组蛋白甲基化酶抑制剂对人淋巴瘤Raji细胞生存、凋亡及细胞周期的影响  

Effects of Histone Methyltransferase Inhibitors on the Survival, Apoptosis and Cell Cycle of Raji Cells

作　　者：肖丽容[1] 侯宸 郑仕洁 郭波[2] 陈大年[1,2] 闫乃红[1,2] XIAO Lirong;HOU Chen;ZHENG Shi-jie;GUO Bo;CHEN Da-nian;YAN Nai-hong(Research Laboratory of Ophthalmology,West China Hospital,Sichuan University,Chengdu 610041,China;Department of Ophthalmology,West China Hospital,Sichuan University,Chengdu 610041,China)

机构地区：[1]四川大学华西医院眼科学研究室,成都610041 [2]四川大学华西医院眼科,成都610041

出　　处：《四川大学学报（医学版）》2019年第3期305-310,共6页Journal of Sichuan University(Medical Sciences)

基　　金：国家自然科学基金(No.81371024)资助

摘　　要：目的研究组蛋白甲基化酶Zeste基因增强子同源物2 (enhancer of zeste homolog 2, EZH2)的3种抑制剂(UNC1999、DZNep及GSK343)对人B细胞非霍奇金淋巴瘤Raji细胞的影响。方法 PCR扩增EZH2基因16(Y641)和18(A677)外显子,Sanger测序检测其突变情况;蛋白质免疫印迹法(Western blot)检测正常成年人淋巴细胞及Raji细胞EZH2的表达;使用不同浓度的UNC1999、DZNep及GSK343处理Raji细胞,CCK-8技术检测Raji细胞的生存情况;流式细胞术检测Raji细胞的凋亡情况和细胞周期变化;Western blot检测药物处理后EZH2及H3K27 me3的表达。结果 Sanger测序显示Raji细胞不携带EZH2基因Y641和A677突变位点。Western blot显示,相比正常成年人淋巴细胞,Raji细胞EZH2蛋白高表达。CCK-8法结果显示UNC1999、DZNep及GSK343对Raji细胞均有抑制生存的作用, DZNep抑制能力最弱。流式细胞术凋亡检测显示UNC1999、DZNep及GSK343促进Raji细胞的凋亡,UNC1999作用强于GSK343与DZNep;3种抑制剂均阻滞Raji细胞于G_1/G_0期,UNC1999组细胞周期阻滞最显著。Western blot显示UNC1999和GSK343抑制EZH2组蛋白甲基化酶的活性,降低H3K27 me3的表达。结论 EZH2抑制剂可通过降低H3K27 me3的表达抑制Raji细胞生存,促进细胞凋亡,改变细胞周期。UNC1999作用效果优于GSK343及DZNep,且EZH2抑制剂对Raji细胞的作用不依赖于EZH2的突变。Objective To determine the effects of three histone methylase inhibitors UNC1999,DZNep and GSK343on the survival,apoptosis and cell cycle of non-hodgkin′s lymphoma Raji cells.Methods PCR amplified 16and 18exons of enhancer of zeste homolog 2(EZH2)gene were detected.The expression of EZH2in normal adult lymphocytes and Raji cells was detected by Western blot.The Raji cells were treated by UNC1999,DZNep and GSK343,followed by CCK-8assays analyzing cell survival,flow cytometry detecting cell apoptosis and cell cycle,and Western blot detecting the expressions of EZH2and H3K27me3.Results The Sanger sequencing results showed that the Raji cells did not carry Y641and A677mutation sites of EZH2.The Western blot results showed high expressions of EZH2in the Raji cells.The results of CCK-8showed that UNC1999,DZNep and GSK343inhibited cell survival,and the weakest effect was from DZNep.The flow cytometric assay showed that UNC1999,DZNep and GSK343promoted apoptosis of the Raji cells,and the effect of UNC1999was stronger than that of GSK343and DZNep.The cell cycle was arrested at phase G1/G0after treatment of the Raji cells with the three inhibitors,with UNC1999triggering the most significant changes.The Western blot showed that UNC1999 and GSK343inhibited the histone methylase activity of EZH2and significantly reduced the expression of H3K27 me3.Conclusion EZH2inhibitors can inhibit cell survival,promote cell apoptosis and arrest cell cycle at phase G1/G0of Raji cells through reducing the expression of H3K27me3.UNC1999has a stronger effect than GSK343 and DZNep.

关 键 词：淋巴瘤Raji细胞 EZH2抑制剂 细胞凋亡 细胞周期 

分 类 号：R733.1[医药卫生—肿瘤]