小分子干扰RNA对胃癌MGC803细胞E2F-1表达及其生物学行为的影响

The effects of short interfering RNAs on E2F-1 gene expression and biological behaviors of gastric cancer cell line MGC803

目的观察小分子干扰RNA（siRNA）对胃癌MGC803细胞E2F-1基因表达及其生物学行为的影响。方法将实验组（E2F-1-siRNA）和阴性对照组（negative control-siRNA）转染进MGC803细胞，48h后采用实时定量聚合酶链反应（PCR）及Western blot技术分别检测E2F-1mRNA及蛋白的表达，噻唑蓝（M3T）比色法检测MGC803细胞增殖的变化，流式细胞仪检测MGC803细胞周期变化。结果E2F-1-siRNA有效抑制胃癌细胞E2F-1mRNA和蛋白的表达并且影响MGC803细胞的增殖，与阴性对照组及未转染组细胞比较mRNA降低了84．8％和85．3％（F=14．35，P〈0．05），蛋白降低了77．2％和79．6％，MGC803细胞增殖分别抑制了69．0％和81．6％，差异有统计学意义（F=170．18，P〈0．05）；同时E2F-1-siRNA促使更多MGC803细胞DNA含量聚集于G2／M期附近，G2／M期DNA含量比例为（54．98±3．46）％，与阴性对照组（44．70±3．82）％和未转染组（31．47±2．12）％比较，差异有统计学意义（F=88．90，P〈0．05）。结论E2F-1-siRNA可以有效抑制人胃癌MGC803细胞E2F-1 mRNA及蛋白的表达，使G1期细胞的DNA含量下调，细胞分裂停滞在G2／M期附近，抑制胃癌细胞的增殖。

Objective To investigate the effects of RNA interference （RNAi） targeting E2F-1 on biological behaviors of gastric cancer cell line MGC803. Methods E2F-1-siRNA or negative control-siR- NA was transfected into gastric cancer cell line MGC803 cells. Forty-eight h later,the expression of E2F-1 mRNA and protein in gastric cancer cell line MGC803 was detected by real-time quantitative PCR and Western blot respectively. Cell cycle was examined by flow cytometry, and the proliferative rate of MGC803 cells tested by MTT assay. Results E2F-1-siRNA effectively inhibited the expression of E2F-1 mRNA and protein and influenced proliferation of MGC803 cells. As compared with negative control and nontransfected groups ,the expression levels of E2F-1 mRNA and protein were decreased by 84.8%, 85.3% （P 〈 0. 05 ）, and 77.2%, 79.6% , and proliferation of MGC803 cells was suppressed by 69.0% and 81.6% （P 〈 0.05） in E2F-1-siRNA transfection group, respectively. E2F-1-siRNA promoted accumulation of more DNA of MGC803 cells at G2/M phase, and the rate of DNA content in GJM phase in E2F-1- siRNA transfection group was （ 54.98 ± 3.46） %, which was significantly higher than （43.07 ± 3.82） % in negative control group, and （31.47 ± 2.12 ）% in non-transfection group （P 〈 0.05 ）. Conclusion E2F-1 gene repression by RNAi can decrease the DNA content in G1 phase of MGC803 cells, which can arrest the cells in G2/M phase and inhibit the proliferation of MGCS03 cells.