胰岛素样生长因子结合蛋白7过表达对人乳腺癌细胞系-7增殖的影响及其机制研究

Effect of IGFBP7 overexpression on proliferation of breast cancer cell line MCF-7

目的探究过表达胰岛素样生长因子结合蛋白7(IGFBP7)对人乳腺癌细胞系-7(MCF-7)增殖的影响及其机制。方法采用LipofectamineTM2000将p IRES2-Zs Green1-IGFBP7质粒或p IRES2-Zs Green1空白质粒转染进MCF-7乳腺癌细胞,并用荧光显微镜鉴定细胞转染;实时荧光定量聚合酶链反应(Real-time PCR)对转染48 h后IGFBP7蛋白进行定量;细胞凋亡实验检测转染24、48和72 h后各组细胞增殖情况;Western bolt检测转染48 h后各组细胞(蛋白激酶B/雷帕霉素靶蛋白)(AKT/m TOR)信号通路相关蛋白的表达。结果 Real-time PCR的检测结果提示经过IGFBP7转染的细胞IGFBP7 m RNA表达水平明显升高;细胞凋亡实验结果发现,与空白处理组和空白质粒转染组比较,IGFBP7过表达组细胞增殖能力明显降低;Western bolt检测结果发现,AKT蛋白的磷酸化水平明显下降,且其下游的m TOR表达明显下降(P<0.05)。结论IGFBP7过表达对MCF-7乳腺癌细胞的增殖具有抑制效果,可能是通过对AKT/m TOR信号通路的抑制达到对MCF-7的抑制。

Objective To investigate the effect of insulin-like growth factor binding protein 7（IGFBP7） on proliferation of human breast cancer cell line MCF-7 and its mechanism. Methods Plasmid p IRES2-Zs Green1-IGFBP7 or empty plasmid p IRES2-Zs Green1 was transfected into MCF-7 cells using LipofectamineTM2000 and the cell transfection efficiency was examined by fluorescence microscopy. Real-time fluorescent quantitative PCR（RTFQPCR） was used to quantify the expression of IGFBP7. MTT was performed to evaluate the effect of IGFBP7 on proliferation and apoptosis of MCF-7 cells at 24, 48 and 72 hours after transfection. The expression of relevant proteins of AKT/m TOR signaling pathway was analyzed by Western blot. Results The IGFBP7 m RNA level increased significantly after IGFBP7 transfection. Compared with the controls, cell proliferation noticeably decreased in the IGFBP7-transfected group. Western bolt analysis indicated that the AKT phosphorylation was down-regulated, and the m TOR level dropped markedly（P〈0.05）. Conclusions Overexpression of IGFBP7 could down-regulate the proliferation of MCF-7 cells, possibly via inhibiting the AKT/m TOR signaling pathway.