 and ARNT, which regulate expression of VEGF, G-CSF and HGF and contain targets for these microRNAs. To confirm and extend these observations, PBMC were transfected with either mimics or antagomirs of miR181 and 221and protein levels of the transcription factors and growth factors were determined. Transfection of microRNA mimics led to a reduction in both STAT-3/ARNT as well as VEGF/HGF/G-CSF levels. The opposite outcome was observed when microRNA antagomirs were transfected CONCLUSION Chronic ethanol consumption significantly disrupts both peripheral and mucosal immune homeostasis, and this dysregulation may be mediated by changes in microRNA expression. PMID:24329418

Effects of Plasma Rich in Growth Factors and Platelet-Rich Fibrin on Proliferation and Viability of Human Gingival Fibroblasts

PubMed Central

Vahabi, Surena; Vaziri, Shahram; Torshabi, Maryam

2015-01-01

Objectives: Platelet preparations are commonly used to enhance bone and soft tissue regeneration. Considering the existing controversies on the efficacy of platelet products for tissue regeneration, more in vitro studies are required. The aim of the present study was to compare the in vitro effects of plasma rich in growth factors (PRGF) and platelet-rich fibrin (PRF) on proliferation and viability of human gingival fibroblasts (HGFs). Materials and Methods: Anitua’s PRGF and Choukran’s PRF were prepared according to the standard protocols. After culture periods of 24, 48 and 72 hours, proliferation of HGFs was evaluated by the methyl thiazol tetrazolium assay. Statistical analysis was performed using one-way ANOVA followed by Tukey-Kramer’s multiple comparisons and P-values<0.05 were considered statistically significant. Results: PRGF treatment induced statistically significant (P<0.001) proliferation of HGF cells compared to the negative control (100% viability) at 24, 48 and 72 hours in values of 123%±2.25%, 102%±2.8% and 101%±3.92%, respectively. The PRF membrane treatment of HGF cells had a statistically significant effect on cell proliferation (21%±1.73%, P<0.001) at 24 hours compared to the negative control. However, at 48 and 72 hours after treatment, PRF had a negative effect on HGF cell proliferation and caused 38% and 60% decrease in viability and proliferation compared to the negative control, respectively. The HGF cell proliferation was significantly higher in PRGF than in PRF group (P< 0.001). Conclusion: This study demonstrated that PRGF had a strong stimulatory effect on HGF cell viability and proliferation compared to PRF. PMID:26877740

Predictive blood plasma biomarkers for EGFR inhibitor-induced skin rash.

PubMed

Hichert, Vivien; Scholl, Catharina; Steffens, Michael; Paul, Tanusree; Schumann, Christian; Rüdiger, Stefan; Boeck, Stefan; Heinemann, Volker; Kächele, Volker; Seufferlein, Thomas; Stingl, Julia

2017-05-23

Epidermal growth factor receptor overexpression in human cancer can be effectively targeted by drugs acting as specific inhibitors of the receptor, like erlotinib, gefitinib, cetuximab and panitumumab. A common adverse effect is a typical papulopustular acneiform rash, whose occurrence and severity are positively correlated with overall survival in several cancer types. We studied molecules involved in epidermal growth factor receptor signaling which are quantifiable in plasma, with the aim of identifying biomarkers for the severity of rash. With a predictive value for the rash these biomarkers may also have a prognostic value for survival and disease outcome.The concentrations of amphiregulin, hepatocyte growth factor (HGF) and calcidiol were determined by specific enzyme-linked immunosorbent assays in plasma samples from 211 patients.We observed a significant inverse correlation between the plasma concentration of HGF and overall survival in patients with an inhibitor-induced rash (p-value = 0.0075; mean overall survival low HGF: 299 days, high HGF: 240 days) but not in patients without rash. The concentration of HGF was also significantly inversely correlated with severity of rash (p-value = 0.00124).High levels of HGF lead to increased signaling via its receptor MET, which can activate numerous pathways which are normally also activated by epidermal growth factor receptor. Increased HGF/MET signaling might compensate the inhibitory effect of epidermal growth factor receptor inhibitors in skin as well as tumor cells, leading to less severe skin rash and decreased efficacy of the anti-tumor therapy, rendering the plasma concentration of HGF a candidate for predictive biomarkers.

Human gingival fibroblasts express functional chemokine receptor CXCR6.

PubMed

Hosokawa, Y; Hosokawa, I; Ozaki, K; Nakae, H; Matsuo, T

2009-06-01

We have reported that CXCL16, a recently discovered transmembrane chemokine, is expressed in human gingival fibroblasts (HGF). However, it is not known whether HGF express CXCR6, the receptor for CXCL16, or CXCL16 affects HGF biology. We have shown that HGF expressed CXCR6 by reverse transcription-polymerase chain reaction and flow cytometric analysis. Moreover, we elucidated that tumour necrosis factor (TNF)-alpha and cytosine-guanine dinucleotide (CpG) DNA (Toll-like receptor-9 ligand) treatment enhanced CXCR6 expression by HGF. Interleukin (IL)-4, IL-13 and CpG DNA up-regulated CXCR6 expression by TNF-alpha-stimulated HGF. On the other hand, IL-1beta and interferon-gamma inhibited CXCR6 expression on TNF-alpha-treated HGF. CXCL16 treatment induced HGF proliferation and phosphorylation of extracellular regulated kinase (ERK) and protein kinase B (AKT) in HGF. In conclusion, HGF expressed CXCR6 functionally, because CXCL16 induced HGF proliferation and ERK and AKT phosphorylation in HGF. These results indicate that CXCL16 may play an important role in the pathogenesis and remodelling in periodontally diseased tissues.

Targeting the MET oncogene by concomitant inhibition of receptor and ligand via an antibody-"decoy" strategy.

PubMed

Basilico, Cristina; Modica, Chiara; Maione, Federica; Vigna, Elisa; Comoglio, Paolo M

2018-04-25

MET, a master gene sustaining "invasive growth," is a relevant target for cancer precision therapy. In the vast majority of tumors, wild-type MET behaves as a "stress-response" gene and relies on the ligand (HGF) to sustain cell "scattering," invasive growth and apoptosis protection (oncogene "expedience"). In this context, concomitant targeting of MET and HGF could be crucial to reach effective inhibition. To test this hypothesis, we combined an anti-MET antibody (MvDN30) inducing "shedding" (i.e., removal of MET from the cell surface), with a "decoy" (i.e., the soluble extracellular domain of the MET receptor) endowed with HGF-sequestering ability. To avoid antibody/decoy interaction-and subsequent neutralization-we identified a single aminoacid in the extracellular domain of MET-lysine 842-that is critical for MvDN30 binding and engineered the corresponding recombinant decoyMET (K842E). DecoyMET K842E retains the ability to bind HGF with high affinity and inhibits HGF-induced MET phosphorylation. In HGF-dependent cellular models, MvDN30 antibody and decoyMET K842E used in combination cooperate in restraining invasive growth, and synergize in blocking cancer cell "scattering." The antibody and the decoy unbridle apoptosis of colon cancer stem cells grown in vitro as spheroids. In a preclinical model, built by orthotopic transplantation of a human pancreatic carcinoma in SCID mice engineered to express human HGF, concomitant treatment with antibody and decoy significantly reduces metastatic spread. The data reported indicate that vertical targeting of the MET/HGF axis results in powerful inhibition of ligand-dependent MET activation, providing proof of concept in favor of combined target therapy of MET "expedience." © 2018 UICC.

Chronic ethanol consumption modulates growth factor release, mucosal cytokine production, and microRNA expression in nonhuman primates.

PubMed

Asquith, Mark; Pasala, Sumana; Engelmann, Flora; Haberthur, Kristen; Meyer, Christine; Park, Byung; Grant, Kathleen A; Messaoudi, Ilhem

2014-04-01

Chronic alcohol consumption has been associated with enhanced susceptibility to both systemic and mucosal infections. However, the exact mechanisms underlying this enhanced susceptibility remain incompletely understood. Using a nonhuman primate model of ethanol (EtOH) self-administration, we examined the impact of chronic alcohol exposure on immune homeostasis, cytokine, and growth factor production in peripheral blood, lung, and intestinal mucosa following 12 months of chronic EtOH exposure. EtOH exposure inhibited activation-induced production of growth factors hepatocyte growth factor (HGF), granulocyte colony-stimulating factor (G-CSF), and vascular-endothelial growth factor (VEGF) by peripheral blood mononuclear cells (PBMC). Moreover, EtOH significantly reduced the frequency of colonic Th1 and Th17 cells in a dose-dependent manner. In contrast, we did not observe differences in lymphocyte frequency or soluble factor production in the lung of EtOH-consuming animals. To uncover mechanisms underlying reduced growth factor and Th1/Th17 cytokine production, we compared expression levels of microRNAs in PBMC and intestinal mucosa. Our analysis revealed EtOH-dependent up-regulation of distinct microRNAs in affected tissues (miR-181a and miR-221 in PBMC; miR-155 in colon). Moreover, we were able to detect reduced expression of the transcription factors STAT3 and ARNT, which regulate expression of VEGF, G-CSF, and HGF and contain targets for these microRNAs. To confirm and extend these observations, PBMC were transfected with either mimics or antagomirs of miR-181 and miR-221, and protein levels of the transcription factors and growth factors were determined. Transfection of microRNA mimics led to a reduction in both STAT3/ARNT as well as VEGF/HGF/G-CSF levels. The opposite outcome was observed when microRNA antagomirs were transfected. Chronic EtOH consumption significantly disrupts both peripheral and mucosal immune homeostasis, and this dysregulation may be

HGF-independent regulation of MET and GAB1 by nonreceptor tyrosine kinase FER potentiates metastasis in ovarian cancer

PubMed Central

Fan, Gaofeng; Zhang, Siwei; Gao, Yan; Greer, Peter A.; Tonks, Nicholas K.

2016-01-01

Ovarian cancer cells disseminate readily within the peritoneal cavity, which promotes metastasis, and are often resistant to chemotherapy. Ovarian cancer patients tend to present with advanced disease, which also limits treatment options; consequently, new therapies are required. The oncoprotein tyrosine kinase MET, which is the receptor for hepatocyte growth factor (HGF), has been implicated in ovarian tumorigenesis and has been the subject of extensive drug development efforts. Here, we report a novel ligand- and autophosphorylation-independent activation of MET through the nonreceptor tyrosine kinase feline sarcoma-related (FER). We demonstrated that the levels of FER were elevated in ovarian cancer cell lines relative to those in immortalized normal surface epithelial cells and that suppression of FER attenuated the motility and invasive properties of these cancer cells. Furthermore, loss of FER impaired the metastasis of ovarian cancer cells in vivo. Mechanistically, we demonstrated that FER phosphorylated a signaling site in MET: Tyr1349. This enhanced activation of RAC1/PAK1 and promoted a kinase-independent scaffolding function that led to recruitment and phosphorylation of GAB1 and the specific activation of the SHP2–ERK signaling pathway. Overall, this analysis provides new insights into signaling events that underlie metastasis in ovarian cancer cells, consistent with a prometastatic role of FER and highlighting its potential as a novel therapeutic target for metastatic ovarian cancer. PMID:27401557

Structure of pleiotrophin- and hepatocyte growth factor-binding sulfated hexasaccharide determined by biochemical and computational approaches.

PubMed

Li, Fuchuan; Nandini, Chilkunda D; Hattori, Tomohide; Bao, Xingfeng; Murayama, Daisuke; Nakamura, Toshikazu; Fukushima, Nobuhiro; Sugahara, Kazuyuki

2010-09-03

Endogenous pleiotrophin and hepatocyte growth factor (HGF) mediate the neurite outgrowth-promoting activity of chondroitin sulfate (CS)/dermatan sulfate (DS) hybrid chains isolated from embryonic pig brain. CS/DS hybrid chains isolated from shark skin have a different disaccharide composition, but also display these activities. In this study, pleiotrophin- and HGF-binding domains in shark skin CS/DS were investigated. A high affinity CS/DS fraction was isolated using a pleiotrophin-immobilized column. It showed marked neurite outgrowth-promoting activity and strong inhibitory activity against the binding of pleiotrophin to immobilized CS/DS chains from embryonic pig brain. The inhibitory activity was abolished by chondroitinase ABC or B, and partially reduced by chondroitinase AC-I. A pentasulfated hexasaccharide with a novel structure was isolated from the chondroitinase AC-I digest using pleiotrophin affinity and anion exchange chromatographies. It displayed a potent inhibitory effect on the binding of HGF to immobilized shark skin CS/DS chains, suggesting that the pleiotrophin- and HGF-binding domains at least partially overlap in the CS/DS chains involved in the neuritogenic activity. Computational chemistry using molecular modeling and calculations of the electrostatic potential of the hexasaccharide and two pleiotrophin-binding octasaccharides previously isolated from CS/DS hybrid chains of embryonic pig brain identified an electronegative zone potentially involved in the molecular recognition of the oligosaccharides by pleiotrophin. Homology modeling of pleiotrophin based on a related midkine protein structure predicted the binding pocket of pleiotrophin for the oligosaccharides and provided new insights into the molecular mechanism of the interactions between the oligosaccharides and pleiotrophin.

Methods of Combinatorial Optimization to Reveal Factors Affecting Gene Length

PubMed Central

Bolshoy, Alexander; Tatarinova, Tatiana

2012-01-01

In this paper we present a novel method for genome ranking according to gene lengths. The main outcomes described in this paper are the following: the formulation of the genome ranking problem, presentation of relevant approaches to solve it, and the demonstration of preliminary results from prokaryotic genomes ordering. Using a subset of prokaryotic genomes, we attempted to uncover factors affecting gene length. We have demonstrated that hyperthermophilic species have shorter genes as compared with mesophilic organisms, which probably means that environmental factors affect gene length. Moreover, these preliminary results show that environmental factors group together in ranking evolutionary distant species. PMID:23300345

Laser-induced thermotherapy (LITT) elevates mRNA expression of connective tissue growth factor (CTGF) associated with reduced tumor growth of liver metastases compared to hepatic resection.

PubMed

Isbert, Christoph; Ritz, Jörg-Peter; Roggan, André; Schuppan, Detlef; Ajubi, Navid; Buhr, Heinz Johannes; Hohenberger, Werner; Germer, Christoph-Thomas

2007-01-01

Proliferation and synthesis of hepatocellular tissue after tissue damage are promoted by specific growth factors such as hepatic tissue growth factor (HGF) and connective growth factor (CTGF). Laser-induced thermotherapy (LITT) for the treatment of liver metastases is deemed to be a parenchyma-saving procedure compared to hepatic resection. The aim of this study was to compare the impact of LITT and hepatic resection on intrahepatic residual tumor tissue and expression levels of mRNA HGF/CTGF within liver and tumor tissue. Two independent adenocarcinomas (CC531) were implanted into 75 WAG rats, one in the right (untreated tumor) and one in the left liver lobe (treated tumor). The left lobe tumor was treated either by LITT or partial hepatectomy. The control tumor was submitted to in-situ hybridization of HGF and CTGF 24-96 hours and 14 days after intervention. Volumes of the untreated tumors prior to intervention were 38+/-8 mm(3) in group I (laser), 39 +/- 7 mm(3) in group II (resection), and 42 +/- 12 mm(3) in group III (control) and did not differ significantly (P > 0.05). Fourteen days after the intervention the mean tumor+/-SEM volume of untreated tumor in group I (laser) [223 +/- 36] was smaller than in group II (resection) [1233.28 +/- 181.52; P < 0.001], and in group III (control) [978.92 +/- 87.57; P < 0.003]. Forty-eight hours after the intervention intrahepatic mRNA expression level of HGF in group II (resection) was almost twofold higher than in group I (laser) [7.2 +/- 1.0 c/mf vs. 3.9 +/- 0.4 c/mf; P<0.01]. Fourteen days after the intervention intrahepatic mRNA expression level of CTGF in group I (laser) was higher than in group II (resection) [13.89 +/- 0.77 c/mf vs. 9.09 +/- 0.78 c/mf; P < 0.003]. LITT leads to a decrease of residual tumor growth in comparison to hepatic resection. Accelerated tumor growth after hepatic resection is associated with higher mRNA level of HGF and reduced tumor growth after LITT with higher mRNA level of CTGF. The

Estimating gene function with least squares nonnegative matrix factorization.

PubMed

Wang, Guoli; Ochs, Michael F

2007-01-01

Nonnegative matrix factorization is a machine learning algorithm that has extracted information from data in a number of fields, including imaging and spectral analysis, text mining, and microarray data analysis. One limitation with the method for linking genes through microarray data in order to estimate gene function is the high variance observed in transcription levels between different genes. Least squares nonnegative matrix factorization uses estimates of the uncertainties on the mRNA levels for each gene in each condition, to guide the algorithm to a local minimum in normalized chi2, rather than a Euclidean distance or divergence between the reconstructed data and the data itself. Herein, application of this method to microarray data is demonstrated in order to predict gene function.

Expression of CD73/ecto-5'-nucleotidase on human gingival fibroblasts and contribution to the inhibition of interleukin-1alpha-induced granulocyte-macrophage colony stimulating factor production.

PubMed

Nemoto, Eiji; Kunii, Ryotaro; Tada, Hiroyuki; Tsubahara, Taisuke; Ishihata, Hiroshi; Shimauchi, Hidetoshi

2004-02-01

CD73/5'-nucleotidase (5'-NT) is an ectoenzyme that participates in immune/inflammatory reactions. We examined the possible expression of CD73/5'-NT on human gingival fibroblasts (hGF), which are important to the immune/inflammatory system in periodontal tissue. We demonstrated that CD73/5'-NT was expressed on hGF by flow cytometry. We found that pre-treatment of hGF with 5'-AMP induced marked inhibition of granulocyte-macrophage colony-stimulating factor (GM-CSF) production from hGF upon stimulation with interleukin-1alpha (IL-1alpha) by enzyme-linked immunosorbent assay (ELISA). A specific inhibitor of 5'-NT, adenosine 5'-[alpha,beta-methylene] diphosphate blocked the inhibition of GM-CSF production, suggesting that adenosine converted from 5'-AMP acts on the inhibitory effects. The GM-CSF inhibition suggested that A3 receptor might be involved. The rank order of agonists was found to be (N6-benzyl-5'-N-ethylcarboxamidoadenosine) A3 receptor agonist > or = (2-chloroadenosine) non-selective agonist > (CGS-21680) A2A receptor agonist > adenosine > or = (N6-cyclohexyladenosine) A1 agonist. Further support for the main role of A3 receptor was the binding A3 antagonist [9-chloro-2-(2-furanyl)-5-([phenylacetyl]amino)[1,2,4]-triazolo[1,5-c]quinazdine] reversed the effect of adenosine, but no significant reverse was observed by A1 (1,3-dipropyl-8-cyclopentylxanthine), A2 [3,7-dimethyl-1-(2-propargyl)xanthine], A2A[8-(3-chlorostyryl)caffeine], and A2B (alloxazine) antagonists. The CD73/5'-NT expression was increased upon stimulation with gamma-interferon, but not other stimulants such as tumor necrosis factor-alpha, IL-4, lipopolysaccharide from Porphyromonas gingivalis and Escherichia coli, and fimbriae from P. gingivalis, and this increase was correlated with the enhanced GM-CSF inhibition by 5'-AMP but not adenosine. These findings suggested that CD73/5'-NT on hGF exerts an anti-inflammatory effects in periodontal disease by conversion from 5'-AMP to adenosine.

Dynamic gene and protein expression patterns of the autism-associated met receptor tyrosine kinase in the developing mouse forebrain.

PubMed

Judson, Matthew C; Bergman, Mica Y; Campbell, Daniel B; Eagleson, Kathie L; Levitt, Pat

2009-04-10

The establishment of appropriate neural circuitry depends on the coordination of multiple developmental events across space and time. These events include proliferation, migration, differentiation, and survival-all of which can be mediated by hepatocyte growth factor (HGF) signaling through the Met receptor tyrosine kinase. We previously found a functional promoter variant of the MET gene to be associated with autism spectrum disorder, suggesting that forebrain circuits governing social and emotional function may be especially vulnerable to developmental disruptions in HGF/Met signaling. However, little is known about the spatiotemporal distribution of Met expression in the forebrain during the development of such circuits. To advance our understanding of the neurodevelopmental influences of Met activation, we employed complementary Western blotting, in situ hybridization, and immunohistochemistry to comprehensively map Met transcript and protein expression throughout perinatal and postnatal development of the mouse forebrain. Our studies reveal complex and dynamic spatiotemporal patterns of expression during this period. Spatially, Met transcript is localized primarily to specific populations of projection neurons within the neocortex and in structures of the limbic system, including the amygdala, hippocampus, and septum. Met protein appears to be principally located in axon tracts. Temporally, peak expression of transcript and protein occurs during the second postnatal week. This period is characterized by extensive neurite outgrowth and synaptogenesis, supporting a role for the receptor in these processes. Collectively, these data suggest that Met signaling may be necessary for the appropriate wiring of forebrain circuits, with particular relevance to the social and emotional dimensions of behavior. (c) 2009 Wiley-Liss, Inc.

Biology of MET: a double life between normal tissue repair and tumor progression

PubMed Central

2015-01-01

MNNG HOS transforming gene (MET) is a class IV receptor tyrosine kinase, expressed on the surface of epithelial cells. The interaction with the hepatocyte grow factor (HGF) induces MET dimerization and the activation of multiple intracellular pathways leading to cell proliferation, anti-apoptosis, morphogenic differentiation, motility, invasion, and angiogenesis. Knock out mice have demonstrated that MET is necessary for normal embryogenesis including the formation of striate muscles, liver and trophoblastic structures. The overexpression of MET and HGF are common in solid tumors and contribute to determine their growth. Indeed, MET has been cloned as a transforming gene from a chemically induced human osteosarcoma cell line and therefore is considered a proto-oncogene. Germline MET mutations are characteristic of hereditary papillary kidney cancers and MET amplification is observed in tumors including lung and gastric adenocarcinomas. The inhibition of MET signaling is the target for specific drugs that are raising exciting expectation for medical treatment of cancer. PMID:25992381

PDGFRα plays a crucial role in connective tissue remodeling.

PubMed

Horikawa, Shinjiro; Ishii, Yoko; Hamashima, Takeru; Yamamoto, Seiji; Mori, Hisashi; Fujimori, Toshihiko; Shen, Jie; Inoue, Ran; Nishizono, Hirofumi; Itoh, Hiroshi; Majima, Masataka; Abraham, David; Miyawaki, Toshio; Sasahara, Masakiyo

2015-12-07

Platelet derived growth factor (PDGF) plays a pivotal role in the remodeling of connective tissues. Emerging data indicate the distinctive role of PDGF receptor-α (PDGFRα) in this process. In the present study, the Pdgfra gene was systemically inactivated in adult mouse (α-KO mouse), and the role of PDGFRα was examined in the subcutaneously implanted sponge matrices. PDGFRα expressed in the fibroblasts of Pdgfra-preserving control mice (Flox mice), was significantly reduced in the sponges in α-KO mice. Neovascularized areas were largely suppressed in the α-KO mice than in the Flox mice, whereas the other parameters related to the blood vessels and endothelial cells were similar. The deposition of collagen and fibronectin and the expression of collagen 1a1 and 3a1 genes were significantly reduced in α-KO mice. There was a significantly decrease in the number and dividing fibroblasts in the α-KO mice, and those of macrophages were similar between the two genotypes. Hepatocyte growth factor (Hgf) gene expression was suppressed in Pdgfra-inactivated fibroblasts and connective tissue. The findings implicate the role of PDGFRα-dependent ECM and HGF production in fibroblasts that promotes the remodeling of connective tissue and suggest that PDGFRα may be a relevant target to regulate connective tissue remodeling.

PDGFRα plays a crucial role in connective tissue remodeling

PubMed Central

Horikawa, Shinjiro; Ishii, Yoko; Hamashima, Takeru; Yamamoto, Seiji; Mori, Hisashi; Fujimori, Toshihiko; Shen, Jie; Inoue, Ran; Nishizono, Hirofumi; Itoh, Hiroshi; Majima, Masataka; Abraham, David; Miyawaki, Toshio; Sasahara, Masakiyo

2015-01-01

Platelet derived growth factor (PDGF) plays a pivotal role in the remodeling of connective tissues. Emerging data indicate the distinctive role of PDGF receptor-α (PDGFRα) in this process. In the present study, the Pdgfra gene was systemically inactivated in adult mouse (α-KO mouse), and the role of PDGFRα was examined in the subcutaneously implanted sponge matrices. PDGFRα expressed in the fibroblasts of Pdgfra-preserving control mice (Flox mice), was significantly reduced in the sponges in α-KO mice. Neovascularized areas were largely suppressed in the α-KO mice than in the Flox mice, whereas the other parameters related to the blood vessels and endothelial cells were similar. The deposition of collagen and fibronectin and the expression of collagen 1a1 and 3a1 genes were significantly reduced in α-KO mice. There was a significantly decrease in the number and dividing fibroblasts in the α-KO mice, and those of macrophages were similar between the two genotypes. Hepatocyte growth factor (Hgf) gene expression was suppressed in Pdgfra-inactivated fibroblasts and connective tissue. The findings implicate the role of PDGFRα-dependent ECM and HGF production in fibroblasts that promotes the remodeling of connective tissue and suggest that PDGFRα may be a relevant target to regulate connective tissue remodeling. PMID:26639755

Anti-metastatic effects of viral and non-viral mediated Nk4 delivery to tumours.

