Expression Of Hepatocyte Growth Factor Activator Research Articles

Mechanisms of PAK1-Induced Skeletal Muscle Myokines in Improving Islet β-Cell Function

The critical role of skeletal muscle in regulating metabolic homeostasis is well-acknowledged. The p21-activated kinase (PAK1), a prominent signaling molecule in skeletal muscle, has garnered attention for its potential modulatory role in glucose and mitochondrial metabolism. By applying conditioned media derived from PAK1-enriched L6 Glut4 myc myotubes or myoblasts to INS-1 832/13 clonal β-cells, we confirmed that a muscle-derived circulating factor(s) could enhance β-cell function. Furthermore, our mass spectrometry analysis in the conditioned media (CM) collected from PAK1 enriched L6.GLUT4myc myotubes revealed elevated levels of one protein in particular, hepatocyte growth factor activator (HGFAC), which is categorized as a secreted myokine. HGFAC is secreted into the circulation, where, upon activation, it can act in a paracrine or endocrine fashion to proteolytically cleave and activate HGF. It has been reported that HGF has a vital role in islet β-cell mass and function and HNFα, a transcription factor that is expressed in pancreatic β-cells regulates β-cell mass or function by modulating HGFAC expression. Thus, we hypothesize that PAK1-enriched skeletal muscle releases HGFAC into the serum-free conditioned medium and the medium carrying HGFAC when applied to islet β-cells enhances insulin secretion. Elevated levels of HGFAC protein were detected in the cells and their associated CM from PAK1-enriched skeletal myotubes, as compared with that of GFP-transduced control myotubes, using sandwich ELISA to quantify HGFAC. Notably, these cells were incubated in a serum-free medium, a means to eliminate serum factors that might impact HGFAC levels or detection capacity. Given this, we tested the effect of this serum-free CM collected from PAK1-enriched L6.GLUT4myc myotubes application to species-matched (rat) clonal β-cells functional capacity. Indeed, CM from PAK1-enriched myotubes significantly enhanced glucose-stimulated insulin secretion (GSIS), as compared that of the control CM. The presence of elevated HGFAC levels in PAK1-enriched cells, and the beneficial impact of this HGFAC-enriched CM upon beta cell function, reveals a new mechanism by which PAK1-enrichment in skeletal muscle imparts improved overall whole-body glucose homeostasis. further hints at a possible intricate relationship between muscle-pancreas crosstalk in the regulation of islet β-cell insulin secretion. Future investigations into possible autocrine functions of PAK1-enriched skeletal muscle are underway. This research was funded by the National Institutes of Health to D.C.T. (DK067912, DK112917, and DK102233), and an American Heart Association Postdoctoral fellowship to R.B. (#905770). This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

205-LB: HGFAC Is a ChREBP-Regulated Hepatokine That Enhances Glucose and Lipid Homeostasis

