Age-related nitration/dysfunction of myogenic stem cell activator HGF

Mechanical perturbation triggers activation of resident myogenic stem cells to enter the cell cycle through a cascade of events including hepatocyte growth factor (HGF) release from its extracellular tethering and the subsequent presentation to signaling-receptor c-met. Here, we show that with aging, extracellular HGF undergoes tyrosine-residue (Y) nitration and loses c-met binding, thereby disturbing muscle homeostasis. Biochemical studies demonstrated that nitration/dysfunction is specific to HGF among other major growth factors and is characterized by its locations at Y198 and Y250 in c-met-binding domains. Direct-immunofluorescence microscopy of lower hind limb muscles from three age groups of rat, provided direct in vivo evidence for age-related increases in nitration of ECM-bound HGF, preferentially stained for anti-nitrated Y198 and Y250-HGF mAbs (raised in-house) in fast IIa and IIx myofibers. Overall, findings highlight inhibitory impacts of HGF nitration on myogenic stem cell dynamics, pioneering a cogent discussion for better understanding age-related muscle atrophy and impaired regeneration with fibrosis (including sarcopenia and frailty).     

Mast cell and neutrophil peptidases attack an inactivation segment in hepatocyte growth factor to generate NK4-like antagonists

Hepatocyte growth factor (HGF) is a plasminogen-like protein with an alpha chain linked to a trypsin-like beta chain without peptidase activity. The interaction of HGF with c-met, a receptor tyrosine kinase expressed by many cells, is important in cell growth, migration, and formation of endothelial and epithelial tubes. Stimulation of c-met requires two-chain, disulfide-linked HGF. Portions of an alpha chain containing an N-terminal segment and four kringle domains (NK4) antagonize HGF activity. Until now, no physiological pathway for generating NK4 was known. Here we show that chymases, which are chymotryptic peptidases secreted by mast cells, hydrolyze HGF, thereby abolishing scatter factor activity while generating an NK4-like antagonist of HGF scatter factor activity. Thus, chymase interferes with HGF directly by destroying active protein and indirectly by generating an antagonist. The site of hydrolysis, Leu480, lies in the alpha chain on the N-terminal side of the cysteine linking the alpha and beta chains. This site appears to be specific for HGF because chymase does not hydrolyze other plasminogen-like proteins, such as macrophage-stimulating protein and plasminogen itself. Mast cell/neutrophil cathepsin G and neutrophil elastase generate similar fragments of HGF by cleaving near the chymase site. Mast cell and neutrophil peptidases are secreted during tissue injury, infection, ischemia, and allergic inflammation, where they may oppose HGF effects on epithelial repair. Thus, HGF possesses an "inactivation segment" that serves as an Achilles' heel attacked by inflammatory proteases. This work reveals a potential physiological pathway for inactivation of HGF and generation of NK4-like antagonists.     

Immunohistochemical expression of HGF, c-MET and transcription factor STAT3 in colorectal tumors

By immunohistochemistry, we have investigated the expression of hepatocyte growth factor (HGF), HGF-R or c-met and the transcritor factor STAT3 in a series of 80 colorectal tumours (40 adenomas and 40 adenocarcinomas). The expression of HGF, c-met and STAT3 was revealed in 40/40 (100%) of adenomas and in 26/40 (65%) of adenocarcinomas; the remaining 14/40 (35%) carcinomas expressed c-met but failed to express HGF and STAT3. Positive immunoreaction score was defined through the number of stained cells: low (1-10%), moderate (11-50%) and high (>51%). In adenomas, the HGF immunoreaction was high in 33 (82.5%) and moderate in 7 (17.5%); the c-met staining was high in 3 (7.5%) and moderate in 37 (92.5%); and the STAT3 reactivity was high in 25 (62.5%) and moderate in 15(37.5%). In carcinomas, the HGF immunoreaction was moderate in 21 (80.7%) and low in 5 (19.2%); the c-met staining was high in 14 (35%), moderate in 25 (62.5) and low in 1 (2.5%); and the STAT3 reactivity was moderate in 17 (65.3%) and low in 9 (34.6%). In both type of lesions, HGF and c-met showed a membranous and cytoplasmic location. In adenomas, STAT3 was detected in cytoplasm and nucleus and in carcinomas it was limited to cytoplasm. While the HGF/c-met/STAT3 expression in adenomas was significantly different from carcinomas (c2 = 17, p < 0.0001), no correlation was found among HGF, c-met, or STAT3 immunostaining with histotype or degree of dysplasia in adenomas and the same for histotype, grading or staging in carcinomas. These features, suggesting a role of the HGF/c-met/STAT3 signal in colon tumorigenesis, indicate that a reduced expression of HGF and c-met is associated to progression of adenoma into carcinoma.     

