异柠檬酸脱氢酶1对替莫唑胺干预下U87细胞细胞凋亡的影响  

Effect of Isocitrate Dehydrogenase 1 on Temozolomide-Induced Apoptosis of Glioma U87 Cells

作　　者：王举波[1] 权瑜[1] 吕健[1] 董丹凤[2] WANG Jubo;QUAN Yu;LV Jian;DONG Danfeng(Department of Neurosurgery, the Second Affiliated Hospital of Xi′an Jiaotong University,Xi′an 710004, Shaanxi, China;Department of Oncology, the First Affiliated Hospital of Xi′an Jiaotong University, Xi′an 710061, Shaanxi, China)

机构地区：[1]西安交通大学第二附属医院神经外科,陕西西安710004 [2]西安交通大学第一附属医院肿瘤内科,陕西西安710061

出　　处：《贵州医科大学学报》2019年第8期892-897,共6页Journal of Guizhou Medical University

基　　金：国家自然科学基金青年科学基金项目(81801121);西安交通大学基本科研业务自由探索项目(xjj2018136)

摘　　要：目的:探讨人异柠檬酸脱氢酶1(mIDH1)基因突变对替莫唑胺(TMZ)干预下脑胶质瘤U87细胞凋亡的影响。方法:用基因重组技术构建真核表达载体mIDH1/wIDH1,转染技术构建目的基因的U87细胞的稳转细胞系,采用流式细胞术检测TMZ干预24h后的细胞凋亡率;构建裸鼠移植瘤模型,采用免疫组织化学法(Westernblot)检测mIDH1对TMZ干预U87细胞24h时及TMZ连续灌胃5d时裸鼠移植瘤相关凋亡蛋白Caspase-3、Caspase-9、Bax及Bcl-2的表达水平。结果:成功构建胶质瘤稳转细胞系,流式细胞术结果显示,mIDH1能提高TMZ作用后U87细胞的凋亡率,也能上调TMZ干预24h时U87细胞及TMZ连续灌胃5d时裸鼠移植瘤中Caspase-3、Caspase-9、Bax蛋白表达水平,下调Bcl-2蛋白表达水平(P<0.05)。结论:mIDH1通过上调TMZ作用的U87细胞、裸鼠移植瘤组织的Caspase-3、Caspase-9、Bax表达、下调Bcl-2表达促进胶质瘤细胞凋亡。Objective: To investigate the effect of human isocitrate dehydrogenase 1 (IDH1) gene mutation (mutIDH1) on the apoptosis of glioma U87 cells induced by temozolomide (TMZ). Methods: Human wild type IDH1 cDNA was amplified by PCR and cloned into pCMV expression vector.Mutated IDH1(mutIDH1)was generated using a mutagenesis kit.The recombinant vectors expressing IDH1 or mutIDH1 gene were transfected into U87 cells to establish stable cell lines.Flow cytometry was used to detect the TMZ-induced U87 cell apoptosis.Glioma-bearing nude mice were treated with TMZ for 5 days.Immunohistochemistry and Western blot were used to detect the expression levels of caspase-3,Caspase-9,Bax and Bcl-2 in glioma. Results: IDH1or mutIDH1 were stably expressed in U87 cells.The mutation of IDH1 enhanced TMZ-induced apoptosis compared to wild type IDH1.Moreover,the mutation of IDH1 upregulated expression levels of proapoptotic protein such as Caspase-3,Caspase-9,Bax,while downregulated Bcl-2 expression level ( P <0.05). Conclusion: mutIDH1 can syngerize TMZ-induced apoptosis by up-regulating Caspase-3,Caspase-9 and Bax expression in vivo,and down-regulating Bcl-2 expression.

关 键 词：胶质瘤细胞 异柠檬酸脱氢酶1 凋亡 细胞 凋亡相关蛋白 替莫唑胺 

分 类 号：R739.41[医药卫生—肿瘤]