姜黄素抗胶质瘤作用的分子机制研究  

Molecular mechanism of antiglioma effect of curcumin

作　　者：胡科[1] 贾东佩[1] 任应国[1] 白方会 张保朝[2] Hu Ke;Jia Dongpei;Ren Yingguo;Bai Fanghui;Zhang Baochao(Department of Neurorehabilitation,Nanyang Central Hospital,Nanyang 473000,China;Department of Neurology,Nanyang Central Hospital,Nanyang 473000,China)

机构地区：[1]南阳市中心医院神经康复科,473000 [2]南阳市中心医院神经内科,473000

出　　处：《中华神经医学杂志》2021年第2期141-152,共12页Chinese Journal of Neuromedicine

摘　　要：目的探讨姜黄素抗胶质瘤作用的分子机制。方法(一)细胞实验:(1)取对数生长期的U251MG、SHG-44细胞,分别分为姜黄素组与对照组、阴性对照小干扰RNA(siRNA)组与H19 siRNA组、阴性对照siRNA+姜黄素组与H19 siRNA+姜黄素组、H19 siRNA+阴性对照抑制物组与H19 siRNA+miR-491-5p抑制物组、H19 siRNA+阴性对照抑制物+姜黄素组与H19 siRNA+miR-491-5p抑制物+姜黄素组、miR-491-5p模拟物+空白质粒+姜黄素组与miR-491-5p模拟物+同源盒基因A9(HOXA9)过表达质粒+姜黄素组,各组细胞分别予10μmol/L姜黄素或阴性对照siRNA、H19 siRNA转染或miRNA抑制物、miR-491-5p抑制物共转染或miR-491-5p模拟物+空白质粒、miR-491-5p模拟物+HOXA9过表达质粒共转染等不同处理。采用实时荧光定量PCR(qRT-PCR)法检测各组细胞中H19、miR-491-5p、HOXA9 mRNA的表达水平,采用细胞计数试剂(CCK)-8法检测各组细胞的增殖率,采用流式细胞术检测各组细胞的凋亡率,采用平板克隆法检测各组细胞的克隆形成数,采用Transwell小室实验检测各组细胞的迁移情况,采用Western blotting法检测各组细胞中HOXA9蛋白的表达水平。(2)取对数生长期的293T细胞,分别分为阴性对照模拟物+野生型H19组与miR-491-5p模拟物+野生型H19组、阴性对照模拟物+野生型HOXA93'-UTR组与miR-491-5p模拟物+野生型HOXA93'-UTR组,各组细胞分别予阴性对照miRNA模拟物、miR-491-5p模拟物与野生型H19、野生型HOXA93'-UTR质粒载体共转染等不同处理。采用双荧光素酶报告实验检测各组细胞的荧光素酶活性。(二)病例标本实验:收集南阳市中心医院神经外科自2017年5月至2019年5月手术切除且经病理检查确诊为胶质瘤的30例组织标本(胶质瘤组)及同期行内减压术获得的30例正常脑组织标本(正常组),采用qRT-PCR法检测各组标本中H19、miR-491-5p、HOXA9 mRNA的表达水平,采用Western blotting法检测各组标本中HOXA9蛋白的表达水平。(�Objective To investigate the molecular mechanism of antiglioma effect of curcumin.Methods Cell experiment:(1)U251MG and SHG-44 cells at logarithmic growth phase were treated with 10μmol/L curcumin(curcumin group)or same volume of dimethyl sulfoxide solution(control group);cells were transfected with negative control small interfering RNA(siRNA)and long non-coding RNA(lncRNA)H19 siRNA(negative control siRNA group and H19 siRNA group);cells were transfected with negative control siRNA and H19 siRNA,respectively,and then,they were treated with 10μmol/L curcumin(negative control siRNA+curcumin group and H19 siRNA+curcumin group);the H19 siRNA was co-transfected with negative control miR inhibitor or miR-491-5p inhibitor into these cells(H19 siRNA+negative control inhibitor group and H19 siRNA+miR-4915p inhibitor group);H19 siRNA+negative control miR inhibitor or H19 siRNA+miR-491-5p inhibitor were co-transfected into the cells,and then,they were treated with 10μmol/L curcumin(H19 siRNA+negative control inhibitor+curcumin group and H19 siRNA+miR-491-5p inhibitor+curcumin group);the cells were co-transfected with miR-491-5p mimic+blank plasmid or miR-491-5p mimic+HOXA9 overexpression plasmid,and then they were treated with 10μmol/L curcumin(miR-491-5p mimic+blank plasmid+curcumin group and miR-491-5p mimic+HOXA9 overexpression plasmid+curcumin group);real-time fluorescent quantitative PCR(qRT-PCR)was used to detect the mRNA expressions of H19,miR-491-5p,and HOXA9;CCK-8 assay was used to detect the cell proliferation;flow cytometry was used to detect the cell apoptosis;plate cloning method was employed to detect the number of cell clone formation;Transwell assay was used to detect the cell migration;and the HOXA9 protein expression was measured by Western blotting.(2)The 293T cells at the logarithmic growth phase were chosen;the negative control miRNA mimics or miR-491-5p mimics combined with wild-type H19,wild-type HOXA93'-UTR plasmid vectors were co-transfected into the cells,respectively(negative control mimic+wi

关 键 词：姜黄素 神经胶质瘤 长链非编码RNA H19 miR-491-5p 同源盒基因A9 细胞特性 

分 类 号：R285[医药卫生—中药学]