摘要
目的探索microRNA let-7a-3在结直肠癌(CRC)中的表达状态、生物学功能及可能的机制。方法使用联合亚硫酸氢钠限制性内切酶分析法(COBRA)或亚硫酸氢盐测序法(BSP)分析let-7a-3启动子内的DNA甲基化状态。使用实时定量聚合酶链反应(RT-PCR)检测CRC细胞系中let-7a-3和RAB11FIP2的表达。使用流式细胞仪和锚定-非依赖性生长试验检测let-7a-3表达上调时CRC细胞凋亡水平及集落形成能力。结果let-7a-3基因甲基化在CRC组织中很常见,let-7a-3的甲基化状态与组织来源有关(P<0.001)。去甲基化剂5-氮杂-2-脱氧胞苷(5-aza-dC)可以诱导let-7a-3的表达。let-7a-3表达上调时促进细胞凋亡并抑制细胞集落形成能力(P<0.001)。let-7a-3通过靶向RAB11FIP2基因3’-UTR区域中的同源DNA区域抑制RAB11FIP2的表达。结论CRC中let-7a-3甲基化是一种常见现象,let-7a-3过表达可诱导细胞凋亡,抑制锚定非依赖性生长,提示let-7a-3可能是结直肠癌治疗的潜在生物标志物。
Objective To explore the expression status,biological function and possible mechanism of microRNA let-7a-3 in colorectal cancer(CRC).Methods The DNA methylation status within the let-7a-3 promoter was analyzed using combined bisulfite restriction analysis(COBRA)or bisulfite sequencing PCR(BSP).Expression levels of let-7a-3 and RAB11FIP2 in CRC cell lines were detected using real-time quantitative polymerase chain reaction(RT-PCR).Flow cytometry and anchorage-independent growth assay were used to detect the level of apoptosis and colony-forming ability of CRC cells when the expression of let-7a-3 was up-regulated.Results The methylation of let-7a-3 gene was common in CRC tissues,and the methylation status of let-7a-3 was related to the tissue origin(P<0.001).The demethylating agent 5-aza-2’-deoxycytidine(5-aza-dC)can induce the expression of let-7a-3.When let-7a-3 expression was upregulated,it promoted apoptosis and inhibited the ability of colony formation(P<0.001).Let-7a-3 inhibits RAB11FIP2 expression by targeting a homologous DNA region in the 3’-UTR region of the RAB11FIP2 gene.Conclusion Let-7a-3 methylation is a common phenomenon in CRC.Overexpression of let-7a-3 can induce cell apoptosis and inhibit anchorage-independent growth,suggesting that let-7a-3 may be a potential biomarker for colorectal cancer therapy.
作者
张利苹
董文杰
李静文
陈璐璐
ZHANG Liping;DONG Wenjie;LI Jingwen;CHEN Lulu(Department of Oncology,the First Affiliated Hospital of Zhengzhou University,Zhengzhou 450052,China)
出处
《河南医学研究》
CAS
2023年第2期197-203,共7页
Henan Medical Research
基金
国家自然科学基金资助项目(81672424)。
