[Small interfering RNA-mediated nuclear factor-kappaB P65 suppression induces apoptosis of hepatic carcinoma SMMC-7721 cells].
Chen Huang, Jiayi Yao, Zongfang Li, Liying Liu, Lei Ni, Tusheng Song
Abstract
Chen Huang, Jiayi Yao, Zongfang Li, Liying Liu, Lei Ni, Tusheng Song
Abstract
OBJECTIVE: To study the mechanism of hepatic carcinoma cell apoptosis induced by small interfering RNA (siRNA)-mediated nuclear factor-kappaB (NF-kappaB) P65 silencing. METHODS: Hepatic carcinoma SMMC-7721 cells were exposed to liposome-mediated transfection with NF-kappaB P65 siRNA synthesized by in vitro transcription, and the cells with empty liposome transfection and those without particular treatment served as the control groups. The expression of NF-kappaB P65 in the cells was detected by Western blotting, the cell viability examined by MTT assay, and the cell apoptosis assessed by flow cytometry. Immunohistochemistry was used to examine the expressions of Bcl-2 and Bax. RESULTS: siRNA transfection significantly inhibited the expression of NF-kappaB P65 in SMMC-7721cells, with inhibition rates of 64.74% compared with the untreated cells and of 34.52% compared with the liposome-treated cells. The siRNA-treated SMMC-7721 cells also exhibited significant decrease in cell proliferation by 33.39% and 27.23% in comparison with the untreated and liposome-treated cells, respectively. NF-kappaB P65 siRNA induced obvious cell apoptosis with down-regulated Bcl-2 and up-regulated Bax expressions. CONCLUSION: NF-kappaB p65 siRNA can induce SMMC-7721 cell apoptosis via the Bcl-2/Bax pathway.
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OBJECTIVE: To study the mechanism of hepatic carcinoma cell apoptosis induced by small interfering RNA (siRNA)-mediated nuclear factor-kappaB (NF-kappaB) P65 silencing. METHODS: Hepatic carcinoma SMMC-7721 cells were exposed to liposome-mediated transfection with NF-kappaB P65 siRNA synthesized by in vitro transcription, and the cells with empty liposome transfection and those without particular treatment served as the control groups. The expression of NF-kappaB P65 in the cells was detected by Western blotting, the cell viability examined by MTT assay, and the cell apoptosis assessed by flow cytometry. Immunohistochemistry was used to examine the expressions of Bcl-2 and Bax. RESULTS: siRNA transfection significantly inhibited the expression of NF-kappaB P65 in SMMC-7721cells, with inhibition rates of 64.74% compared with the untreated cells and of 34.52% compared with the liposome-treated cells. The siRNA-treated SMMC-7721 cells also exhibited significant decrease in cell proliferation by 33.39% and 27.23% in comparison with the untreated and liposome-treated cells, respectively. NF-kappaB P65 siRNA induced obvious cell apoptosis with down-regulated Bcl-2 and up-regulated Bax expressions. CONCLUSION: NF-kappaB p65 siRNA can induce SMMC-7721 cell apoptosis via the Bcl-2/Bax pathway.
Key concepts: Transfection, Small interfering RNA, Apoptosis, Viability assay, Molecular biology, Gene silencing, Flow cytometry, MTT assay