敲低膜联蛋白A5对人胃癌细胞系MKN-45增殖的作用

Annexin A5 Knockdown Prompts Proliferation of Gastric Cancer MKN-45 Cells

目的探讨膜联蛋白A5低表达对人胃癌MKN-45细胞增殖和凋亡的影响。方法将胃癌MKN-45细胞分为siRNA干扰组、阴性对照组和空白对照组。采用RNA干扰技术将靶向膜联蛋白A5的siRNA和阴性对照siRNA脂质体转染法转染细胞,转染48 h后采用qRT-PCR和Western blot法分别在mRNA水平和蛋白水平进行抑制效率的鉴定,空白对照组不予任何处理。应用MTT、平板克隆形成法检测膜联蛋白A5低表达对胃癌MKN-45细胞增殖的影响,流式细胞术检测细胞凋亡情况。Western blot法检测增殖核抗原PCNA和周期相关蛋白CyclinD1的表达。结果靶向膜联蛋白A5的siRNA转染后,胃癌MKN-45细胞中膜联蛋白A5的表达较阴性对照和空白对照组明显降低(P<0.05)。MTT和克隆形成实验可见siRNA干扰组细胞增殖活力和集落形成能力较阴性对照和空白对照组显著增高(P<0.05);流式细胞术显示细胞凋亡率无明显改变;PCNA和Cyclin D1的表达显著上调。结论膜联蛋白A5低表达能促进胃癌细胞的增殖。

Objective To investigate the effect of annexin A5 knockdown on the proliferation and apoptosis of gastric cancer MKN-45 cells. Methods The gastric cancer MKN-45 cells were divided into three groups： siRNA interference group transfected with siRNA targeting to ANXA5, negative control group transfected with scrambled siRNA and blank group without any treatment. After 48 h of transfection, qRT-PCR and Western blot were used to detect the suppression of ANXA5 on both mRNA and protein levels. MTT and clone formation assays were used to detect the effect of low annexin A5 expression on the proliferation of gastric cancer MKN-45 cells. Cell apoptosis was analyzed by flow cytometry. Western blot was performed to detect the expression of PCNA and Cyclin D1. Results After transfection with siRNA targeting to annexin A5, the expression of annexin A5 in gastric cancer MKN-45 cells was significantly lower than those in the negative and blank control groups（P〈0.05）. The results of MTT and clone formulation assays showed that cell proliferation activity and colony-forming ability of siRNA interference group were significantly higher than those of negative and blank control groups（P〈0.05）, whereas cell apoptosis was not affected statistically（P〉0.05）. The expression of PCNA and Cyclin D1 were significantly increased. Conclusion Low annexin A5 expression could promote the proliferation of gastric cancer cells.