Oct4基因沉默对结直肠癌细胞凋亡的诱导作用及机制研究

目的 探讨Oct4基因沉默对人结直肠癌SW480细胞凋亡的影响及其可能机制.方法 将对数生长期SW480细胞分为3组.①空白对照(Con)组:不转染病毒；②阴性对照(NC)组:转染阴性对照病毒；③RNA干扰(RNAi)组:转染Oct4-shRNA慢病毒载体.采用实时荧光定量PCR检测Oct4 mRNA表达；采用Western blotting检测Oct4、p-Akt 蛋白表达；采用流式细胞仪测定Oct4+及CD44+细胞比例变化及肿瘤细胞凋亡情况.结果 与NC组比较,RNAi组Oct4 的mRNA(分别为1.00和0.19±0.02)和蛋白(分别为0.032±0.004和0.007±0.001)表达量均有明显下降(P＜0.0l),Oct4+细胞比例(分别为91.53％和10.70％)和CD44+细胞比例(分别为59.69％和23.58％)明显减少,p-Akt蛋白表达明显受抑(0.11+0.03和0.03±0.01),细胞凋亡明显增加(分别为l2.1％±1.8％和43.2％±4.5％,P＜0.01).结论 沉默Oct4基因可明显减少SW480细胞中Oct4+及CD44+细胞比例,增加细胞凋亡,其作用可能与PI3K/Akt通路有关.

Apoptosis induced by Octamer 4 gene silencing and its potential mechanism in colorectal cancer

Objective To investigate the effect of Oct4 gene silencing on apoptosis in human colorectal cancer SW480 cell lines and its possible mechanism.Methods SW480480 cells at the late logarithmic phase were divided into 3 groups.(1) control group(Con): no virus transfection;(2) negative control group(NC): transfected with negative control viruse;(3) RNA interference group(RNAi): transfected with Oct4-shRNA lentiviral vector plasmids.In the above three groups,the Oct4 mRNA level was assayed by real-time quantitative PCR,the protein levels of Oct4 and p-Akt were determined by Western blotting,and the proportion of Oct4+ and CD44+ cells and apoptosis of the tumor were analyzed by flow cytometry.Results Compared with those in the NC group,the Oct4 mRNA levels(1.00 vs 0.19±0.02),the Oct4 protein levels(0.032±0.004 vs 0.007±0.001),the p-Akt protein levels(0.11±0.03 vs 0.03±0.01),the proportion of Oct4+ cells(91.53% vs 10.70%) and the proportion of CD44+ cells(59.69% vs 23.58%) in the RNAi group were significantly reduced,while apoptosis of cells in the RNAi group was significantly increased(43.2%±4.5% vs 12.1%±1.8%,P<0.01).Conclusion Oct4 gene silencing could significantly reduce the proportion of Oct4+ and CD44+ cells,and obviously increase the rate of apoptosis in SW480 cells,and the latter may be related to the PI3K/Akt pathway.