The tyrosine kinase receptor c-met,its cognate ligand HGF and the tyrosine kinase receptor trasducers STAT3, PI3K and RHO in thyroid nodules associated with Hashimoto's thyroiditis: an immunohisto-chemical characterization

Hepatocyte growth factor (HGF) exerts proliferative activities in thyrocytes upon binding to its tyrosine kinase receptor c-met and is also expressed in benign thyroid nodules as well as in Hashimoto''s thyroiditis (HT).The simultaneous expression of HGF/c-met and three trasducers of tyrosine kinase receptors (STAT3, PI3K, RHO) in both the nodular and extranodular tissues were studied by immunohistochemistry in 50 benign thyroid nodules (NGs), 25 of which associated with HT. The ligand/tyrosine kinase receptor pair HGF/c-met and the two trasducers PI3K and RHO were expressed in NGs, regardless of association with HT, with a higher positive cases percentage in HT-associated NGs compared to not HT-associated NGs (25/25 or 100% vs 7/25 or 28%; P<0.001). HGF, PI3K and RHO expression was only stromal (fibroblasts and endothelial cells), in all 32 reactive NGs, while c-met localization was consistently epithelial (thyrocyes). Immunoreactions for HGF, c-met, PI3K and RHO were also apparent in the extra-nodular tissue of HT specimens, where HGF and PI3K were expressed not only in stromal cells but also in thyrocyes along with the c-met. Finally, a positive correlation was observed between the proportion of HGF, c-met, PI3K follicular cells and the grade of lymphoid aggregates in HT. In conclusion, HGF, c-met, PI3K are much more frequently and highly expressed in HT compared to NGs, and among all NGs in those present in the context of HT. A paracrine effect of HFG/c-met on nodule development, based on the prevalent stromal expression, may be suggested along with a major role of HGF/c-met and PI3K in HT. Finally, the expression of such molecules in HT may be regulated by lymphoid infiltrate.Key words: HGF/c-met signaling, PI3K, RHO, Hashimoto''s thyroiditis, thyroid nodules.     

Calcium influx through a possible coupling of cation channels impacts skeletal muscle satellite cell activation in response to mechanical stretch

When skeletal muscle is stretched or injured, satellite cells, resident myogenic stem cells positioned beneath the basal lamina of mature muscle fibers, are activated to enter the cell cycle. This signaling pathway is a cascade of events including calcium-calmodulin formation, nitric oxide (NO) radical production by NO synthase, matrix metalloproteinase activation, release of hepatocyte growth factor (HGF) from the extracellular matrix, and presentation of HGF to the receptor c-met, as demonstrated by assays of primary cultures and in vivo experiments. Here, we add evidence that two ion channels, the mechanosensitive cation channel (MS channel) and the long-lasting-type voltage-gated calcium-ion channel (L-VGC channel), mediate the influx of extracellular calcium ions in response to cyclic stretch in satellite cell cultures. When applied to 1-h stretch cultures with individual inhibitors for MS and L-VGC channels (GsMTx-4 and nifedipine, respectively) or with a less specific inhibitor (gadolinium chloride, Gd), satellite cell activation and upstream HGF release were abolished, as revealed by bromodeoxyuridine-incorporation assays and Western blotting of conditioned media, respectively. The inhibition was dose dependent with a maximum at 0.1 μM (GsMTx-4), 10 μM (nifedipine), or 100 μM (Gd) and canceled by addition of HGF to the culture media; a potent inhibitor for transient-type VGC channels (NNC55-0396, 100 μM) did not show any significant inhibitory effect. The stretch response was also abolished when calcium-chelator EGTA (1.8 mM) was added to the medium, indicating the significance of extracellular free calcium ions in our present activation model. Finally, cation/calcium channel dependencies were further documented by calcium-imaging analyses on stretched cells; results clearly demonstrated that calcium ion influx was abolished by GsMTx-4, nifedipine, and EGTA. Therefore, these results provide an additional insight that calcium ions may flow in through L-VGC channels by possible coupling with adjacent MS channel gating that promotes the local depolarization of cell membranes to initiate the satellite cell activation cascade.     

