摘要
目的探讨靶向Bcl-XL的shRNA对人结肠癌Lovo细胞的基因下调效应和细胞生长的抑制作用。方法构建靶向Bcl-XL的shRNA载体质粒,然后转染入结肠癌Lovo细胞。用RT-PCR和Western Blot方法检测Bcl-XL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Bcl-XL的shRNA能够明显下调Bcl-XL mRNA和蛋白质表达(P<0.05),其抑制率分别是41.24%和45.26%。MTT实验显示转染特异性shRNA的BCL实验组细胞在转染后呈现随着时间延长而进行性生长抑制的现象。在转染后72 h,BCL组的细胞存活率为47.8%,与空白对照组相比有显著性降低,该组细胞还显示出明显的G_0/G_1阻滞和明显增高的凋亡率(P<0.05)。结论靶向Bcl-XL的特异性shRNA对结肠癌Lovo细胞具有下调Bcl-XL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 shRNA interference against Bcl-XL on gene expression and cell proliferation of the human colon cancer cell line Lovo.Methods The shRNA plasmid vector against Bcl-XL was constructed and transfected into Lovo cells with Lipofectamine^(TM) 2000.Down-regulation of Bcl-XL expression was detected by RT-PCR and western blot analysis.Cell proliferation inhibition was determined by MTT assay.The effect of these shRNAs on cell cycle distribution and apoptosis was examined by flow cytometry.Results The shRNA vector targeted against Bcl-XL was successfully constructed and efficiently suppressed the expression of Bcl-XL mRNA and protein(P<0.05).The expression inhibition rates were 41.24%and 45.26%at mRNA and protein level respectively.MTT assay indicated that the specific shRNA resulted in significant inhibition of cell growth on a time-dependent manner.At 72h after transfection,the cell viability of BCL group was 47.8%.significantly lower than that of blank control group(P<0.05).Moreover,the cancer cells showed significant G_0/G_1 arrest and increased apoptosis at 72 hour after transfection. Conclusions The specific shRNAs targeted against Bcl-XL can down-regulate the expression of Bcl-XL gene,increase apoptosis and inhibit the growth of Lovo cells.
出处
《消化肿瘤杂志（电子版）》
2009年第2期97-100,共4页
Journal of Digestive Oncology(Electronic Version)
基金
广东省科技厅科研基金资助项目(2007B030702012)
