Gli-1 siRNA induced apoptosis in Huh7 cells.
OBJECTIVE
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.


CONCLUSIONS
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.
Chen
Xi-Lin
XL
Department of Hepatobiliary Surgery, The First Affiliated Hospital of Sun Yat-Sen University, Guangzhou 510080, Guangdong Province, China.
Cao
Liang-Qi
LQ
She
Miao-Rong
MR
Wang
Qian
Q
Huang
Xiao-Hui
XH
Fu
Xin-Hui
XH
D017209:Apoptosis / Q000502:physiology*; D006528:Carcinoma, Hepatocellular / Q000473:pathology* / Q000628:therapy; D045744:Cell Line, Tumor; D019254:Genes, bcl-2 / Q000502:physiology; D015316:Genetic Therapy / Q000379:methods; D022781:Hepatocytes / Q000166:cytology / Q000502:physiology*; D006801:Humans; D008113:Liver Neoplasms / Q000473:pathology* / Q000628:therapy; D034741:RNA, Small Interfering / Q000494:pharmacology; D014157:Transcription Factors / Q000235:genetics*; D000071676:Zinc Finger Protein GLI1
C400551:GLI1 protein, human; D034741:RNA, Small Interfering; D014157:Transcription Factors; D000071676:Zinc Finger Protein GLI1