Tetrandrine inhibits activation of rat cultured hepatic stellate cell and signaling of transforming growth factor β

Objective To investigate the effect of various concentrations of tetrandrine on activation of cultured quiescent rat hepatic stellate cell(HSC) and signaling of transforming growth factor β(TGF-β).Methods HSC was isolated from rat by in situ perfusion of livers and 18% Nycondenz gradient centrifugation,and primarily cultured on uncoated plastic for 48 h. Then, the HSC was subjected to tetrandrine (0.25 mg/L, 0.5 mg/L, 1.0 mg/L, 2.0 mg/L, respectively) treatment. Cell morphology was observed under inverted microscope. α-smooth muscle actin(α-SMA) expression was analyzed by immunocytochemical staining and image analysis system. Laminin(LN) and type Ⅲ procollagen (PCⅢ) in the supernatant were determined by radioimmunoassay. TGF-β_1 mRNA, Smad 7 mRNA and Smad 7 protein were analyzed with reverse transcription polymerase chain reaction (RT-PCR) and Western blot. Results Tetrandrine in the concentration of 0.25-2.0 mg/L could prevent morphological transformation of HSC from quiescent phenotype to the activated form whereas the α-SMA、LN and PCⅢ expressions were inhibited. RT-PCR showed decreased expression of TGF-β_1 mRNA with an inhibition rate of 87.85%(P0.01). It was significantly negative correlating with increased Smad 7 mRNA(r=-0.755, P0.01),which was 2.4-5.8 times that of the control, and up-regulation of Smad 7 protein by tetrandrine(1.0 mg/L) was confirmed by Western blot as well.Conclusions Tetrandrine exerts direct inhibiting effect on the activation of cultured rat hepatic stellate cells,and upregulates the expression of Smad7 which in turn blocks the expression and signaling of TGF-β_1.