The HGF/SF-induced phosphorylation of paxillin, matrix adhesion, and invasion of prostate cancer cells were suppressed by NK4, an HGF/SF variant

Hepatocyte growth factor/scatter factor (HGF/SF) plays a crucial role in cancer cell migration, matrix adhesion, invasion, and angiogenesis, via the phosphorylation of the c-met tyrosine kinase. This study examined the ability of NK4, a recently discovered HGF/SF variant, to inhibit the influence of HGF/SF on cell-matrix interaction, paxillin phosphorylation, and invasion of prostate cancer cells. HGF/SF was shown to dramatically enhance tumour cell motility, invasion, cell-matrix adhesion, together with an increase in the degree of paxillin phosphorylation and formation of focal adhesion complexes. However, these HGF/SF-induced effects were suppressed by the presence of NK4. NK4 effectively inhibited the degree of HGF/SF-induced paxillin phosphorylation and matrix adhesion. As a consequence, the matrix invasion of these prostate cancer cells was also suppressed by NK4. In conclusion, this study shows that these HGF/SF-enhanced events, which are critical steps in metastasis, can be inhibited through the addition of NK4, thus warranting further in vivo studies on the implication of NK4 as a potential antimetastasis agent in prostate cancer.     

Engineering the NK1 fragment of hepatocyte growth factor/scatter factor as a MET receptor antagonist

The growth and motility factor hepatocyte growth factor/scatter factor (HGF/SF) and its receptor MET, the tyrosine kinase encoded by the c-MET proto-oncogene, exert major roles in cancer invasion and metastasis and are key targets for therapy. NK1 is an alternative spliced variant of HGF/SF that consists of the N-terminal (N) and first kringle (K1) domains and has partial agonistic activity. NK1 crystallises as a head-to-tail dimer with an extensive inter-protomeric interface resulting from contacts between the two short interdomain linkers and reciprocal contacts between the N and K1 domains. Here we show that a subset of mutants at the NK1 dimer interface, such as the linker mutants Y124A or N127A or the kringle mutant V140A:I142A, bind the MET receptor with affinities comparable to wild-type NK1 but fail to assemble a dimeric, signalling competent NK1-MET complex. These NK1 variants have no detectable agonistic activity on, behave as bona fide receptor antagonists by blocking cell migration and DNA synthesis in target cells and have strong prospects as therapeutics for human cancer.     

Intracellular signaling cascades triggered by the NK1 fragment of hepatocyte growth factor in human prostate epithelial cell line PNT1A

