联合顺铂提高对替莫唑胺耐药的多形性胶质母细胞瘤细胞药物敏感性的研究

In combination with cisplatin to increase durg sensitivity of temozolomide-resistant glioblastoma multiforme cells

目的探讨多形性胶质母细胞瘤(glioblastoma multiforme,GBM)细胞中O6甲基鸟嘌呤DNA甲基转移酶(O6methylguanine-DNA-methyltransferase,MGMT)的甲基化状态在替莫唑胺(temozolomide,TMZ)耐药机制中的作用,并检测TMZ与周期非特异性化疗药顺铂(cisplatin,CDDP)联合应用对GBM细胞的增殖抑制效应。方法将胶质瘤细胞系接种于DMEM培养基(含10%胎牛血清),MTT检测细胞增殖,甲基化特异性PCR(methylation specific polymerase chain reaction,MS-PCR)检测MGMT甲基化状态,Hoechst33342/PI检测凋亡,Chou-Talalay软件分析联合用药的机制。结果 U87MG细胞不具有MG-MT活性,无MGMT蛋白表达;T98G细胞具有MGMT活性,有MGMT蛋白表达;有MGMT活性的T98G细胞抵抗TMZ诱导的凋亡;MGMT抑制剂O6-BG提高了T98G细胞对TMZ敏感性;TMZ与CDDP联合用药效果在T98G细胞中更明显。结论无MGMT活性的U87MG细胞对TMZ更敏感;有MGMT活性的T98G细胞对TMZ和CDDP的联合化疗更敏感。

Objective To investigate O^6 methylguanine-DNA-methyltransferase（MGMT） meth- ylation status mediated temozolomide（TMZ） resistant and evaluate the anti-proliferation effect of TMZ in combination with cell cycle non-specific agent cisplatin（CDDP）. Methods Glio- blastoma multiforme to be inoculated in DMEM （ 10% fetal bovine serum）. MTr assay was used to determine cell proliferation, methylation-specifie polymerase chain reaction （MS-PCR） was applied to detect the methylation status of MGMT gene, Hoechst 33342/PI fluorescent stai- ning was employed to detect the apoptosis, and mechanism of combine medication was evalua- ted by Chou-Talalay method. Results MGMT was methylated in U87MG which not expressingMGMT protein while incompletely methylated in T98G which expressing MGMT protein. T98G was less sensitive to apoptosis induction following TMZ treatment, while O^6-BG improved the sensitivity of T98G cells to TMZ, and Chou-Talalay analysis showed that TMZ combination with CDDP was more sensitive in T98G than U87MG when IC50. Conclusion U87MG which MG- MT methylated is more sensitive to TMZ. The combination of TMZ and CDDP can effectively inhibit T98G that MGMT is incompletely methylated.