Effect of interfering RNA targeting plasminogen activator inhibitor-1 on the biological characters of hepatic stellate cells / 中华消化杂志

Objective To investigate the effect of synthetic small interfering RNA (siRNA) targe- ting plasminogen activator inhibitor-1 (PAI-1) on the biological characters of hepatic stellate cells (HSCs).Methods Synthetic siRNA targeting PAI-1 was transfected into HSC-T6 by lipofectamine package.Negative siRNA transfection and no transfection were used as negative control and blank con- trol respectively.After incubation with siRNA,total RNA and protein of HSC-T6 cells were extracted. The expression of PAI-1 gene was detected by reverse transcription-polymerase chain reaction and immuno- cytofluorescent.The proliferation of HSC-T6 was determined using MTT assay.HSC-T6 cycle and apoptosis were measured by flow cytometry.Content of typeⅠandⅢcollagen in the supernatants were determined by enzyme linked immunosorbent assay.Results The expressions of PAI-1 mRNA and pro- tein were markedly down-regulated in siRNA-transfected HSC-T6.The proliferation of HSC-T6 was inhibited by transfection of siRNA.The level of typeⅠcollagen in the supernatants decreased by (20.71?8.40)% and (37.97?6.40)%,repectively(F=42.69,P=0.0001) compared to those in the negative control at 48 and 72 hours.The level of typeⅢcollagen decreased by (35.98?4.60)% (F=105.52,P= 0.0001) at 72 hours.Conclusion Inhibition of PA-1 by siRNA may have a potential effect in prevention and treatment of hepatic fibrosis by inhibiting proliferation,promoting apoptosis of stellate cells.Therefore,it decreases the synthesis and the secretions of typeⅠandⅢcollagen.

Effect of reduced glutathione on the proliferation,oxidative stress and transforming growth factor?1 expression of human hepatic stellate cells / 中华消化杂志

Objective To investigate the impact of reduced glutathione(GSH) on the prolifera- tion,oxidative stress and transforming growth factor?1(TGF-?1) expression of human hepatocytes and hepatic stellate cells(HSCs)(LX-2 cell line).Methods Human hepatocytes and HSCs were incubated with various concentrations of GSH(0.5—50 mmol/L or 0.5—10 mmol/L).The effects of GSH on the proliferation of hepatocytes and HSCs were studied by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphennyhera- zolium bromide colorimetric assay.Human hepatocytes and HSCs were co-cultured with GSH and ferric nitrilotriacetic acid,superoxide dismutase (SOD) activity and malondialdehyde (MDA) contents were detected.HSCs were incubated with high(5.0 mmol/L),media(2.5 mmol/L) and low (0.5 mmol/L) concentrations of GSH,the expressions of TGF-?1 mRNA and protein were detected by ELISA and real- time PCR.Results In concentration ranged from 2.5 to 10 mmol/L,the GSH could promote the pro- liferation of hepatocytes but no HSCs,significantly increased the activity of SOD and decrease the con- tents of MDA in hepatocytes and HSCs,and inhibited the expression of TGF-?1 in HSCs.Conclusions GSH can not only promote the proliferation of hepatocytes,but also protect hepatocytes and HSCs from oxidative stress,and inhibit the secretion of TGF-?1 in HSCs.GSH may play a role in hepatocellular protection,antioxidation and anti-fibrosis.