Hepatocyte Growth Factor (HGF)/c-MET signaling has an emerging role in promoting cell proliferation, survival, migration, wound repair and branching in a variety of cell types. HGF plays a crucial role as a mediator of stromal–epithelial interactions in the normal prostate but the precise biological function of HGF/c-Met interaction in the normal prostate and in prostate cancer is not clear. HGF has two naturally occurring splice variants and NK1, the smallest of these HGF variants, consists of the HGF amino terminus through the first kringle domain. We evaluated the intracellular signaling cascades and the morphological changes triggered by NK1 in human prostate epithelial cell line PNT1A which shows molecular and biochemical properties close to the normal prostate epithelium. We demonstrated that these cells express a functional c-MET, and cell exposure to NK1 induces the phosphorylation of tyrosines 1313/1349/1356 residues of c-MET which provide docking sites for signaling molecules. We observed an increased phosphorylation of ERK1/2, Akt, c-Src, p125FAK, SMAD2/3, and STAT3, down-regulation of the expression of epithelial cell–cell adhesion marker E-cadherin, and enhanced expression levels of mesenchymal markers vimentin, fibronectin, vinculin, α-actinin, and α-smooth muscle actin. This results in cell proliferation, in the appearance of a mesenchymal phenotype, in morphological changes resembling cell scattering and in wound healing. Our findings highlight the function of NK1 in non-tumorigenic human prostatic epithelial cells and provide a picture of the signaling pathways triggered by NK1 in a unique cell line.     

Matrix metalloproteinase-2 mediates stretch-induced activation of skeletal muscle satellite cells in a nitric oxide-dependent manner

When skeletal muscle is stretched or injured, myogenic satellite cells are activated to enter the cell cycle. This process depends on nitric oxide (NO) production, release of hepatocyte growth factor (HGF) from the extracellular matrix, and presentation of HGF to the c-met receptor. Matrix metalloproteinases (MMPs), a large family of zinc-dependent endopeptidases, mediate HGF release from the matrix and this step in the pathway is downstream from NO synthesis [Yamada, M., Tatsumi, R., Kikuiri, T., Okamoto, S., Nonoshita, S., Mizunoya, W., et al. (2006). Matrix metalloproteinases are involved in mechanical stretch-induced activation of skeletal muscle satellite cells. Muscle Nerve, 34, 313-319]. Experiments reported herein provide evidence that MMP2 may be involved in the NO-dependent release of HGF in vitro. Whole lysate analyses of satellite cells demonstrated the presence of MMP2 mRNA and the protein. When rat satellite cells were treated with 30 microM sodium nitroprusside a NO donor or mechanical cyclic stretch for 2h period, inactive proMMP2 (72 kDa) was converted into 52-kDa form and this processing was abolished by adding a NO synthase inhibitor l-NAME (10 microM) to the stretch culture. The 52-kDa species was also generated by treatment of the recombinant MMP2 protein with 1 microM NOC-7 that can spontaneously release NO under physiological conditions without any cofactor, and its activating activity was demonstrated by applying the NOC-7-treated MMP2 to satellite cell culture. HGF release was detected in NOC-7-MMP2-conditioned media by western blotting; very little HGF was found in media that were generated from cultures receiving NOC-7-treated MMP2 (10 ng/ml) plus 250 ng/ml tissue inhibitor-1 of metalloproteinases. Therefore, results from these experiments provide evidence that NO-activated MMP2 may cause release of HGF from the extracellular matrix of satellite cells and contribute to satellite cell activation.     

Reconstitution of Mammary Gland Development In Vitro: Requirement of c-met and c-erbB2 Signaling for Branching and Alveolar Morphogenesis

We have established a cell culture system that reproduces morphogenic processes in the developing mammary gland. EpH4 mouse mammary epithelial cells cultured in matrigel form branched tubules in the presence of hepatocyte growth factor/scatter factor (HGF/SF), the ligand of the c-met tyrosine kinase receptor. In contrast, alveolar structures are formed in the presence of neuregulin, a ligand of c-erbB tyrosine kinase receptors. These distinct morphogenic responses can also be observed with selected human mammary carcinoma tissue in explant culture. HGF/SF-induced branching was abrogated by the PI3 kinase inhibitors wortmannin and {"type":"entrez-nucleotide","attrs":{"text":"LY294002","term_id":"1257998346","term_text":"LY294002"}}LY294002. In contrast, neuregulin- induced alveolar morphogenesis was inhibited by the MAPK kinase inhibitor PD98059. The c-met–mediated response could also be evoked by transfection of a c-met specific substrate, Gab1, which can activate the PI3 kinase pathway. An activated hybrid receptor that contained the intracellular domain of c-erbB2 receptor suffices to induce alveolar morphogenesis, and was observed in the presence of tyrosine residues Y1028, Y1144, Y1201, and Y1226/27 in the substrate-binding domain of c-erbB2. Our data demonstrate that c-met and c-erbB2 signaling elicit distinct morphogenic programs in mammary epithelial cells: formation of branched tubules relies on a pathway involving PI3 kinase, whereas alveolar morphogenesis requires MAPK kinase.     

