文章摘要
人胃癌细胞Caveolin-1基因的表达与DNA甲基化修饰之间的关系研究
The Interaction Between Expression and Methylation Status of Caveolin-1 gene in Human Gastric Cancer Cells
投稿时间:2020-12-30  修订日期:2021-04-16
DOI:
中文关键词: 小窝蛋白-1  ?亚型  胃癌  甲基化修饰  启动子区  CpG岛  转录调控  基因表达
英文关键词: Caveolin-1  subtype ?  gastric cancer  methylation  promoter domain  CpG island  transcription  gene expression
基金项目:浙江省医药卫生科技计划项目青年人才计划(2019RC228和2019RC229);浙江省自然科学(LQ14H160014, LY21H030002);国家自然科学(81503297, 81400594, 81603340, 81773945和81770535)
作者单位邮编
毛立祺 浙江中医药大学附属第一医院浙江省消化道疾病病理生理研究重点实验室 杭州 310006
戴春艳 浙江中医药大学附属第一医院浙江省消化道疾病病理生理研究重点实验室 杭州 
金威洋 杭州师范大学生命与环境科学学院 杭州 
傅宇斐 浙江中医药大学附属第一医院浙江省消化道疾病病理生理研究重点实验室 杭州 
陈喆 浙江中医药大学附属第一医院浙江省消化道疾病病理生理研究重点实验室 杭州 
吕宾 浙江中医药大学附属第一医院浙江省消化道疾病病理生理研究重点实验室 
王曦 浙江中医药大学附属第一医院浙江省消化道疾病病理生理研究重点实验室 杭州 310006
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中文摘要:
      [目的]观察胃癌细胞系中Caveolin-1(Cav-1)基因不同亚型的表达水平,以及探讨影响其基因转录调控的甲基化修饰途径。[方法]选择表达不同水平Cav-1基因的胃癌细胞系,应用实时荧光定量PCR反应检测不同胃癌细胞系中Cav-1基因两个亚型a和b的mRNA水平,之后利用CpG岛在线预测软件对其启动子区和第一外显子区进行CpG岛分布情况进行分析,最后应用甲基化荧光定量PCR反应对其甲基化水平进行检测。[结果]qPCR结果显示,人胃癌细胞(MGC-803)中的Cav-1以及Cav-1a的表达分别是人正常胃粘膜上皮细胞(GES1)的1.782±0.489倍和2.604±0.945倍;另外,人胃癌细胞AGS中的Cav-1以及Cav-1a的表达则仅是GES1细胞的0.017±0.002倍和0.0005±0.0.0003倍,差异均有统计学意义(P<0.05);然而,相对于GES1细胞,MGC-803细胞和AGS细胞中Cav-1b的mRNA表达并无明显改变。利用CpG岛在线预测软件分析结果表明,人Cav-1a基因的启动子区存在一个174bp大小的CpG岛,包含了17个CG位点。甲基化qPCR检测发现,Cav-1a基因启动子区-158~-77区域处于去甲基化状态,则Cav-1a的mRNA表达水平较高,相反,该区域处于高度甲基化状态,则其mRNA表达较低。[结论]人胃癌细胞中Cav-1的表达主要与其a亚型的表达相关,与b亚型无关;并且其启动子区CpG岛内-158~-77区域的甲基化修饰程度与其mRNA转录调控水平密切相关。
英文摘要:
      [Objective] To observe the expression of different transcript variants Caveolin-1 (Cav-1) gene in human gastric cancer cells, and to investigate the epigenetic mechanism. [Methods] The gastric cancer cell lines expressing different levels of Cav-1 gene were selected and the mRNA levels of two subtypes of Cav-1 gene were detected by real time quantitative PCR (real time qPCR). Then the status of CpG island within the promoter region and the first exon region of Cav-1 gene was analyzed by the software of MethPrimer-Design Primers of Methylation PCRs. [Results] qPCR showed that the expression of Cav-1 and Cav-1 in human gastric cancer cells (MGC-803) was 1.782±0.489 folds and 2.604±0.945 folds respectively, compared with the normal gastric epithelial cells (GES1), whereas the expression of Cav-1 and Cav-1 in the other human gastric cancer cells (AGS) was only 0.017±0.002 and 0.0005±0.0.0003 folds of GES1 cells (P<0.05). However, there was no change of Cav-1 mRNA expression in MGC-803 and AGS cells. The analysis of CpG Island status prediction software showed that there was a CpG island located between -250~-77bp relative to +1 within the promoter region of Cav-1 gene, which was 174bp in length and contained 17 CG dinucleotides. The qMethyl-PCR assay showed that the demethylation of the region between -158~-77bp of Cav-1 gene was associated with the high level of its mRNA expression. Meanwhile, the methylation of this region was related with the low level. [Conclusion] The expression of Cav-1 in human gastric cancer cells is related to the expression of subtype a and has no correlation with the subtype, and the methylation of -158~-77 region within the promoter functioned as the major regulation domain of Cav-1 transcription activity.
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