苯妥英钠对化疗耐药胶质母细胞瘤多药耐药相关蛋白1及O6-甲基鸟嘌呤-DNA-甲基转移酶基因表达的影响  

Influence of Phenytoin on expression of multidrug resistance associated protein and O6-methylguanine-DNA methytransferase gene in glioblastoma cell lines

作　　者：王峻[1] 刘仁忠[1] 徐亚青[2] 连海伟[1] 吕华荣[3] 

机构地区：[1]武汉大学人民医院神经外科,430060 [2]武汉大学人民医院医院感染管理办公室,430060 [3]黄石市中心医院神经外科

出　　处：《中华实验外科杂志》2015年第11期2718-2721,共4页Chinese Journal of Experimental Surgery

基　　金：2014年湖北省自然科学基金面上项目（2014CFB421）

摘　　要：目的 观察苯妥英钠对胶质母细胞瘤细胞株8-MG-BA及胶质瘤细胞株H4细胞内多药耐药相关蛋白1（MRP1）及O6-甲基鸟嘌呤-DNA-甲基转移酶（MGMT）基因表达的影响.方法 将体外培养的胶质瘤细胞株分为用药组及对照组.以梯度浓度苯妥英钠作用作用24、48、96 h后,反转录-聚合酶链反应（RT-PCR）测定细胞中MRP1、MGMT mRNA的表达;Western blot测定细胞中MRP1、MGMT蛋白的水平.结果 （1）在8-MG-BA细胞中,可显示出明显的MGMT mRNA和MRP1mRNA条带;（2）5 mg/L苯妥英钠处理的8-MG-BA细胞中MRP1 mRNA表达量24h时减少,48 h内较明显（P＜0.05）,但96h时恢复到处理前水平（P＞0.05）.在10、20 mg/L苯妥英钠作用下,MRP1 mRNA的表达量在24h后明显减少,并持续到96 h（P ＜0.05）.在苯妥英钠5 mg/L组中MGMT mRNA的表达在3个时间点差异均无统计学意义（P＞0.05）,在10、20 mg/L苯妥英钠作用下,24h出现降低,48 h降低更明显,96 h仍维持低表达的状态（P＜0.01）;（3）苯妥英钠处理后的细胞中MRP1及MGMT蛋白的表达与mRNA表达水平的改变呈现出时间上的一致性.结论 苯妥英钠可引起8-MG-BA细胞的MRP1、MGMT的mRNA及蛋白产物表达的下调.Objective To investigate the gene expression of multidrug resistance associated protein （MRP1） and O6-methylguanine-DNA methytransferase （MGMT） in glioblastoma cell lines 8-MG-BA and H4 after incubation with phenytoin （PHT） in vitro.Methods We divided the cultured 8-MG-BA and H4 cells into drug group and control group.Reverse transcriptase-polymerase chain reaction （RT-PCR） and Western blotting were used respectively to observe the alteration of MRP1 and MGMT expression in 8-MG-BA cells after treatment with PHT at different time points （24, 48, and 96 h）.Results （1） A specific MRP1 band of 257 bp and a specific MGMT band of 230 bp were visualized by RT-PCR analysis in 8-MG-BA cells, while the protein bands of MRP1 and MGMT also visualized by Western blotting in 8-MG-BA cells.（2） In 5 mg/L PHT group, the expression level of MRP1 mRNA in 8-MG-BA cells decreased in 24 h and 48 h （P 〈 0.05）, and that was raised to a normal level in 96 h （P〉0.05）.In 10 and 20 mg/L PHT group, MRP1 mRNA level decreased remarkably in 24 and 48 h, and that had no significant change in 96 h （P 〈 0.05）.The expression level of MGMT mRNA had no significant change in 5 mg/L PHT group （P 〉 0.05）.In 10 and 20 mg/L PHT group, the MGMT mRNA expression was lowered remarkably （P 〈 0.01）.The down-regulation of MGMT mRNA had a PHT dose-dependent manner.（3） The expression of MRP1 and MGMT proteins in 8-MG-BA cells had similar tendency to mRNA after stimulation by PHT.Conclusion PHT can decrease the expression of MRP1 and MGMT genes.The down-regulations of MRP1 and MGMT have a dose-dependent manner.The time that MGMT down-regulated is later than that of MRP1.

关 键 词：苯妥英钠 多药耐药相关蛋白 胶质母细胞瘤 

分 类 号：R733.7[医药卫生—肿瘤]