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目的:探讨小干扰RNA降低环氧化酶-2(cyclooxygenase-2,COX-2)基因对肝癌细胞HepG2增殖与细胞外信号调节蛋白激酶(ERK)表达的影响.方法:将人肝癌细胞株HepG2细胞分为4组:COX-2 siRNA干预组、阴性对照siRNA组、空脂质体组、空白对照组.脂质体转染法将COX-2 siRNA转染入HepG2细胞;MTT法测定转染后24,48,72h的增殖抑制率;流式细胞仪测定转染后24h的细胞周期分布;半定量RT-PCR与Western blot检测转染后24h肝癌细胞COX-2和ERK1/2的表达.结果:COX-2 siRNA干预组的增殖抑制率明显高于阴性对照组和空脂质体转染组(67.08%vs2.45%,1.56%,均P<0.01);COX-2 siRNA干预组G1期细胞明显增多,与空白组、空脂质体组及阴性对照组相比,均具有显著差异(72.80%vs50.27%,50.97%,53.13%,均P<0.05);RT-PCR显示:COX-2siRNA干预组COX-2,ERK1,ERK2 mRNA表达(0.58,0.32,0.48)明显降低,与空白组(0.93,0.76,0.72)、空脂质体组(0.89,0.64,0.67)及阴性对照组(0.83,0.71,0.65)比较,差异具有统计学意义(均P<0.05),Western blot显示以上各项指标表达趋势与RT-PCR相同.结论:小干扰RNA可降低肝癌细胞HepG2的COX-2基因表达,抑制肝癌细胞的生长.COX-2促进肝癌生长可能与ERK1/2通路调控有关.
Objective: To investigate the effect of small interfering RNA (RNAi) on cyclooxygenase-2 (COX-2) gene on the proliferation of HepG2 cells and the expression of extracellular signal-regulated protein kinase (ERK) The cells were divided into 4 groups: COX-2 siRNA intervention group, negative control siRNA group, empty liposome group and blank control group.COX-2 siRNA was transfected into HepG2 cells by lipofection method and transfected into HepG2 cells by MTT assay 24 h, 48 h and 72 h after transfection. The cell cycle distribution was determined by flow cytometry at 24 h after transfection. The expression of COX-2 and ERK1 / 2 was detected by semi-quantitative RT-PCR and Western blot. Results: The proliferation inhibition rate of COX-2 siRNA group was significantly higher than that of negative control group and empty liposome transfection group (67.08% vs 2.45%, 1.56%, both P <0.01) (72.80% vs50.27%, 50.97%, 53.13%, both P <0.05). Compared with the blank group, the empty liposome group and the negative control group, COX-2siRNA intervention group COX-2, ERK1, ERK2 mRNA expression (0.58,0.32,0.48) was significantly lower than the blank group (0.93,0.76,0.72), empty liposome group (0.89,0.64,0.67) and negative control Group (0.83, 0.71, 0.65 (All P <0.05) .Western blot showed that the expression of all the above indexes was the same as that of RT-PCR.Conclusion Small interfering RNA can reduce the expression of COX-2 in HepG2 cells and inhibit the expression of COX-2 in hepatocellular carcinoma cells COX-2 promotes the growth of hepatocellular carcinoma may be related to the regulation of ERK1 / 2 pathway.