摘要
目的:探讨伊马替尼(商品名:格列卫)对K562及其耐药细胞株K562/AO2细胞色素P4503A亚家族多肽5(CYP3A5)基因转录、蛋白表达及活性的调控。方法:采用实时聚合酶链反应(real-timePCR)、蛋白印迹(Westernblot)和高效液相色谱(HPLC)法检测CYP3A5基因转录、蛋白表达和活性水平。同时将CYP3A5重组质粒稳定转染有CYP3A5基础转录的K562细胞,噻唑蓝(MTT)法测定伊马替尼的半数抑制浓度(IC50)值,观察CYP3A5基因的过表达是否介导白血病细胞对伊马替尼敏感性的改变。结果:K562细胞经伊马替尼作用24h后CYP3A5mRNA转录增强,蛋白表达和活性均增加,与未加药细胞相比,差异有统计学意义(P<0.05、P<0.01、P<0.01);K562/AO2细胞经伊马替尼作用24h后,CYP3A5基因转录、蛋白表达和活性未见改变,经伊马替尼作用36h后,CYP3A5基因转录增强、蛋白表达和活性均增加,与未加药细胞相比,差异有统计学意义(P<0.01、P<0.01、P<0.05)。K562细胞稳定转染CYP3A5重组质粒后对伊马替尼表现出明显耐受(耐药倍数为1.318倍)。结论:伊马替尼可诱导K562及其耐药细胞株K562/AO2CYP3A5基因转录、蛋白表达和活性的增高。
Objective To investigate the modulation effect of Glivec on transcription, protein expression and activity of CYP3A5 gene in K562 and its drug resistant K562/AO2 cell lines. Methods CYP3A5 mRNA, protein expression and activity were detected by real-time PCR, Western blot and HPLC assay. K562 cells having CYP3A5 basic transcription were stably transfected with CYP3A5 recombinant plasmid, and MTT assay was used to determine the half maximal inhibitory concentration （IC50） of Gilvec and to observe whether the overexpression of CYP3A5 gene mediated the alteration of leukemia cell sensitivity to Gilvec. Results Transcription of CYP3A5 mRNA, protein expression and activity in K562 cells increased significantly after treated with Gilvec for 24 h when compared with that not treated with Gilvec （P0.05, P0.01, P0.01）. Transcription of CYP3A5 mRNA, protein expression and activity in K562/AO2 cells increased only after 36 h treatment with Gilvec （P0.01, P0.01, P0.05）and no changes were observed after 24 h treatment with Gilvec. Significant resistance to Glivec （1.318 times） was observed when the K562 cells were stably transfected with CPY3A5 recombinant plasmid. Conclusions Gilvec could induce the increase of transcription of CYP3A5 gene, protein expression and activity in K562 and its drug resistant K562/AO2 cells.
出处
《诊断学理论与实践》
2010年第5期469-472,共4页
Journal of Diagnostics Concepts & Practice
基金
国家自然科学基金项目(30400183)
