摘要
目的探讨靶向CTNNB1的shRNA对人结肠癌SW480细胞的基因下调效应和细胞增殖的抑制作用。方法构建靶向CTNNB1的shRNA载体质粒,然后转染入结肠癌SW480细胞。用RT-PCR和Western blotting方法检测CTNNB1的mRNA和蛋白的表达情况。MTT实验评价转染后各组细胞的增殖情况,并用流式细胞仪检测各组细胞的周期分布和凋亡情况。结果靶向CTNNB1的shRNA能够明显下调CTNNB1 mRNA和蛋白质表达(P<0.05),其抑制率分别是43.87%和45.16%。MTT实验提示CTN组细胞在转染后呈现随着时间延长而进行性增殖抑制的现象。在转染后72h,CTN组的细胞存活率为46.4%,与空白对照组相比有显著性降低(P<0.05)。而流式细胞仪显示CTN组细胞有明显G0/G1周期阻滞和凋亡率增高(P<0.05)。结论靶向CTNNB1的特异性shRNA对结肠癌SW480细胞具有下调CTNNB1基因表达,促进细胞凋亡并且显著抑制细胞增殖的效应。
Objective To observe the inhibitory effect of CTNNB1 gene expression and cell proliferation of the human colon cancer cell line SW480 caused by CTNNB 1-targeted ShRNA. Methods The shRNA against CTNNB1 was constructed and transfected into SW480 cells. The down-regulations of CTNNB1 expres- sions were detected by RT-PCR and Western blotting analysis. The cell proliferation was determined by soft agar colony formation assay. The effect of these shRNAs on cell cycle distribution and apoptosis was examined by flow cytometry. Results The CTNNBI-shRNA efficiently suppressed the expression of CTNNB1 mRNA and pro- tein, P〈0.05. The expression inhibition rates were 43.87% and 45.16% at the mRNA and protein level respec- tively. The MTY assay indicated that the specific shRNA resulted in significant inhibition of cell growth on the culture plates on a time-dependent manner. Seventy-two hours after transfection, the cell viability of CTN group was 46.4%, which was of statistical significance when compared with that of blank control group, P〈0.05. More- over, the cancer cells showed significant G0/G1 arrest and increased apoptosis in the CTN group by flow cytome- try, P〈0.05. Conclusion The specific CTNNBI-shRNA may down-regulate the expression of CTNNB1 gene, increase apoptosis and inhibit the growth of SW480 cells.
出处
《中华普通外科学文献（电子版）》
2011年第2期12-15,共4页
Chinese Archives of General Surgery(Electronic Edition)
基金
广东省科技厅科研基金资助项目(2010B080701038)
作者简介
通信作者：钟诗龙，Email：txzz2001@gmail．com