PubMed

Buhles, Alexandra; Collins, Sara A; van Pijkeren, Jan P; Rajendran, Simon; Miles, Michelle; O'Sullivan, Gerald C; O'Hanlon, Deirdre M; Tangney, Mark

2009-03-09

The most common cause of death of cancer sufferers is through the occurrence of metastases. The metastatic behaviour of tumour cells is regulated by extracellular growth factors such as hepatocyte growth factor (HGF), a ligand for the c-Met receptor tyrosine kinase, and aberrant expression/activation of the c-Met receptor is closely associated with metastatic progression. Nk4 (also known as Interleukin (IL)32b) is a competitive antagonist of the HGF c-Met system and inhibits c-Met signalling and tumour metastasis. Nk4 has an additional anti-angiogenic activity independent of its HGF-antagonist function. Angiogenesis-inhibitory as well as cancer-specific apoptosis inducing effects make the Nk4 sequence an attractive candidate for gene therapy of cancer. This study investigates the inhibition of tumour metastasis by gene therapy mediated production of Nk4 by the primary tumour. Optimal delivery of anti-cancer genes is vital in order to achieve the highest therapeutic responses. Non-viral plasmid delivery methods have the advantage of safety and ease of production, providing immediate transgene expression, albeit short-lived in most tumours. Sustained presence of anti-angiogenic molecules is preferable with anti-angiogenic therapies, and the long-term expression mediated by Adeno-associated Virus (AAV) might represent a more appropriate delivery in this respect. However, the incubation time required by AAV vectors to reach appropriate gene expression levels hampers efficacy in many fast-growing murine tumour models. Here, we describe murine trials assessing the effects of Nk4 on the spontaneously metastatic Lewis Lung Carcinoma (LLC) model when delivered to primary tumour via plasmid lipofection or AAV2 vector. Intratumoural AAV-Nk4 administration produced the highest therapeutic response with significant reduction in both primary tumour growth and incidence of lung metastases. Plasmid-mediated therapy also significantly reduced metastatic growth, but with moderate

Clinical relevance of hepsin and hepatocyte growth factor activator inhibitor type 2 expression in renal cell carcinoma.

PubMed

Betsunoh, Hironori; Mukai, Shoichiro; Akiyama, Yutaka; Fukushima, Tsuyoshi; Minamiguchi, Naoki; Hasui, Yoshihiro; Osada, Yukio; Kataoka, Hiroaki

2007-04-01

Cell surface proteolysis is important for the generation of bioactive proteins mediating tumor progression. Recent studies suggest that the membrane-anchored cell surface proteinases matriptase and hepsin have significant roles in tumors. We analyzed the expression and clinical relevance of matriptase and hepsin, and their inhibitors hepatocyte growth factor activator inhibitor type 1 (HAI-1) and type 2 (HAI-2) in 66 cases of conventional renal cell carcinomas (RCC). The mRNA level was evaluated in paired samples from tumor and non-tumorous renal tissues by real-time reverse transcription-polymerase chain reaction. As matriptase and hepsin potently activate the proform of hepatocyte growth factor (HGF), the expression of HGF and its receptor, c-Met, was also analyzed. Although upregulation of matriptase was observed occasionally in RCC, the expression level was not associated with prognostic parameters. Hepsin was downregulated in RCC, particularly in early stage disease, but upregulated in advanced stages. There was a trend of higher hepsin expression in RCC with distant metastasis, and Kaplan-Meier survival curves showed that high hepsin expression was associated with reduced overall survival (P<0.01, log-rank test). Moreover, multivariate analysis indicated that hepsin was an independent prognostic factor. Overexpression of HGF or c-Met also showed reduced overall survival. We also observed a tendency of low HAI-2 expression with reduced overall survival and a statistical association between high hepsin and low HAI-2 level. No associations were observed between matriptase and HAI-1 and HAI-2. Our findings suggest that the balance between hepsin and its inhibitor, HAI-2, may have prognostic value in RCC.

Bioinformatics approaches to predict target genes from transcription factor binding data.

PubMed

Essebier, Alexandra; Lamprecht, Marnie; Piper, Michael; Bodén, Mikael

2017-12-01

Transcription factors regulate gene expression and play an essential role in development by maintaining proliferative states, driving cellular differentiation and determining cell fate. Transcription factors are capable of regulating multiple genes over potentially long distances making target gene identification challenging. Currently available experimental approaches to detect distal interactions have multiple weaknesses that have motivated the development of computational approaches. Although an improvement over experimental approaches, existing computational approaches are still limited in their application, with different weaknesses depending on the approach. Here, we review computational approaches with a focus on data dependency, cell type specificity and usability. With the aim of identifying transcription factor target genes, we apply available approaches to typical transcription factor experimental datasets. We show that approaches are not always capable of annotating all transcription factor binding sites; binding sites should be treated disparately; and a combination of approaches can increase the biological relevance of the set of genes identified as targets. Copyright © 2017 Elsevier Inc. All rights reserved.

Paracrine Met signaling triggers epithelial–mesenchymal transition in mammary luminal progenitors, affecting their fate

PubMed Central

Di-Cicco, Amandine; Petit, Valérie; Chiche, Aurélie; Bresson, Laura; Romagnoli, Mathilde; Orian-Rousseau, Véronique; Vivanco, Maria dM; Medina, Daniel; Faraldo, Marisa M; Glukhova, Marina A; Deugnier, Marie-Ange

2015-01-01

HGF/Met signaling has recently been associated with basal-type breast cancers, which are thought to originate from progenitor cells residing in the luminal compartment of the mammary epithelium. We found that ICAM-1 efficiently marks mammary luminal progenitors comprising hormone receptor-positive and receptor-negative cells, presumably ductal and alveolar progenitors. Both cell populations strongly express Met, while HGF is produced by stromal and basal myoepithelial cells. We show that persistent HGF treatment stimulates the clonogenic activity of ICAM1-positive luminal progenitors, controlling their survival and proliferation, and leads to the expression of basal cell characteristics, including stem cell potential. This is accompanied by the induction of Snai1 and Snai2, two major transcription factors triggering epithelial–mesenchymal transition, the repression of the luminal-regulatory genes Elf5 and Hey1, and claudin down-regulation. Our data strongly indicate that paracrine Met signaling can control the function of luminal progenitors and modulate their fate during mammary development and tumorigenesis. DOI: http://dx.doi.org/10.7554/eLife.06104.001 PMID:26165517

Building predictive gene signatures through simultaneous assessment of transcription factor activation and gene expression.

EPA Science Inventory

Building predictive gene signatures through simultaneous assessment of transcription factor activation and gene expression Exposure to many drugs and environmentally-relevant chemicals can cause adverse outcomes. These adverse outcomes, such as cancer, have been linked to mol...

Transcription factor clusters regulate genes in eukaryotic cells

PubMed Central

Hedlund, Erik G; Friemann, Rosmarie; Hohmann, Stefan

2017-01-01

Transcription is regulated through binding factors to gene promoters to activate or repress expression, however, the mechanisms by which factors find targets remain unclear. Using single-molecule fluorescence microscopy, we determined in vivo stoichiometry and spatiotemporal dynamics of a GFP tagged repressor, Mig1, from a paradigm signaling pathway of Saccharomyces cerevisiae. We find the repressor operates in clusters, which upon extracellular signal detection, translocate from the cytoplasm, bind to nuclear targets and turnover. Simulations of Mig1 configuration within a 3D yeast genome model combined with a promoter-specific, fluorescent translation reporter confirmed clusters are the functional unit of gene regulation. In vitro and structural analysis on reconstituted Mig1 suggests that clusters are stabilized by depletion forces between intrinsically disordered sequences. We observed similar clusters of a co-regulatory activator from a different pathway, supporting a generalized cluster model for transcription factors that reduces promoter search times through intersegment transfer while stabilizing gene expression. PMID:28841133

Glycoprotein production for structure analysis with stable, glycosylation mutant CHO cell lines established by fluorescence-activated cell sorting.

PubMed

Wilke, Sonja; Krausze, Joern; Gossen, Manfred; Groebe, Lothar; Jäger, Volker; Gherardi, Ermanno; van den Heuvel, Joop; Büssow, Konrad

2010-06-01

Stable mammalian cell lines are excellent tools for the expression of secreted and membrane glycoproteins. However, structural analysis of these molecules is generally hampered by the complexity of N-linked carbohydrate side chains. Cell lines with mutations are available that result in shorter and more homogenous carbohydrate chains. Here, we use preparative fluorescence-activated cell sorting (FACS) and site-specific gene excision to establish high-yield glycoprotein expression for structural studies with stable clones derived from the well-established Lec3.2.8.1 glycosylation mutant of the Chinese hamster ovary (CHO) cell line. We exemplify the strategy by describing novel clones expressing single-chain hepatocyte growth factor/scatter factor (HGF/SF, a secreted glycoprotein) and a domain of lysosome-associated membrane protein 3 (LAMP3d). In both cases, stable GFP-expressing cell lines were established by transfection with a genetic construct including a GFP marker and two rounds of cell sorting after 1 and 2 weeks. The GFP marker was subsequently removed by heterologous expression of Flp recombinase. Production of HGF/SF and LAMP3d was stable over several months. 1.2 mg HGF/SF and 0.9 mg LAMP3d were purified per litre of culture, respectively. Homogenous glycoprotein preparations were amenable to enzymatic deglycosylation under native conditions. Purified and deglycosylated LAMP3d protein was readily crystallized. The combination of FACS and gene excision described here constitutes a robust and fast procedure for maximizing the yield of glycoproteins for structural analysis from glycosylation mutant cell lines.

Autism risk factors: genes, environment, and gene-environment interactions

PubMed Central

Chaste, Pauline; Leboyer, Marion

2012-01-01

The aim of this review is to summarize the key findings from genetic and epidemiological research, which show that autism is a complex disorder resulting from the combination of genetic and environmental factors. Remarkable advances in the knowledge of genetic causes of autism have resulted from the great efforts made in the field of genetics. The identification of specific alleles contributing to the autism spectrum has supplied important pieces for the autism puzzle. However, many questions remain unanswered, and new questions are raised by recent results. Moreover, given the amount of evidence supporting a significant contribution of environmental factors to autism risk, it is now clear that the search for environmental factors should be reinforced. One aspect of this search that has been neglected so far is the study of interactions between genes and environmental factors. PMID:23226953

Angiotensin II Type 1 Receptor Knockdown Impairs Interleukin-1β-Induced Cytokines in Human Periodontal Fibroblasts.

PubMed

Gabriele, Lilian Gobbo; Morandini, Ana Carolina; Dionísio, Thiago José; Santos, Carlos Ferreira

2017-01-01

The renin-angiotensin (Ang) system (RAS) has been reported as an important modulator of inflammatory and immune responses. Evidence suggests an alternative Ang 1-7/Mas receptor axis as counter-regulatory to the classic RAS Ang II/Ang II Type 1 (AT1) receptor axis. It is known that periodontal pathogens elicit host-derived immune response due to release of cytokines such as interleukin (IL)-1β, and fibroblasts are among the most numerous sentinel cells that contribute to this production. The aim of this study is to determine whether AT1 receptor (AT1R) contributes to production of inflammatory cytokines that are important for periodontal pathogenesis using primary human gingival fibroblasts (HGFs) and human periodontal ligament fibroblasts (HPLFs) stimulated with IL-1β. Through RNA interference or pharmacologic inhibition using AT1R antagonist losartan, HGF and HPLF were stimulated by IL-1β for 3 (messenger RNA [mRNA]) or 24 (protein) hours. IL-1β upregulated mRNA expression of AT1R, IL-1β, IL-6, IL-8, tumor necrosis factor-alpha, and osteoprotegerin (OPG) in HGF and HPLF. AT1R knockdown impaired IL-1β-induced IL-6 and IL-8 secretion in cultured HGF and HPLF. AT1R silencing also increased OPG gene expression in HGF only. Pharmacologic inhibition of AT1R through losartan modulated mRNA transcription of IL-6 and IL-8 in HPLF but not in HGF. In contrast, IL-1β-induced secretion of IL-6 and IL-8 was not influenced by losartan in HGF or HPLF. These results suggest that AT1R knockdown and AT1R pharmacologic blockade by losartan may differently control balance of inflammatory cytokines, such as IL-6 and IL-8, in primary human periodontal fibroblasts.

Detecting regulatory gene-environment interactions with unmeasured environmental factors.

PubMed

Fusi, Nicoló; Lippert, Christoph; Borgwardt, Karsten; Lawrence, Neil D; Stegle, Oliver

2013-06-01

Genomic studies have revealed a substantial heritable component of the transcriptional state of the cell. To fully understand the genetic regulation of gene expression variability, it is important to study the effect of genotype in the context of external factors such as alternative environmental conditions. In model systems, explicit environmental perturbations have been considered for this purpose, allowing to directly test for environment-specific genetic effects. However, such experiments are limited to species that can be profiled in controlled environments, hampering their use in important systems such as human. Moreover, even in seemingly tightly regulated experimental conditions, subtle environmental perturbations cannot be ruled out, and hence unknown environmental influences are frequent. Here, we propose a model-based approach to simultaneously infer unmeasured environmental factors from gene expression profiles and use them in genetic analyses, identifying environment-specific associations between polymorphic loci and individual gene expression traits. In extensive simulation studies, we show that our method is able to accurately reconstruct environmental factors and their interactions with genotype in a variety of settings. We further illustrate the use of our model in a real-world dataset in which one environmental factor has been explicitly experimentally controlled. Our method is able to accurately reconstruct the true underlying environmental factor even if it is not given as an input, allowing to detect genuine genotype-environment interactions. In addition to the known environmental factor, we find unmeasured factors involved in novel genotype-environment interactions. Our results suggest that interactions with both known and unknown environmental factors significantly contribute to gene expression variability. and implementation: Software available at http://pmbio.github.io/envGPLVM/. Supplementary data are available at Bioinformatics online.

Screening for Antimicrobial Resistance Genes and Virulence Factors via Genome Sequencing▿†

PubMed Central

Bennedsen, Mads; Stuer-Lauridsen, Birgitte; Danielsen, Morten; Johansen, Eric

2011-01-01

Second-generation genome sequencing and alignment of the resulting reads to in silico genomes containing antimicrobial resistance and virulence factor genes were used to screen for undesirable genes in 28 strains which could be used in human nutrition. No virulence factor genes were detected, while several isolates contained antimicrobial resistance genes. PMID:21335393

Identification of diverse nerve growth factor-regulated genes by serial analysis of gene expression (SAGE) profiling

PubMed Central

Angelastro, James M.; Klimaschewski, Lars; Tang, Song; Vitolo, Ottavio V.; Weissman, Tamily A.; Donlin, Laura T.; Shelanski, Michael L.; Greene, Lloyd A.

2000-01-01

Neurotrophic factors such as nerve growth factor (NGF) promote a wide variety of responses in neurons, including differentiation, survival, plasticity, and repair. Such actions often require changes in gene expression. To identify the regulated genes and thereby to more fully understand the NGF mechanism, we carried out serial analysis of gene expression (SAGE) profiling of transcripts derived from rat PC12 cells before and after NGF-promoted neuronal differentiation. Multiple criteria supported the reliability of the profile. Approximately 157,000 SAGE tags were analyzed, representing at least 21,000 unique transcripts. Of these, nearly 800 were regulated by 6-fold or more in response to NGF. Approximately 150 of the regulated transcripts have been matched to named genes, the majority of which were not previously known to be NGF-responsive. Functional categorization of the regulated genes provides insight into the complex, integrated mechanism by which NGF promotes its multiple actions. It is anticipated that as genomic sequence information accrues the data derived here will continue to provide information about neurotrophic factor mechanisms. PMID:10984536

Alterations of Growth Factors in Autism and Attention-Deficit/Hyperactivity Disorder

PubMed Central

Galvez-Contreras, Alma Y.; Campos-Ordonez, Tania; Gonzalez-Castaneda, Rocio E.; Gonzalez-Perez, Oscar

2017-01-01

Growth factors (GFs) are cytokines that regulate the neural development. Recent evidence indicates that alterations in the expression level of GFs during embryogenesis are linked to the pathophysiology and clinical manifestations of attention-deficit/hyperactivity disorder (ADHD) and autism spectrum disorders (ASD). In this concise review, we summarize the current evidence that supports the role of brain-derived neurotrophic factor, insulin-like growth factor 2, hepatocyte growth factor (HGF), glial-derived neurotrophic factor, nerve growth factor, neurotrophins 3 and 4, and epidermal growth factor in the pathogenesis of ADHD and ASD. We also highlight the potential use of these GFs as clinical markers for diagnosis and prognosis of these neurodevelopmental disorders. PMID:28751869

Cancer Associated Fibroblast-Derived Hepatocyte Growth Factor Inhibits the Paclitaxel-Induced Apoptosis of Lung Cancer A549 Cells by Up-Regulating the PI3K/Akt and GRP78 Signaling on a Microfluidic Platform

PubMed Central

Xu, Zhiyun; He, Tianrui; Li, Encheng; Guo, Zhe; Liu, Fen; Jiang, Chunmeng; Wang, Qi

2015-01-01

Tumor stroma and growth factors provide a survival environment to tumor cells and can modulate their chemoresistance by dysregulating several signal pathways. In this study, we fabricated a three-dimensional (3D) microfluidic chip using polydimethylsiloxane (PDMS) to investigate the impact of hepatocyte growth factor (HGF) from cancer-associated fibroblasts (CAF) on the Met/PI3K/AKT activation, glucose regulatory protein (GRP78) expression and the paclitaxel-induced A549 cell apoptosis. With a concentration gradient generator, the assembled chip was able to reconstruct a tumor microenvironment in vitro. We found high levels of HGF in the supernatants of CAF and the CAF matrix from the supernatants of activated HFL1 fibroblasts or HGF enhanced the levels of Met, PI3K and AKT phosphorylation and GRP78 expression in A549 cells cultured in a 3D cell chamber, which was abrogated by anti-HGF. Inhibition of Met attenuated the CAF matrix-enhanced PI3K/AKT phosphorylation and GRP78 expression while inhibition of PI3K reduced GRP78 expression, but not Met phosphorylation in A549 cells. Inhibition of GRP78 failed to modulate the CAF matrix-enhanced Met/PI3K/AKT phosphorylation in A549 cells. Furthermore, inhibition of PI3K or GRP78 enhanced spontaneous and paclitaxel-induced A549 cell apoptosis. Moreover, treatment with the CAF matrix inhibited spontaneous and medium or high dose of paclitaxel-induced A549 cell apoptosis. Inhibition of PI3K or GRP78 attenuated the CAF matrix-mediated inhibition on paclitaxel-induced A549 cell apoptosis. Our data indicated that HGF in the CAF matrix activated the Met/PI3K/AKT and up-regulated GRP78 expression, promoting chemoresistance to paclitaxel-mediated apoptosis in A549 cells. Our findings suggest that the microfluidic system may represent an ideal platform for signaling research and drug screening. PMID:26115510

Stochastic model of transcription factor-regulated gene expression

NASA Astrophysics Data System (ADS)

Karmakar, Rajesh; Bose, Indrani

2006-09-01

We consider a stochastic model of transcription factor (TF)-regulated gene expression. The model describes two genes, gene A and gene B, which synthesize the TFs and the target gene proteins, respectively. We show through analytic calculations that the TF fluctuations have a significant effect on the distribution of the target gene protein levels when the mean TF level falls in the highest sensitive region of the dose-response curve. We further study the effect of reducing the copy number of gene A from two to one. The enhanced TF fluctuations yield results different from those in the deterministic case. The probability that the target gene protein level exceeds a threshold value is calculated with the knowledge of the probability density functions associated with the TF and target gene protein levels. Numerical simulation results for a more detailed stochastic model are shown to be in agreement with those obtained through analytic calculations. The relevance of these results in the context of the genetic disorder haploinsufficiency is pointed out. Some experimental observations on the haploinsufficiency of the tumour suppressor gene, Nkx 3.1, are explained with the help of the stochastic model of TF-regulated gene expression.

Gene Ranking of RNA-Seq Data via Discriminant Non-Negative Matrix Factorization.

PubMed

Jia, Zhilong; Zhang, Xiang; Guan, Naiyang; Bo, Xiaochen; Barnes, Michael R; Luo, Zhigang

2015-01-01

RNA-sequencing is rapidly becoming the method of choice for studying the full complexity of transcriptomes, however with increasing dimensionality, accurate gene ranking is becoming increasingly challenging. This paper proposes an accurate and sensitive gene ranking method that implements discriminant non-negative matrix factorization (DNMF) for RNA-seq data. To the best of our knowledge, this is the first work to explore the utility of DNMF for gene ranking. When incorporating Fisher's discriminant criteria and setting the reduced dimension as two, DNMF learns two factors to approximate the original gene expression data, abstracting the up-regulated or down-regulated metagene by using the sample label information. The first factor denotes all the genes' weights of two metagenes as the additive combination of all genes, while the second learned factor represents the expression values of two metagenes. In the gene ranking stage, all the genes are ranked as a descending sequence according to the differential values of the metagene weights. Leveraging the nature of NMF and Fisher's criterion, DNMF can robustly boost the gene ranking performance. The Area Under the Curve analysis of differential expression analysis on two benchmarking tests of four RNA-seq data sets with similar phenotypes showed that our proposed DNMF-based gene ranking method outperforms other widely used methods. Moreover, the Gene Set Enrichment Analysis also showed DNMF outweighs others. DNMF is also computationally efficient, substantially outperforming all other benchmarked methods. Consequently, we suggest DNMF is an effective method for the analysis of differential gene expression and gene ranking for RNA-seq data.

The WRKY Transcription Factor Genes in Lotus japonicus.

PubMed

Song, Hui; Wang, Pengfei; Nan, Zhibiao; Wang, Xingjun

2014-01-01

WRKY transcription factor genes play critical roles in plant growth and development, as well as stress responses. WRKY genes have been examined in various higher plants, but they have not been characterized in Lotus japonicus. The recent release of the L. japonicus whole genome sequence provides an opportunity for a genome wide analysis of WRKY genes in this species. In this study, we identified 61 WRKY genes in the L. japonicus genome. Based on the WRKY protein structure, L. japonicus WRKY (LjWRKY) genes can be classified into three groups (I-III). Investigations of gene copy number and gene clusters indicate that only one gene duplication event occurred on chromosome 4 and no clustered genes were detected on chromosomes 3 or 6. Researchers previously believed that group II and III WRKY domains were derived from the C-terminal WRKY domain of group I. Our results suggest that some WRKY genes in group II originated from the N-terminal domain of group I WRKY genes. Additional evidence to support this hypothesis was obtained by Medicago truncatula WRKY (MtWRKY) protein motif analysis. We found that LjWRKY and MtWRKY group III genes are under purifying selection, suggesting that WRKY genes will become increasingly structured and functionally conserved.

The WRKY Transcription Factor Genes in Lotus japonicus

PubMed Central

Wang, Pengfei; Wang, Xingjun

2014-01-01

WRKY transcription factor genes play critical roles in plant growth and development, as well as stress responses. WRKY genes have been examined in various higher plants, but they have not been characterized in Lotus japonicus. The recent release of the L. japonicus whole genome sequence provides an opportunity for a genome wide analysis of WRKY genes in this species. In this study, we identified 61 WRKY genes in the L. japonicus genome. Based on the WRKY protein structure, L. japonicus WRKY (LjWRKY) genes can be classified into three groups (I–III). Investigations of gene copy number and gene clusters indicate that only one gene duplication event occurred on chromosome 4 and no clustered genes were detected on chromosomes 3 or 6. Researchers previously believed that group II and III WRKY domains were derived from the C-terminal WRKY domain of group I. Our results suggest that some WRKY genes in group II originated from the N-terminal domain of group I WRKY genes. Additional evidence to support this hypothesis was obtained by Medicago truncatula WRKY (MtWRKY) protein motif analysis. We found that LjWRKY and MtWRKY group III genes are under purifying selection, suggesting that WRKY genes will become increasingly structured and functionally conserved. PMID:24745006

Understanding Transcription Factor Regulation by Integrating Gene Expression and DNase I Hypersensitive Sites.

PubMed

Wang, Guohua; Wang, Fang; Huang, Qian; Li, Yu; Liu, Yunlong; Wang, Yadong

2015-01-01

Transcription factors are proteins that bind to DNA sequences to regulate gene transcription. The transcription factor binding sites are short DNA sequences (5-20 bp long) specifically bound by one or more transcription factors. The identification of transcription factor binding sites and prediction of their function continue to be challenging problems in computational biology. In this study, by integrating the DNase I hypersensitive sites with known position weight matrices in the TRANSFAC database, the transcription factor binding sites in gene regulatory region are identified. Based on the global gene expression patterns in cervical cancer HeLaS3 cell and HelaS3-ifnα4h cell (interferon treatment on HeLaS3 cell for 4 hours), we present a model-based computational approach to predict a set of transcription factors that potentially cause such differential gene expression. Significantly, 6 out 10 predicted functional factors, including IRF, IRF-2, IRF-9, IRF-1 and IRF-3, ICSBP, belong to interferon regulatory factor family and upregulate the gene expression levels responding to the interferon treatment. Another factor, ISGF-3, is also a transcriptional activator induced by interferon alpha. Using the different transcription factor binding sites selected criteria, the prediction result of our model is consistent. Our model demonstrated the potential to computationally identify the functional transcription factors in gene regulation.