NK1, a Natural Splice Variant of Hepatocyte Growth Factor/Scatter Factor, Is a Partial Agonist In Vivo

Hepatocyte growth factor/scatter factor (HGF/SF) is a potent mitogen, motogen, and morphogen for epithelial cells expressing its tyrosine kinase receptor, the c-met proto-oncogene product, and is required for normal development in the mouse. Inappropriate stimulation of Met signal transduction induces aberrant morphogenesis and oncogenesis in mice and has been implicated in human cancer. NK1 is a naturally occurring HGF/SF splice variant composed of only the amino terminus and first kringle domain. While the biological activities of NK1 have been controversial, in vitro data suggest that it may have therapeutic value as an HGF/SF antagonist. Here, we directly test this hypothesis in vivo by expressing mouse NK1 in transgenic mice and comparing the consequent effects with those observed for mice carrying an HGF/SF transgene. Despite robust expression, NK1 did not behave as an HGF/SF antagonist in vivo. Instead, NK1-transgenic mice displayed most of the phenotypic characteristics associated with HGF/SF-transgenic mice, including enlarged livers, ectopic skeletal-muscle formation, progressive renal disease, aberrant pigment cell localization, precocious mammary lobuloalveolar development, and the appearance of mammary, hepatocellular, and melanocytic tumors. And like HGF/SF-transgenic livers, NK1 livers had higher levels of tyrosine-phosphorylated complexes associated with Met, suggesting that the mechanistic basis for the effects of NK1 overexpression in vivo was autocrine activation of Met. We conclude that NK1 acts in vivo as a partial agonist. As such, the efficacy of NK1 as a therapeutic HGF/SF antagonist must be seriously questioned.     

Effect of hepatocyte growth factor on methionine adenosyltransferase genes and growth is cell density-dependent in HepG2 cells

Hepatocyte growth factor (HGF) is a potent hepatocyte mitogen but its effect in liver cancer is conflicting. Methionine adenosyltransferase (MAT) is an essential enzyme encoded by two genes (MAT1A and MAT2A), while a third gene (MAT2beta) encodes for a subunit that regulates the MAT2A-encoded isoenzyme. MAT1A is silenced while MAT2A and MAT2beta are induced in hepatocellular carcinoma (HCC). The current work examined expression of HGF/c-met in HCC and whether HGF regulates MAT genes and growth in HepG2 cells. We found the mRNA levels of HGF and c-met are markedly increased in HCC. To study the influence of cell density, HepG2 cells were plated under high-density (HD) or low-density (LD) and treated with HGF (10 ng/ml). Cell density had a dramatic effect on MAT1A expression, being nearly undetectable at LD to a ninefold induction under HD. Cell density also determined the effect of HGF. At HD, HGF increased the mRNA levels of p21 and p27, while lowering the levels of MAT genes, cyclin A, and c-met. At LD, HGF increased the mRNA levels of cyclin A, MAT2A, MAT2beta, and c-met. Consistently, HGF inhibits growth under HD but stimulates growth under LD. HGF induced sustained high ERK activation under HD as compared to LD. In summary, HGF induces genes favoring growth and is mitogenic when HepG2 cells are plated under LD; however, the opposite occurs under HD. This involves cell density-dependent differences in HGF-induced ERK activation. This may explain why HGF is mitogenic only when there is loss of cell-cell contact in vivo.     

