Effects Of Exogenous Transforming Growth Factor-β1on Early Expression Of Smad3and Smad7in Hepatic Stellate Cell

Objective To investigate the effects of exogenous transforming growth factor-β1(TGF-β1) on the expression of TGF-β/Smad in hepatic stellate cell (HSC) of rat.Methods (1) HSCs were treated with/without exogenous TGF-β1(10ng/ml), and the mRNA expression of factors in TGF-β/Smad signaling pathway were detected by Real Time PCR at2h.(2)The same method was used to detect the mRNA expression of Smad7induced by exogenous TGF-β1at different time points in HSCs.(3)The negative control plasmid (ctrl) and siRNA-Smad3plasmid (siRNA-Smad3) were respectively transfected into HSCs, according to whether or not the two groups were exposed to exogenous TGF-β1(10ng/ml), they were divided into two parts:(+),(-), the expressions of Smad3and Smad7mRNA were detected by Real Time PCR.(4)Western-blot was used to detect the protein synthesis of Smad3or Smad7at different time points in HSCs.Results (1)Exogenous TGF-β1up-regulated Smad7expression obviously (2.990±0.101, t=-33.962,P=0.001), but had no effect on the mRNA expressions of TGF-βRⅠ, TGF-βRⅡ, Smad3, Smad4and Smad6(P>0.05).(2) After treated by exogenous TGF-β1, Smad7mRNA expression level increased and reached its peak at2h (2.99folds versus control), and it slowly declined.(3)The expression of Smad3mRNA decreased in siRNA-Smad3group, compared with ctrl (0.532±0.169, t=4.810,P=0.041). The mRNA expression of Smad7decreased in siRNA-Smad3(-) group and siRNA-Smad3(+) group, compared with those in ctrl(-) and ctrl(+)(F=9.787,P=0.02).(4) No significant difference about the protein expression of Smad3was found at different time points (P>0.05). But the protein expression of Smad7was time-dependent in accord with the results of PCR.Conclusions Exogenous TGF-β1could increase the high expression of Smad7quickly in an early phase; Smad3is probably a crucial regulator for increasing smad7expression.