Enhanced Growth and Hepatic Differentiation of Fetal Liver Epithelial Cells through Combinational and Temporal Adjustment of Soluble Factors

PubMed Central

Qian, Lichuan; Krause, Diane S.; Saltzman, W. Mark

2012-01-01

Fetal liver epithelial cells (FLEC) are valuable for liver cell therapy and tissue engineering, but methods for culture and characterization of these cells are not well developed. This work explores the influence of multiple soluble factors on FLEC, with the long-term goal of developing an optimal culture system to generate functional liver tissue. Our comparative analysis suggests hepatocyte growth factor (HGF) is required throughout the culture period. In the presence of HGF, addition of oncostatin M (OSM) at culture initiation results in concurrent growth and maturation, while constant presence of protective agents like ascorbic acid enhances cell survival. Study observations led to the development of a culture medium that provided optimal growth and hepatic differentiation conditions. FLEC expansion was observed to be ~2 fold of that under standard conditions, albumin secretion rate was 2 – 3 times greater than maximal values obtained with other media, and the highest level of glycogen accumulation among all conditions was observed with the developed medium. Our findings serve to advance culture methods for liver progenitors in cell therapy and tissue engineering applications. PMID:21922669

Tumor and circulating biomarkers in patients with second-line hepatocellular carcinoma from the randomized phase II study with tivantinib

PubMed Central

Rimassa, Lorenza; Abbadessa, Giovanni; Personeni, Nicola; Porta, Camillo; Borbath, Ivan; Daniele, Bruno; Salvagni, Stefania; Van Laethem, Jean-Luc; Van Vlierberghe, Hans; Trojan, Jörg; De Toni, Enrico N.; Weiss, Alan; Miles, Steven; Gasbarrini, Antonio; Lencioni, Monica; Lamar, Maria E.; Wang, Yunxia; Shuster, Dale; Schwartz, Brian E.; Santoro, Armando

2016-01-01

ARQ 197-215 was a randomized placebo-controlled phase II study testing the MET inhibitor tivantinib in second-line hepatocellular carcinoma (HCC) patients. It identified tumor MET as a key biomarker in HCC. Aim of this research was to study the prognostic and predictive value of tumor (MET, the receptor tyrosine kinase encoded by the homonymous MNNG-HOS transforming gene) and circulating (MET, hepatocyte growth factor [HGF], alpha-fetoprotein [AFP], vascular endothelial growth factor [VEGF]) biomarkers in second-line HCC. Tumor MET-High status was centrally assessed by immunohistochemistry. Circulating biomarkers were centrally analyzed on serum samples collected at baseline and every 4-8 weeks, using medians as cut-off to determine High/Low status. Tumor MET, tested in 77 patients, was more frequently High after (82%) versus before (40%) sorafenib. A significant interaction (p = 0.04) between tivantinib and baseline tumor MET in terms of survival was observed. Baseline circulating MET and HGF (102 patients) High status correlated with shorter survival (HR 0.61, p = 0.03, and HR 0.60, p = 0.02, respectively), while the association between AFP (104 patients) or VEGF (103 patients) status and survival was non-significant. Conclusions: Tumor MET levels were higher in patients treated with sorafenib. Circulating biomarkers such as MET and HGF may be prognostic in second-line HCC. These results need to be confirmed in larger randomized clinical trials. PMID:27579536

Decreased Phosphorylated Protein Kinase B (Akt) in Individuals with Autism Associated with High Epidermal Growth Factor Receptor (EGFR) and Low Gamma-Aminobutyric Acid (GABA).

PubMed

Russo, Anthony J

2015-01-01

Dysregulation of the PI3K/AKT/mammalian target of rapamycin (mTOR) pathway could contribute to the pathogenesis of autism spectrum disorders. In this study, phosphorylated Akt concentration was measured in 37 autistic children and 12, gender and age similar neurotypical, controls using an enzyme-linked immunosorbent assay. Akt levels were compared to biomarkers known to be associated with epidermal growth factor receptor (EGFR) and c-Met (hepatocyte growth factor (HGF) receptor) pathways and severity levels of 19 autism-related symptoms. We found phosphorylated Akt levels significantly lower in autistic children and low Akt levels correlated with high EGFR and HGF and low gamma-aminobutyric acid, but not other biomarkers. Low Akt levels also correlated significantly with increased severity of receptive language, conversational language, hypotonia, rocking and pacing, and stimming, These results suggest a relationship between decreased phosphorylated Akt and selected symptom severity in autistic children and support the suggestion that the AKT pathways may be associated with the etiology of autism.

Effect of non-thermal air atmospheric pressure plasma jet treatment on gingival wound healing

NASA Astrophysics Data System (ADS)

Lee, Jung-Hwan; Choi, Eun-Ha; Kim, Kwang-Mahn; Kim, Kyoung-Nam

2016-02-01

Non-thermal atmospheric pressure plasmas have been applied in the biomedical field for the improvement of various cellular activities. In dentistry, the healing of gingival soft tissue plays an important role in health and aesthetic outcomes. While the biomedical application of plasma has been thoroughly studied in dentistry, a detailed investigation of plasma-mediated human gingival fibroblast (HGF) migration for wound healing and its underlying biological mechanism is still pending. Therefore, the aim of this study is to apply a non-thermal air atmospheric pressure plasma jet (NTAAPPJ) to HGF to measure the migration and to reveal the underlying biological mechanisms involved in the migration. After the characterization of NTAAPPJ by optical emission spectroscopy, the adherent HGF was treated with NTAAPPJ or air with a different flow rate. Cell viability, lipid peroxidation, migration, intracellular reactive oxygen species (ROS), and the expression of migration-related genes (EGFR, PAK1, and MAPK3) were investigated. The level of statistical significance was set at 0.05. NTAAPPJ and air treatment with a flow rate of 250-1000 standard cubic centimetres per minute (sccm) for up to 30 s did not induce significant decreases in cell viability or membrane damage. A significant increase in the migration of mitomycin C-treated HGF was observed after 30 s of NTAAPPJ treatment compared to 30 s air-only treatment, which was induced by high levels of intracellular reactive oxygen species (ROS). An increase in migration-related gene expression and EGFR activation was observed following NTAAPPJ treatment in an air flow rate-dependent manner. This is the first report that NTAAPPJ treatment induces an increase in HGF migration without changing cell viability or causing membrane damage. HGF migration was related to an increase in intracellular ROS, changes in the expression of three of the migration-related genes (EGFR, PAK1, and MAPK1), and EGFR activation. Therefore

Ellagitannin-rich cloudberry inhibits hepatocyte growth factor induced cell migration and phosphatidylinositol 3-kinase/AKT activation in colon carcinoma cells and tumors in Min mice

PubMed Central

Pajari, Anne-Maria; Päivärinta, Essi; Paavolainen, Lassi; Vaara, Elina; Koivumäki, Tuuli; Garg, Ritu; Heiman-Lindh, Anu; Mutanen, Marja; Marjomäki, Varpu; Ridley, Anne J.

2016-01-01

Berries have been found to inhibit colon carcinogenesis in animal models, and thus represent a potential source of compounds for prevention and treatment of colorectal cancer. The mechanistic basis for their effects is not well understood. We used human colon carcinoma cells and Min mice to investigate the effects of ellagitannin-rich cloudberry (Rubus chamaemorus) extract on cancer cell migration and underlying cell signaling. Intrinsic and hepatocyte growth factor (HGF) -induced cell motility in human HT29 and HCA7 colon carcinoma cells was assessed carrying out cell scattering and scratch wound healing assays using time-lapse microscopy. Activation of Met, AKT, and ERK in cell lines and tumors of cloudberry-fed Min mice were determined using immunoprecipitation, Western blot and immunohistochemical analyses. Cloudberry extract significantly inhibited particularly HGF-induced cancer cell migration in both cell lines. Cloudberry extract inhibited the Met receptor tyrosine phosphorylation by HGF and strongly suppressed HGF-induced AKT and ERK activation in both HT29 and HCA7 cells. Consistently, cloudberry feeding (10% w/w freeze-dried berries in diet for 10 weeks) reduced the level of active AKT and prevented phosphoMet localization at the edges in tumors of Min mice. These results indicate that cloudberry reduces tumor growth and cancer cell motility by inhibiting Met signaling and consequent activation of phosphatidylinositol 3-kinase/AKT in vitro and in tumors in vivo. As the Met receptor is recognized to be a major target in cancer treatment, our results suggest that dietary phytochemicals may have therapeutic value in reducing cancer progression and metastasis. PMID:27270323

In vitro differentiation of rat bone marrow mesenchymal stem cells into hepatocytes.

PubMed

Feng, Zhihui; Li, Changying; Jiao, Shuxian; Hu, Bin; Zhao, Lin

2011-01-01

To investigate the mechanism and regulation of differentiation from bone marrow mesenchymal stem cells (BMSCs) into hepatocytes and to find a new source for therapies of hepatic diseases. We isolated BMSCs for subsequent differentiation in the presence of hepatocyte growth factor (HGF) or beta-nerve growth factor (beta-NGF). Cell morphology was observed and cell surface phenotypings were detected by flow cytometry. a1-antitrypsin (AAT) expression of the hepatocytes was confirmed by immunocytochemistry and albumin expression was validated by real time PCR and western blotting. The expression of high-affinity nerve growth factor receptor (TrkA) and the activation of Erk pathway were detected by western blotting. Hepatocyte functional activity was confirmed by uptake of indocyanine green (ICG) assay. Small round cells appeared in the presence of HGF on day 10 or beta-NGF on day 12. Differentiated cells expressed albumin and had functional characteristics of hepatocytes, such as uptake of ICG. BMSCs were positive for TrkA. HGF and beta-NGF significantly upregulated the protein levels of phospho-Erk. BMSCs could differentiate into hepatocytes in the differentiation media including HGF or beta-NGF. Combination of HGF and beta-NGF significantly increased the efficiency of hepatic differentiation.

MiR-375 inhibits the hepatocyte growth factor-elicited migration of mesenchymal stem cells by downregulating Akt signaling.

PubMed

He, Lihong; Wang, Xianyao; Kang, Naixin; Xu, Jianwei; Dai, Nan; Xu, Xiaojing; Zhang, Huanxiang

2018-04-01

The migration of mesenchymal stem cells (MSCs) is critical for their use in cell-based therapies. Accumulating evidence suggests that microRNAs are important regulators of MSC migration. Here, we report that the expression of miR-375 was downregulated in MSCs treated with hepatocyte growth factor (HGF), which strongly stimulates the migration of these cells. Overexpression of miR-375 decreased the transfilter migration and the migration velocity of MSCs triggered by HGF. In our efforts to determine the mechanism by which miR-375 affects MSC migration, we found that miR-375 significantly inhibited the activation of Akt by downregulating its phosphorylation at T308 and S473, but had no effect on the activity of mitogen-activated protein kinases. Further, we showed that 3'phosphoinositide-dependent protein kinase-1 (PDK1), an upstream kinase necessary for full activation of Akt, was negatively regulated by miR-375 at the protein level. Moreover, miR-375 suppressed the phosphorylation of focal adhesion kinase (FAK) and paxillin, two important regulators of focal adhesion (FA) assembly and turnover, and decreased the number of FAs at cell periphery. Taken together, our results demonstrate that miR-375 inhibits HGF-elicited migration of MSCs through downregulating the expression of PDK1 and suppressing the activation of Akt, as well as influencing the tyrosine phosphorylation of FAK and paxillin and FA periphery distribution.

Quantitative gene expression deregulation in mantle-cell lymphoma: correlation with clinical and biologic factors.

PubMed

Kienle, Dirk; Katzenberger, Tiemo; Ott, German; Saupe, Doreen; Benner, Axel; Kohlhammer, Holger; Barth, Thomas F E; Höller, Sylvia; Kalla, Jörg; Rosenwald, Andreas; Müller-Hermelink, Hans Konrad; Möller, Peter; Lichter, Peter; Döhner, Hartmut; Stilgenbauer, Stephan

2007-07-01

There is evidence for a direct role of quantitative gene expression deregulation in mantle-cell lymphoma (MCL) pathogenesis. Our aim was to investigate gene expression associations with other pathogenic factors and the significance of gene expression in a multivariate survival analysis. Quantitative expression of 20 genes of potential relevance for MCL prognosis and pathogenesis were analyzed using real-time reverse transcriptase polymerase chain reaction and correlated with clinical and genetic factors, tumor morphology, and Ki-67 index in 65 MCL samples. Genomic losses at the loci of TP53, RB1, and P16 were associated with reduced transcript levels of the respective genes, indicating a gene-dosage effect as the pathomechanism. Analysis of gene expression correlations between the candidate genes revealed a separation into two clusters, one dominated by proliferation activators, another by proliferation inhibitors and regulators of apoptosis. Whereas only weak associations were identified between gene expression and clinical parameters or blastoid morphology, several genes were correlated closely with the Ki-67 index, including the short CCND1 variant (positive correlation) and RB1, ATM, P27, and BMI (negative correlation). In multivariate survival analysis, expression levels of MYC, MDM2, EZH2, and CCND1 were the strongest prognostic factors independently of tumor proliferation and clinical factors. These results indicate a pathogenic contribution of several gene transcript levels to the biology and clinical course of MCL. Genes can be differentiated into factors contributing to proliferation deregulation, either by enhancement or loss of inhibition, and proliferation-independent factors potentially contributing to MCL pathogenesis by apoptosis impairment.

Gene therapy for therapeutic angiogenesis in peripheral arterial disease - a systematic review and meta-analysis of randomized, controlled trials.

PubMed

Hammer, Alexandra; Steiner, Sabine

2013-09-01

Beyond pharmacological, endovascular and surgical treatment strategies for peripheral arterial disease (PAD), therapeutic angiogenesis has been advocated to relieve symptoms and support limb salvage, in particular in patients with critical limb ischemia. We aimed to systematically review randomized controlled trials (RCTs) of gene therapy in PAD. A systematic search of electronic databases was performed to identify RCTs studying local administration of pro-angiogenic growth factors (VEGF, FGF, HGF, Del-1, HIF-1alpha) using plasmid or viral gene transfer by intra-arterial or intra-muscular injections. Outcomes of interest comprised all-cause mortality, amputations, ulcer healing, walking distance and ankle-brachial index. If feasible, standard meta-analysis should be performed with subgroup analysis for claudicants and patients with critical limb ischemia (CLI). The systematic search yielded 12 RCTs for analysis from 1163 citations. In total, 1494 patients (29 % females) were included with the majority suffering from CLI (64 %). Various endpoints were improved by single studies, but none by a majority of studies. Meta-analysis showed neither a significant benefit nor harm for gene therapy when synthesizing data for all-cause mortality (OR 0.88, 95 % CI 0.62 - 1.26) amputations (OR 0.64, 95 % CI 0.31 - 1.31) or ulcer healing (OR 1.79, 95 % CI 0.8 - 4.01). No differences were seen between patients with intermittent claudication or CLI. Despite promising results in single studies, no clear benefit could be identified for gene therapy in PAD patients, irrespective of disease severity.

PreImplantation factor (PIF*) promotes embryotrophic and neuroprotective decidual genes: effect negated by epidermal growth factor.

PubMed

Duzyj, Christina M; Paidas, Michael J; Jebailey, Lellean; Huang, Jing Shun; Barnea, Eytan R

2014-01-01

Intimate embryo-maternal interaction is paramount for pregnancy success post-implantation. The embryo follows a specific developmental timeline starting with neural system, dependent on endogenous and decidual factors. Beyond altered genetics/epigenetics, post-natal diseases may initiate at prenatal/neonatal, post-natal period, or through a continuum. Preimplantation factor (PIF) secreted by viable embryos promotes implantation and trophoblast invasion. Synthetic PIF reverses neuroinflammation in non-pregnant models. PIF targets embryo proteins that protect against oxidative stress and protein misfolding. We report of PIF's embryotrophic role and potential to prevent developmental disorders by regulating uterine milieu at implantation and first trimester. PIF's effect on human implantation (human endometrial stromal cells (HESC)) and first-trimester decidua cultures (FTDC) was examined, by global gene expression (Affymetrix), disease-biomarkers ranking (GeneGo), neuro-specific genes (Ingenuity) and proteins (mass-spectrometry). PIF co-cultured epidermal growth factor (EGF) in both HESC and FTDC (Affymetrix) was evaluated. In HESC, PIF promotes neural differentiation and transmission genes (TLX2, EPHA10) while inhibiting retinoic acid receptor gene, which arrests growth. PIF promotes axon guidance and downregulates EGF-dependent neuroregulin signaling. In FTDC, PIF promotes bone morphogenetic protein pathway (SMAD1, 53-fold) and axonal guidance genes (EPH5) while inhibiting PPP2R2C, negative cell-growth regulator, involved in Alzheimer's and amyotrophic lateral sclerosis. In HESC, PIF affects angiotensin via beta-arrestin, transforming growth factor-beta (TGF-β), notch, BMP, and wingless-int (WNT) signaling pathways that promote neurogenesis involved in childhood neurodevelopmental diseases-autism and also affected epithelial-mesenchymal transition involved in neuromuscular disorders. In FTDC, PIF upregulates neural development and hormone signaling, while

PreImplantation factor (PIF*) promotes embryotrophic and neuroprotective decidual genes: effect negated by epidermal growth factor

PubMed Central

2014-01-01

Background Intimate embryo-maternal interaction is paramount for pregnancy success post-implantation. The embryo follows a specific developmental timeline starting with neural system, dependent on endogenous and decidual factors. Beyond altered genetics/epigenetics, post-natal diseases may initiate at prenatal/neonatal, post-natal period, or through a continuum. Preimplantation factor (PIF) secreted by viable embryos promotes implantation and trophoblast invasion. Synthetic PIF reverses neuroinflammation in non-pregnant models. PIF targets embryo proteins that protect against oxidative stress and protein misfolding. We report of PIF’s embryotrophic role and potential to prevent developmental disorders by regulating uterine milieu at implantation and first trimester. Methods PIF’s effect on human implantation (human endometrial stromal cells (HESC)) and first-trimester decidua cultures (FTDC) was examined, by global gene expression (Affymetrix), disease-biomarkers ranking (GeneGo), neuro-specific genes (Ingenuity) and proteins (mass-spectrometry). PIF co-cultured epidermal growth factor (EGF) in both HESC and FTDC (Affymetrix) was evaluated. Results In HESC, PIF promotes neural differentiation and transmission genes (TLX2, EPHA10) while inhibiting retinoic acid receptor gene, which arrests growth. PIF promotes axon guidance and downregulates EGF-dependent neuroregulin signaling. In FTDC, PIF promotes bone morphogenetic protein pathway (SMAD1, 53-fold) and axonal guidance genes (EPH5) while inhibiting PPP2R2C, negative cell-growth regulator, involved in Alzheimer’s and amyotrophic lateral sclerosis. In HESC, PIF affects angiotensin via beta-arrestin, transforming growth factor-beta (TGF-β), notch, BMP, and wingless-int (WNT) signaling pathways that promote neurogenesis involved in childhood neurodevelopmental diseases—autism and also affected epithelial-mesenchymal transition involved in neuromuscular disorders. In FTDC, PIF upregulates neural development

Stability-driven nonnegative matrix factorization to interpret spatial gene expression and build local gene networks.

PubMed

Wu, Siqi; Joseph, Antony; Hammonds, Ann S; Celniker, Susan E; Yu, Bin; Frise, Erwin

2016-04-19

Spatial gene expression patterns enable the detection of local covariability and are extremely useful for identifying local gene interactions during normal development. The abundance of spatial expression data in recent years has led to the modeling and analysis of regulatory networks. The inherent complexity of such data makes it a challenge to extract biological information. We developed staNMF, a method that combines a scalable implementation of nonnegative matrix factorization (NMF) with a new stability-driven model selection criterion. When applied to a set ofDrosophilaearly embryonic spatial gene expression images, one of the largest datasets of its kind, staNMF identified 21 principal patterns (PP). Providing a compact yet biologically interpretable representation ofDrosophilaexpression patterns, PP are comparable to a fate map generated experimentally by laser ablation and show exceptional promise as a data-driven alternative to manual annotations. Our analysis mapped genes to cell-fate programs and assigned putative biological roles to uncharacterized genes. Finally, we used the PP to generate local transcription factor regulatory networks. Spatially local correlation networks were constructed for six PP that span along the embryonic anterior-posterior axis. Using a two-tail 5% cutoff on correlation, we reproduced 10 of the 11 links in the well-studied gap gene network. The performance of PP with theDrosophiladata suggests that staNMF provides informative decompositions and constitutes a useful computational lens through which to extract biological insight from complex and often noisy gene expression data.

Multiple conversion between the genes encoding bacterial class-I release factors

PubMed Central

Ishikawa, Sohta A.; Kamikawa, Ryoma; Inagaki, Yuji

2015-01-01

Bacteria require two class-I release factors, RF1 and RF2, that recognize stop codons and promote peptide release from the ribosome. RF1 and RF2 were most likely established through gene duplication followed by altering their stop codon specificities in the common ancestor of extant bacteria. This scenario expects that the two RF gene families have taken independent evolutionary trajectories after the ancestral gene duplication event. However, we here report two independent cases of conversion between RF1 and RF2 genes (RF1-RF2 gene conversion), which were severely examined by procedures incorporating the maximum-likelihood phylogenetic method. In both cases, RF1-RF2 gene conversion was predicted to occur in the region encoding nearly entire domain 3, of which functions are common between RF paralogues. Nevertheless, the ‘direction’ of gene conversion appeared to be opposite from one another—from RF2 gene to RF1 gene in one case, while from RF1 gene to RF2 gene in the other. The two cases of RF1-RF2 gene conversion prompt us to propose two novel aspects in the evolution of bacterial class-I release factors: (i) domain 3 is interchangeable between RF paralogues, and (ii) RF1-RF2 gene conversion have occurred frequently in bacterial genome evolution. PMID:26257102

Synergistic effect of factor VII gene polymorphisms causing mild factor VII deficiency in a case of severe factor X deficiency.

PubMed

Deshpande, Rutuja; Ghosh, Kanjaksha; Shetty, Shrimati

2017-01-01

Congenital combined deficiency of coagulation factors VII and X are mainly attributed to large deletions involving both the genes in chromosome 13 or occasionally due to the coincidental occurrence of independently occurring mutations. We report the molecular basis of congenital combined deficiency of factors VII and X in a 6-year-old female child. Direct DNA sequencing of both factor VII (F7) and factor X (F10) genes showed a novel homozygous missense mutation p.Cys90Tyr (c.307G>A) in exon 4 of F10. No mutations were detected in F7; however, the patient was homozygous for three polymorphic alleles known to be associated with reduced factor VII levels. The present case illustrates the synergistic effect of multiple polymorphisms resulting in phenotypic factor VII deficiency in the absence of a pathogenic mutation.

c-Met must translocate to the nucleus to initiate calcium signals.

PubMed

Gomes, Dawidson A; Rodrigues, Michele A; Leite, M Fatima; Gomez, Marcus V; Varnai, Peter; Balla, Tamas; Bennett, Anton M; Nathanson, Michael H

2008-02-15

Hepatocyte growth factor (HGF) is important for cell proliferation, differentiation, and related activities. HGF acts through its receptor c-Met, which activates downstream signaling pathways. HGF binds to c-Met at the plasma membrane, where it is generally believed that c-Met signaling is initiated. Here we report that c-Met rapidly translocates to the nucleus upon stimulation with HGF. Ca(2+) signals that are induced by HGF result from phosphatidylinositol 4,5-bisphosphate hydrolysis and inositol 1,4,5-trisphosphate formation within the nucleus rather than within the cytoplasm. Translocation of c-Met to the nucleus depends upon the adaptor protein Gab1 and importin beta1, and formation of Ca(2+) signals in turn depends upon this translocation. HGF may exert its particular effects on cells because it bypasses signaling pathways in the cytoplasm to directly activate signaling pathways in the nucleus.

Tumour Necrosis Factor-alpha and Nuclear Factor-kappa B Gene Variants in Sepsis.

