摘要
目的探讨HBx小发夹RNA(shRNA)对HepG2.2.15细胞中基质金属蛋白酶2(MMP-2)表达的影响。方法采用psiHBV/X质粒转染HepG2.2.15细胞,反转录PCR评估沉默效率;MTT法检测转染后HepG2.2.15细胞增殖情况;实时定量PCR(qRT-PCR)检测MMP-2 mRNA表达;Western blot法检测MMP-2蛋白表达的变化。结果反转录PCR检测HBx基因的沉默效率为53.6%;MTT检测结果显示转染24、48、72 h后HepG2.2.15细胞的增殖受抑制,与对照组比较,差异有统计学意义(P<0.05);psiHBV/X质粒转染HepG2.2.15细胞后,qRT-PCR和Western blot检测结果显示,HepG2.2.15细胞中MMP-2基因的mRNA水平和蛋白水平表达分别有不同程度的下调,与对照组比较差异有统计学意义(P<0.05)。结论 RNA干扰技术抑制HepG2.2.15细胞中HBx基因的表达,可抑制细胞增殖,下调HepG2.2.15细胞中MMP-2的表达。
Objective To observe the effects of HBx shRNA on MMP-2 expression in human hepatocellular carcinoma cell HepG2.2. 15. Methods HepG2.2. 15 cells were transfected with psiHBV/X plasmid using UpofectamineTM 2000. The silencing efficacy was evaluated by reverse transcription PCR （RT-PCR） detecting the expression of HBx gene. The proliferation of HepG2.2.15 cells was examined by MTT assay. The expression levels of MMP-2 mRNA and protein were assayed by real-time quantitative PCR （qRT-PCR） and Western blotting, respectively. Results RT-PCR showed that the efficiency of RNA interference of HBx gene was 53.6%. Ml-r detection revealed that the cell proliferation was inhibited at 24, 48, and 72 hours after the transfection （0.388 ± 0.087, 0.623 ± 0.016, 0.997 ±0.036） and the differences had statistically significance as compared with the control group （0.436 ±0.027, 0.731± 0.017, 1. 105 ± 0.051 ） （ P 〈 0.05）. The qRT-PCR and Western blotting showed that the expression of MMP-2 decreased at both mRNA and protein levels, and the differences had statistically significance as compared with the control group （ P 〈 0.05 ）. Conclusion HBx gene RNA interference can inhibit the cell proliferation and down-regulate the expression of MMP-2 in HepG2.2. 15 cells.
出处
《细胞与分子免疫学杂志》
CAS
CSCD
北大核心
2014年第4期371-374,共4页
Chinese Journal of Cellular and Molecular Immunology
基金
贵州省联合基金(黔科合J字LKZ[2011]33号)
遵义市红花岗区科技基金[(2010)16]
