摘要
目的观察外源性线粒体融合素基因-2(Mfn2)对乳腺癌细胞(MCF-7)增殖的影响,探讨其抑制乳腺癌细胞产生增殖的机制。方法将含有Mfn2cDNA的质粒在阳离子聚合物的介导下在体外转染MCF-7,蛋白印记法(Western blot)检测绿色荧光蛋白(GFP)、p21^wafl、CDK2蛋白的表达,细胞计数测定Mfn2对MCF-7细胞增殖的影响,DNA直方图分析法测定Mfn2对MCF-7周期分布的影响,用Cyclins/DNA双参数流式细胞术对CyclinA进行标记,Cellquest软件分析。结果转染Mfn2基因的MCF-7细胞可以稳定高表达GFP蛋白;细胞计数测定表明转染Mfn2cDNA后,MCF-7细胞增殖明显受抑,DNA直方图分析法测定显示细胞停滞于S期;转染质粒组S期细胞占42.70%,转染空质粒组和空白对照组分别为17.15%、19.58%(P〈0.05);用Cyclins/DNA双参数流式细胞术检测转染质粒组CyclinA较对照两组明显升高;Westernblot显示转染质粒组p21高表达,磷酸化CDK2低表达。结论转染Mfn2基因可以明显抑制MCF-7细胞的增殖,并使细胞停滞于S期。
Objective To study the role and mechanism of mitofusin-2 gene in cell proliferation of human breast carcinoma cell line MCF-7 after in vitro transfer. Methods pEGFP Mfn2 was transferred by sofast in vitro. The expression of GFP was detected by Western blot, and the cell proliferation of MCF-7 by cell counting. Cell cycle of MCF-7 was observed by DNA analytical method,and the expression of Cyclin A examined by Cyclins/DNA flow cytometry, and the expression of p21^walf, CDK2 by Western blot. Re- suits The stable expression of GFP protein in MCF-7cells was confirmed by Western blot. Mfn2 could also induce cell cycle arrest (S) by DNA analytical method. The percentage of S phase cells was 42.70 in thansfer group,whereas in control groups it was 17.15 and 19.58 respectively (P 〈0.05). Mfn2 could induce the expression of Cyclin A protein obviously ,and the expression of p21 was high and phosphorylated CDK2 was low. Conclusion Mfn2 can strongly inhibit cell proliferation in MCF-7 cell line, and result in cell arrest in S phase.
出处
《中华实验外科杂志》
CAS
CSCD
北大核心
2008年第1期9-11,共3页
Chinese Journal of Experimental Surgery
基金
卫生部临床学科重点项目:“临床肿瘤的细胞周期诊断与分析(三)”
“973”计划子项目(2004CB518705)
关键词
乳腺癌
融合素基因
增殖
Breast carcinoma
Mitofusin gene
Proliferation