PubMed

Acar, Leyla; Atalan, Nazan; Karagedik, E Hande; Ergen, Arzu

2018-01-20

The humoral system is activated and various cytokines are released due to infections in tissues and traumatic damage. Nuclear factor-kappa B dimers are encoded by nuclear factor-kappa B genes and regulate transcription of several crucial proteins of inflammation such as tumour necrosis factor-alpha. To investigate the possible effect of polymorphisms on tumour necrosis factor-alpha serum levels with clinical and prognostic parameters of sepsis by determining the nuclear factor-kappa B-1-94 ins/del ATTG and tumour necrosis factor-alpha (-308 G/A) gene polymorphisms and tumour necrosis factor-alpha serum levels. Case-control study. Seventy-two patients with sepsis and 104 healthy controls were included in the study. In order to determine the polymorphisms of nuclear factor-kappa B-1-94 ins/del ATTG and tumour necrosis factor-alpha (-308 G/A), polymerase chain reaction-restriction fragment length polymorphism analysis was performed and serum tumour necrosis factor-alpha levels were determined using an enzyme-linked immunosorbent assay. We observed no significant differences in tumour necrosis factor-alpha serum levels between the study groups. In the patient group, an increase in the tumour necrosis factor-alpha serum levels in patients carrying the tumour necrosis factor-alpha (-308 G/A) A allele compared to those without the A allele was found to be statistically significant. Additionally, an increase in the tumour necrosis factor-alpha serum levels in patients carrying tumour necrosis factor-alpha (-308 G/A) AA genotype compared with patients carrying the AG or GG genotypes was statistically significant. No significant differences were found in these 2 polymorphisms between the patient and control groups (p>0.05). Our results showed the AA genotype and the A allele of the tumour necrosis factor-alpha (-308 G/A) polymorphism may be used as a predictor of elevated tumour necrosis factor-alpha levels in patients with sepsis.

Regulation of endogenous human gene expression by ligand-inducible TALE transcription factors.

PubMed

Mercer, Andrew C; Gaj, Thomas; Sirk, Shannon J; Lamb, Brian M; Barbas, Carlos F

2014-10-17

The construction of increasingly sophisticated synthetic biological circuits is dependent on the development of extensible tools capable of providing specific control of gene expression in eukaryotic cells. Here, we describe a new class of synthetic transcription factors that activate gene expression in response to extracellular chemical stimuli. These inducible activators consist of customizable transcription activator-like effector (TALE) proteins combined with steroid hormone receptor ligand-binding domains. We demonstrate that these ligand-responsive TALE transcription factors allow for tunable and conditional control of gene activation and can be used to regulate the expression of endogenous genes in human cells. Since TALEs can be designed to recognize any contiguous DNA sequence, the conditional gene regulatory system described herein will enable the design of advanced synthetic gene networks.

Stability-driven nonnegative matrix factorization to interpret spatial gene expression and build local gene networks

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wu, Siqi; Joseph, Antony; Hammonds, Ann S.

Spatial gene expression patterns enable the detection of local covariability and are extremely useful for identifying local gene interactions during normal development. The abundance of spatial expression data in recent years has led to the modeling and analysis of regulatory networks. The inherent complexity of such data makes it a challenge to extract biological information. We developed staNMF, a method that combines a scalable implementation of nonnegative matrix factorization (NMF) with a new stability-driven model selection criterion. When applied to a set of Drosophila early embryonic spatial gene expression images, one of the largest datasets of its kind, staNMF identifiedmore » 21 principal patterns (PP). Providing a compact yet biologically interpretable representation of Drosophila expression patterns, PP are comparable to a fate map generated experimentally by laser ablation and show exceptional promise as a data-driven alternative to manual annotations. Our analysis mapped genes to cell-fate programs and assigned putative biological roles to uncharacterized genes. Finally, we used the PP to generate local transcription factor regulatory networks. Spatially local correlation networks were constructed for six PP that span along the embryonic anterior-posterior axis. Using a two-tail 5% cutoff on correlation, we reproduced 10 of the 11 links in the well-studied gap gene network. In conclusion, the performance of PP with the Drosophila data suggests that staNMF provides informative decompositions and constitutes a useful computational lens through which to extract biological insight from complex and often noisy gene expression data.« less

Stability-driven nonnegative matrix factorization to interpret spatial gene expression and build local gene networks

DOE PAGES

Wu, Siqi; Joseph, Antony; Hammonds, Ann S.; ...

2016-04-06

Spatial gene expression patterns enable the detection of local covariability and are extremely useful for identifying local gene interactions during normal development. The abundance of spatial expression data in recent years has led to the modeling and analysis of regulatory networks. The inherent complexity of such data makes it a challenge to extract biological information. We developed staNMF, a method that combines a scalable implementation of nonnegative matrix factorization (NMF) with a new stability-driven model selection criterion. When applied to a set of Drosophila early embryonic spatial gene expression images, one of the largest datasets of its kind, staNMF identifiedmore » 21 principal patterns (PP). Providing a compact yet biologically interpretable representation of Drosophila expression patterns, PP are comparable to a fate map generated experimentally by laser ablation and show exceptional promise as a data-driven alternative to manual annotations. Our analysis mapped genes to cell-fate programs and assigned putative biological roles to uncharacterized genes. Finally, we used the PP to generate local transcription factor regulatory networks. Spatially local correlation networks were constructed for six PP that span along the embryonic anterior-posterior axis. Using a two-tail 5% cutoff on correlation, we reproduced 10 of the 11 links in the well-studied gap gene network. In conclusion, the performance of PP with the Drosophila data suggests that staNMF provides informative decompositions and constitutes a useful computational lens through which to extract biological insight from complex and often noisy gene expression data.« less

Transforming growth factor-beta inhibits the expression of clock genes.

PubMed

Gast, Heidemarie; Gordic, Sonja; Petrzilka, Saskia; Lopez, Martin; Müller, Andreas; Gietl, Anton; Hock, Christoph; Birchler, Thomas; Fontana, Adriano

2012-07-01

Disturbances of sleep-wake rhythms are an important problem in Alzheimer's disease (AD). Circadian rhythms are regulated by clock genes. Transforming growth factor-beta (TGF-β) is overexpressed in neurons in AD and is the only cytokine that is increased in cerebrospinal fluid (CSF). Our data show that TGF-β2 inhibits the expression of the clock genes Period (Per)1, Per2, and Rev-erbα, and of the clock-controlled genes D-site albumin promoter binding protein (Dbp) and thyrotroph embryonic factor (Tef). However, our results showed that TGF-β2 did not alter the expression of brain and muscle Arnt-like protein-1 (Bmal1). The concentrations of TGF-β2 in the CSF of 2 of 16 AD patients and of 1 of 7 patients with mild cognitive impairment were in the dose range required to suppress the expression of clock genes. TGF-β2-induced dysregulation of clock genes may alter neuronal pathways, which may be causally related to abnormal sleep-wake rhythms in AD patients. © 2012 New York Academy of Sciences.

Cancer-associated fibroblasts drive glycolysis in a targetable signaling loop implicated in head and neck squamous cell carcinoma progression.

PubMed

Kumar, Dhruv; New, Jacob; Vishwakarma, Vikalp; Joshi, Radhika; Enders, Jonathan; Lin, Fangchen; Dasari, Sumana; Gutierrez, Wade R; Leef, George; Ponnurangam, Sivapriya; Chavan, Hemantkumar; Ganaden, Lydia; Thornton, Mackenzie M; Dai, Hongying; Tawfik, Ossama; Straub, Jeffrey; Shnayder, Yelizaveta; Kakarala, Kiran; Tsue, Terance Ted; Girod, Douglas A; Van Houten, Bennett; Anant, Shrikant; Krishnamurthy, Partha; Thomas, Sufi Mary

2018-05-16

Despite aggressive therapies, head and neck squamous cell carcinoma (HNSCC) is associated with a less than 50% 5-year survival rate. Late stage HNSCC frequently consists of up to 80% cancer-associated fibroblasts (CAF). We previously reported that CAF-secreted hepatocyte growth factor (HGF) facilitates HNSCC progression, however very little is known about the role of CAFs in HNSCC metabolism. Here we demonstrate that CAF-secreted HGF increases extracellular lactate levels in HNSCC via upregulation of glycolysis. CAF-secreted HGF induced basic fibroblast growth factor (bFGF) secretion from HNSCC. CAFs were more efficient than HNSCC in using lactate as a carbon source. HNSCC-secreted bFGF increased mitochondrial oxidative phosphorylation (OXPHOS) and HGF secretion from CAFs. Combined inhibition of c-Met and FGFR significantly inhibited CAF-induced HNSCC growth in vitro and in vivo (p<0.001). Our cumulative findings underscore reciprocal signaling between CAF and HNSCC involving bFGF and HGF. This contributes to metabolic symbiosis and a targetable therapeutic axis involving c-Met and FGFR. Copyright ©2018, American Association for Cancer Research.

Genomewide analysis of TCP transcription factor gene family in Malus domestica.

PubMed

Xu, Ruirui; Sun, Peng; Jia, Fengjuan; Lu, Longtao; Li, Yuanyuan; Zhang, Shizhong; Huang, Jinguang

2014-12-01

Teosinte branched 1/cycloidea/proliferating cell factor 1 (TCP) proteins are a large family of transcriptional regulators in angiosperms. They are involved in various biological processes, including development and plant metabolism pathways. In this study, a total of 52 TCP genes were identified in apple (Malus domestica) genome. Bioinformatic methods were employed to predicate and analyse their relevant gene classification, gene structure, chromosome location, sequence alignment and conserved domains of MdTCP proteins. Expression analysis from microarray data showed that the expression levels of 28 and 51 MdTCP genes changed during the ripening and rootstock-scion interaction processes, respectively. The expression patterns of 12 selected MdTCP genes were analysed in different tissues and in response to abiotic stresses. All of the selected genes were detected in at least one of the tissues tested, and most of them were modulated by adverse treatments indicating that the MdTCPs were involved in various developmental and physiological processes. To the best of our knowledge, this is the first study of a genomewide analysis of apple TCP gene family. These results provide valuable information for studies on functions of the TCP transcription factor genes in apple.

Trophic factors from adipose tissue-derived multi-lineage progenitor cells promote cytodifferentiation of periodontal ligament cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Sawada, Keigo; Takedachi, Masahide, E-mail: takedati@dent.osaka-u.ac.jp; Yamamoto, Satomi

Stem and progenitor cells are currently being investigated for their applicability in cell-based therapy for periodontal tissue regeneration. We recently demonstrated that the transplantation of adipose tissue-derived multi-lineage progenitor cells (ADMPCs) enhances periodontal tissue regeneration in beagle dogs. However, the molecular mechanisms by which transplanted ADMPCs induce periodontal tissue regeneration remain to be elucidated. In this study, trophic factors released by ADMPCs were examined for their paracrine effects on human periodontal ligament cell (HPDL) function. ADMPC conditioned medium (ADMPC-CM) up-regulated osteoblastic gene expression, alkaline phosphatase activity and calcified nodule formation in HPDLs, but did not significantly affect their proliferative response.more » ADMPCs secreted a number of growth factors, including insulin-like growth factor binding protein 6 (IGFBP6), hepatocyte growth factor and vascular endothelial growth factor. Among these, IGFBP6 was most highly expressed. Interestingly, the positive effects of ADMPC-CM on HPDL differentiation were significantly suppressed by transfecting ADMPCs with IGFBP6 siRNA. Our results suggest that ADMPCs transplanted into a defect in periodontal tissue release trophic factors that can stimulate the differentiation of HPDLs to mineralized tissue-forming cells, such as osteoblasts and cementoblasts. IGFBP6 may play crucial roles in ADMPC-induced periodontal regeneration. - Highlights: • ADMPC-derived humoral factors stimulate cytodifferentiation of HPDLs. • ADMPCs secret growth factors including IGFBP6, VEGF and HGF. • IGFBP6 is involved in the promotion effect of ADMPC-CM on HPDL cytodifferentiation.« less

EBF factors drive expression of multiple classes of target genes governing neuronal development.

PubMed

Green, Yangsook S; Vetter, Monica L

2011-04-30

Early B cell factor (EBF) family members are transcription factors known to have important roles in several aspects of vertebrate neurogenesis, including commitment, migration and differentiation. Knowledge of how EBF family members contribute to neurogenesis is limited by a lack of detailed understanding of genes that are transcriptionally regulated by these factors. We performed a microarray screen in Xenopus animal caps to search for targets of EBF transcriptional activity, and identified candidate targets with multiple roles, including transcription factors of several classes. We determined that, among the most upregulated candidate genes with expected neuronal functions, most require EBF activity for some or all of their expression, and most have overlapping expression with ebf genes. We also found that the candidate target genes that had the most strongly overlapping expression patterns with ebf genes were predicted to be direct transcriptional targets of EBF transcriptional activity. The identification of candidate targets that are transcription factor genes, including nscl-1, emx1 and aml1, improves our understanding of how EBF proteins participate in the hierarchy of transcription control during neuronal development, and suggests novel mechanisms by which EBF activity promotes migration and differentiation. Other candidate targets, including pcdh8 and kcnk5, expand our knowledge of the types of terminal differentiated neuronal functions that EBF proteins regulate.

Inhibition of attachment of oral bacteria to immortalized human gingival fibroblasts (HGF-1) by tea extracts and tea components

PubMed Central

2013-01-01

Background Tea has been suggested to promote oral health by inhibiting bacterial attachment to the oral cavity. Most studies have focused on prevention of bacterial attachment to hard surfaces such as enamel. Findings This study investigated the effect of five commercial tea (green, oolong, black, pu-erh and chrysanthemum) extracts and tea components (epigallocatechin gallate and gallic acid) on the attachment of five oral pathogens (Streptococcus mutans ATCC 25175, Streptococcus mutans ATCC 35668, Streptococcus mitis ATCC 49456, Streptococcus salivarius ATCC 13419 and Actinomyces naeslundii ATCC 51655) to the HGF-1 gingival cell line. Extracts of two of the teas (pu-erh and chrysanthemum) significantly (p < 0.05) reduced attachment of all the Streptococcus strains by up to 4 log CFU/well but effects of other teas and components were small. Conclusions Pu-erh and chrysanthemum tea may have the potential to reduce attachment of oral pathogens to gingival tissue and improve the health of oral soft tissues. PMID:23578062

Inhibition of attachment of oral bacteria to immortalized human gingival fibroblasts (HGF-1) by tea extracts and tea components.

PubMed

Wang, Yi; Chung, Felicia F L; Lee, Sui M; Dykes, Gary A

2013-04-11

Tea has been suggested to promote oral health by inhibiting bacterial attachment to the oral cavity. Most studies have focused on prevention of bacterial attachment to hard surfaces such as enamel. This study investigated the effect of five commercial tea (green, oolong, black, pu-erh and chrysanthemum) extracts and tea components (epigallocatechin gallate and gallic acid) on the attachment of five oral pathogens (Streptococcus mutans ATCC 25175, Streptococcus mutans ATCC 35668, Streptococcus mitis ATCC 49456, Streptococcus salivarius ATCC 13419 and Actinomyces naeslundii ATCC 51655) to the HGF-1 gingival cell line. Extracts of two of the teas (pu-erh and chrysanthemum) significantly (p < 0.05) reduced attachment of all the Streptococcus strains by up to 4 log CFU/well but effects of other teas and components were small. Pu-erh and chrysanthemum tea may have the potential to reduce attachment of oral pathogens to gingival tissue and improve the health of oral soft tissues.

Reinke's edema: investigations on the role of MIB-1 and hepatocyte growth factor.

PubMed

Artico, M; Bronzetti, E; Ionta, B; Bruno, M; Greco, A; Ruoppolo, G; De Virgilio, A; Longo, L; De Vincentiis, M

2010-07-08

Reinke's edema is a benign disease of the human vocal fold, which mainly affects the sub-epithelial layer of the vocal fold. Microscopic observations show a strongly oedematous epithelium with loosened intercellular junctions, a disruption of the extracellular connections between mucosal epithelium and connective tissue, closely adherent to the thyroarytenoid muscle. Thickening of the basal layer of epithelium, known as Reinke's space, high deposition of fibronectin and chronic inflammatory infiltration it is also visible. We analyzed, together with the hepatocyte growth factor (HGF), the expression level of MIB-1 in samples harvested from patients affected by Reinke's edema, in order to define its biological role and consider it as a possible prognostic factor in the follow-up after surgical treatment. We observed a moderate expression of HGF in the lamina propria of the human vocal fold and in the basal membrane of the mucosal epithelium. Our finding suggests that this growth factor acts as an antifibrotic agent in Reinke's space and affects the fibronectin deposition in the lamina propria. MIB-1, on the contrary, showed a weak expression in the basement membrane of the mucosal epithelium and a total absence in the lamina propria deep layer, thus suggesting that only the superficial layer is actively involved in the reparatory process with a high regenerative capacity, together with a high deposition of fibronectin. The latter is necessary for the cellular connections reconstruction, after the inflammatory infiltration.

Reinke's Edema: investigations on the role of MIB-1 and hepatocyte growth factor

PubMed Central

Artico, M.; Bronzetti, E.; Ionta, B.; Bruno, M.; Greco, A.; Ruoppolo, G.; De Virgilio, A.; Longo, L.; De Vincentiis, M.

2010-01-01

Reinke's edema is a benign disease of the human vocal fold, which mainly affects the sub-epithelial layer of the vocal fold. Microscopic observations show a strongly oedematous epithelium with loosened intercellular junctions, a disruption of the extracellular connections between mucosal epithelium and connective tissue, closely adherent to the thyroarytenoid muscle. Thickening of the basal layer of epithelium, known as Reinke's space, high deposition of fibronectin and chronic inflammatory infiltration it is also visible. We analyzed, together with the hepatocyte growth factor (HGF), the expression level of MIB-1 in samples harvested from patients affected by Reinke's edema, in order to define its biological role and consider it as a possible prognostic factor in the follow-up after surgical treatment. We observed a moderate expression of HGF in the lamina propria of the human vocal fold and in the basal membrane of the mucosal epithelium. Our finding suggests that this growth factor acts as an anti - fibrotic agent in Reinke's space and affects the fibronectin deposition in the lamina propria. MIB-1, on the contrary, showed a weak expression in the basement membrane of the mucosal epithelium and a total absence in the lamina propria deep layer, thus suggesting that only the superficial layer is actively involved in the reparatory process with a high regenerative capacity, together with a high deposition of fibronectin. The latter is necessary for the cellular connections reconstruction, after the inflammatory infiltration. PMID:20819770

Inhibition of neuropathic hyperalgesia by intrathecal bone marrow stromal cells is associated with alteration of multiple soluble factors in cerebrospinal fluid.

PubMed

Fischer, Gregory; Wang, Fei; Xiang, Hongfei; Bai, Xiaowen; Yu, Hongwei; Hogan, Quinn H

2017-09-01

Injury-induced neuropathic pain remains a serious clinical problem. Recent studies indicate that bone marrow stromal cells (BMSCs) effectively attenuate chronic neuropathic pain in animal models. Here, we examined the therapeutic effect of intrathecal administration of BMSCs isolated from young (1-month-old) rats on pain hypersensitivity induced by tibial nerve injury. Cerebrospinal fluid (CSF) was collected and analyzed to examine the effect of BMSC administration on the expression of 67 soluble factors in CSF. A sustained remission in injury-induced mechanical hyperalgesia was observed in BMSC-treated rats but not in control animals. Engrafted BMSCs were observed in spinal cords and dorsal root ganglia at 5 weeks after cell injection. Injury significantly decreased the levels of six soluble factors in CSF: intercellular adhesion molecule 1 (ICAM-1), interleukin-1β (IL-1β), IL-10, hepatocyte growth factor (HGF), Nope protein, and neurogenic locus notch homolog protein 1 (Notch-1). Intrathecal BMSCs significantly attenuated the injury-induced reduction of ICAM-1, IL-1β, HGF, IL-10, and Nope. This study adds to evidence supporting the use of intrathecal BMSCs in pain control and shows that this effect is accompanied by the reversal of injury-induced reduction of multiple CSF soluble factors. Our findings suggest that these soluble factors may be potential targets for treating chronic pain.

Liposomal gene transfer of keratinocyte growth factor improves wound healing by altering growth factor and collagen expression.

PubMed

Pereira, Clifford T; Herndon, David N; Rocker, Roland; Jeschke, Marc G

2007-05-15

Growth factors affect the complex cascade of wound healing; however, interaction between different growth factors during dermal and epidermal regeneration are still not entirely defined. In the present study, we thought to determine the interaction between keratinocyte growth factor (KGF) administered as liposomal cDNA with other dermal and epidermal growth factors and collagen synthesis in an acute wound. Rats received an acute wound and were divided into two groups to receive weekly subcutaneous injections of liposomes plus the Lac-Z gene (0.22 microg, vehicle), or liposomes plus the KGF cDNA (2.2 microg) and Lac-Z gene (0.22 microg). Histological and immunohistochemical techniques were used to determine growth factor, collagen expression, and dermal and epidermal structure. KGF cDNA increased insulin-like growth factor-I (IGF-I), insulin-like growth factor binding protein-3 (IGFBP-3), and fibroblast growth factor (FGF), decreased transforming growth factor-beta (TGF-beta), while it had no effect on platelet-derived growth factor (PDGF) levels in the wound. KGF cDNA significantly increased collagen Type IV at both the wound edge as well as the wound bed, while it had no effect on collagen Type I and III. KGF cDNA increased re-epithelialization, improved dermal regeneration, and increased neovascularization. Exogenous administered KGF cDNA causes increases in IGF-I, IGF-BP3, FGF, and collagen IV and decreases TGF-beta concentration. KGF gene transfer accelerates wound healing without causing an increase in collagen I or III.

Prognostic factors and genes associated with endometrial cancer based on gene expression profiling by bioinformatics analysis.

PubMed

Zhang, Ying; Zhang, Wei; Li, Xinglan; Li, Dapeng; Zhang, Xiaoling; Yin, Yajie; Deng, Xiangyun; Sheng, Xiugui

2016-06-01

Endometrial cancer (EC) is the most prevalent malignancy worldwide. Although several efforts had been made to explore the molecular mechanism responsible for EC progression, it is still not fully understood. To evaluate the clinical characteristics and prognostic factors of patients with EC, and further to search for novel genes associated with EC progression. We recruited 328 patients with EC and analyzed prognostic factors using Cox proportional hazard regression model. Further, a gene expression profile of EC was used to identify the differentially expressed genes (DEGs) between normal samples and tumor samples. Subsequently, Kyoto Encyclopedia of Genes and Genomes pathway enrichment analysis ( http://www.genome.jp/kegg/ ) for DEGs were performed, and then protein-protein interaction (PPI) network of DEGs as well as the subnetwork of PPI were constructed with plug-in, MCODE by mapping DEGs into the Search Tool for the Retrieval of Interacting Genes database. Our results showed that body mass index (BMI), hypertension, myometrial invasion, pathological type, and Glut4 positive expression were prognostic factors in EC (P < 0.05). Bioinformatics analysis showed that upregulated DEGs were associated with cell cycle, and downregulated DEGs were related to MAPK pathway. Meanwhile, PPI network analysis revealed that upregulated CDK1 and CCNA2 as well as downregulated JUN and FOS were listed in top two nodes with high degrees. Patients with EC should be given more focused attentions in respect of pathological type, BMI, hypertension, and Glut4-positive expression. In addition, CDK1, CCNA2, JUN, and FOS might play important roles in EC development.

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.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Carmona-Rodriguez, Bruno; Alvarez-Perez, Marco Antonio; Narayanan, A. Sampath

We recently presented evidence showing that a human cementoblastoma-derived protein, named Cementum Protein 1 (CEMP1) may play a role as a local regulator of cementoblast differentiation and cementum-matrix mineralization. This protein was shown to be expressed by cementoblasts and progenitor cells localized in the periodontal ligament. In this study we demonstrate that transfection of CEMP1 into human gingival fibroblasts (HGF) induces mineralization and expression of bone and cementum-matrix proteins. The transfected HGF cells had higher alkaline phosphatase activity and proliferation rate and they expressed genes for alkaline phosphatase, bone sialoprotein, osteocalcin, osteopontin, the transcription factor Runx2/Cbfa1, and cementum attachment proteinmore » (CAP). They also produced biological-type hydroxyapatite. These findings indicate that the CEMP1 might participate in differentiation and mineralization of nonosteogenic cells, and that it might have a potential function in cementum and bone formation.« less

Identifying Stress Transcription Factors Using Gene Expression and TF-Gene Association Data

PubMed Central

Wu, Wei-Sheng; Chen, Bor-Sen

2007-01-01

Unicellular organisms such as yeasts have evolved to survive environmental stresses by rapidly reorganizing the genomic expression program to meet the challenges of harsh environments. The complex adaptation mechanisms to stress remain to be elucidated. In this study, we developed Stress Transcription Factor Identification Algorithm (STFIA), which integrates gene expression and TF-gene association data to identify the stress transcription factors (TFs) of six kinds of stresses. We identified some general stress TFs that are in response to various stresses, and some specific stress TFs that are in response to one specific stress. The biological significance of our findings is validated by the literature. We found that a small number of TFs may be sufficient to control a wide variety of expression patterns in yeast under different stresses. Two implications can be inferred from this observation. First, the adaptation mechanisms to different stresses may have a bow-tie structure. Second, there may exist extensive regulatory cross-talk among different stress responses. In conclusion, this study proposes a network of the regulators of stress responses and their mechanism of action. PMID:20066130

Naturally occurring mutations in the human 5-lipoxygenase gene promoter that modify transcription factor binding and reporter gene transcription.

