CAJ | 학술논문

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Lipofectamine2000介导转染人胃癌细胞MKN28；实时定量聚合酶链反应(Real-time PCR)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-β-半乳糖苷酶(SA-β-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(MTT)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 EZH2 siRNA能有效抑制MKN28细胞中EZH2 mRNA表达,促进阿霉素诱导的MKN28细胞衰老和SAHF形成,EZH2 siRNA+阿霉素组细胞SA-β-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EZH2 siRNA能阻滞MKN28细胞周期于G0/G1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 EZH2 siRNA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.
목적 관찰소간우RNA(siRNA)침묵zeste기인증강자동원물2(EZH2)대아매소유도위암세포MKN28쇠로적영향.방법 체외화학합성EZH2기인특이성소간우RNA,지질체Lipofectamine2000개도전염인위암세포MKN28；실시정량취합매련반응(Real-time PCR)검측특이성siRNA대EZH2기인mRNA수평적침묵효과；채용아매소유도세포쇠로,쇠로상관-β-반유당감매(SA-β-gal)염색검측각조세포쇠로정황,격광공취초현미경검측각조세포포핵중쇠로상관이염색질조(SAHF)적형성.류식세포의화새서람실험(MTT)검측siRNA대세포주기화생장적영향,병비교재아매소유도하각조세포적상응변화.결과 EZH2 siRNA능유효억제MKN28세포중EZH2 mRNA표체,촉진아매소유도적MKN28세포쇠로화SAHF형성,EZH2 siRNA+아매소조세포SA-β-gal염색양성솔화SAHF형성비례명현고우대조조,분별위(87.88±2.66)％、(96.27±1.71)％화(42.01±7.50)％、(51.72±2.97)％,차EZH2 siRNA능조체MKN28세포주기우G0/G1기,비례위(47.41±10.68)％,억제세포증식,병촉진아매소적조체세포주기화항증식효응.결론 EZH2 siRNA가유효억제위암세포MKN28중EZH2표체,촉진아매소유도적위암MKN28세포쇠로,억제세포증식.
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 cells via Lipofectamine 2000.The expression level of EZH2 mRNA was detected by using real-time quantitative polymerase chain reaction (Real-time PCR).The doxorubicin-induced cellular senescence of transfected MKN28 cells was analyzed by senescence associated-β-galactosidase (SA-β-gal) stain and the formation of senescence-associated heterochromatic foci (SAHF) was measured by laser confocal microscopy.Cell cycle and cellular proliferation of transfected MKN28 cells were measured by flow cytometry and MTT assay respectively,with or without the induction of doxorubicin.Results SiRNA can effectively suppress the expression 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-β-gal and SAHF formation proportion 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.