Carbohydrate Responsive Element-Binding Protein (ChREBP) is a carbohydrate sensing transcription factor that regulates metabolic gene programs. Genetic variants in the ChREBP locus associate with diverse metabolic traits in humans, including circulating triglycerides (TG). The transcriptional targets that mediate its pleiotropic physiological effects are poorly understood. To identify novel ChREBP-regulated hepatokines that contribute to its effects on metabolism, we integrated ChIP-seq for ChREBP in mouse liver with human GWAS data for TGs and identified Hepatocyte Growth Factor Activator (HGFAC) as a promising candidate. HGFAC is a circulating zymogen that activates Hepatocyte Growth Factor (HGF), a multifunction cytokine involved in cellular growth, motility, and morphogenesis. High fructose feeding increased circulating HGFAC by 1.8-fold (p<0.001) and this was abrogated in liver specific ChREBP KO mice. Similar increases were seen in mice fed high sucrose/high fat diet and in obese Zucker rats (2.6-fold, p<0.001). In humans, expression of hepatic ChREBP transcriptional targets associated with HGFAC expression (Pearson r= 0.66, p <0.0001), and its expression was elevated in patients with obesity and T2D (2.7-fold, p<0.001). Putative loss of function variants in HGFAC associated with hypertriglyceridemia in humans, and deletion of HGFAC in mice increased circulating TGs (1.4-fold, p<0.005), and caused glycerol and glucose intolerance despite a 40% reduction in hepatic TG content (p<0.001). RNA-seq indicated that HGFAC regulates hepatic metabolism through modulation of hepatic PPARG expression. Reciprocal effects on PPARG expression and metabolic phenotypes were observed following adenovirus HGFAC overexpression in mice. Moreover, HGF induces PPARG express 1.3-fold in HepG2 cells (p<0.0001). In conclusion, ChREBP responds to carbohydrate overnutrition and regulates systemic glucose and lipid metabolism via an HGFAC-HGF-PPARG signaling axis. Disclosure A. Sargsyan: None. I. Astapova: None. L. T. Tsai: None. M. A. Herman: Research Support; Self; Eli Lilly and Company. L. Doridot: Employee; Spouse/Partner; Eli Lilly and Company. S. A. Hannou: None. W. Tong: None. H. Srinivasan: None. R. Ivison: None. R. Monn: None. H. H. Kou: None. J. Haldeman: None. Funding American Diabetes Association (1-19-PDF-088 to A.S.)

HGFAC expression decreased in liver cancer and its low expression correlated with DNA hypermethylation and poor prognosis.

As the activator of hepatocyte growth factor (HGF), hepatocyte growth factor activator (HGFAC) has been previously reported to involve in liver, kidney, and stomach regeneration in response to injury and several types of cancers. This study aimed to explore the expression of HGFAC and the correlation between decreased HGFAC and DNA methylation in liver cancer and its clinical significance. The molecular profiling of HGFAC in liver cancer was analyzed using genomic, transcriptomic, immunohistochemistry, and patient survival information from databases, including oncomine, HPA, COSMIC, UCSC Xena, MPRESS, and Kaplan-Meier plotter. It was found that HGFAC expression at transcriptional and translational level decreased in liver cancer compared with normal tissues. There is a negative correlation between the HGFAC expression and methylation around the promoter region which indicated that HGFAC expression might be controlled by DNA methylation. Moreover, the decreased HGFAC is associated with significantly short over survival time in liver cancer patients. In conclusion, decreased HGFAC expression in liver cancer associated with poor survival outcome. Hypermethylation around the promoter region contributed to the decreased expression of HGFAC. HGFAC may be a useful and predictive biomarker for the prognosis of liver cancer patients. To the best of our knowledge, this is the first report describing the relationship between HGFAC DNA methylation and its expression.

Hepatocyte Growth Factor System in the Mouse Uterus: Variation Across the Estrous Cycle and Regulation by 17-Beta-Estradiol and Progesterone1

Hepatocyte growth factor (HGF) and its receptor MET have been implicated in uterine development, pregnancy, and endometrial disorders, such as endometriosis and carcinoma. In vitro studies have shown that HGF acts as a mitogen, motogen, and morphogen on endometrial epithelial cells. However, the expression and regulation of HGF and MET in the uteri of different species remain obscure. The present study aimed to investigate the changes of HGF, MET, and HGF activator (HGFA) expression in the uterine endometrium during the estrous cycle in mice and to explore estrogen and progesterone regulation of their expression. MKI67 immunostaining was conducted to examine the association between HGF/MET expression and endometrial cell proliferation. Endometrial epithelial and stromal cells both expressed HGF, HGFA, and MET, but the cell type-specific patterns changed during the cycle. Estrogen and progesterone differentially regulated HGF, MET, and HGFA expression. Progesterone up-regulated their expression in the stroma and down-regulated their expression in the luminal epithelium, whereas 17-beta-estradiol down-regulated their expression in the glandular epithelium. The pattern of HGF/MET overall correlated with that of MKI67. In conclusion, HGF, HGFA, and MET expression in mouse uterus changes during the estrous cycle in a stage-, cell type-, and compartment-specific manner under the influence of estrogen and progesterone. HGF likely plays a role in cyclic endometrial remodeling, such as cell proliferation via autocrine/paracrine mechanisms in mouse uterus.

