小干扰RNA沉默zeste基因增强子同源物2对阿霉素诱导胃癌细胞MKN28衰老的影响  

Effects of small interfering RNA targeting enhancer of zeste homolog 2 on doxorubicin-induced cel-lular senescence of gastric cancer ceil line MKN28 cells

作　　者：白洁一[1] 马牧原[1] 蔡明[1] 许飞[1] 陈俊华[1] 韩高雄[1] 帅晓明[1] 陶凯雄[1] 

机构地区：[1]华中科技大学同济医学院附属协和医院胃肠外科Ⅱ,武汉430022

出　　处：《中华实验外科杂志》2013年第2期214-216,共3页Chinese Journal of Experimental Surgery

基　　金：湖北省自然科学基金资助项目（2010CDB07706）

摘　　要：目的观察小干扰RNA（siRNA）沉默zeste基因增强子同源物2（EZH2）对阿霉素诱导胃癌细胞MKN28衰老的影响。方法体外化学合成EZH2基因特异性小干扰RNA，脂质体Lipo-fectamine2000介导转染人胃癌细胞MKN28；实时定量聚合酶链反应（Real—timePCR）检测特异性siRNA对EZH2基因mRNA水平的沉默效果；采用阿霉素诱导细胞衰老，衰老相关．p．半乳糖苷酶（SA—B．gal）染色检测各组细胞衰老情况，激光共聚焦显微镜检测各组细胞胞核中衰老相关异染色质灶（SAHF）的形成。流式细胞仪和噻唑蓝实验（MTF）检测siRNA对细胞周期和生长的影响，并比较在阿霉素诱导下各组细胞的相应变化。结果EZH2siRNA能有效抑制MKN28细胞中EZH2mRNA表达，促进阿霉素诱导的MKN28细胞衰老和SAHF形成，EZH2siRNA＋阿霉素组细胞SA-13-gal染色阳性率和SAHF形成比例明显高于对照组，分别为（87．88±2．66）％、（96．27±1．71）％和（42．01±7．50）％、（51．72±2．97）％，且EZH2siRNA能阻滞MKN28细胞周期于Go／G．期，比例为（47．41±10．68）％，抑制细胞增殖，并促进阿霉素的阻滞细胞周期和抗增殖效应。结论EZH2siRNA可有效抑制胃癌细胞MKN28中EZH2表达，促进阿霉素诱导的胃癌MKN28细胞衰老，抑制细胞增殖。Objective To study the effect of small interfering RNA （siRNA） targeting enhancer of zeste homolog 2 （EZH2） on doxorubicin-induced cellular senescence of gastric cancer cell line MKN28 cells. Methods The siRNA targeting EZH2 was constructed and transfected into MKN28 ceils via Lipo- fectamine 2000. The expression level of EZH2 mRNA was detected by using real-time quantitative polymer- ase chain reaction （Real-time PCR）. The doxorubicin-induced cellular senescence of transfected MKN28 cells was analyzed by senescence associated-13-galactosidase （SA-B-gal） stain and the formation of senes- cence-associated heterochromatic foci （SAHF） was measured by laser confocal microscopy. Cell cycle and cellular proliferation of transfected MKN28 ceils were measured by flow cytometry and MTT assay respec- tively, with or without the induction of doxorubicin. Results SiRNA can effectively suppress the expres- sion of EZH2 mRNA, promote the doxorubicin-induced cellular senescence and formation of SAHF in MKN28 cells. In siRNA ＋ doxorubicin group, the cells positive for SA-13-gal and SAHF formation propor- tion were increased more significantly than control group, （87.88 ± 2. 66）% and （96. 27 ±1.71 ）% vs. （42. 01 ±7.50）% and （51.72 ± 2. 97）%, respectively. In transfected MKN28 cells, the proportion of cells in G0/G1 phase was increased to （47.41± 10. 68）%, and cellular proliferation inhibited, and this effect was more notable under the induction of doxorubicin. Conclusion SiRNA can effectively suppress the expression of EZH2, improve the progress of doxorubicin-induced cellular senescence and inhibite the cellular proliferation of MKN28 cells.

关 键 词：胃癌 zeste基因增强子同源物2 小干扰RNA 细胞衰老 

分 类 号：R73-36[医药卫生—肿瘤]