Runx3基因对HepG2细胞耐药性的影响及可能机制.docVIP

Runx3基因对HepG2细胞耐药性的影响及可能机制 郭元红，陈伟庆，房殿亮（400010重庆，重庆医科大学附属第二医院消化内科） [摘要] 目的 探讨Runx3基因对HepG2细胞耐药性的影响及可能机制。方法 取在1.6ug/ml顺铂（CDDP）下生长良好的HepG2耐药细胞(HepG2/CDDP-1.6)，建立耐药HepG2/CDDP的动态变化模型HepG2/CDDP/2.0，并予以甲基转移酶抑制剂5-氮-2，-脱氧胞苷（5-Aza-CdR）处理；另外，将Runx3-shRNA表达载体转染HepG2/ CDDP-1.6细胞，RT-PCR检测处理及转染前后细胞Runx3 mRNA的表达。MTT法检测细胞的增值活性，流式细胞仪分析细胞周期，Hoechst 33258检测凋亡，Western Blot检测P-gp表达的变化。结果 在动态变化模型中随CDDP诱导时间的延长，Runx3 mRNA的表达逐渐降低。5-Aza-CdR处理后Runx3 mRNA的表达增加，细胞生长受到明显抑制，S期细胞逐渐增加, 凋亡细胞明显增多，细胞内P-gp的表达减少。转染Runx3-shRNA载体后HepG2/CDDP-1.6细胞对CDDP的耐受性增强，G1期细胞增加，P-gp的表达增加。结论 通过对Runx3基因表达的初步调控，提示Runx3基因的表达可抑制HepG2/CDDP耐药的形成，提高HepG2/CDDP细胞对化疗药物的敏感性。 [关键词] Runx3；耐药；肝癌；DNA甲基化；细胞周期；RNA干扰 Possible involvement of RUNX3 activity in the drug resistance against hepatocellular carcinoma cells Guo Yuanhong,Chen Weiqing,Fang Dianliang(Department of Digestive Diseases, The Second Affiliated Hospital of Chongqing Medical University, Chongqing,400010,China) [Abstract] Objective To investigate the effect of Runx3 activity during drug resistance against hepatocellular carcinoma cells,and further characterize the the possible mechanisms. Methods HepG2/cisplatin(CDDP) kinetic anti-cancer drug resistance model was constructed using anti-cancer drug inducing method and treated with 5-Aza-2,-deoxycytidine(5-Aza-CdR) for different periods of time.Runx3-shRNA expression vectors were transfected into drug-resistant HepG2 cells.Reverse transcription-polymerase chain reaction(RT-PCR) was used to measure Runx3 messenger RNA(mRNA) expression level before and after treatment.The cell viability was evaluated by MTT assy and apotosis was analyzed using Hoechst 33258. Flow cytometry(FCM) was used to detect the cell cycle distribution,Western blot was used to assess the changes in the expression levels of P-gp、Runx3.Results We observed that the expression of Runx3 mRNA decreased with the increasing drug resistance against HepG2 cells.However,it increased gradually after treated with 5-Aza-CdR, cell proliferation was inhibited obviously