Protein C inhibitor regulates hepatocyte growth factor activator-mediated liver regeneration in mice

Background:We recently reported that human protein C inhibitor (PCI), a major inhibitor of activated protein C (APC), inhibits hepatocyte growth factor activator (HGFA) by forming HGFA–PCI complexes in vitro. In...

Augmented expression of hepatocytes growth factor activator inhibitor type 1 (HAI-1) in intrahepatic small bile ducts in primary biliary cirrhosis

The repair system of damaged biliary mucosa was not fully clarified so far in primary biliary cirrhosis (PBC). Given that related factors of the hepatocyte growth factor (HGF) such as HGF activator (HGFA) and HGFA inhibitor type 1 (HAI-1) participate in the repair of injured gastrointestinal mucosa, we investigated the involvement of the HGF/HGFA/HAI-1 system in PBC and control livers. The expression of HGFA, HAI-1, and c-Met was examined in PBC livers (n=24), diseased livers (control, n=30), and normal livers (n=15) by immunohistochemistry and semiquantitative reverse transcriptase-polymerase chain reaction. We examined the expression of HGFA, HAI-1, and c-Met, and the effect of HGF administration on cell proliferation and wound healing, and HAI expression in cultured mouse biliary epithelial cells (BECs). HAI-1 expression was faint in control livers, whereas it was significantly augmented in damaged small bile ducts, bile ductules, and periportal hepatocytes in PBC (p<0.05). HGFA and c-Met were homogeneously expressed in BECs in PBC and control livers. HAI-1 expression was increased at the front of wound healing and the treatment with HGF-enhanced HAI-1 expression, cell proliferation, and wound healing in cultured BECs. HGF/HGFA/HAI-1 system may participate in biliary mucosal repair as reported in gastrointestinal mucosal repair.

Hepatocyte Growth Factor Activator Inhibitor Types 1 and 2 are Expressed by Tubular Epithelium in Kidney and Down-regulated in Renal Cell Carcinoma

Hepatocyte growth factor (HGF) and its receptor MET have been implicated in kidney development and renal cell carcinoma (RCC) progression. HGF is secreted as an inactive proform and it must be activated to initiate MET signaling. HGF activator (HGFA) activates pro-HGF in injured tissue. We evaluated the expression of HGFA and its endogenous inhibitors HAI-1 and HAI-2 in normal kidney and RCC. We examined the gene expression of HGFA, HAI-1, HAI-2, HGF and MET in a normal kidney by laser captured microdissection, followed by reverse transcriptase-polymerase chain reaction. We also quantified the mRNA levels of these proteins in 14 RCC cases by real-time reverse transcriptase-polymerase chain reaction. HAI-1 and HAI-2 were abundant in the normal kidney. The uriniferous tubules showed the highest levels of HAI-1 and HAI-2 mRNA. HGFA was hardly detectable in the normal kidney. However, in the kidney with RCC a low but distinct level of HGFA mRNA became detectable in the tumor and adjacent renal tissue. The HAI-1 mRNA level was significantly and consistently down-regulated in RCC relative to normal tissue. HAI-2 mRNA was also significantly low in the advanced stage of RCC. MET was up-regulated in most cases of RCC. HAI-1 and HAI-2 were expressed in renal tubular epithelial cells. The expression of the 2 HAIs was significantly down-regulated in RCC, whereas HGFA expression was enhanced in the diseased kidney, suggesting an imbalance between HAI and its target proteinases, including HGFA, in favor of proteinase activities in RCC.

Hepatocyte growth factor (HGF) system in gingiva: HGF activator expression by gingival epithelial cells.

