异柠檬酸脱氢酶1突变体对U87细胞生物节律基因表达的影响  

Effects of isocitrate dehydrogenase 1 mutants on the expression of biorhythm gene in U87 cells

作　　者：武艳玮 王世雄 徐辉 于佳翔 马杰 刘焕 曹相玫 Wu Yanwei;Wang Shixiong;Xu Hui;Yu Jiaxiang;Ma Jie;Liu Huan;Cao Xiangmei(Department of Pathology,School of Basic Medicine,Ningxia Medical University,Ningxia Medical University,Yinchuan 750004;Teaching Reform Class 1,Grade 2016,School of Clinical Medicine,Ningxia Medical University,Ningxia Medical University,Yinchuan 750004;Basic Medicine Class of 2018,School of Basic Medicine,Ningxia Medical University,Ningxia Medical University,Yinchuan 750004;Biotechnology Class of 2017,School of Basic Medical Science,Ningxia Medical University,Yinchuan 750004;Department of Basic Medicine,Xi an Medical University,Xi an 710021,China)

机构地区：[1]宁夏医科大学基础医学院病理学系,银川75004 [2]宁夏医科大学临床医学院,银川75004 [3]宁夏医科大学基础医学院,银川75004 [4]西安医学院基础医学部病理生理学教研室,西安710021

出　　处：《神经解剖学杂志》2020年第5期519-524,共6页Chinese Journal of Neuroanatomy

基　　金：国家自然科学基金(81560501;81601637);宁夏自然科学基金(2020AAC03185;2020AAC03351)。

摘　　要：目的:研究异柠檬酸脱氢酶1(IDH1)野生型与突变型(mut IDH1)对人胶质瘤细胞系U87节律基因的影响。方法:分别用IDH1和mut IDH1重组慢病毒感染U87细胞,应用荧光倒置显微镜观察感染效率;流式细胞术检测细胞周期分布;Western Blot检测U87细胞内IDH1、mut IDH1、Bmal1、Clock、Smad通路相关分子的表达水平。结果:成功感染IDH1和mut IDH1重组慢病毒,当感染复数(MOI)为20∶1时,感染效率最高。流式细胞术检测细胞周期分布提示IDH1基因突变使U87细胞G1期比例增高(野生型组62.00%,突变型组72.31%),S期比例降低(野生型组29.82%,突变型组19.39%)。Western Blot结果显示IDH1基因突变使得正调节节律分子Bmal1与Clock的蛋白表达明显降低,以及Smad通路相关分子Smad2、Smad3、Smad2-3的蛋白表达降低,pSmad2、p-Smad3、Smad4的蛋白表达升高。结论:IDH1基因可能通过TGF-β/Smad通路来改变U87细胞的节律基因。Objective: To study the effect of isocitrate dehydrogenase 1(IDH1) wild type and mutated type(mut IDH1) on U87 rhythm gene in human glioma cell line.Methods: U87 cells were infected with IDH1 and mut IDH1 recombinant lentivirus,and fluorescence inversion microscope was used to observe the infection efficiency;Cell cycle distribution was detected by flow cytometry;Western Blot was used to detect the protein expression levels of IDH1,mut IDH1,Bmal1,Clock,and genes related to Smad pathway in U87 cells.Results: Successfully infected with IDH1 and mut IDH1 recombinant lentivirus,and the infection efficiency was highest when multiplicity of infection(MOI) 20∶1.The cell cycles distribution detected by flow cytometry indicated that IDH1 gene mutation increased the G1 phase ratio of U87 cells(62.00% in the wild type group,72.31% in the mutant group) and decreased the S phase ratio(29.82% in the wild type group,19.39% in the mutant group).Western Blot results showed that IDH1 gene mutation reduced the protein expression of positively regulated rhythm gene Bmal1 and Clock,decreased the protein expression of Smad pathway related proteins Smad2,Smad3,and Smad2-3,and increased the protein expression of p-Smad2,p-Smad3,and Smad4.Conclusion: IDH1 gene may alter the rhythm gene of U87 cells through TGF-β/Smad pathway.

关 键 词：异柠檬酸脱氢酶1 突变 生物节律 胶质瘤 SMAD U87细胞 

分 类 号：R739.41[医药卫生—肿瘤]