| Objective:To isolate HSC from rat live and culture in vitro, investigate the effect of tryptase on HSC and the role played by tryptase during the development of liver fibrosis.Methods:Primary rat hepatic stellate cells (HSC) were isolated from healthy rat liver, cultured in vitro and identified by cell morphology,immunocytochemistry and fluorescence microscope. The identified HSC were used in this study. The cultured HSC were randomly divided into different concentrations of tryptase-treated groups(1ng/ml,10ng/ml,100ng/ml) and control group(0ng/ml). The cell proliferation were examined by methylthio tetrazole(MTT) after 36 hours. The expressions of collagen I mRNA,protease activated receptor-2(PAR-2) mRNA and peroxisome proliferated-activated receptor gamma ( PPAR-γ) mRNA were examined by semi-quantitive reverse transcriptase polymerase chain reaction (RT-PCR) after 36 hours.Resμlts:1,Immunoctyochemistry assay showed that primary cultured HSC had positive staining of desmin. The cultured cells showed fusiform shape under microscope. Blue-green fluorescent can be observed in HSC under condition of 328nm wavelength. These data proved that the cultured cells were HSC.2,Tryptase stimulated HSC proliferation in dose-depended manner at the range of 1ng/ml to 100ng/ml. The proliferation of cultured cell stimulated with 100ng/ml tryptase was 1.64-fold higher than that of control group(P<0.05).3,Tryptase-treated HSC express higher PAR-2 mRNA and collagen I mRNA than control HSC. These effects were showd in dose-depended manner. the expression of PAR-2 mRNA and collagen I mRNA were 1.32 fold and 1.29 fold respectively compared with control group. The expression of PPAR-γmRNA was significant less than that of control group(P<0.05).Conclusion:1,Tryptase could stimulate HSC proliferation , stimulate the expression of PAR-2,collagen I and suppress the expression of PPAR-γ. These results suggest that tryptase excreted by the mast cells has effect on the occurrence and development of liver fibrosis. 2,Tryptase promotes HSC proliferation and activation through the PAR-2 path. |
