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目的采用RNA干扰技术(RNAi)沉默癌胚抗原相关细胞黏附分子1(CEACAM1)基因的表达,观察沉默效果及沉默CEACAM1表达后对SHG44人胶质瘤细胞增殖和凋亡的影响。方法设计并化学合成针对CEACAM1的3对小干扰RNA(siRNA),脂质体法瞬时转染SHG44细胞。流式细胞术(FCM)检测转染效率,采用实时定量PCR(qRT-PCR)检测siRNA转染前后SHG44细胞中CEACAM1 mRNA的表达,Western blot法检测CEACAM1蛋白的表达。CCK-8法检测SHG44细胞细胞增殖活性,annexin V-FITC/PI染色结合FCM检测SHG44细胞凋亡,Western blot法检测cleaved caspase-3和裂解型多聚腺苷酸二磷酸核糖聚合酶(cleaved PARP)的变化。结果 CEACAM1 siRNA转染效率达85%。与空白对照组和阴性对照组比较,qRT-PCR及Western blot结果显示,3组特异性siRNA转染48 h后,在mRNA和蛋白水平上CEACAM1的表达均降低,以CEACAM1-siRNA3效果最明显。siRNA组SHG44细胞的增殖能力较正常对照组明显下降,凋亡细胞比例增加。沉默CEACAM1表达可以上调cleaved caspase-3和cleaved PARP的表达。结论沉默CEACAM1基因表达可有效抑制SHG44胶质瘤细胞的增殖,促进细胞凋亡。
Objective To silence the expression of carcinoembryonic antigen-related cell adhesion molecule 1 (CEACAM1) by RNAi and observe the effect of silencing CEACAM1 silencing on the proliferation and apoptosis of SHG44 human glioma cells. Methods Three pairs of small interfering RNA (siRNA) targeting CEACAM1 were designed and synthesized. SHG44 cells were transiently transfected by liposome. The transfection efficiency was detected by flow cytometry (FCM). The expression of CEACAM1 mRNA in SHG44 cells was detected by real-time quantitative PCR (qRT-PCR). The expression of CEACAM1 protein was detected by Western blot. The proliferation of SHG44 cells was detected by CCK-8 assay. The apoptosis of SHG44 cells was detected by annexin V-FITC / PI staining and FCM. Cleaved caspase-3 and cleaved PARP )The change. Results CEACAM1 siRNA transfection efficiency of 85%. Compared with the blank control group and the negative control group, the results of qRT-PCR and Western blot showed that the expression of CEACAM1 at the mRNA and protein levels decreased after 3 hours of specific siRNA transfection and the effect of CEACAM1-siRNA3 was the most obvious. siRNA group SHG44 cell proliferation than the normal control group decreased significantly, the proportion of apoptotic cells increased. Silencing CEACAM1 expression up-regulates cleaved caspase-3 and cleaved PARP expression. Conclusion Silencing CEACAM1 gene expression can effectively inhibit the proliferation of SHG44 glioma cells and promote apoptosis.