PubMed

In, K H; Asano, K; Beier, D; Grobholz, J; Finn, P W; Silverman, E K; Silverman, E S; Collins, T; Fischer, A R; Keith, T P; Serino, K; Kim, S W; De Sanctis, G T; Yandava, C; Pillari, A; Rubin, P; Kemp, J; Israel, E; Busse, W; Ledford, D; Murray, J J; Segal, A; Tinkleman, D; Drazen, J M

1997-03-01

Five lipoxygenase (5-LO) is the first committed enzyme in the metabolic pathway leading to the synthesis of the leukotrienes. We examined genomic DNA isolated from 25 normal subjects and 31 patients with asthma (6 of whom had aspirin-sensitive asthma) for mutations in the known transcription factor binding regions and the protein encoding region of the 5-LO gene. A family of mutations in the G + C-rich transcription factor binding region was identified consisting of the deletion of one, deletion of two, or addition of one zinc finger (Sp1/Egr-1) binding sites in the region 176 to 147 bp upstream from the ATG translation start site where there are normally 5 Sp1 binding motifs in tandem. Reporter gene activity directed by any of the mutant forms of the transcription factor binding region was significantly (P < 0.05) less effective than the activity driven by the wild type transcription factor binding region. Electrophoretic mobility shift assays (EMSAs) demonstrated the capacity of wild type and mutant transcription factor binding regions to bind nuclear extracts from human umbilical vein endothelial cells (HUVECs). These data are consistent with a family of mutations in the 5-LO gene that can modify reporter gene transcription possibly through differences in Sp1 and Egr-1 transactivation.

[Long distance-PCR for detection of factor VIII gene inversion in patients with severe hemophilia A].

PubMed

Ding, Pei-Fang; Sun, Wei-Sheng; Wang, Qin-You; Liu, De-Chun; Zhang, Xue-Qin; Teng, Bin; Shen, Fa-Kui

2003-08-01

The aim of current study was to detect intron 22 inversion of factor VIII gene in severe hemophilia A (HA) patients and screen the carriers of the gene inversion. Fifty-five cases of severe HA were involved and factor VIII gene inversion was detected and identified by long distance-PCR (LD-PCR) and 0.6% agarose gel electrophoresis. The 11 kb and 12 kb bands indicate the factor VIII gene inversion and non-inversion, respectively. Occurring of both 11 kb and 12 kb bands indicates a carrier of the inversion. The results showed that factor VIII gene inversion existed in 22 out of 55 cases, which accounted for about 40% of total detected patients. Five carriers of factor VIII gene inversion were diagnosed from the members in 15 families. In conclusion, LD-PCR assay is a simple, rapid and accurate method for detection of factor VIII gene inversion, and this approach is helpful in screening, carrier testing, and prenatal diagnosis of severe hemophilia A.

Tyrosine residues 654 and 670 in {beta}-cat enin are crucial in regulation of Met-{beta}-catenin interactions

DOE Office of Scientific and Technical Information (OSTI.GOV)

Zeng, Gang; Apte, Udayan; Micsenyi, Amanda

2006-11-01

{beta}-catenin, a key component of the canonical Wnt pathway, is also regulated by tyrosine phosphorylation that regulates its association to E-cadherin. Previously, we reported its association with the hepatocyte growth factor (HGF) receptor Met at the membrane. HGF induced Met-{beta}-catenin dissociation and nuclear translocation of {beta}-catenin, which was tyrosine-phosphorylation-dependent. Here, we further investigate the Met-{beta}-catenin interaction by selectively mutating several tyrosine residues, alone or in combination, in {beta}-catenin. The mutants were subcloned into FLAG-CMV vector and stably transfected into rat hepatoma cells, which were treated with HGF. All single or double-mutant-transfected cells continued to show HGF-induced nuclear translocation of FLAG-{beta}-cateninmore » except the mutations affecting 654 and 670 simultaneously (Y654/670F), which coincided with the lack of formation of {beta}-catenin-TCF complex and DNA synthesis, in response to the HGF treatment. In addition, the Y654/670F-transfected cells also showed no phosphorylation of {beta}-catenin or dissociation from Met in response to HGF. Thus, intact 654 and 670 tyrosine residues in {beta}-catenin are crucial in HGF-mediated {beta}-catenin translocation, activation and mitogenesis.« less

Epidermal growth factor gene is a newly identified candidate gene for gout.

PubMed

Han, Lin; Cao, Chunwei; Jia, Zhaotong; Liu, Shiguo; Liu, Zhen; Xin, Ruosai; Wang, Can; Li, Xinde; Ren, Wei; Wang, Xuefeng; Li, Changgui

2016-08-10

Chromosome 4q25 has been identified as a genomic region associated with gout. However, the associations of gout with the genes in this region have not yet been confirmed. Here, we performed two-stage analysis to determine whether variations in candidate genes in the 4q25 region are associated with gout in a male Chinese Han population. We first evaluated 96 tag single nucleotide polymorphisms (SNPs) in eight inflammatory/immune pathway- or glucose/lipid metabolism-related genes in the 4q25 region in 480 male gout patients and 480 controls. The SNP rs12504538, located in the elongation of very-long-chain-fatty-acid-like family member 6 gene (Elovl6), was found to be associated with gout susceptibility (Padjusted = 0.00595). In the second stage of analysis, we performed fine mapping analysis of 93 tag SNPs in Elovl6 and in the epidermal growth factor gene (EGF) and its flanking regions in 1017 male patients gout and 1897 healthy male controls. We observed a significant association between the T allele of EGF rs2298999 and gout (odds ratio = 0.77, 95% confidence interval = 0.67-0.88, Padjusted = 6.42 × 10(-3)). These results provide the first evidence for an association between the EGF rs2298999 C/T polymorphism and gout. Our findings should be validated in additional populations.

A review of the influence of growth factors and cytokines in in vitro human keratinocyte migration.

PubMed

Peplow, Philip V; Chatterjee, Marissa P

2013-04-01

Keratinocyte migration from the wound edge is a crucial step in the reepithelization of cutaneous wounds. Growth factors and cytokines, released from cells that invade the wound matrix, play an important role, and several in vitro assays have been performed to elucidate this. The purposes of this study were to review in vitro human studies on keratinocyte migration to identify those growth factors or cytokines that stimulate keratinocyte migration and whether these assays might serve as a screening procedure prior to testing combinations of growth factors or cytokines to promote wound closure in vivo. Research papers investigating effect of growth factors and cytokines on human keratinocyte migration in vitro were retrieved from library sources, PubMed databases, reference lists of papers, and searches of relevant journals. Fourteen different growth factors and cytokines enhanced migration in scratch wound assay and HGF together with TGF-β, and IGF-1 with EGF, were more stimulatory than either growth factor alone. HGF with TGF-β1 had a greater chemokinetic effect than either growth factor alone in transmigration assay. TGF-β1, FGF-7, FGF-2 and AGF were chemotactic to keratinocytes. EGF, TGF-α, IL-1α, IGF and MGSA enhanced cell migration on ECM proteins. Many growth factors and cytokines enhanced migration of keratinocytes in vitro, and certain combinations of growth factors were more stimulatory than either alone. These and other combinations that stimulate keratinocyte migration in vitro should be tested for effect on wound closure and repair in vivo. The scratch wound assay provides a useful, inexpensive and easy-to-perform screening method for testing individual or combinations of growth factors or cytokines, or growth factors combined with other modalities such as laser irradiation, prior to performing wound healing studies with laboratory animals. Copyright © 2013 Elsevier Ltd. All rights reserved.

The Chapel Hill hemophilia A dog colony exhibits a factor VIII gene inversion

PubMed Central

Lozier, Jay N.; Dutra, Amalia; Pak, Evgenia; Zhou, Nan; Zheng, Zhili; Nichols, Timothy C.; Bellinger, Dwight A.; Read, Marjorie; Morgan, Richard A.

2002-01-01

In the Chapel Hill colony of factor VIII-deficient dogs, abnormal sequence (ch8, for canine hemophilia 8, GenBank no. AF361485) follows exons 1–22 in the factor VIII transcript in place of exons 23–26. The canine hemophilia 8 locus (ch8) sequence was found in a 140-kb normal dog genomic DNA bacterial artificial chromosome (BAC) clone that was completely outside the factor VIII gene, but not in BAC clones containing the factor VIII gene. The BAC clone that contained ch8 also contained a homologue of F8A (factor 8 associated) sequence, which participates in a common inversion that causes severe hemophilia A in humans. Fluorescence in situ hybridization analysis indicated that exons 1–26 normally proceed sequentially from telomere to centromere at Xq28, and ch8 is telomeric to the factor VIII gene. The appearance of an “upstream” genomic sequence element (ch8) at the end of the aberrant factor VIII transcript suggested that an inversion of genomic DNA replaced factor VIII exons 22–26 with ch8. The F8A sequence appeared also in overlapping normal BAC clones containing factor VIII sequence. We hypothesized that homologous recombination between copies of canine F8A inside and outside the factor VIII gene had occurred, as in human hemophilia A. High-resolution fluorescent in situ hybridization on hemophilia A dog DNA revealed a pattern consistent with this inversion mechanism. We also identified a HindIII restriction fragment length polymorphism of F8A fragments that distinguished hemophilia A, carrier, and normal dogs' DNA. The Chapel Hill hemophilia A dog colony therefore replicates the factor VIII gene inversion commonly seen in humans with severe hemophilia A. PMID:12242334

The association of environmental, individual factors, and dopamine pathway gene variation with smoking cessation.

PubMed

Li, Suyun; Wang, Qiang; Pan, Lulu; Yang, Xiaorong; Li, Huijie; Jiang, Fan; Zhang, Nan; Han, Mingkui; Jia, Chongqi

2017-09-01

This study aimed to examine whether dopamine (DA) pathway gene variation were associated with smoking cessation, and compare the relative importance of infulence factors on smoking cessation. Participants were recruited from 17 villages of Shandong Province, China. Twenty-five single nucleotide polymorphisms in 8 DA pathway genes were genotyped. Weighted gene score of each gene was used to analyze the whole gene effect. Logistic regression was used to calculate odds ratios (OR) of the total gene score for smoking cessation. Dominance analysis was employed to compare the relative importance of individual, heaviness of smoking, psychological and genetic factors on smoking cessation. 415 successful spontaneous smoking quitters served as the cases, and 404 unsuccessful quitters served as the controls. A significant negative association of total DA pathway gene score and smoking cessation was observed (p < 0.001, OR: 0.25, 95% CI 0.16-0.38). Dominance analysis showed that the most important predictor for smoking cessation was heaviness of smoking score (42%), following by individual (40%), genetic (10%) and psychological score (8%). In conclusion, although the DA pathway gene variation was significantly associated with successful smoking cessation, heaviness of smoking and individual factors had bigger effect than genetic factors on smoking cessation.

Ultrasound Irradiation Combined with Hepatocyte Growth Factor Accelerate the Hepatic Differentiation of Human Bone Marrow Mesenchymal Stem Cells.

PubMed

Li, Fan; Liu, Yang; Cai, Yingyu; Li, Xin; Bai, Min; Sun, Ting; Du, Lianfang

2018-05-01

This study investigated the impact of ultrasound (US) irradiation on the hepatic differentiation of human bone marrow mesenchymal stem cells (hBMSCs) induced by hepatocyte growth factor (HGF) and the possible mechanisms. We treated hBMSCs, using HGF with and without US irradiation. Cell viability and stem cell surface markers were analyzed. Hepatocyte-like cell markers and functional markers including α-fetoprotein (αFP/AFP), cytokeratin 18 (CK18), albumin (ALB) and glycogen content were analyzed at the time point of day 1, 3 and 5 after treatment. The involvement of Wnt/β-catenin signaling pathway was evaluated as well. The results showed that the US treatment at 1.0 W/cm 2 or 1.5 W/cm 2 for 30 s or 60 s conditions yielded favorable cell viability and engendered stem cell differentiation. At day 5, the expressions of AFP, CK18, ALB and the glycogen content were significantly elevated in the US-treated group at both messenger ribonucleic acid and protein levels (all p <0.05), in comparison with HGF and control groups. Among all the US treated groups, the expression levels of specific hepatic markers in the (1.5 W/cm 2 for 60 s) group were the highest. Furthermore, Wnt1, β-Catenin, c-Myc and Cyclin D1 were significantly increased after US irradiation (all p <0.05), and the enhancements of c-Myc and Cyclin D1 could be obviously impaired by the inhibitor ICG-001 (p <0.05, p <0.05), in accordance with decreased ALB and CK18 expression and glycogen content (all p <0.05). In conclusion, US irradiation was able to promote the hBMSCs' differentiation mediated by HGF in vitro safely, easily and controllably. The activation of Wnt/β-catenin signaling pathway was involved in this process. US irradiation could serve as a potentially beneficial tool for the research and application of stem cell differentiation. Copyright © 2018 World Federation for Ultrasound in Medicine and Biology. Published by Elsevier Inc. All rights reserved.

Arabidopsis ensemble reverse-engineered gene regulatory network discloses interconnected transcription factors in oxidative stress.

PubMed

Vermeirssen, Vanessa; De Clercq, Inge; Van Parys, Thomas; Van Breusegem, Frank; Van de Peer, Yves

2014-12-01

The abiotic stress response in plants is complex and tightly controlled by gene regulation. We present an abiotic stress gene regulatory network of 200,014 interactions for 11,938 target genes by integrating four complementary reverse-engineering solutions through average rank aggregation on an Arabidopsis thaliana microarray expression compendium. This ensemble performed the most robustly in benchmarking and greatly expands upon the availability of interactions currently reported. Besides recovering 1182 known regulatory interactions, cis-regulatory motifs and coherent functionalities of target genes corresponded with the predicted transcription factors. We provide a valuable resource of 572 abiotic stress modules of coregulated genes with functional and regulatory information, from which we deduced functional relationships for 1966 uncharacterized genes and many regulators. Using gain- and loss-of-function mutants of seven transcription factors grown under control and salt stress conditions, we experimentally validated 141 out of 271 predictions (52% precision) for 102 selected genes and mapped 148 additional transcription factor-gene regulatory interactions (49% recall). We identified an intricate core oxidative stress regulatory network where NAC13, NAC053, ERF6, WRKY6, and NAC032 transcription factors interconnect and function in detoxification. Our work shows that ensemble reverse-engineering can generate robust biological hypotheses of gene regulation in a multicellular eukaryote that can be tested by medium-throughput experimental validation. © 2014 American Society of Plant Biologists. All rights reserved.

Association between serum ligands and the skin toxicity of anti-epidermal growth factor receptor antibody in metastatic colorectal cancer.

PubMed

Takahashi, Naoki; Yamada, Yasuhide; Furuta, Koh; Nagashima, Kengo; Kubo, Akiko; Sasaki, Yusuke; Shoji, Hirokazu; Honma, Yoshitaka; Iwasa, Satoru; Okita, Natsuko; Takashima, Atsuo; Kato, Ken; Hamaguchi, Tetsuya; Shimada, Yasuhiro

2015-05-01

Skin toxicity is a known clinical signature used to predict the prognosis of anti-epidermal growth factor receptor (EGFR) antibody treatment in metastatic colorectal cancer (mCRC). There are no biological markers to predict skin toxicity before anti-EGFR antibody treatment in mCRC patients. Between August 2008 and August 2011, pretreatment serum samples were obtained from KRAS wild-type (WT) patients who received anti-EGFR antibody treatment. Serum levels of ligands were measured by ELISA. A total of 103 KRAS WT patients were enrolled in the study. Progression-free survival and overall survival of patients with a high grade (grade 2-3) of skin toxicity were significantly longer than those with a low grade (grade 0-1) of skin toxicity (median progression-free survival, 6.4 months vs 2.4 months, P < 0.001; median overall survival, 14.6 months vs 7.1 months, P = 0.006). There were significant differences in distribution of serum levels of epiregulin (EREG), amphiregulin (AREG), and hepatocyte growth factor (HGF) between groups of low/high grade of skin toxicity (P < 0.048, P < 0.012, P < 0.012, respectively). In addition, serum levels of HGF, EREG, and AREG were inversely proportional to grades of skin toxicity as determined by the Cochran-Armitage test (P = 0.019, P = 0.047, P = 0.021, respectively). Our study indicated that serum levels such as HGF, EREG, and AREG may be significant markers to predict the grade of skin toxicity and the prognosis of anti-EGFR antibody treatment, which contribute to improvement of the management of skin toxicity and survival time in mCRC patients. © 2015 The Authors. Cancer Science published by Wiley Publishing Asia Pty Ltd on behalf of Japanese Cancer Association.

Aberrant RNA splicing in cancer; expression changes and driver mutations of splicing factor genes.

PubMed

Sveen, A; Kilpinen, S; Ruusulehto, A; Lothe, R A; Skotheim, R I

2016-05-12

Alternative splicing is a widespread process contributing to structural transcript variation and proteome diversity. In cancer, the splicing process is commonly disrupted, resulting in both functional and non-functional end-products. Cancer-specific splicing events are known to contribute to disease progression; however, the dysregulated splicing patterns found on a genome-wide scale have until recently been less well-studied. In this review, we provide an overview of aberrant RNA splicing and its regulation in cancer. We then focus on the executors of the splicing process. Based on a comprehensive catalog of splicing factor encoding genes and analyses of available gene expression and somatic mutation data, we identify cancer-associated patterns of dysregulation. Splicing factor genes are shown to be significantly differentially expressed between cancer and corresponding normal samples, and to have reduced inter-individual expression variation in cancer. Furthermore, we identify enrichment of predicted cancer-critical genes among the splicing factors. In addition to previously described oncogenic splicing factor genes, we propose 24 novel cancer-critical splicing factors predicted from somatic mutations.

Engineering synthetic TALE and CRISPR/Cas9 transcription factors for regulating gene expression.

PubMed

Kabadi, Ami M; Gersbach, Charles A

2014-09-01

Engineered DNA-binding proteins that can be targeted to specific sites in the genome to manipulate gene expression have enabled many advances in biomedical research. This includes generating tools to study fundamental aspects of gene regulation and the development of a new class of gene therapies that alter the expression of endogenous genes. Designed transcription factors have entered clinical trials for the treatment of human diseases and others are in preclinical development. High-throughput and user-friendly platforms for designing synthetic DNA-binding proteins present innovative methods for deciphering cell biology and designing custom synthetic gene circuits. We review two platforms for designing synthetic transcription factors for manipulating gene expression: Transcription activator-like effectors (TALEs) and the RNA-guided clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 system. We present an overview of each technology and a guide for designing and assembling custom TALE- and CRISPR/Cas9-based transcription factors. We also discuss characteristics of each platform that are best suited for different applications. Copyright © 2014 Elsevier Inc. All rights reserved.

Resveratrol regulates gene transcription via activation of stimulus-responsive transcription factors.

PubMed

Thiel, Gerald; Rössler, Oliver G

2017-03-01

Resveratrol (trans-3,4',5-trihydroxystilbene), a polyphenolic phytoalexin of grapes and other fruits and plants, is a common constituent of our diet and of dietary supplements. Many health-promoting benefits have been connected with resveratrol in the treatment of cardiovascular diseases, cancer, diabetes, inflammation, neurodegeneration, and diseases connected with aging. To explain the pleiotropic effects of resveratrol, the molecular targets of this compound have to be identified on the cellular level. Resveratrol induces intracellular signal transduction pathways which ultimately lead to changes in the gene expression pattern of the cells. Here, we review the effect of resveratrol on the activation of the stimulus-responsive transcription factors CREB, AP-1, Egr-1, Elk-1, and Nrf2. Following activation, these transcription factors induce transcription of delayed response genes. The gene products of these delayed response genes are ultimately responsible for the changes in the biochemistry and physiology of resveratrol-treated cells. The activation of stimulus-responsive transcription factors may explain many of the intracellular activities of resveratrol. However, results obtained in vitro may not easily be transferred to in vivo systems. Copyright © 2016 Elsevier Ltd. All rights reserved.

Origin and Evolution of the Sponge Aggregation Factor Gene Family

PubMed Central

Grice, Laura F.; Gauthier, Marie E.A.; Roper, Kathrein E.; Fernàndez-Busquets, Xavier; Degnan, Sandie M.

2017-01-01

Although discriminating self from nonself is a cardinal animal trait, metazoan allorecognition genes do not appear to be homologous. Here, we characterize the Aggregation Factor (AF) gene family, which encodes putative allorecognition factors in the demosponge Amphimedon queenslandica, and trace its evolution across 24 sponge (Porifera) species. The AF locus in Amphimedon is comprised of a cluster of five similar genes that encode Calx-beta and Von Willebrand domains and a newly defined Wreath domain, and are highly polymorphic. Further AF variance appears to be generated through individualistic patterns of RNA editing. The AF gene family varies between poriferans, with protein sequences and domains diagnostic of the AF family being present in Amphimedon and other demosponges, but absent from other sponge classes. Within the demosponges, AFs vary widely with no two species having the same AF repertoire or domain organization. The evolution of AFs suggests that their diversification occurs via high allelism, and the continual and rapid gain, loss and shuffling of domains over evolutionary time. Given the marked differences in metazoan allorecognition genes, we propose the rapid evolution of AFs in sponges provides a model for understanding the extensive diversification of self–nonself recognition systems in the animal kingdom. PMID:28104746

Naturally occurring mutations in the human 5-lipoxygenase gene promoter that modify transcription factor binding and reporter gene transcription.

PubMed Central

In, K H; Asano, K; Beier, D; Grobholz, J; Finn, P W; Silverman, E K; Silverman, E S; Collins, T; Fischer, A R; Keith, T P; Serino, K; Kim, S W; De Sanctis, G T; Yandava, C; Pillari, A; Rubin, P; Kemp, J; Israel, E; Busse, W; Ledford, D; Murray, J J; Segal, A; Tinkleman, D; Drazen, J M

1997-01-01

Five lipoxygenase (5-LO) is the first committed enzyme in the metabolic pathway leading to the synthesis of the leukotrienes. We examined genomic DNA isolated from 25 normal subjects and 31 patients with asthma (6 of whom had aspirin-sensitive asthma) for mutations in the known transcription factor binding regions and the protein encoding region of the 5-LO gene. A family of mutations in the G + C-rich transcription factor binding region was identified consisting of the deletion of one, deletion of two, or addition of one zinc finger (Sp1/Egr-1) binding sites in the region 176 to 147 bp upstream from the ATG translation start site where there are normally 5 Sp1 binding motifs in tandem. Reporter gene activity directed by any of the mutant forms of the transcription factor binding region was significantly (P < 0.05) less effective than the activity driven by the wild type transcription factor binding region. Electrophoretic mobility shift assays (EMSAs) demonstrated the capacity of wild type and mutant transcription factor binding regions to bind nuclear extracts from human umbilical vein endothelial cells (HUVECs). These data are consistent with a family of mutations in the 5-LO gene that can modify reporter gene transcription possibly through differences in Sp1 and Egr-1 transactivation. PMID:9062372

Opposite cytokine synthesis by fibroblasts in contact co-culture with osteosarcoma cells compared with transwell co-cultures.

PubMed

David, Manu S; Kelly, Elizabeth; Zoellner, Hans

2013-04-01

We recently reported exchange of membrane and cytoplasm during contact co-culture between human Gingival Fibroblasts (h-GF) and SAOS-2 osteosarcoma cells, a process we termed 'cellular sipping' to reflect the manner in which cells become morphologically diverse through uptake of material from the opposing cell type, independent of genetic change. Cellular sipping is increased by Tumor Necrosis Factor-α (TNF-α), and we here show for the first time altered cytokine synthesis in contact co-culture supporting cellular sipping compared with co-culture where h-GF and SAOS-2 were separated in transwells. SAOS-2 had often undetectably low cytokine levels, while Interleukin-6 (IL-6), Granulocyte Colony Stimulating Factor (G-CSF) and Granulocyte-Macrophage Colony Stimulating Factor (GM-CSF) were secreted primarily by TNF-α stimulated h-GF and basic Fibroblast Growth Factor (FGF) was prominent in h-GF lysates (p < 0.001). Contact co-cultures permitting cellular sipping had lower IL-6, G-CSF and GM-CSF levels, as well as higher lysate FGF levels compared with TNF-α treated h-GF alone (p < 0.05). The opposite was the case for co-cultures in transwells, with increased IL-6, G-CSF and GM-CSF levels (p < 0.03) and no clear difference in FGF. We thus demonstrate significant phenotypic change in cultures where cellular sipping occurs, potentially contributing to tumor inflammatory responses. Crown Copyright © 2013. Published by Elsevier Ltd. All rights reserved.

[Effect of human oviductal embryotrophic factors on gene expression of mouse preimplantation embryos].

PubMed

Yao, Yuan-Qing; Lee, Kai-Fai; Xu, Jia-Seng; Ho, Pak-Chung; Yeung, Shu-Biu

2007-09-01

To investigate the effect of embryotrophic factors (ETF) from human oviductal cells on gene expression of mouse early developmental embryos and discuss the role of fallopian tube in early development of embryos. ETF was isolated from conditioned medium of human oviductal cell line by sequential liquid chromatographic systems. Mouse embryos were treated by ETF in vitro. Using differential display RT-PCR, the gene expression of embryos treated by ETF was compared with embryos without ETF treatment. The differentially expressed genes were separated, re-amplified, cloned and sequenced. Gene expression profiles of embryos with ETF treatment was different from embryos without this treatment. Eight differentially expressed genes were cloned and sequenced. These genes functioned in RNA degradation, synthesis, splicing, protein trafficking, cellular differentiation and embryo development. Embryotrophic factors from human oviductal cells affect gene expression of early developmental embryos. The human oviductal cells play wide roles in early developmental stages of embryos.

