【摘要】 目的探讨胶质瘤细胞异柠檬酸脱氢酶1(IDH1)基因突变对替莫唑胺(TMZ)细胞毒性的影响。方法将培养的U87胶质瘤细胞分组转染含空载体(空载体组)、野生型IDH1基因(野生组)和R132H突变型IDH1基因(突变组;IDH1第132位的精氨酸被组氨酸所取代)质粒,利用液相色谱串联质谱法检测细胞上清液中2-羟基戊二酸(2-HG)水平、噻唑蓝法检测细胞增殖活性、实时荧光定量PCR法和免疫印迹法检测细胞caspase 3 mRNA和蛋白质表达;将含三种不同质粒的U87细胞接种大鼠建立荷瘤大鼠模型并观察大鼠经TMZ治疗后60 d存活率。结果与空载体组和野生组比较,突变组细胞上清液中2-HG水平、细胞caspase 3 mRNA和蛋白质表达水平均显著升高(P<0.05),而细胞增殖活性显著下降(P<0.05);TMZ治疗后60 d,突变组大鼠存活率显著升高(P<0.05);空载体组和野生组之间均无统计学差异(P<0.05)。结论 IDH1 R132H基因能够增加TMZ对神经胶质瘤细胞的细胞毒性作用并提高其治疗荷瘤大鼠的存活率。更多还原

【Abstract】 ObjectiveTo investigate the effect of mutant isocitrate dehydrogenase 1(IDH1) gene in the glioma cells on the cytotoxicity of temozolomide(TMZ).MethodsCultured U87 glioma cells were transfected with recombinant plasmids containing IDH1wild type and IDH1 R132H mutant type gene respectively. Then the levels of 2-hydroxyglutarate(2-HG) were detected by liquid chromatography tandem mass spectrometry(LC-MS/MS). Moreover, the expressions of cysteine-containing aspartate-specific proteases(caspase) 3 in the glioma cells were analyzed by real-time PCR and immunoblotting technique. In addition, tumor-bearing rat models treated with different IDH1 gene were made to observe the survival rate over 60 days after TMZ treatment.ResultsThe levels of 2-HG were significantly higher in the glioma cells transfected with IDH1 R132H plasmids than those in the glioma cells of the control group(P<0.05). After TMZ treatment, the expressions of caspase 3 were significantly up-regulated in the glioma cells transfected with IDH1R132H gene compared to the control group(P<0.05), and there was insignificant difference in the levels of caspase 3 expression between the glioma cells transfected with IDH1 wild gene(experimental group) and the control group(P>0.05). The survival rate the rats treated with IDH1 gene(experimental group) over 60 days after TMZ treatment was significantly higher than that in the control group(P<0.05).ConclusionsIDH1 R132H gene may augment the cytotoxicity effect of TMZ on glioma cells and increase the survival rate in rats with tumors.更多还原