Effects Of Hepatocyte Growth Factor On TGF-β1 Triggered Tubular Epithelial-Myofibroblast Transdifferentiation And Expression Of CTGF In Vitro

OBJECTIVE: Renal interstitial Fibrosis is believed to be the reliable andimportant hallmark in various chronic renal diseases which eventually lead to end-stage renal failure. As a key profibrogenic cytokine ,TGF- β1 is involved in the pathogenesis of renal interstitial fibrosis. TGF- β1 may induce tubular epithelial-myofibroblast transdifferentiation, which plays an important role in renal interstitial fibrosis. Hepatocyte growth factor, a multifunctional cytokine, is a negative regulator of organ fibrosis(such as liver cirrohsis and pulmonary fibrosis). The object of our experiment is to obser/e the effects of hepatocyte growth factor on TGF- 3 1 triggered tubular epithelial-myofibroblast transdifferentiation and on the expression of connective tissue growth factor.METHODS: Cultured NRK52E cells were divided into 8 groups which include control group, HGF-treated with different doses, TGF-β1 -treated group , TGF-β1 and HGF-control with different doses. The morphology of transdilferentiate tubular cells was observed under phase-contrast microscopy and scanning electron microscopy, α- SMA was assessed by immunohistochemistry and semiquantified by mean intergrated opiticaldensity(IOD). The level of fibronectin(FN) in the culture supernatant was measured by ELISA . CTGFmRNA expression was examined by RT-PCR .RESULTS: Dose of 10ng/ml of TGF-β1 caused transdifferentiation ofNRK52E cells into myofibroblast-like cells. Scanning electron microscopy and phase-cntrast microscopy identified TGF-β1-induced morphological changes as a loss of apical-basal polarity and microvilli, cell hypertrophy and the development of an elongated appearance. Phenptypically, TGF- β 1-induced TEMT was characterized by expression of α-SMA shown by irnrnunohistochemistry. TGF-β1 was also shown to stimulate the secretion of FN in cultured supernatant and the CTGFmRNA expression of NRK52E cells. There was no significant difference between HGF-treated groups and control in α-SMA immunostaining and the level of FN, except that CTGFmRNA expression was slightly increased in HGF-treated groups . The addition of HGF inhibited TGF-β1-induced TEMT, the secfetion of FN and the CTGF expression of NRK52E cells. There was a significant correlation between the expression of CTGF and the expression of α-SMA.CONCLUSIONS: 1. TGF- β1 could induce tubular epithelial-myofibroblast transdifferentiation and increased the level of FN in culturesupernatant and the expression of CTGF. The results suggest that tubularepithelial to myofibroblast transdifTerentiation may play an important role inthe pathogenesis of renal interstitial fibrosis .2. HGF down-regulated TEMT and FN secretion triggered by TGF-β1, whichimplies that HGF could participate in renal interstitial fibrosis as a negativeregulator.3. The negative regulation of transdifferentiation of HGF may be partially achieved by attenuation of CTGF expression.