p53抑制剂对热化疗损伤结肠上皮细胞p53Bax和MDM2表达的影响.doc

p53抑制剂对热化疗损伤结肠上皮细胞p53、 Bax和MDM2表达的影响 p53抑制剂对热化疗损伤结肠上皮细胞p53、 Bax和MDM2表达的影响 作者：张安平 刘宝华 张连阳 童卫东 何渝军 付涛 作者单位：400042 重庆，第三军医大学大坪医院野战外科研究所普通外科 【摘要】 目的 探讨p53抑制剂PFTΑ(pifithrin alpha，PFTΑ)对热化疗诱导原代培养结肠上皮细胞凋亡及p53靶基因Bax和MDM2表达的影响。方法 原代培养结肠上皮细胞分为正常对照组、热化疗组和PFTΑ加热化疗组，运用顺铂联合温热(43 ℃)处理结肠上皮细胞30 min，对比加入不同浓度的PFTΑ后，Annexin VFITC/PI染色，流式细胞仪检测细胞凋亡。MTT法分析PFTΑ对热化疗杀伤DLD1细胞的影响。Western blot检测结肠上皮细胞的p53、Bax和MDM2蛋白表达。结果 热化疗导致结肠上皮细胞发生凋亡，Bax和MDM2蛋白表达明显高于对照组。PFTΑ作用于顺铂联合温热处理结肠上皮细胞后，细胞凋亡率下降呈剂量依赖性，同时p53在结肠上皮细胞胞核/胞质表达比例下降，Bax和MDM2的表达逐渐降低。热化疗杀伤肿瘤细胞的效应并未受到低剂量PFTΑ影响。结论 PFTΑ对热化疗损伤结肠上皮细胞具有保护作用，其机制可能与改变p53核转位和抑制促凋亡基因Bax和MDM2的表达相关，抑制p53可作为克服热化疗抗肿瘤治疗副效应的新策略。 【关键词】 基因表达 p53抑制剂 热化疗 结肠肿瘤 p53 specific inhibitor, pifithrin Α, influencing expressions of p53, Bax and MDM2 in colonic epithelial cells injured by hyperthermic chemotherapy ZHANG Anping, LIU Baohua, ZHANG Lianyang, TONG Weidong, HE Yujun, FU Tao. Department of General Surgery, Research Institute of Surgery, Daping Hospital, Third Military Medical University, Chongqing 400042, China 【Abstract】 Objective To investigate the effect of a p53 specific inhibitor, pifithrin alpha (PFTΑ), on apoptosis of colonic epithelial cells and expressions of p53 target genes Bax and MDM2 in colonic epithelial cells injured by hyperthermic chemotherapy. Methods The primarily cultured colonic epithelial cells were divided into control group, hyperthermic chemotherapy group (HTC group) and PFTΑ plus hyperthermic chemotherapy group (PFTΑ plus HTC group). After the colonic epithelial cells were treated with hyperthermia at 43 ℃ and cisplatin at 10 mg/L for 30 minutes and added with PFTΑ at different concentrations, apoptosis rate was detected by using flow cytometry with Annexin VFITC and PI double labeling method. The influence of PFTΑ on antitumor potency of hyperthermic chemotherapy was analyzed with using methabenzthiazuron (MTT) assay. The expressions of p53, Bax and MDM2 were measured by Western blot. Re