摘要
AIM： To investigate the effects of Gli-1 small interference RNA （siRNA） on Huh7 cells, and the change of Bcl-2 expression in Huh7 cells. METHODS： Human hepatocellular carcinoma cells Huh7 were used. Cell viability was analyzed by 3-（4, 5-Dimethylthiazol-2-yl）-2, 5-diphenyl tetrazolium bromide （MTT） assay. The expressions of Gli-1 and Bcl-2 family members were detected by RT-PCR and Western blot. Apoptosis was detected by Flow cytometry using propidium iodide, measured by Hoechst 33258 staining using Advanced Fluorescence Microscopy and caspase-3 enzymatic assay. Cell growth was analyzed after treatment with Gli-1 siRNA and 5-fluorouracil （5-Fu）. RESULTS： Inhibition of Gli-1 mRNA in Huh7 cells through Gli-1 siRNA reduced cell viability. Gli-1 siRNA treatment also induced apoptosis by three criteria, increase in the sub-G1 cell cycle fraction, nuclear condensation, a morphologic change typical of apoptosis, and activation of caspase-3. Gli-1 siRNA was also able to down-regulate Bcl-2. However, Gli-1 siRNA resulted in no significant changes in Bcl-xl, Bax, Bad, and Bid. Furthermore, Gli-1 siRNA increased the cytotoxic effect of 5-Fu on Huh7 cell. CONCLUSION： Down-regulation of Bcl-2 plays an important role in apoptosis induced by Gli-1 siRNA in HCC cells. Combination Gli-1 siRNA with chemotherapeutic drug could represent a more promising strategy against HCC. The effects of the strategies need further investigation in vivo and may have potential clinical application.
AIM: To investigate the effects of Gli-1 small interference RNA (siRNA) on Huh7 cells,and the change of Bcl-2 expression in Huh7 cells. METHODS: Human hepatocellular carcinoma cells Huh7 were used. Cell viability was analyzed by 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay. The expressions of Gli-1 and Bcl-2 family members were detected by RT-PCR and Western blot. Apoptosis was detected by Flow cytometry using propidium iodide,measured by Hoechst 33258 staining using Advanced Fluorescence Microscopy and caspase-3 enzymatic assay. Cell growth was analyzed after treatment with Gli-1 siRNA and 5-? uorouracil (5-Fu). RESULTS: Inhibition of Gli-1 mRNA in Huh7 cells through Gli-1 siRNA reduced cell viability. Gli-1 siRNA treatment also induced apoptosis by three criteria,increase in the sub-G1 cell cycle fraction,nuclear condensation,a morphologic change typical of apoptosis,and activation of caspase-3. Gli-1 siRNA was also able to down-regulate Bcl-2. However,Gli-1 siRNA resulted in no significant changes in Bcl-xl,Bax,Bad,and Bid. Furthermore,Gli-1 siRNA increased the cytotoxic effect of 5-Fu on Huh7 cell. CONCLUSION: Down-regulation of Bcl-2 plays an important role in apoptosis induced by Gli-1 siRNA in HCC cells. Combination Gli-1 siRNA with chemotherapeutic drug could represent a more promising strategy against HCC. The effects of the strategies need further investigation in vivo and may have potential clinical application.
基金
The National Science Foundation of China, No. 30672053