Epidermal growth factor gene is a newly identified candidate gene for gout

PubMed Central

Han, Lin; Cao, Chunwei; Jia, Zhaotong; Liu, Shiguo; Liu, Zhen; Xin, Ruosai; Wang, Can; Li, Xinde; Ren, Wei; Wang, Xuefeng; Li, Changgui

2016-01-01

Chromosome 4q25 has been identified as a genomic region associated with gout. However, the associations of gout with the genes in this region have not yet been confirmed. Here, we performed two-stage analysis to determine whether variations in candidate genes in the 4q25 region are associated with gout in a male Chinese Han population. We first evaluated 96 tag single nucleotide polymorphisms (SNPs) in eight inflammatory/immune pathway- or glucose/lipid metabolism-related genes in the 4q25 region in 480 male gout patients and 480 controls. The SNP rs12504538, located in the elongation of very-long-chain-fatty-acid-like family member 6 gene (Elovl6), was found to be associated with gout susceptibility (Padjusted = 0.00595). In the second stage of analysis, we performed fine mapping analysis of 93 tag SNPs in Elovl6 and in the epidermal growth factor gene (EGF) and its flanking regions in 1017 male patients gout and 1897 healthy male controls. We observed a significant association between the T allele of EGF rs2298999 and gout (odds ratio = 0.77, 95% confidence interval = 0.67–0.88, Padjusted = 6.42 × 10−3). These results provide the first evidence for an association between the EGF rs2298999 C/T polymorphism and gout. Our findings should be validated in additional populations. PMID:27506295

Nerve Growth Factor Gene Therapy Activates Neuronal Responses in Alzheimer’s Disease

PubMed Central

Tuszynski, Mark H.; Yang, Jennifer H.; Barba, David; U, H S.; Bakay, Roy; Pay, Mary M.; Masliah, Eliezer; Conner, James M.; Kobalka, Peter; Roy, Subhojit; Nagahara, Alan H.

2016-01-01

IMPORTANCE Alzheimer’s disease (AD) is the most common neurodegenerative disorder, and lacks effective disease modifying therapies. In 2001 we initiated a clinical trial of Nerve Growth Factor (NGF) gene therapy in AD, the first effort at gene delivery in an adult neurodegenerative disorder. This program aimed to determine whether a nervous system growth factor prevents or reduces cholinergic neuronal degeneration in AD patients. We present post-mortem findings in 10 subjects with survival times ranging from 1 to 10 years post-treatment. OBJECTIVE To determine whether degenerating neurons in AD retain an ability to respond to a nervous system growth factor delivered after disease onset. DESIGN, SETTING, AND PARTICIPANTS 10 patients with early AD underwent NGF gene therapy using either ex vivo or in vivo gene transfer. The brains of all eight patients in the first Phase 1 ex vivo trial and two patients in a subsequent Phase 1 in vivo trial were examined. MAIN OUTCOME MEASURES Brains were immunolabeled to evaluate in vivo gene expression, cholinergic neuronal responses to NGF, and activation of NGF-related cell signaling. In two cases, NGF protein levels were measured by ELISA. RESULTS Degenerating neurons in the AD brain respond to NGF. All patients exhibited a trophic response to NGF, in the form of axonal sprouting toward the NGF source. Comparing treated and non-treated sides of the brain in three patients that underwent unilateral gene transfer, cholinergic neuronal hypertrophy occurred on the NGF-treated side (P>0.05). Activation of cellular signaling and functional markers were present in two patients that underwent AAV2-mediated NGF gene transfer. Neurons exhibiting tau pathology as well as neurons free of tau expressed NGF, indicating that degenerating cells can be infected with therapeutic genes with resulting activation of cell signaling. No adverse pathological effects related to NGF were observed. CONCLUSIONS AND RELEVANCE These findings indicate that

AAV5-Factor VIII Gene Transfer in Severe Hemophilia A.

PubMed

Rangarajan, Savita; Walsh, Liron; Lester, Will; Perry, David; Madan, Bella; Laffan, Michael; Yu, Hua; Vettermann, Christian; Pierce, Glenn F; Wong, Wing Y; Pasi, K John

2017-12-28

Patients with hemophilia A rely on exogenous factor VIII to prevent bleeding in joints, soft tissue, and the central nervous system. Although successful gene transfer has been reported in patients with hemophilia B, the large size of the factor VIII coding region has precluded improved outcomes with gene therapy in patients with hemophilia A. We infused a single intravenous dose of a codon-optimized adeno-associated virus serotype 5 (AAV5) vector encoding a B-domain-deleted human factor VIII (AAV5-hFVIII-SQ) in nine men with severe hemophilia A. Participants were enrolled sequentially into one of three dose cohorts (low dose [one participant], intermediate dose [one participant], and high dose [seven participants]) and were followed through 52 weeks. Factor VIII activity levels remained at 3 IU or less per deciliter in the recipients of the low or intermediate dose. In the high-dose cohort, the factor VIII activity level was more than 5 IU per deciliter between weeks 2 and 9 after gene transfer in all seven participants, and the level in six participants increased to a normal value (>50 IU per deciliter) that was maintained at 1 year after receipt of the dose. In the high-dose cohort, the median annualized bleeding rate among participants who had previously received prophylactic therapy decreased from 16 events before the study to 1 event after gene transfer, and factor VIII use for participant-reported bleeding ceased in all the participants in this cohort by week 22. The primary adverse event was an elevation in the serum alanine aminotransferase level to 1.5 times the upper limit of the normal range or less. Progression of preexisting chronic arthropathy in one participant was the only serious adverse event. No neutralizing antibodies to factor VIII were detected. The infusion of AAV5-hFVIII-SQ was associated with the sustained normalization of factor VIII activity level over a period of 1 year in six of seven participants who received a high dose, with

Enhanced actions of insulin-like growth factor-I and interferon-alpha co-administration in experimental cirrhosis.

PubMed

Tutau, Federico; Rodríguez-Ortigosa, Carlos; Puche, Juan Enrique; Juanarena, Nerea; Monreal, Iñigo; García Fernández, María; Clavijo, Encarna; Castilla, Alberto; Castilla-Cortázar, Inma

2009-01-01

Cirrhosis is a diffuse process of hepatic fibrosis and regenerative nodule formation. The liver is the major source of circulating insulin-like growth factor-I (IGF-I) whose plasma levels are diminished in cirrhosis. IGF-I supplementation has been shown to induce beneficial effects in cirrhosis, including antifibrogenic and hepatoprotective effects. On other hand, interferon-alpha (IFN-alpha) therapy seems to suppress the progression of hepatic fibrosis. The aim of this study was to investigate the effect of the co-administration of IGF-I+IFN-alpha to Wistar rats with CCl(4)-induced cirrhosis, exploring liver function tests, hepatic lipid peroxidation and histopathology. The mechanisms underlying the effects of these agents were studied by reverse transcription-polymerase chain reaction, determining the expression of some factors [hepatocyte growth factor (HGF), transforming growth factor-beta (TGF-beta), alpha-smooth muscle actin, collagen, tissular inhibitor of metalloproteinases-1 and pregnane X receptor (PXR)] involved in fibrogenesis, fibrolysis and/or hepatoprotection. Both IGF-I and IFN-alpha exerted significant effects on fibrogenesis. IGF-I significantly increased serum albumin and HGF whereas IFN-alpha-therapy did not. The inhibition of TGF-beta expression was only observed by the effect of IFN-alpha-therapy. In addition, only the co-administration of IGF-I and IFN-alpha was able to increase the PXR. The combined therapy with both factors improved liver function tests, hepatic lipid peroxidation and reduced fibrosis, inducing a relevant histological improvement, reducing fibrosis and recovering hepatic architecture. The co-administration IGF-I+IFN enhanced all the beneficial effects observed with each factor separately, showing an additive action on histopathology and PXR expression, which is involved in the inhibition of fibrogenesis.

Virulence factors genes of Staphylococcus spp. isolated from caprine subclinical mastitis.

PubMed

Salaberry, Sandra Renata Sampaio; Saidenberg, André Becker Simões; Zuniga, Eveline; Melville, Priscilla Anne; Santos, Franklin Gerônimo Bispo; Guimarães, Ednaldo Carvalho; Gregori, Fábio; Benites, Nilson Roberti

2015-08-01

The aim of this study was to investigate genes involved in adhesion expression, biofilm formation, and enterotoxin production in isolates of Staphylococcus spp. from goats with subclinical mastitis and associate these results with the staphylococcal species. One hundred and twenty-four isolates were identified and polymerase chain reaction (PCR) was performed to detect the following genes: cna, ebpS, eno, fib, fnbA, fnbB, bap, sea, seb, sec, sed and see. The most commonly Staphylococcus species included S. epidermidis, S. lugdunensis, S. chromogenes, S. capitis ss capitis and S. intermedius. With the exception of fnbB, the genes were detected in different frequencies of occurrence in 86.3% of the Staphylococcus spp. isolates. Eno (73.2%) and bap (94.8%) were more frequently detected in coagulase-negative staphylococci (CNS); ebpS (76%), fib (90.9%) and fnbA (87%) were the most frequent genes in coagulase-positive staphylococci (CPS). Regarding enterotoxins, genes sed (28.2%) and see (24.2%) had a higher frequency of occurrence; sec gene was more frequently detected in CPS (58.8%). There was no association between the presence of the genes and the Staphylococcus species. Different virulence factors genes can be detected in caprine subclinical mastitis caused by CNS and CPS. The knowledge of the occurrence of these virulence factors is important for the development of effective control and prevention measures of subclinical mastitis caused by CNS and CPS in goats. Copyright © 2015 Elsevier Ltd. All rights reserved.

Appl1 Is Dispensable for Mouse Development, and Loss of Appl1 Has Growth Factor-selective Effects on Akt Signaling in Murine Embryonic Fibroblasts*

PubMed Central

Tan, Yinfei; You, Huihong; Wu, Chao; Altomare, Deborah A.; Testa, Joseph R.

2010-01-01

The adaptor protein APPL1 (adaptor protein containing pleckstrin homology (PH), phosphotyrosine binding (PTB), and leucine zipper motifs) was first identified as a binding protein of AKT2 by yeast two-hybrid screening. APPL1 was subsequently found to bind to several membrane-bound receptors and was implicated in their signal transduction through AKT and/or MAPK pathways. To determine the unambiguous role of Appl1 in vivo, we generated Appl1 knock-out mice. Here we report that Appl1 knock-out mice are viable and fertile. Appl1-null mice were born at expected Mendelian ratios, without obvious phenotypic abnormalities. Moreover, Akt activity in various fetal tissues was unchanged compared with that observed in wild-type littermates. Studies of isolated Appl1−/− murine embryonic fibroblasts (MEFs) showed that Akt activation by epidermal growth factor, insulin, or fetal bovine serum was similar to that observed in wild-type MEFs, although Akt activation by HGF was diminished in Appl1−/− MEFs. To rule out a possible redundant role played by the related Appl2, we used small interfering RNA to knock down Appl2 expression in Appl1−/− MEFs. Unexpectedly, cell survival was unaffected under normal culture conditions, and activation of Akt was unaltered following epidermal growth factor stimulation, although Akt activity did decrease further after HGF stimulation. Furthermore, we found that Appl proteins are required for HGF-induced cell survival and migration via activation of Akt. Our studies suggest that Appl1 is dispensable for development and only participate in Akt signaling under certain conditions. PMID:20040596

Factors affecting expression of the recF gene of Escherichia coli K-12.

PubMed

Sandler, S J; Clark, A J

1990-01-31

This report describes four factors which affect expression of the recF gene from strong upstream lambda promoters under temperature-sensitive cIAt2-encoded repressor control. The first factor was the long mRNA leader sequence consisting of the Escherichia coli dnaN gene and 95% of the dnaA gene and lambda bet, N (double amber) and 40% of the exo gene. When most of this DNA was deleted, RecF became detectable in maxicells. The second factor was the vector, pBEU28, a runaway replication plasmid. When we substituted pUC118 for pBEU28, RecF became detectable in whole cells by the Coomassie blue staining technique. The third factor was the efficiency of initiation of translation. We used site-directed mutagenesis to change the mRNA leader, ribosome-binding site and the 3 bp before and after the translational start codon. Monitoring the effect of these mutational changes by translational fusion to lacZ, we discovered that the efficiency of initiation of translation was increased 30-fold. Only an estimated two- or threefold increase in accumulated levels of RecF occurred, however. This led us to discover the fourth factor, namely sequences in the recF gene itself. These sequences reduce expression of the recF-lacZ fusion genes 100-fold. The sequences responsible for this decrease in expression occur in four regions in the N-terminal half of recF. Expression is reduced by some sequences at the transcriptional level and by others at the translational level.

Problem-Based Test: The Effect of Fibroblast Growth Factor on Gene Expression

ERIC Educational Resources Information Center

Szeberenyi, Jozsef

2011-01-01

This paper shows the results of an experiment in which the effects of fibroblast growth factor (FGF), actinomycin D (Act D; an inhibitor of transcription), and cycloheximide (CHX; an inhibitor of translation) were studied on the expression of two genes: a gene called "Fnk" and the gene coding for glyceraldehyde-3-phosphate dehydrogenase (GAPDH).…

BAG-1 enhances cell-cell adhesion, reduces proliferation and induces chaperone-independent suppression of hepatocyte growth factor-induced epidermal keratinocyte migration

DOE Office of Scientific and Technical Information (OSTI.GOV)

Hinitt, C.A.M.; Wood, J.; Lee, S.S.

2010-08-01

Cell motility is important in maintaining tissue homeostasis, facilitating epithelial wound repair and in tumour formation and progression. The aim of this study was to determine whether BAG-1 isoforms regulate epidermal cell migration in in vitro models of wound healing. In the human epidermal cell line HaCaT, endogenous BAG-1 is primarily nuclear and increases with confluence. Both transient and stable p36-Bag-1 overexpression resulted in increased cellular cohesion. Stable transfection of either of the three human BAG-1 isoforms p36-Bag-1 (BAG-1S), p46-Bag-1 (BAG-1M) and p50-Bag-1 (BAG-1L) inhibited growth and wound closure in serum-containing medium. However, in response to hepatocyte growth factor (HGF)more » in serum-free medium, BAG-1S/M reduced communal motility and colony scattering, but BAG-1L did not. In the presence of HGF, p36-Bag-1 transfectants retained proliferative response to HGF with no change in ERK1/2 activation. However, the cells retained E-cadherin localisation at cell-cell junctions and exhibited pronounced cortical actin. Point mutations in the BAG domain showed that BAG-1 inhibition of motility is independent of its function as a chaperone regulator. These findings are the first to suggest that BAG-1 plays a role in regulating cell-cell adhesion and suggest an important function in epidermal cohesion.« less

Origin and Evolution of the Sponge Aggregation Factor Gene Family.

PubMed

Grice, Laura F; Gauthier, Marie E A; Roper, Kathrein E; Fernàndez-Busquets, Xavier; Degnan, Sandie M; Degnan, Bernard M

2017-05-01

Although discriminating self from nonself is a cardinal animal trait, metazoan allorecognition genes do not appear to be homologous. Here, we characterize the Aggregation Factor (AF) gene family, which encodes putative allorecognition factors in the demosponge Amphimedon queenslandica, and trace its evolution across 24 sponge (Porifera) species. The AF locus in Amphimedon is comprised of a cluster of five similar genes that encode Calx-beta and Von Willebrand domains and a newly defined Wreath domain, and are highly polymorphic. Further AF variance appears to be generated through individualistic patterns of RNA editing. The AF gene family varies between poriferans, with protein sequences and domains diagnostic of the AF family being present in Amphimedon and other demosponges, but absent from other sponge classes. Within the demosponges, AFs vary widely with no two species having the same AF repertoire or domain organization. The evolution of AFs suggests that their diversification occurs via high allelism, and the continual and rapid gain, loss and shuffling of domains over evolutionary time. Given the marked differences in metazoan allorecognition genes, we propose the rapid evolution of AFs in sponges provides a model for understanding the extensive diversification of self-nonself recognition systems in the animal kingdom. © The Author 2017. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.

WRKY transcription factor genes in wild rice Oryza nivara

PubMed Central

Xu, Hengjian; Watanabe, Kenneth A.; Zhang, Liyuan; Shen, Qingxi J.

2016-01-01

The WRKY transcription factor family is one of the largest gene families involved in plant development and stress response. Although many WRKY genes have been studied in cultivated rice (Oryza sativa), the WRKY genes in the wild rice species Oryza nivara, the direct progenitor of O. sativa, have not been studied. O. nivara shows abundant genetic diversity and elite drought and disease resistance features. Herein, a total of 97 O. nivara WRKY (OnWRKY) genes were identified. RNA-sequencing demonstrates that OnWRKY genes were generally expressed at higher levels in the roots of 30-day-old plants. Bioinformatic analyses suggest that most of OnWRKY genes could be induced by salicylic acid, abscisic acid, and drought. Abundant potential MAPK phosphorylation sites in OnWRKYs suggest that activities of most OnWRKYs can be regulated by phosphorylation. Phylogenetic analyses of OnWRKYs support a novel hypothesis that ancient group IIc OnWRKYs were the original ancestors of only some group IIc and group III WRKYs. The analyses also offer strong support that group IIc OnWRKYs containing the HVE sequence in their zinc finger motifs were derived from group Ia WRKYs. This study provides a solid foundation for the study of the evolution and functions of WRKY genes in O. nivara. PMID:27345721

GC[Formula: see text]NMF: A Novel Matrix Factorization Framework for Gene-Phenotype Association Prediction.

PubMed

Zhang, Yaogong; Liu, Jiahui; Liu, Xiaohu; Hong, Yuxiang; Fan, Xin; Huang, Yalou; Wang, Yuan; Xie, Maoqiang

2018-04-24

Gene-phenotype association prediction can be applied to reveal the inherited basis of human diseases and facilitate drug development. Gene-phenotype associations are related to complex biological processes and influenced by various factors, such as relationship between phenotypes and that among genes. While due to sparseness of curated gene-phenotype associations and lack of integrated analysis of the joint effect of multiple factors, existing applications are limited to prediction accuracy and potential gene-phenotype association detection. In this paper, we propose a novel method by exploiting weighted graph constraint learned from hierarchical structures of phenotype data and group prior information among genes by inheriting advantages of Non-negative Matrix Factorization (NMF), called Weighted Graph Constraint and Group Centric Non-negative Matrix Factorization (GC[Formula: see text]NMF). Specifically, first we introduce the depth of parent-child relationships between two adjacent phenotypes in hierarchical phenotypic data as weighted graph constraint for a better phenotype understanding. Second, we utilize intra-group correlation among genes in a gene group as group constraint for gene understanding. Such information provides us with the intuition that genes in a group probably result in similar phenotypes. The model not only allows us to achieve a high-grade prediction performance, but also helps us to learn interpretable representation of genes and phenotypes simultaneously to facilitate future biological analysis. Experimental results on biological gene-phenotype association datasets of mouse and human demonstrate that GC[Formula: see text]NMF can obtain superior prediction accuracy and good understandability for biological explanation over other state-of-the-arts methods.

Transcription factor trapping by RNA in gene regulatory elements.

PubMed

Sigova, Alla A; Abraham, Brian J; Ji, Xiong; Molinie, Benoit; Hannett, Nancy M; Guo, Yang Eric; Jangi, Mohini; Giallourakis, Cosmas C; Sharp, Phillip A; Young, Richard A

2015-11-20

Transcription factors (TFs) bind specific sequences in promoter-proximal and -distal DNA elements to regulate gene transcription. RNA is transcribed from both of these DNA elements, and some DNA binding TFs bind RNA. Hence, RNA transcribed from regulatory elements may contribute to stable TF occupancy at these sites. We show that the ubiquitously expressed TF Yin-Yang 1 (YY1) binds to both gene regulatory elements and their associated RNA species across the entire genome. Reduced transcription of regulatory elements diminishes YY1 occupancy, whereas artificial tethering of RNA enhances YY1 occupancy at these elements. We propose that RNA makes a modest but important contribution to the maintenance of certain TFs at gene regulatory elements and suggest that transcription of regulatory elements produces a positive-feedback loop that contributes to the stability of gene expression programs. Copyright © 2015, American Association for the Advancement of Science.

Arabidopsis Ensemble Reverse-Engineered Gene Regulatory Network Discloses Interconnected Transcription Factors in Oxidative Stress[W

PubMed Central

Vermeirssen, Vanessa; De Clercq, Inge; Van Parys, Thomas; Van Breusegem, Frank; Van de Peer, Yves

2014-01-01

The abiotic stress response in plants is complex and tightly controlled by gene regulation. We present an abiotic stress gene regulatory network of 200,014 interactions for 11,938 target genes by integrating four complementary reverse-engineering solutions through average rank aggregation on an Arabidopsis thaliana microarray expression compendium. This ensemble performed the most robustly in benchmarking and greatly expands upon the availability of interactions currently reported. Besides recovering 1182 known regulatory interactions, cis-regulatory motifs and coherent functionalities of target genes corresponded with the predicted transcription factors. We provide a valuable resource of 572 abiotic stress modules of coregulated genes with functional and regulatory information, from which we deduced functional relationships for 1966 uncharacterized genes and many regulators. Using gain- and loss-of-function mutants of seven transcription factors grown under control and salt stress conditions, we experimentally validated 141 out of 271 predictions (52% precision) for 102 selected genes and mapped 148 additional transcription factor-gene regulatory interactions (49% recall). We identified an intricate core oxidative stress regulatory network where NAC13, NAC053, ERF6, WRKY6, and NAC032 transcription factors interconnect and function in detoxification. Our work shows that ensemble reverse-engineering can generate robust biological hypotheses of gene regulation in a multicellular eukaryote that can be tested by medium-throughput experimental validation. PMID:25549671

Upregulation of the coagulation factor VII gene during glucose deprivation is mediated by activating transcription factor 4.

PubMed

Cronin, Katherine R; Mangan, Thomas P; Carew, Josephine A

2012-01-01

Constitutive production of blood coagulation proteins by hepatocytes is necessary for hemostasis. Stressful conditions trigger adaptive cellular responses and delay processing of most proteins, potentially affecting plasma levels of proteins secreted exclusively by hepatocytes. We examined the effect of glucose deprivation on expression of coagulation proteins by the human hepatoma cell line, HepG2. Expression of coagulation factor VII, which is required for initiation of blood coagulation, was elevated by glucose deprivation, while expression of other coagulation proteins decreased. Realtime PCR and ELISA demonstrated that the relative percentage expression +/- SD of steady-state F7 mRNA and secreted factor VII antigen were significantly increased (from 100+/-15% to 188+/-27% and 100+/-8.8% to 176.3+/-17.3% respectively, p<0.001) at 24 hr of treatment. The integrated stress response was induced, as indicated by upregulation of transcription factor ATF4 and of additional stress-responsive genes. Small interfering RNAs directed against ATF4 potently reduced basal F7 expression, and prevented F7 upregulation by glucose deprivation. The response of the endogenous F7 gene was replicated in reporter gene assays, which further indicated that ATF4 effects were mediated via interaction with an amino acid response element in the F7 promoter. Our data indicated that glucose deprivation enhanced F7 expression in a mechanism reliant on prior ATF4 upregulation primarily due to increased transcription from the ATF4 gene. Of five coagulation protein genes examined, only F7 was upregulated, suggesting that its functions may be important in a systemic response to glucose deprivation stress.

Diversification of the insulin-like growth factor 1 gene in mammals.

PubMed

Rotwein, Peter

2017-01-01

Insulin-like growth factor 1 (IGF1), a small, secreted peptide growth factor, is involved in a variety of physiological and patho-physiological processes, including somatic growth, tissue repair, and metabolism of carbohydrates, proteins, and lipids. IGF1 gene expression appears to be controlled by several different signaling cascades in the few species in which it has been evaluated, with growth hormone playing a major role by activating a pathway involving the Stat5b transcription factor. Here, genes encoding IGF1 have been evaluated in 25 different mammalian species representing 15 different orders and ranging over ~180 million years of evolutionary diversification. Parts of the IGF1 gene have been fairly well conserved. Like rat Igf1 and human IGF1, 21 of 23 other genes are composed of 6 exons and 5 introns, and all 23 also contain recognizable tandem promoters, each with a unique leader exon. Exon and intron lengths are similar in most species, and DNA sequence conservation is moderately high in orthologous exons and proximal promoter regions. In contrast, putative growth hormone-activated Stat5b-binding enhancers found in analogous locations in rodent Igf1 and in human IGF1 loci, have undergone substantial variation in other mammals, and a processed retro-transposed IGF1 pseudogene is found in the sloth locus, but not in other mammalian genomes. Taken together, the fairly high level of organizational and nucleotide sequence similarity in the IGF1 gene among these 25 species supports the contention that some common regulatory pathways had existed prior to the beginning of mammalian speciation.