Hepatocyte growth factor (HGF) acts as a mitogen, motogen, morphogen, and anti-apoptotic factor for various kinds of epithelial cells. We previously showed that periodontal ligament and gingival fibroblasts secreted an HGF-like chemoattractant for a gingival epithelial cell line and found that the HGF content of gingival crevicular fluid was well correlated with clinical parameters and interleukin-1beta level. Since HGF is secreted as an inactive form (proHGF), and converted to an active form by serine proteases such as HGF activator (HGFA), extracellular processing of proHGF is presumed to be critical in the regulation of HGF activity. To examine the role of the HGF system in epithelial invasion followed by loss of connective tissue attachment in periodontitis, mRNA expression of HGF, its receptor (c-met) and HGFA in gingival tissues was monitored. Ten gingival biopsies were obtained, and epithelium and connective tissues were separated by enzymatic digestion. The gene expression of HGF and keratinocyte growth factor (KGF) in gingival connective tissue, and c-met, HGFA and KGF receptor (KGFR) in gingival epithelial tissues was monitored using RT-PCR. Furthermore, HGFA protein in the conditioned medium of cultured primary gingival epithelial cells was examined using Western blotting. All the connective tissue samples expressed KGF, and 8 out of 10 samples expressed HGF. All the epithelial samples expressed KGFR and c-met, whereas 5 out of 10 samples expressed HGFA. Protein expression of HGFA by cultured primary gingival epithelial cells was also confirmed. In terms of local production and activation of HGF in gingival tissue, these results suggest that synergistic expression of HGF in connective tissue and HGFA expression in epithelium may contribute to disease progression in periodontitis.

Expression of Hepatocyte Growth Factor Activator and Hepatocyte Growth Factor Activator Inhibitor Type 1 in Human Hepatocellular Carcinoma

Hepatocyte growth factor activator inhibitor type 1 (HAI-1), a Kunitz-type serine protease inhibitor for hepatocyte growth factor activator (HGFA), is responsible for proteolytic activation of hepatocyte growth factor. We examined the expression of HGFA and HAI-1 in liver tissues of chronic liver diseases including hepatocellular carcinoma (HCC). HGFA expression was detected not only in the liver tissues of chronic hepatitis and cirrhosis and in the nontumorous liver tissues surrounding HCC, but also in HCC tissues. On the other hand, none of the liver tissues of hepatitis and cirrhosis and none of the nontumorous tissues surrounding HCC were stained with anti-HAI-1. However, 35% of HCC tissues were stained with anti-HAI-1, and HAI-1 positivity increased as the histological grade decreased and as serum α-fetoprotein increased. Transduction of antisense HAI-1 inhibited the growth of human hepatoma cells. These results suggest the possibility that HAI-1 plays an important role in the progression of HCC.

Localization of hepatocyte growth factor activator inhibitor type 1 in Langhans' cells of human placenta.

Activation of hepatocyte growth factor (HGF) is a crucial limiting step in HGF-induced signaling pathway. The HGF activator inhibitor type 1 (HAI-1) was identified as a potent inhibitor of HGF activator (HGFA), a serine proteinase that is responsible for the activation of HGF in vivo. HAI-1 is an integral membrane Kunitz-type serine proteinase inhibitor, and its mRNA has been reported to be most abundant in the placenta. In this report, specific antibody to HAI-1 was used in an immunohistochemical procedure to determine the localization of HAI-I in human placenta. HAI-1 was expressed in cytotrophoblasts (Langhans' cells) of the double-layered trophoblastic epithelium of chorionic villi tissue, and syncytiotrophoblasts were almost negative. On the other hand, extravillous trophoblasts of cytotrophoblastic columns showed markedly decreased immunoreactivity, and those infiltrating into the superficial decidua membrane of early placenta were hardly stainable. The amnionic epithelial cells were also immunostained intensely. The presence of HAI-1 mRNA was also confirmed in a cultured human cytotrophoblastic cell line. In addition to HAI-1, low but distinct expression of HGFA mRNA was observed in the placenta tissue and cultured cytotrophoblasts by using a sensitive RT-PCR method. Since HGF plays an essential role in the placenta development, expression of HAI-1 and HGFA may have an important regulatory role in the placenta. The localization of HAI-I in the proliferating trophoblastic stem cells (Langhans' cells), but not in syncytiotrophoblasts and extravillous trophoblasts, suggest a possible role of HAI-1 in the proliferation of trophoblasts.