Hepatocyte growth factor is necessary for efficient outgrowth of injured peripheral axons in in vitro culture system and in vivo nerve crush mouse model

Hepatocyte growth factor (HGF) is a neurotrophic factor and its role in peripheral nerves has been relatively unknown. In this study, biological functions of HGF and its receptor c-met have been investigated in the context of regeneration of damaged peripheral nerves. Axotomy of the peripheral branch of sensory neurons from embryonic dorsal root ganglia (DRG) resulted in the increased protein levels of HGF and phosphorylated c-met. When the neuronal cultures were treated with a pharmacological inhibitor of c-met, PHA665752, the length of axotomy-induced outgrowth of neurite was significantly reduced. On the other hand, the addition of recombinant HGF proteins to the neuronal culture facilitated axon outgrowth. In the nerve crush mouse model, the protein level of HGF was increased around the injury site by almost 5.5-fold at 24 h post injury compared to control mice and was maintained at elevated levels for another 6 days. The amount of phosphorylated c-met receptor in sciatic nerve was also observed to be higher than control mice. When PHA665752 was locally applied to the injury site of sciatic nerve, axon outgrowth and injury mediated induction of cJun protein were effectively inhibited, indicating the functional involvement of HGF/c-met pathway in the nerve regeneration process. When extra HGF was exogenously provided by intramuscular injection of plasmid DNA expressing HGF, axon outgrowth from damaged sciatic nerve and cJun expression level were enhanced. Taken together, these results suggested that HGF/c-met pathway plays important roles in axon outgrowth by directly interacting with sensory neurons and thus HGF might be a useful tool for developing therapeutics for peripheral neuropathy.     

Functional characterization of human hepatocyte growth factor mutants obtained by deletion of structural domains.

Human hepatocyte growth factor (hHGF) consists of characteristic structural domains. In this study, we prepared mutant proteins lacking each of these domains and examined their biological activities for stimulation of hepatocyte DNA synthesis, inhibition of Meth A cell growth, and induction of MDCK cell dissociation. We also examined their interactions with the c-met/HGF receptor by competition analysis and by analysis of levels of tyrosine phosphorylation. The mutant proteins lacking the N-terminal, the first kringle, or the second kringle domain were not biologically effective and could not compete with hHGF bound to the c-met/HGF receptor. The results indicate that these domains are necessary for the biological activities of hHGF mediated by binding to the c-met/HGF receptor. The mutant proteins lacking the third or fourth kringle domain moderately retained biological activities and the receptor binding. The relative levels of the tyrosine phosphorylation of the c-met/HGF receptor by these mutant proteins correlated well with the relative potencies of the biological activities when compared with that of the wild-type hHGF. The mutant protein lacking the light chain was not effective in the biological activities and tyrosine phosphorylation of the c-met/HGF receptor, but competed with hHGF bound to the c-met/HGF receptor. These results suggest that the heavy chain plays an important role in the interaction of hHGF with the c-met/HGF receptor and that the light chain is further required for the tyrosine phosphorylation of the c-met/HGF receptor.     

Antagonistic effect of NK4 on HGF/SF induced changes in the transendothelial resistance (TER) and paracellular permeability of human vascular endothelial cells

Hepatocyte growth factor/scatter factor (HGF/SF) is a multi-function cytokine that has been shown to regulate the expression of cell adhesion molecules in human endothelial cells. It is also a key cytokine in the development and progression of cancer, particularly during metastasis. NK4 is a variant of HGF/SF that has already been shown to be antagonistic to HGF/SF. This study shows that HGF/SF decreased transendothelial resistance (TER) and increased paracellular permeability in human vascular endothelial cells can that such effects can be inhibited by addition of the NK4 variant. In addition, HGF/SF-stimulated invasion of endothelium by breast cancer cells was inhibited by the addition of NK4. Western blotting revealed that HGF/SF decreased the protein level, and increased tyrosine phosphorylation of ZO-1, but did not cause a change in level of occludin or claudin-1, both molecules involved in tight junction function. RT-PCR revealed that addition of HGF/SF caused no change in signal for claudin-5 or junctional adhesion molecule (JAM), but there was a decrease in the signal for claudin-1. NK4 was able to prevent the decrease in levels of ZO-1 protein by HGF/SF.     