The regulatory role of heparin on c-Met signaling in hepatocellular carcinoma cells.

PubMed

İşcan, Evin; Güneş, Aysim; Korhan, Peyda; Yılmaz, Yeliz; Erdal, Esra; Atabey, Neşe

2017-06-01

The role of heparin as an anticoagulant is well defined; however, its role in tumorigenesis and tumor progression is not clear yet. Some studies have shown that anticoagulant treatment in cancer patients improve overall survival, however, recent clinical trials have not shown a survival benefit in cancer patients receiving heparin treatment. In our previous studies we have shown the inhibitory effects of heparin on Hepatocyte Growth Factor (HGF)-induced invasion and migration in hepatocellular carcinoma (HCC) cells. In this study, we showed the differential effects of heparin on the behaviors of HCC cells based on the presence or absence of HGF. In the absence of HGF, heparin activated HGF/c-Met signaling and promoted motility and invasion in HCC cells. Heparin treatment led to c-Met receptor dimerization and activated c-Met signaling in an HGF independent manner. Heparin-induced c-Met activation increased migration and invasion through ERK1/2, early growth response factor 1 (EGR1) and Matrix Metalloproteinases (MMP) axis. Interestingly, heparin modestly decreased the proliferation of HCC cells by inhibiting activatory phosphorylation of Akt. The inhibition of c-Met signaling reversed heparin-induced increase in motility and invasion and, proliferation inhibition. Our study provides a new perspective into the role of heparin on c-Met signaling in HCC.

ULTRAPETALA trxG genes interact with KANADI transcription factor genes to regulate Aradopsis Gynoecium patterning

USDA-ARS?s Scientific Manuscript database

Organ formation relies upon precise patterns of gene expression that are under tight spatial and temporal regulation. Transcription patterns are specified by several cellular processes during development, including chromatin remodeling, but little is known about how chromatin remodeling factors cont...

WRKY transcription factor genes in wild rice Oryza nivara.

PubMed

Xu, Hengjian; Watanabe, Kenneth A; Zhang, Liyuan; Shen, Qingxi J

2016-08-01

The WRKY transcription factor family is one of the largest gene families involved in plant development and stress response. Although many WRKY genes have been studied in cultivated rice (Oryza sativa), the WRKY genes in the wild rice species Oryza nivara, the direct progenitor of O. sativa, have not been studied. O. nivara shows abundant genetic diversity and elite drought and disease resistance features. Herein, a total of 97 O. nivara WRKY (OnWRKY) genes were identified. RNA-sequencing demonstrates that OnWRKY genes were generally expressed at higher levels in the roots of 30-day-old plants. Bioinformatic analyses suggest that most of OnWRKY genes could be induced by salicylic acid, abscisic acid, and drought. Abundant potential MAPK phosphorylation sites in OnWRKYs suggest that activities of most OnWRKYs can be regulated by phosphorylation. Phylogenetic analyses of OnWRKYs support a novel hypothesis that ancient group IIc OnWRKYs were the original ancestors of only some group IIc and group III WRKYs. The analyses also offer strong support that group IIc OnWRKYs containing the HVE sequence in their zinc finger motifs were derived from group Ia WRKYs. This study provides a solid foundation for the study of the evolution and functions of WRKY genes in O. nivara. © The Author 2016. Published by Oxford University Press on behalf of Kazusa DNA Research Institute.

A Histologically Distinctive Interstitial Pneumonia Induced by Overexpression of the Interleukin 6, Transforming Growth Factor β1, or Platelet-Derived Growth Factor B Gene

NASA Astrophysics Data System (ADS)

Yoshida, Mitsuhiro; Sakuma, Junko; Hayashi, Seiji; Abe, Kin'ya; Saito, Izumu; Harada, Shizuko; Sakatani, Mitsunoir; Yamamoto, Satoru; Matsumoto, Norinao; Kaneda, Yasufumi; Kishmoto, Tadamitsu

1995-10-01

Interstitial pneumonia is characterized by alveolitis with resulting fibrosis of the interstitium. To determine the relevance of humoral factors in the pathogenesis of interstitial pneumonia, we introduced expression vectors into Wistar rats via the trachea to locally overexpress humoral factors in the lungs. Human interleukin (IL) 6 and IL-6 receptor genes induced lymphocytic alveolitis without marked fibroblast proliferation. In contrast, overexpression of human transforming growth factor β1 or human platelet-derived growth factor B gene induced only mild or apparent cellular infiltration in the alveoli, respectively. However, both factors induced significant proliferation of fibroblasts and deposition of collagen fibrils. These histopathologic changes induced by the transforming growth factor β1 and platelet-derived growth factor B gene are partly akin to those changes seen in lung tissues from patients with pulmonary fibrosis and markedly contrast with the changes induced by overexpression of the IL-6 and IL-6 receptor genes that mimics lymphocytic interstitial pneumonia.

Transient Gene and miRNA Expression Profile Changes of Confluent Human Fibroblast Cells in Space

NASA Technical Reports Server (NTRS)

Zhang, Ye; Lu, Tao; Wong, Michael; Feiveson, Alan; Stodieck, Louis; Karouia, Fathi; Wang, Xiaoyu; Wu, Honglu

2015-01-01

Microgravity or an altered gravity environment from the static 1 gravitational constant has been shown to influence global gene expression patterns and protein levels in cultured cells. However, most of the reported studies conducted in space or using simulated microgravity on the ground have focused on the growth or differentiation of the cells. Whether non-dividing cultured cells will sense the presence of microgravity in space has not been specifically addressed. In an experiment conducted on the International Space Station, confluent human fibroblast cells were fixed after being cultured in space for 3 and 14 days for investigations of gene and miRNA (microRNA) expression profile changes in these cells. A fibroblast is a type of cell that synthesizes the extracellular matrix and collagen, the structural framework for tissues, and plays a critical role in wound healing and other functions. Results of the experiment showed that on Day 3, both the flown and ground cells were still proliferating slowly even though they were confluent, as measured by the expression of the protein Ki-67 positive cells, and the cells in space grew slightly faster. Gene and miRNA expression data indicated activation of NF(sub kappa)B (nuclear factor kappa-light-chain-enhancer of activated B cells) and other growth related pathways involving HGF and VEGF in the flown cells. On Day 14 when the cells were mostly non-dividing, the gene and miRNA expression profiles between the flight and ground samples were indistinguishable. Comparison of gene and miRNA expressions in the Day 3 samples in respect to Day 14 revealed that most of the changes observed on Day 3 were related to cell growth for both the flown and ground cells. Analysis of cytoskeleton changes by immunohistochemistry staining of the cells with antibodies for alpha-tubulin showed no difference between the flight and ground samples. Results of our study suggest that in true non-dividing human fibroblast cells, microgravity in

Hepatocyte Growth Factor Modulates MET Receptor Tyrosine Kinase and β-Catenin Functional Interactions to Enhance Synapse Formation

PubMed Central

Xie, Zhihui; Eagleson, Kathie L.

2016-01-01

MET, a pleiotropic receptor tyrosine kinase implicated in autism risk, influences multiple neurodevelopmental processes. There is a knowledge gap, however, in the molecular mechanism through which MET mediates developmental events related to disorder risk. In the neocortex, MET is expressed transiently during periods of peak dendritic outgrowth and synaptogenesis, with expression enriched at developing synapses, consistent with demonstrated roles in dendritic morphogenesis, modulation of spine volume, and excitatory synapse development. In a recent coimmunoprecipitation/mass spectrometry screen, β-catenin was identified as part of the MET interactome in developing neocortical synaptosomes. Here, we investigated the influence of the MET/β-catenin complex in mouse neocortical synaptogenesis. Western blot analysis confirms that MET and β-catenin coimmunoprecipitate, but N-cadherin is not associated with the MET complex. Following stimulation with hepatocyte growth factor (HGF), β-catenin is phosphorylated at tyrosine142 (Y142) and dissociates from MET, accompanied by an increase in β-catenin/N-cadherin and MET/synapsin 1 protein complexes. In neocortical neurons in vitro, proximity ligation assays confirmed the close proximity of these proteins. Moreover, in neurons transfected with synaptophysin-GFP, HGF stimulation increases the density of synaptophysin/bassoon (a presynaptic marker) and synaptophysin/PSD-95 (a postsynaptic marker) clusters. Mutation of β-catenin at Y142 disrupts the dissociation of the MET/β-catenin complex and prevents the increase in clusters in response to HGF. The data demonstrate a new mechanism for the modulation of synapse formation, whereby MET activation induces an alignment of presynaptic and postsynaptic elements that are necessary for assembly and formation of functional synapses by subsets of neocortical neurons that express MET/β-catenin complex. PMID:27595133

Four Genes of Medicago truncatula Controlling Components of a Nod Factor Transduction Pathway

PubMed Central

Catoira, Romy; Galera, Christine; de Billy, Francoise; Penmetsa, R. Varma; Journet, Etienne-Pascal; Maillet, Fabienne; Rosenberg, Charles; Cook, Douglas; Gough, Clare; Dénarié, Jean

2000-01-01

Rhizobium nodulation (Nod) factors are lipo-chitooligosaccharides that act as symbiotic signals, eliciting several key developmental responses in the roots of legume hosts. Using nodulation-defective mutants of Medicago truncatula, we have started to dissect the genetic control of Nod factor transduction. Mutants in four genes (DMI1, DMI2, DMI3, and NSP) were pleiotropically affected in Nod factor responses, indicating that these genes are required for a Nod factor–activated signal transduction pathway that leads to symbiotic responses such as root hair deformations, expressions of nodulin genes, and cortical cell divisions. Mutant analysis also provides evidence that Nod factors have a dual effect on the growth of root hair: inhibition of endogenous (plant) tip growth, and elicitation of a novel tip growth dependent on (bacterial) Nod factors. dmi1, dmi2, and dmi3 mutants are also unable to establish a symbiotic association with endomycorrhizal fungi, indicating that there are at least three common steps to nodulation and endomycorrhization in M. truncatula and providing further evidence for a common signaling pathway between nodulation and mycorrhization. PMID:11006338

Upregulation of the Coagulation Factor VII Gene during Glucose Deprivation Is Mediated by Activating Transcription Factor 4

PubMed Central

Cronin, Katherine R.; Mangan, Thomas P.; Carew, Josephine A.

2012-01-01

Background Constitutive production of blood coagulation proteins by hepatocytes is necessary for hemostasis. Stressful conditions trigger adaptive cellular responses and delay processing of most proteins, potentially affecting plasma levels of proteins secreted exclusively by hepatocytes. We examined the effect of glucose deprivation on expression of coagulation proteins by the human hepatoma cell line, HepG2. Methodology/Principal Findings Expression of coagulation factor VII, which is required for initiation of blood coagulation, was elevated by glucose deprivation, while expression of other coagulation proteins decreased. Realtime PCR and ELISA demonstrated that the relative percentage expression +/− SD of steady-state F7 mRNA and secreted factor VII antigen were significantly increased (from 100+/−15% to 188+/−27% and 100+/−8.8% to 176.3+/−17.3% respectively, p<0.001) at 24 hr of treatment. The integrated stress response was induced, as indicated by upregulation of transcription factor ATF4 and of additional stress-responsive genes. Small interfering RNAs directed against ATF4 potently reduced basal F7 expression, and prevented F7 upregulation by glucose deprivation. The response of the endogenous F7 gene was replicated in reporter gene assays, which further indicated that ATF4 effects were mediated via interaction with an amino acid response element in the F7 promoter. Conclusions/Significance Our data indicated that glucose deprivation enhanced F7 expression in a mechanism reliant on prior ATF4 upregulation primarily due to increased transcription from the ATF4 gene. Of five coagulation protein genes examined, only F7 was upregulated, suggesting that its functions may be important in a systemic response to glucose deprivation stress. PMID:22848420

Splicing factor gene mutations in the myelodysplastic syndromes: impact on disease phenotype and therapeutic applications.

PubMed

Pellagatti, Andrea; Boultwood, Jacqueline

2017-01-01

Splicing factor gene mutations are the most frequent mutations found in patients with the myeloid malignancy myelodysplastic syndrome (MDS), suggesting that spliceosomal dysfunction plays a major role in disease pathogenesis. The aberrantly spliced target genes and deregulated cellular pathways associated with the commonly mutated splicing factor genes in MDS (SF3B1, SRSF2 and U2AF1) are being identified, illuminating the molecular mechanisms underlying MDS. Emerging data from mouse modeling studies indicate that the presence of splicing factor gene mutations can lead to bone marrow hematopoietic stem/myeloid progenitor cell expansion, impaired hematopoiesis and dysplastic differentiation that are hallmarks of MDS. Importantly, recent evidence suggests that spliceosome inhibitors and splicing modulators may have therapeutic value in the treatment of splicing factor mutant myeloid malignancies. Copyright © 2016 Elsevier Ltd. All rights reserved.

Evaluation of gene expression classification studies: factors associated with classification performance.

PubMed

Novianti, Putri W; Roes, Kit C B; Eijkemans, Marinus J C

2014-01-01

Classification methods used in microarray studies for gene expression are diverse in the way they deal with the underlying complexity of the data, as well as in the technique used to build the classification model. The MAQC II study on cancer classification problems has found that performance was affected by factors such as the classification algorithm, cross validation method, number of genes, and gene selection method. In this paper, we study the hypothesis that the disease under study significantly determines which method is optimal, and that additionally sample size, class imbalance, type of medical question (diagnostic, prognostic or treatment response), and microarray platform are potentially influential. A systematic literature review was used to extract the information from 48 published articles on non-cancer microarray classification studies. The impact of the various factors on the reported classification accuracy was analyzed through random-intercept logistic regression. The type of medical question and method of cross validation dominated the explained variation in accuracy among studies, followed by disease category and microarray platform. In total, 42% of the between study variation was explained by all the study specific and problem specific factors that we studied together.

Synergistic effect of electrical and chemical factors on endocytosis in micro-discharge plasma gene transfection

NASA Astrophysics Data System (ADS)

Jinno, M.; Ikeda, Y.; Motomura, H.; Isozaki, Y.; Kido, Y.; Satoh, S.

2017-06-01

We have developed a new micro-discharge plasma (MDP)-based gene transfection method, which transfers genes into cells with high efficiency and low cytotoxicity; however, the mechanism underlying the method is still unknown. Studies revealed that the N-acetylcysteine-mediated inhibition of reactive oxygen species (ROS) activity completely abolished gene transfer. In this study, we used laser-produced plasma to demonstrate that gene transfer does not occur in the absence of electrical factors. Our results show that both electrical and chemical factors are necessary for gene transfer inside cells by microplasma irradiation. This indicates that plasma-mediated gene transfection utilizes the synergy between electrical and chemical factors. The electric field threshold required for transfection was approximately 1 kV m-1 in our MDP system. This indicates that MDP irradiation supplies sufficient concentrations of ROS, and the stimulation intensity of the electric field determines the transfection efficiency in our system. Gene transfer by plasma irradiation depends mainly on endocytosis, which accounts for at least 80% of the transfer, and clathrin-mediated endocytosis is a dominant endocytosis. In plasma-mediated gene transfection, alterations in electrical and chemical factors can independently regulate plasmid DNA adhesion and triggering of endocytosis, respectively. This implies that plasma characteristics can be adjusted according to target cell requirements, and the transfection process can be optimized with minimum damage to cells and maximum efficiency. This may explain how MDP simultaneously achieves high transfection efficiency with minimal cell damage.

Transcription factors, sucrose, and sucrose metabolic genes interact to regulate potato phenylpropanoid metabolism

PubMed Central

Payyavula, Raja S.; Navarre, Duroy A.

2013-01-01

Much remains unknown about how transcription factors and sugars regulate phenylpropanoid metabolism in tuber crops like potato (Solanum tuberosum). Based on phylogeny and protein similarity to known regulators of phenylpropanoid metabolism, 15 transcription factors were selected and their expression was compared in white, yellow, red, and purple genotypes with contrasting phenolic and anthocyanin profiles. Red and purple genotypes had increased phenylalanine ammonia lyase (PAL) enzyme activity, markedly higher levels of phenylpropanoids, and elevated expression of most phenylpropanoid structural genes, including a novel anthocyanin O-methyltransferase. The transcription factors Anthocyanin1 (StAN1), basic Helix Loop Helix1 (StbHLH1), and StWD40 were more strongly expressed in red and purple potatoes. Expression of 12 other transcription factors was not associated with phenylpropanoid content, except for StMYB12B, which showed a negative relationship. Increased expression of AN1, bHLH1, and WD40 was also associated with environmentally mediated increases in tuber phenylpropanoids. Treatment of potato plantlets with sucrose induced hydroxycinnamic acids, flavonols, anthocyanins, structural genes, AN1, bHLH1, WD40, and genes encoding the sucrose-hydrolysing enzymes SUSY1, SUSY4, and INV2. Transient expression of StAN1 in tobacco leaves induced bHLH1, structural genes, SUSY1, SUSY4, and INV1, and increased phenylpropanoid amounts. StAN1 infiltration into tobacco leaves decreased sucrose and glucose concentrations. In silico promoter analysis revealed the presence of MYB and bHLH regulatory elements on sucrolytic gene promoters and sucrose-responsive elements on the AN1 promoter. These findings reveal an interesting dynamic between AN1, sucrose, and sucrose metabolic genes in modulating potato phenylpropanoids. PMID:24098049

Coordinating Regulation of Gene Expression in Cardiovascular Disease: Interactions between Chromatin Modifiers and Transcription Factors

PubMed Central

Bauer, Ashley J.; Martin, Kathleen A.

2017-01-01

Cardiovascular disease is a leading cause of death with increasing economic burden. The pathogenesis of cardiovascular diseases is complex, but can arise from genetic and/or environmental risk factors. This can lead to dysregulated gene expression in numerous cell types including cardiomyocytes, endothelial cells, vascular smooth muscle cells, and inflammatory cells. While initial studies addressed transcriptional control of gene expression, epigenetics has been increasingly appreciated to also play an important role in this process through alterations in chromatin structure and gene accessibility. Chromatin-modifying proteins including enzymes that modulate DNA methylation, histone methylation, and histone acetylation can influence gene expression in numerous ways. These chromatin modifiers and their marks can promote or prevent transcription factor recruitment to regulatory regions of genes through modifications to DNA, histones, or the transcription factors themselves. This review will focus on the emerging question of how epigenetic modifiers and transcription factors interact to coordinately regulate gene expression in cardiovascular disease. While most studies have addressed the roles of either epigenetic or transcriptional control, our understanding of the integration of these processes is only just beginning. Interrogating these interactions is challenging, and improved technical approaches will be needed to fully dissect the temporal and spatial relationships between transcription factors, chromatin modifiers, and gene expression in cardiovascular disease. We summarize the current state of the field and provide perspectives on limitations and future directions. Through studies of epigenetic and transcriptional interactions, we can advance our understanding of the basic mechanisms of cardiovascular disease pathogenesis to develop novel therapeutics. PMID:28428957

The significance of alternative transcripts for Caenorhabditis elegans transcription factor genes, based on expression pattern analysis

PubMed Central

2013-01-01

Background Sequence-specific DNA-binding proteins, with their paramount importance in the regulation of expression of the genetic material, are encoded by approximately 5% of the genes in an animal’s genome. But it is unclear to what extent alternative transcripts from these genes may further increase the complexity of the transcription factor complement. Results Of the 938 potential C. elegans transcription factor genes, 197 were annotated in WormBase as encoding at least two distinct isoforms. Evaluation of prior evidence identified, with different levels of confidence, 50 genes with alternative transcript starts, 23 with alternative transcript ends, 35 with alternative splicing and 34 with alternative transcripts generated by a combination of mechanisms, leaving 55 that were discounted. Expression patterns were determined for transcripts for a sample of 29 transcription factor genes, concentrating on those with alternative transcript starts for which the evidence was strongest. Seamless fosmid recombineering was used to generate reporter gene fusions with minimal modification to assay expression of specific transcripts while maintaining the broad genomic DNA context and alternative transcript production. Alternative transcription factor gene transcripts were typically expressed with identical or substantially overlapping distributions rather than in distinct domains. Conclusions Increasingly sensitive sequencing technologies will reveal rare transcripts but many of these are clearly non-productive. The majority of the transcription factor gene alternative transcripts that are productive may represent tolerable noise rather than encoding functionally distinct isoforms. PMID:23586691

The strategy of fusion genes construction determines efficient expression of introduced transcription factors.

PubMed

Adamus, Tomasz; Konieczny, Paweł; Sekuła, Małgorzata; Sułkowski, Maciej; Majka, Marcin

2014-01-01

The main goal in gene therapy and biomedical research is an efficient transcription factors (TFs) delivery system. SNAIL, a zinc finger transcription factor, is strongly involved in tumor, what makes its signaling pathways an interesting research subject. The necessity of tracking activation of intracellular pathways has prompted fluorescent proteins usage as localization markers. Advanced molecular cloning techniques allow to generate fusion proteins from fluorescent markers and transcription factors. Depending on fusion strategy, the protein expression levels and nuclear transport ability are significantly different. The P2A self-cleavage motif through its cleavage ability allows two single proteins to be simultaneously expressed. The aim of this study was to compare two strategies for introducing a pair of genes using expression vector system. We have examined GFP and SNAI1 gene fusions by comprising common nucleotide polylinker (multiple cloning site) or P2A motif in between them, resulting in one fusion or two independent protein expressions respectively. In each case transgene expression levels and translation efficiency as well as nuclear localization of expressed protein have been analyzed. Our data showed that usage of P2A motif provides more effective nuclear transport of SNAIL transcription factor than conventional genes linker. At the same time the fluorescent marker spreads evenly in subcellular space.

Factoring nonviral gene therapy into a cure for hemophilia A.

PubMed

Gabrovsky, Vanessa; Calos, Michele P

2008-10-01

Gene therapy for hemophilia A has fallen short of success despite several clinical trials conducted over the past decade. Challenges to its success include vector immunogenicity, insufficient transgene expression levels of Factor VIII, and inhibitor antibody formation. Gene therapy has been dominated by the use of viral vectors, as well as the immunogenic and oncogenic concerns that accompany these strategies. Because of the complexity of viral vectors, the development of nonviral DNA delivery methods may provide an efficient and safe alternative for the treatment of hemophilia A. New types of nonviral strategies, such as DNA integrating vectors, and the success of several nonviral animal studies, suggest that nonviral gene therapy has curative potential and justifies its clinical development.

Patterns of Positive Selection of the Myogenic Regulatory Factor Gene Family in Vertebrates

PubMed Central

Zhao, Xiao; Yu, Qi; Huang, Ling; Liu, Qing-Xin

2014-01-01

The functional divergence of transcriptional factors is critical in the evolution of transcriptional regulation. However, the mechanism of functional divergence among these factors remains unclear. Here, we performed an evolutionary analysis for positive selection in members of the myogenic regulatory factor (MRF) gene family of vertebrates. We selected 153 complete vertebrate MRF nucleotide sequences from our analyses, which revealed substantial evidence of positive selection. Here, we show that sites under positive selection were more frequently detected and identified from the genes encoding the myogenic differentiation factors (MyoG and Myf6) than the genes encoding myogenic determination factors (Myf5 and MyoD). Additionally, the functional divergence within the myogenic determination factors or differentiation factors was also under positive selection pressure. The positive selection sites were more frequently detected from MyoG and MyoD than Myf6 and Myf5, respectively. Amino acid residues under positive selection were identified mainly in their transcription activation domains and on the surface of protein three-dimensional structures. These data suggest that the functional gain and divergence of myogenic regulatory factors were driven by distinct positive selection of their transcription activation domains, whereas the function of the DNA binding domains was conserved in evolution. Our study evaluated the mechanism of functional divergence of the transcriptional regulation factors within a family, whereby the functions of their transcription activation domains diverged under positive selection during evolution. PMID:24651579

In silico mining and PCR-based approaches to transcription factor discovery in non-model plants: gene discovery of the WRKY transcription factors in conifers.

PubMed

Liu, Jun-Jun; Xiang, Yu

2011-01-01

WRKY transcription factors are key regulators of numerous biological processes in plant growth and development, as well as plant responses to abiotic and biotic stresses. Research on biological functions of plant WRKY genes has focused in the past on model plant species or species with largely characterized transcriptomes. However, a variety of non-model plants, such as forest conifers, are essential as feed, biofuel, and wood or for sustainable ecosystems. Identification of WRKY genes in these non-model plants is equally important for understanding the evolutionary and function-adaptive processes of this transcription factor family. Because of limited genomic information, the rarity of regulatory gene mRNAs in transcriptomes, and the sequence divergence to model organism genes, identification of transcription factors in non-model plants using methods similar to those generally used for model plants is difficult. This chapter describes a gene family discovery strategy for identification of WRKY transcription factors in conifers by a combination of in silico-based prediction and PCR-based experimental approaches. Compared to traditional cDNA library screening or EST sequencing at transcriptome scales, this integrated gene discovery strategy provides fast, simple, reliable, and specific methods to unveil the WRKY gene family at both genome and transcriptome levels in non-model plants.

Myeloid Leukemia Factor Acts in a Chaperone Complex to Regulate Transcription Factor Stability and Gene Expression.

