| 毛立祺,戴春艳,金威洋,等.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[J].浙江中医药大学学报,2021,45(9):939-944. |
| 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 |
| DOI：10.16466/j.issn1005-5509.2021.09.001 |
| 中文关键词: 小窝蛋白-1 亚型 胃癌 甲基化 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 regulation 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、2019RC229)；浙江省自然科学基金项目(LQ14H160014、LY21H030002)；国家自然科学基金项目(81503297、81400594、81603340、81773945、81770535) |
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| 中文摘要: |
| [目的]观察胃癌细胞系中小窝蛋白-1(Caveolin-1，Cav-1)基因不同亚型的表达水平，探讨影响其基因转录调控的甲基化修饰途径。[方法]选择Cav-1基因表达水平不同的胃癌细胞系，应用实时荧光定量聚合酶链式反应(Real-time quantitative polymerase chain reaction，Real-time qPCR)检测不同胃癌细胞系中Cav-1基因的两个亚型α和β的mRNA水平，之后利用磷酸胞苷酰(基)鸟苷(cytidylyl phosphate guanosine，CpG)岛在线预测软件对其启动子区和第一外显子区CpG岛分布情况进行分析，最后应用甲基化qPCR对其甲基化水平进行检测。[结果]Real-time qPCR结果显示，人胃癌细胞MGC-803中Cav-1以及Cav-1α的表达分别是人正常胃黏膜上皮细胞GES1的(1.782±0.489)倍和(2.604±0.945)倍，差异均有统计学意义(P＜0.05，P＜0.01)；人胃癌细胞AGS中的Cav-1以及Cav-1α的表达则仅是GES1细胞的(0.017±0.002)倍和(0.0005±0.00003)倍，差异均有统计学意义(P＜0.001，P＜0.001)。相对于GES1细胞，MGC-803细胞和AGS细胞中Cav-1β的mRNA表达差异无统计学意义(P＞0.05)。利用CpG岛在线预测软件分析结果表明，人Cav-1α基因的启动子区存在一个174bp大小的CpG岛，该岛位于相对于转录起始位点(transcription start stecte，TSS)+1的-250～-77区域内，包含了17个CG位点。甲基化qPCR检测发现，Cav-1α基因启动子区-158～-77区域处于去甲基化状态，则Cav-1α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α亚型的表达相关，与β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 polymerase chain reaction(Real-time qPCR). Then the status of cytidylyl phosphate guanosine(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] Real-time qPCR showed that compared with normal gastric epithelial cells GES1， the expression of Cav-1 and Cav-1α in human gastric cancer cells MGC-803 was (1.782±0.489) folds and(2.604±0.945) folds respectively， the difference was statistically significant(P＜0.05，P＜0.01); whereas the expression of Cav-1 and Cav-1α in the other human gastric cancer cells AGS was only (0.017±0.002) and (0.0005±0.00003) folds of GES1 cells(P＜0.001，P＜0.001). However， compared with GES1 cells， there was no significant difference of Cav-1β mRNA expression in MGC-803 and AGS cells(P＞0.05). The analysis of CpG island status prediction software showed that there was a CpG island located between-250～-77bp relative to +1 within transcription start state(TSS) of Cav-1 gene， which was 174bp in length and contained 17 CG dinucleotides. The Methyl-qPCR assay showed that the demethylation of the region between-158～-77 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 α and has no correlation with the subtype β， 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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