Objective To investigate the effect of Derlin-1 gene inhibition mediated by RNA interference on the proliferation of cervi-cal cancer HeLa cells in vitro. Methods Recombinant plasmids for RNA interference of Derlin-1 gene were constructed and transfect-ed into HeLa cells via lipofectamine 2000, which were named as siRNA-Derlin-1 group. Meanwhile, HeLa cells transfected with empty plasmids were chosen as empty plasmid group, and the untreated HeLa cells were selected as blank control group. The mRNA and pro-tein expression of Derlin-1 in the stably transfected cells was detected by RT-PCR and Western blot. The changes of cell proliferation were detected by MTT assay. Enzyme-linked immunosorbent assay( ELISA) was employed to observe the effect of transfection on the apoptosis. The expression of caspase-3 in the transfected cells was also detected by RT-PCR and Western blot. Results Compared with empty plasmids group and blank control group, the mRNA and protein expression levels of the HeLa cells in Derlin-1-siRNA group were significantly decreased(P<0. 05),and the cell proliferation was inhibited(P<0. 05). The apoptosis cells were increased in Der-lin-1-siRNA group and the expression of caspase-3 was effectively promoted compared with empty plasmids group and blank control group(P<0. 05). Conclusion Derlin-1 gene could be highly expressed in cervical cancer and RNA interference-mediated Derlin-1 silencing can strongly inhibit the proliferation and induce apoptosis in HeLa cells.%目的 采用RNAi技术抑制Derlin-1基因的表达,探讨其表达降低后对宫颈癌HeLa细胞生长的影响. 方法 合成针对Derlin-1基因的干扰片段,构建重组质粒,用脂质体法将重组质粒转染人宫颈癌HeLa细胞,并筛选出稳定表达株,作为siRNA-Derlin-1组,选取空质粒转染后的HeLa细胞为空质粒组,未做任何处理的HeLa细胞为空白对照组,采用RT-PCR检测重组质粒对HeLa细胞Derlin-1 mRNA的影响,Western blot检测其对Derlin-1蛋白表达的影响,MTT法分析其对细胞增殖的影响,ELISA法检测其对细胞凋亡的影响,同时检测凋亡因子caspase-3的表达. 结果 与空质粒组和空白对照组相比,siR-NA-Derlin-1组HeLa细胞中Derlin-1 mRNA和蛋白的表达显著降低(P<0. 05),MTT法显示siRNA-Derlin-1组宫颈癌HeLa细胞增殖能力显著低于空质粒组和空白对照组(P<0. 05),同时siRNA-Derlin-1组凋亡细胞增加,凋亡核心蛋白caspase-3表达增加. 结论 抑制内质网相关降解蛋白Derlin-1的表达能有效降低宫颈癌HeLa细胞的增殖,促进凋亡.
展开▼