PubMed

Dyer, Jamie O; Dutta, Arnob; Gogol, Madelaine; Weake, Vikki M; Dialynas, George; Wu, Xilan; Seidel, Christopher; Zhang, Ying; Florens, Laurence; Washburn, Michael P; Abmayr, Susan M; Workman, Jerry L

2017-06-30

Mutations that affect myelodysplasia/myeloid leukemia factor (MLF) proteins are associated with leukemia and several other cancers. However, with no strong homology to other proteins of known function, the role of MLF proteins in the cell has remained elusive. Here, we describe a proteomics approach that identifies MLF as a member of a nuclear chaperone complex containing a DnaJ protein, BCL2-associated anthanogene 2, and Hsc70. This complex associates with chromatin and regulates the expression of target genes. The MLF complex is bound to sites of nucleosome depletion and sites containing active chromatin marks (e.g., H3K4me3 and H3K4me1). Hence, MLF binding is enriched at promoters and enhancers. Additionally, the MLF-chaperone complex functions to regulate transcription factor stability, including the RUNX transcription factor involved in hematopoiesis. Although Hsc70 and other co-chaperones have been shown to play a role in nuclear translocation of a variety of proteins including transcription factors, our findings suggest that MLF and the associated co-chaperones play a direct role in modulating gene transcription. Copyright © 2016 Elsevier Ltd. All rights reserved.

Neurotrophins, growth-factor-regulated genes and the control of energy balance.

PubMed

Salton, Stephen R J

2003-03-01

Neurotrophic growth factors are proteins that control neuronal differentiation and survival, and consequently play important roles in the developing and adult stages of the nervous system. Study of the genes that are regulated by these growth factors has provided insight into the proteins that are critical to the maturation of the nervous system, suggesting that select neurotrophins may play a role in the control of body homeostasis by the brain and peripheral nervous system. Our understanding of the mechanisms of action of neurotrophic growth factors has increased through experimental manipulation of cultured neurons and neuronal cell lines. In particular, the PC12 pheochromocytoma cell line, which displays many properties of adrenal chromaffin cells and undergoes differentiation into sympathetic neuron-like cells when treated with nerve growth factor, has been extensively investigated to identify components of neurotrophin signaling pathways as well as the genes that they regulate. VGF was one of the first neurotrophin-regulated clones identified in NGF-treated PC12 cells. Subsequent studies indicate that the vgf gene is regulated in vivo in the nervous system by neurotrophins, by electrical activity, in response to injury or seizure, and by feeding and the circadian clock. The vgf gene encodes a polypeptide rich in paired basic amino acids; this polypeptide is differentially processed in neuronal and neuroendocrine cells and is released via the regulated secretory pathway. Generation and analysis of knockout mice that fail to synthesize VGF indicate that this protein plays a critical, non-redundant role in the regulation of energy homeostasis, providing a possible link between neurotrophin function in the nervous system and the peripheral control of feeding and metabolic activity. Future experiments should clarify the sites and mechanisms of action of this neurotrophin-regulated neuronal and neuroendocrine protein.

Tumor necrosis factor receptor-associated factor 6 (TRAF6) participates in anti-lipopolysaccharide factors (ALFs) gene expression in mud crab.

PubMed

Sun, Wan-Wei; Zhang, Xin-Xu; Wan, Wei-Song; Wang, Shu-Qi; Wen, Xiao-Bo; Zheng, Huai-Ping; Zhang, Yue-Ling; Li, Sheng-Kang

2017-02-01

Tumor necrosis factor receptor-associated factor 6 (TRAF6) is a key cytoplasm signal adaptor that mediates signals activated by tumor necrosis factor receptor (TNFR) superfamily and the Interleukin-1 receptor/Toll-like receptor (IL-1/TLR) superfamily. The full-length 2492 bp TRAF6 (Sp-TRAF6) from Scylla paramamosain contains 1800 bp of open reading frame (ORF) encoding 598 amino acids, including an N-terminal RING-type zinc finger, two TRAF-type zinc fingers and a conserved C-terminal meprin and TRAF homology (MATH) domain. Multiple alignment analysis shows that the putative amino acid sequence of Sp-TRAf6 has highest identity of 88% with Pt-TRAF6 from Portunus trituberculatus, while the similarity of Sp-TRAF6 with other crustacean sequences was 54-55%. RT-PCR analysis indicated that Sp-TRAF6 transcripts were predominantly expressed in the hepatopancreas and stomach, whereas it was barely detected in the heart and hemocytes in our study. Moreover, Sp-TRAF6 transcripts were significantly up-regulated after Vibrio parahemolyticus and LPS challenges. RNA interference assay was carried out used by siRNA to investigate the genes expression patterns regulated by Sp-TRAF6. The qRT-PCR results showed that silencing Sp-TRAF6 gene could inhibit SpALF1, SpALF2, SpALF5 and SpALF6 expression in hemocytes, while inhibit SpALF1, SpALF3, SpALF4, SpALF5 and SpALF6 expression in hepatopancreas. Taken together, the acute-phase response to immune challenges and the inhibition of SpALFs gene expression indicate that Sp-TRAF6 plays an important role in host defense against pathogen invasions via regulation of ALF gene expression in S. paramamosain. Copyright © 2016. Published by Elsevier Ltd.

Sema4D, the ligand for Plexin B1, suppresses c-Met activation and migration and promotes melanocyte survival and growth

PubMed Central

Soong, Joanne; Chen, Yulin; Shustef, Elina; Scott, Glynis

2011-01-01

Semaphorins are secreted and membrane bound proteins involved in neural pathfinding, organogenesis, and tumor progression, through Plexin and neuropilins receptors. We recently reported that Plexin B1, the Semaphorin 4D receptor, is a tumor suppressor protein for melanoma, in part, through inhibition of the oncogenic c-Met tyrosine kinase receptor. In this report we show that Sema4D is a protective paracrine factor for normal human melanocyte survival in response to ultraviolet irradiation, that it stimulates proliferation, and regulates the activity of the c-Met receptor. c-Met receptor signaling stimulates melanocyte migration, in part through down-regulation of the cell adhesion molecule E-cadherin. Sema4D suppressed activation of c-Met in response to its ligand hepatocyte growth factor (HGF), and partially blocked the suppressive effects of HGF on E-cadherin expression in melanocytes and HGF-dependent migration. These data demonstrate a role for Plexin B1 in maintenance of melanocyte survival and proliferation in the skin, and suggest that Semaphorin 4D and Plexin B1 act cooperatively with HGF and c-Met to regulate c-Met dependent effects in human melanocytes. Because our data show that Plexin B1 is profoundly down-regulated by UVB in melanocytes, loss of Plexin B1 may accentuate HGF dependent effects on melanocytes, including melanocyte migration. PMID:22189792

Sema4D, the ligand for Plexin B1, suppresses c-Met activation and migration and promotes melanocyte survival and growth.

PubMed

Soong, Joanne; Chen, Yulin; Shustef, Elina M; Scott, Glynis A

2012-04-01

Semaphorins are secreted and membrane-bound proteins involved in neural pathfinding, organogenesis, and tumor progression, through Plexin and neuropilin receptors. We recently reported that Plexin B1, the Semaphorin 4D (Sema4D) receptor, is a tumor-suppressor protein for melanoma, which functions, in part, through inhibition of the oncogenic c-Met tyrosine kinase receptor. In this report, we show that Sema4D is a protective paracrine factor for normal human melanocyte survival in response to UV irradiation, and that it stimulates proliferation and regulates the activity of the c-Met receptor. c-Met receptor signaling stimulates melanocyte migration, partly through downregulation of the cell adhesion molecule E-cadherin. Sema4D suppressed activation of c-Met in response to its ligand, hepatocyte growth factor (HGF), and partially blocked the suppressive effects of HGF on E-cadherin expression in melanocytes and HGF-dependent migration. These data demonstrate a role for Plexin B1 in maintenance of melanocyte survival and proliferation in the skin, and suggest that Sema4D and Plexin B1 act cooperatively with HGF and c-Met to regulate c-Met-dependent effects in human melanocytes. Because our data show that Plexin B1 is profoundly downregulated by UVB in melanocytes, loss of Plexin B1 may accentuate HGF-dependent effects on melanocytes, including melanocyte migration.

Identification of transcriptional factors and key genes in primary osteoporosis by DNA microarray.

PubMed

Xie, Wengui; Ji, Lixin; Zhao, Teng; Gao, Pengfei

2015-05-09

A number of genes have been identified to be related with primary osteoporosis while less is known about the comprehensive interactions between regulating genes and proteins. We aimed to identify the differentially expressed genes (DEGs) and regulatory effects of transcription factors (TFs) involved in primary osteoporosis. The gene expression profile GSE35958 was obtained from Gene Expression Omnibus database, including 5 primary osteoporosis and 4 normal bone tissues. The differentially expressed genes between primary osteoporosis and normal bone tissues were identified by the same package in R language. The TFs of these DEGs were predicted with the Essaghir A method. DAVID (The Database for Annotation, Visualization and Integrated Discovery) was applied to perform the GO (Gene Ontology) and KEGG (Kyoto Encyclopedia of Genes and Genomes) pathway enrichment analysis of DEGs. After analyzing regulatory effects, a regulatory network was built between TFs and the related DEGs. A total of 579 DEGs was screened, including 310 up-regulated genes and 269 down-regulated genes in primary osteoporosis samples. In GO terms, more up-regulated genes were enriched in transcription regulator activity, and secondly in transcription factor activity. A total 10 significant pathways were enriched in KEGG analysis, including colorectal cancer, Wnt signaling pathway, Focal adhesion, and MAPK signaling pathway. Moreover, total 7 TFs were enriched, of which CTNNB1, SP1, and TP53 regulated most up-regulated DEGs. The discovery of the enriched TFs might contribute to the understanding of the mechanism of primary osteoporosis. Further research on genes and TFs related to the WNT signaling pathway and MAPK pathway is urgent for clinical diagnosis and directing treatment of primary osteoporosis.

VISIONET: intuitive visualisation of overlapping transcription factor networks, with applications in cardiogenic gene discovery.

PubMed

Nim, Hieu T; Furtado, Milena B; Costa, Mauro W; Rosenthal, Nadia A; Kitano, Hiroaki; Boyd, Sarah E

2015-05-01

Existing de novo software platforms have largely overlooked a valuable resource, the expertise of the intended biologist users. Typical data representations such as long gene lists, or highly dense and overlapping transcription factor networks often hinder biologists from relating these results to their expertise. VISIONET, a streamlined visualisation tool built from experimental needs, enables biologists to transform large and dense overlapping transcription factor networks into sparse human-readable graphs via numerically filtering. The VISIONET interface allows users without a computing background to interactively explore and filter their data, and empowers them to apply their specialist knowledge on far more complex and substantial data sets than is currently possible. Applying VISIONET to the Tbx20-Gata4 transcription factor network led to the discovery and validation of Aldh1a2, an essential developmental gene associated with various important cardiac disorders, as a healthy adult cardiac fibroblast gene co-regulated by cardiogenic transcription factors Gata4 and Tbx20. We demonstrate with experimental validations the utility of VISIONET for expertise-driven gene discovery that opens new experimental directions that would not otherwise have been identified.

MRG1, the product of a melanocyte-specific gene related gene, is a cytokine-inducible transcription factor with transformation activity

PubMed Central

Sun, Hui Bin; Zhu, Yuan Xiao; Yin, Tinggui; Sledge, George; Yang, Yu-Chung

1998-01-01

Identification of cytokine-inducible genes is imperative for determining the mechanisms of cytokine action. A cytokine-inducible gene, mrg1 [melanocyte-specific gene (msg1) related gene], was identified through mRNA differential display of interleukin (IL) 9-stimulated and unstimulated mouse helper T cells. In addition to IL-9, mrg1 can be induced by other cytokines and biological stimuli, including IL-1α, -2, -4, -6, and -11, granulocyte/macrophage colony-stimulating factor, interferon γ, platelet-derived growth factor, insulin, serum, and lipopolysaccharide in diverse cell types. The induction of mrg1 by these stimuli appears to be transient, with induction kinetics similar to other primary response genes, implicating its role in diverse biological processes. Deletion or point mutations of either the Box1 motif (binds Janus kinase 1) or the signal transducer and activator of transcription 3 binding site-containing region within the intracellular domain of the IL-9 receptor ligand binding subunit abolished or greatly reduced mrg1 induction by IL-9, suggesting that the Janus kinase/signal transducer and activator of transcription signaling pathway is required for mrg1 induction, at least in response to IL-9. Transfection of mrg1 cDNA into TS1, an IL-9-dependent mouse T cell line, converted these cells to IL-9-independent growth through a nonautocrine mechanism. Overexpression of mrg1 in Rat1 cells resulted in loss of cell contact inhibition, anchorage-independent growth in soft agar, and tumor formation in nude mice, demonstrating that mrg1 is a transforming gene. MRG1 is a transcriptional activator and may represent a founding member of an additional family of transcription factors. PMID:9811838

Ectoderm gene activation in sea urchin embryos mediated by the CCAAT-binding factor.

PubMed

Li, Xiaotao; Bhattacharya, Chitralekha; Dayal, Sandeep; Maity, Sankar; Klein, William H

2002-05-01

Transcriptional enhancers are short stretches of DNA that function to achieve highly specific patterns of gene expression. To identify the mechanisms by which enhancers achieve their specificity, we made use of an enhancer from the aboral ectoderm-specific spec2a gene of the sea urchin Strongylocentrotus purpuratus. The spec2a enhancer contains five cis-regulatory elements within 78 base pairs that interact with five distinct DNA-binding proteins to confer aboral ectoderm expression. Here, we present an analysis of the sea urchin CCAAT binding factor (CBF), which binds to a CCAAT motif within the spec2a enhancer. S. purpuratus CBF and SpOtx, a ubiquitously expressed factor, act together at closely placed cis-regulatory elements to mediate spec2a transcription in the ectoderm. SpCBF was the sole factor that bound to the spec2a CCAAT element, and two of the three subunits that make up the CBF holoprotein were cloned and shown to have high sequence conservation with their vertebrate orthologs. Based on its involvement in the regulation of several other sea urchin genes, SpCBF appears to be a major transcription factor in the sea urchin embryo for positive regulation of ectoderm gene expression. In addition to its role in vertebrate cell growth and proliferation, our results indicate that CBF also functions at the early stages of germ layer formation, namely ectoderm differentiation.

Alcohol-related Genes Show an Enrichment of Associations with a Persistent Externalizing Factor

PubMed Central

Ashenhurst, James R.; Harden, K. Paige; Corbin, William R.; Fromme, Kim

2016-01-01

Research using twins has found that much of the variability in externalizing phenotypes – including alcohol and drug use, impulsive personality traits, risky sex and property crime – is explained by genetic factors. Nevertheless, identification of specific genes and variants associated with these traits has proven to be difficult, likely because individual differences in externalizing are explained by many genes of small individual effect. Moreover, twin research indicates that heritable variance in externalizing behaviors is mostly shared across the externalizing spectrum rather than specific to any behavior. We use a longitudinal, “deep phenotyping” approach to model a general externalizing factor reflecting persistent engagement in a variety of socially problematic behaviors measured at eleven assessment occasions spanning early adulthood (ages 18 to 28). In an ancestrally homogenous sample of non-Hispanic Whites (N = 337), we then tested for enrichment of associations between the persistent externalizing factor and a set of 3,281 polymorphisms within 104 genes that were previously identified as associated with alcohol-use behaviors. Next we tested for enrichment among domain-specific factors (e.g., property crime) composed of residual variance not accounted for by the common factor. Significance was determined relative to bootstrapped empirical thresholds derived from permutations of phenotypic data. Results indicated significant enrichment of genetic associations for persistent externalizing, but not for domain-specific factors. Consistent with twin research findings, these results suggest that genetic variants are broadly associated with externalizing behaviors rather than unique to specific behaviors. General Scientific Summary This study shows that variation in 104 genes is associated with socially problematic “externalizing” behavior, including substance misuse, property crime, risky sex, and aspects of impulsive personality. Importantly, this

MAPK pathway inhibition induces MET and GAB1 levels, priming BRAF mutant melanoma for rescue by hepatocyte growth factor

PubMed Central

Caenepeel, Sean; Cooke, Keegan; Wadsworth, Sarah; Huang, Guo; Robert, Lidia; Moreno, Blanca Homet; Parisi, Giulia; Cajulis, Elaina; Kendall, Richard; Beltran, Pedro; Ribas, Antoni; Coxon, Angela; Hughes, Paul E.

2017-01-01

Therapeutic resistance is a major obstacle to achieving durable clinical responses with targeted therapies, highlighting a need to elucidate the underlying mechanisms responsible for resistance and identify strategies to overcome this challenge. An emerging body of data implicates the tyrosine kinase MET in mediating resistance to BRAF inhibitors in BRAFV600E mutant melanoma. In this study we observed a dominant role for the HGF/MET axis in mediating resistance to BRAF and MEK inhibitors in models of BRAFV600E and NRAS mutant melanoma. In addition, we showed that MAPK pathway inhibition induced rapid increases in MET and GAB1 levels, providing novel mechanistic insight into how BRAFV600E mutant melanoma is primed for HGF-mediated rescue. We also determined that tumor-derived HGF, not systemic HGF, may be required to convey resistance to BRAF inhibition in vivo and that resistance could be reversed following treatment with AMG 337, a selective MET inhibitor. In summary, these findings support the clinical evaluation of MET-directed targeted therapy to circumvent resistance to BRAF and MEK inhibitors in BRAFV600E mutant melanoma. In addition, the induction of MET following treatment with BRAF and MEK inhibitors has the potential to serve as a predictive biomarker for identifying patients best suited for MET inhibitor combination therapy. PMID:28147313

oPOSSUM: identification of over-represented transcription factor binding sites in co-expressed genes

PubMed Central

Ho Sui, Shannan J.; Mortimer, James R.; Arenillas, David J.; Brumm, Jochen; Walsh, Christopher J.; Kennedy, Brian P.; Wasserman, Wyeth W.

2005-01-01

Targeted transcript profiling studies can identify sets of co-expressed genes; however, identification of the underlying functional mechanism(s) is a significant challenge. Established methods for the analysis of gene annotations, particularly those based on the Gene Ontology, can identify functional linkages between genes. Similar methods for the identification of over-represented transcription factor binding sites (TFBSs) have been successful in yeast, but extension to human genomics has largely proved ineffective. Creation of a system for the efficient identification of common regulatory mechanisms in a subset of co-expressed human genes promises to break a roadblock in functional genomics research. We have developed an integrated system that searches for evidence of co-regulation by one or more transcription factors (TFs). oPOSSUM combines a pre-computed database of conserved TFBSs in human and mouse promoters with statistical methods for identification of sites over-represented in a set of co-expressed genes. The algorithm successfully identified mediating TFs in control sets of tissue-specific genes and in sets of co-expressed genes from three transcript profiling studies. Simulation studies indicate that oPOSSUM produces few false positives using empirically defined thresholds and can tolerate up to 50% noise in a set of co-expressed genes. PMID:15933209

In Vivo Gene Therapy of Hemophilia B: Sustained Partial Correction in Factor IX-Deficient Dogs

NASA Astrophysics Data System (ADS)

Kay, Mark A.; Rothenberg, Steven; Landen, Charles N.; Bellinger, Dwight A.; Leland, Frances; Toman, Carol; Finegold, Milton; Thompson, Arthur R.; Read, M. S.; Brinkhous, Kenneth M.; Woo, Savio L. C.

1993-10-01

The liver represents a model organ for gene therapy. A method has been developed for hepatic gene transfer in vivo by the direct infusion of recombinant retroviral vectors into the portal vasculature, which results in the persistent expression of exogenous genes. To determine if these technologies are applicable for the treatment of hemophilia B patients, preclinical efficacy studies were done in a hemophilia B dog model. When the canine factor IX complementary DNA was transduced directly into the hepatocytes of affected dogs in vivo, the animals constitutively expressed low levels of canine factor IX for more than 5 months. Persistent expression of the clotting. factor resulted in reductions of whole blood clotting and partial thromboplastin times of the treated animals. Thus, long-term treatment of hemophilia B patients may be feasible by direct hepatic gene therapy in vivo.

Synergistic activation of the chicken mim-1 gene by v-myb and C/EBP transcription factors.

PubMed Central

Burk, O; Mink, S; Ringwald, M; Klempnauer, K H

1993-01-01

The retroviral oncogene v-myb encodes a transcriptional activator which is responsible for the activation of the mim-1 gene in myelomonocytic cells transformed by v-myb. The mim-1 promoter contains several myb consensus binding sites and has previously been shown to be regulated directly by v-myb. Here we report that the mim-1 gene is activated synergistically by v-myb and different C/EBP transcription factors. We have cloned a chicken C/EBP-related gene that is highly expressed in myeloid cells and identified it as the chicken homolog of C/EBP beta. A dominant-negative variant of chicken C/EBP beta interferes with the v-myb induced activation of the mim-1 gene in these cells, suggesting that C/EBP beta or another C/EBP transcription factor is required for the activation of mim-1 by v-myb. We found that C/EBP beta and other C/EBP transcription factors confer to fibroblasts the ability to induce the mim-1 gene in the presence of v-myb. Finally we show that, in contrast to v-myb, c-myb synergizes with C/EBP transcription factors only at low concentrations of c-myb protein. Our results suggest a role for C/EBP beta, and possibly for other C/EBP transcription factors, in v-myb function and in myeloid-specific gene activation. Images PMID:8491193

Transcription factor CREB is involved in CaSR-mediated cytoskeleton gene expression.

PubMed

Huang, Shuaishuai; Ren, Yu; Wang, Ping; Li, Yanyuan; Wang, Xue; Zhuang, Haihui; Fang, Rong; Wang, Yuduo; Liu, Ningsheng; Hehir, Michael; Zhou, Jeff X

2015-03-01

Our previous studies illustrated that a steady increase of intracellular calcium concentration ([Ca2+]i) was important for maintaining microtubules (MTs) rearrangement in apoptotic cells. However, little is known about the effect of calcium sensing receptor (CaSR)-mediated increase in [Ca2+]i on cytoskeleton gene expression. We examined the impact of taxol or CaSR agonist/antagonist on the regulation of [Ca2+]i concentration, cytoskeleton arrangement, phosphorylated CREB and cytoskeleton gene expressions in HeLa cells with dominant negative plasmid of CREB (PM). This study demonstrated that Gdcl3 (a specific CaSR agonist) evoked a rapid increase of [Ca2+]i, formed a rigid bundle of MTs which surrounded the nucleus and decreased the cytoskeleton gene expressions in HeLa cells. These effects were rescued by addition of NPS2390 (a specific CaSR antagonist). Moreover, CaSR activity affected cytoskeleton gene expression through transcription factor CREB. Histoscores of pCREB immunoreactivity in tissues of cervical adenocarcinoma, renal clear cell carcinoma, and diffuse large B-cell lymphoma were markedly increased compared with non malignant tissue. These data demonstrate, for the first time, that CaSR-mediated increase in [Ca2+]i probably modulate cytoskeleton organization and gene expression via transcription factor. © 2014 Wiley Periodicals, Inc.

Bone marrow-derived mesenchymal stem cells effectively regenerate fibrotic liver in bile duct ligation rat model

PubMed Central

Elswefy, Sahar E; Rashed, Laila A; Younis, Nahla N; Shaheen, Mohamed A; Ghanim, Amal MH

2016-01-01

Mesenchymal stem cells (MSCs) have attracted lots of attention for the treatment of acute liver failure and end-stage liver diseases. This study aimed at investigating the fundamental mechanism by which bone marrow-derived MSCs (BM-MSCs) induce liver regeneration of fibrotic liver in rats. Rats underwent bile duct ligation (BDL) surgery and four weeks later they were treated with either BM-MSCs (3 × 106 cells /rat, once, tail vein injection) or silymarin (100 mg/kg, daily, orally) for four weeks. Liver function tests and hepatic oxidative stress were determined. Hepatic injury and fibrosis were assessed by H and E, Sirus red staining and immunohistochemical expression of α-smooth muscle actin (α-SMA). Hepatocyte growth factor (HGF) and the gene expression of cytokeratin-19 (CK-19) and matrix metalloproteinase-2 (MMP-2) in liver tissue were determined. BDL induced cholestatic liver injury characterized by elevated ALT and AST activities, bilirubin and decreased albumin. The architecture damage was staged as Metavir score: F3, A3. Fibrosis increased around proliferating bile duct as indicated by sirus red staining and α-SMA immunostaining. Fibrogenesis was favored over fibrolysis and confirmed by decreased HGF with increased expression of CK-19, but decreased MMP-2 expression. BM-MSCs treatment restored deteriorated liver functions and restored the histological changes, resolved fibrosis by improving liver regenerative capabilities (P < 0.001), increases in HGF and MMP-2 mRNA and downregulating CK-19 mRNA. Sliymarin, however, induced similar but less prominent effects compared to BM-MSCs. In conclusion, liver regenerative capabilities can be stimulated by BM-MSCs via augmentation of HGF that subsequently up-regulate MMP-2 mRNA while downregulating CK-19 mRNA. PMID:26811102