Hepatocyte growth factor activates several transduction pathways in rat pancreatic acini

The receptor of hepatocyte growth factor (HGF), c-met induces different physiological responses in several cell types. Little is known about the role of HGF in exocrine pancreas. However, abnormal HGF signaling has been strongly implicated in pancreatic tumorigenesis and association of HGF with pancreatitis has been demonstrated. We have studied the presence of c-met and activation of their intracellular pathways associated in rat pancreatic acini in comparison with cholecystokinin (CCK) and epidermal growth factor (EGF). C-met expression in rat exocrine pancreas was identified by immunohistochemistry and immunoprecipitation followed by Western analysis. Tyrosine phosphorylation of c-met is strongly stimulated as well as kinase pathways leading to ERK1/2 cascade. HGF, but not CCK or EGF, selectively caused a consistent increase in the amount of p85 regulatory subunit of PI3-K present in anti-phosphotyrosine immunoprecipitates. Downstream of PI3-K, HGF increased Ser473 phosphorylation of Akt selectively, as CCK or EGF did not affect it. HGF selectively stimulated tyrosine phosphorylation of phosphatase PTP1D. HGF failed to promote the well-known CCK effects in pancreatic acini such as amylase secretion and intracellular calcium mobilization. Although HGF shares activation of ERK1/2 with CCK, we demonstrate that it promotes the selective activation of intracellular pathways not regulated by CCK or EGF. Our results suggest that HGF is an in vivo stimulus of pancreatic acini and provide novel insight into the transduction pathways and effects of c-met/HGF in normal pancreatic acinar cells.     

c-met 在大鼠胰岛β细胞INS-1 胰岛素分泌中的作用

目的:从c-met对胰岛β细胞增殖,细胞周期、糖耐受和对GLUT2的表达影响三个方面探讨c-met在胰岛β细胞功能的影响及相关机制。方法:在大鼠胰岛β细胞系INS-1中运用RNA干扰技术(RNAi)抑制HGF的特异性受体c-met蛋白的表达,检测其在正常的生理状况下对成熟的胰岛β细胞增殖以及功能维持的作用。结果:c-met蛋白对成熟的胰岛β细胞的增殖与周期并没有显著影响,但对于β细胞的功能维持具有重要意义。结论:通过调节GLUT2蛋白来维持β细胞的胰岛素分泌功能,有助于进一步阐明HGF/c-met通路在胰岛β细胞功能损伤的分子机制,从而为糖尿病的预防和治疗提供新的理论依据。     

A novel role of hepatocyte growth factor as an immune regulator through suppressing dendritic cell function

Hepatocyte growth factor (HGF) plays an important role in many biological events such as angiogenesis, cell proliferation, anti-fibrosis and antiapoptosis. It is well known that HGF promotes tumor progression and suppresses development of fibrosis after tissue injury. In contrast, its role in immune-mediated disorders has not been fully clarified. In the present study, we examined the role of HGF in Ag-specific immune response using in vitro studies and an experimental model of allergic airway inflammation. We first confirmed that dendritic cells (DCs) expressed the receptor for HGF, c-met, which was not expressed in T cells. Treatment with HGF both in vitro and in vivo potently suppressed DC functions such as Ag-presenting capacity, thus down-regulating Ag-induced Th1- and Th2-type immune responses. Exogenous administration of the HGF expression plasmid into Ag-primed mice markedly suppressed the development of airway eosinophilia and airway hyperresponsiveness, which was induced by Ag inhalation, with suppression of the Ag-presenting capacity of DCs in the lung. HGF exhibited these immunosuppressive effects without up-regulation of IL-10 or TGF-beta. We also found that expression of endogenous HGF in the lung significantly increased following Ag sensitization and inhalation challenges. Finally, neutralization of endogenous HGF in vivo significantly increased airway eosinophilia and airway hyperresponsiveness with up-regulation of the Ag-presenting capacity of DCs in the lung. These results demonstrated a novel, significant, and possibly therapeutic role of HGF as a potent regulator in immune-mediated disorders such as asthma.