组蛋白H3K27甲基化对人高分化鼻咽癌细胞系CNE-1增殖的抑制效应  

Inhibitory effect of histone H3K27 methylation on proliferation of human well-differentiated nasopharyngeal carcinoma cell line CNE-1

作　　者：蔡波 邢娟娟[1] 胡蓉[2] 任欣 王榕[2] 吴恬宁 郭菲[1] 陈少卿[4] CAI Bo;XING Juan-juan;HU Rong;REN Xin;WANG Rong;WU Tian-ning;GUO Fei;CHEN Shao-qing(Burn Center, the First Affiliated Hospital of Nanchang University, Nanchang 330006;Jiangxi Medical College,Nanchang University, Nanchang 330006;the Affiliated High School of Nanchang University, Nanchang 330029;Department of Oncology, the First Affiliated Hospital of Nanchang University, Nanchang 330006, China)

机构地区：[1]南昌大学第一附属医院烧伤中心,江西南昌330006 [2]南昌大学江西医学院,江西南昌330006 [3]南昌大学附属中学,江西南昌330029 [4]南昌大学第一附属医院肿瘤科,江西南昌330006

出　　处：《基础医学与临床》2020年第12期1613-1618,共6页Basic and Clinical Medicine

基　　金：国家自然科学基金(81972445);江西省自然科学基金(20161BAB205241)。

摘　　要：目的分析特异性阻断组蛋白H3K27me3/2去甲基化反应对人高分化鼻咽癌CNE-1细胞增殖能力的调控作用及其分子机制。方法体外培养人鼻咽癌细胞系CNE-1,运用组蛋白H3K27me3/2去甲基化酶抑制剂GSK-J4特异性阻断组蛋白H3K27me3/2去甲基化反应。用含0.25、0.5、1.0、2.0和4.0 mmol/L的GSK-J4培养基(10%FBS的RPMI-1640培养液)作用于CNE-1细胞作为实验组,同时设溶剂对照(DMSO,二甲基亚砜)。激光共聚焦技术和流式细胞计量术分析细胞内组蛋白H3K27me3的表达水平;实时荧光定量PCR技术检测细胞内cyclinD1和Ki-67 mRNA的表达水平;用CCK-8法检测细胞的增殖活性;平板集落实验检测细胞集落形成能力;流式细胞计量术检测细胞周期的分布。结果实验组细胞内组蛋白H3K27me3表达水平较对照组明显增高(P<0.05);实验组细胞内cyclinD1和Ki-67 mRNA表达水平较对照组明显下调(P<0.05);CNE-1细胞活力随GSK-J4作用浓度的增加逐渐降低,呈浓度依赖性;实验组细胞增殖活力和集落形成能力较对照组明显下降(P<0.05);实验组G0/G1期细胞百分数较对照组增多,S期减少(P<0.05)。结论上调CNE-1细胞内组蛋白H3K27的甲基化修饰水平可以诱导细胞周期阻滞,抑制细胞增殖,作用机制可能与下调cyclinD1和Ki-67 mRNA水平相关。Objective To investigate the effects of inhibiting histone H3K27me3/2 demethylase on proliferation of well-differentiated human nasopharyngeal carcinoma cell in vitro and explore the underlying mechanisms.MethodsHuman nasopharyngeal carcinoma cell line CNE-1 was cultured in vitro,and histone H3K27me3/2 demethylation was specifically blocked by histone H3K27me3/2 demethylase inhibitor GSK-J4.The cells were incubated with 0.25,0.5,1.0,2.0,4.0 mmol/L GSK-J4 as the experimental groups,while DMSO was applied as control group.Immunofluorescence assay and flow cytometry were used to analyze the expression of intracellular histone H3K27me3.Real-time quantitative PCR was applied to detect the expression of cyclinD1 and Ki-67 mRNA in cells.Cell counting kit-8(CCK-8)was used to evaluate cell proliferation.The cloning of CNE-1 cells was observed by colony formation assay.Flow cytometry was also used to detect cell cycle distribution.Results The expression level of histone H3K27me3 in the experimental group was significantly higher than that in the control group(P<0.05);cyclinD1 and Ki-67 mRNA expression levels in the experimental groups were down-regulated compared with the control group(P<0.05);The cell viability of CNE-1 cells decreased gradually with the increase of GSK-J4 concentration.The cell proliferation and colony formation in the experimental group were significantly decreased than those in the control group(P<0.05);cell cycle was blocked in G0/G1 phase,the percentage of cells in G0/G1 phase was increased and the S phase decreased in the experimental group(P<0.05).Conclusions Up-regulation of methylation of histone H3K27 in CNE-1 cells induces cell cycle arrest and inhibits cell proliferation.The mechanism may be related to down-regulation of cyclinD1 and Ki-67 mRNA levels.

关 键 词：组蛋白H3K27 CNE-1 增殖 细胞周期 去甲基化酶抑制剂GSK-J4 

分 类 号：R34[医药卫生—基础医学]