人食管癌顺铂耐药细胞系的建立及耐药相关基因的筛选

Establishment of a cisplatin-induced human esophageal carcinoma drug resistant cell line and screening of drug resistance-related genes

目的:建立人食管癌顺铂耐药细胞系Ec9706/cDDP,并初步探讨其耐药机制。方法:采用顺铂(cDDP)中等浓度、间歇冲击作用方法历时9mon诱导人食管癌细胞系Ec9706,建立相应耐药细胞系,采用MTT法测定耐药细胞系对cD-DP的耐药指数,用RT-PCR半定量方法测定亲本细胞及耐药细胞耐药相关基因如DNA聚合酶(polβ)、多药耐药相关基因(MRP)、谷胱甘肽转移酶(GST)、二氢叶酸还原酶(DHFR)、拓扑异构酶(TPO)和多药耐药基因(mdr1)等基因的mRNA表达水平。结果:人食管癌顺铂耐药细胞系Ec9706/cDDP的耐药指数为15.7,该细胞系中polβ、DHFR、GST基因mRNA的表达比亲本细胞显著增加,TPO的表达降低,而mdr1、MRP表达的增加无显著统计学意义。结论:成功建立了人食管癌顺铂耐药细胞系Ec9706/cDDP,其耐药性的产生与polβ、GST、DHFR、TPO等基因的异常表达有关,而与mdr1、MRP表达的关系不密切。

Objective ：To establish a eisplatin -induced human esophageal carcinoma drug -resistant cell line Ec9706/cDDP and study the mechanism of the acquired drug resistance. Methods： The drug - resistant cell line Ec9706/cDDP was established in culture by pulse exposing Ec9706 parent cells to moderate concentration of cisplatin （cDDP） over a period of 9 months. The resistance index to cDDP was determined by MTr cytotoxicity assay. The expression of polβ,mdr1 ,MRP,LRP,DHFR and GST mRNA was analyzed by RT - PCIL Results： The resistance index of Ec9706/cDDP cell was 15.7. Compared with Ec9706 parent cells, the expression of polβ, DHFR and GST mRNA in Ec9706/cDDP cell was increased. However, the expression of TPO was decreased and there were no changes of expression of mdrl, MRP. Conclusion： A human esophageal carcinoma drug - resistant cell line Ec9706/cDDP is established successfully and the mechanism of drug resistance is related to expression of polβ,DHFR, （GT and TPO.