miR-150-5p靶向GHSR调控结直肠癌细胞HT-29凋亡的研究  

Apoptosis of colorectal cancer cell line HT-29 regulated by miR-150-5p targeting GHSR

作　　者：李旭彤[1] 吴宝音[1] 曹双军[1] 李晋[1] 齐富贵 LI Xu-tong;WU Bao-yin;CAO Shuang-jun;LI Jin;QI Fu-gui(Department of General Surgery,Liangxiang Hospital,Fangshan District,Beijing 102401,China)

机构地区：[1]北京市房山区良乡医院普通外科,北京102401

出　　处：《中国现代普通外科进展》2020年第9期683-687,共5页Chinese Journal of Current Advances in General Surgery

摘　　要：目的:探讨miR-150-5p是否靶向GHSR调控结直肠癌细胞HT-29的凋亡。方法:TargetScan在线分析miR-150-5p与GHSR的相关性;将GHSR的3端非编码区克隆进pMir-Glo miRNA载体,利用双荧光素酶报告系统检测miR-150-5p是否靶向调控GHSR;在缺氧以及过表达或敲低miR-150-5p的情况下,通过Western Blot检测GHSR/AMPK/Sirt1/PGC-1α/UCP2通路蛋白表达量和细胞凋亡标志蛋白的表达量。使用Annexin-V染色检测结直肠癌细胞HT-29的凋亡水平。结果:miR-150-5p靶向GHSR的3’-UTR的66-72的位置。缺氧条件下,敲低miR-150-5p后,GHSR、Sirt1、UCP2、p-AMPK、PGC-1α的表达量和细胞凋亡相关蛋白BCL2的表达量均上升(P<0.05)、BAX、Cleaved caspase3表达量和细胞凋亡水平均下降(P<0.05);缺氧条件下,过表达miR-150-5p后,GHSR、Sirt1、UCP2、p-AMPK、PGC-1α的表达量和细胞凋亡相关蛋白BCL2的表达量均下降(P<0.05)、BAX和Cleaved caspase3表达量上升(P<0.05),细胞凋亡水平上升(P<0.05)。结论:缺氧条件下miR-150-5p通过靶向GHSR的mRNA调控GHSR/AMPK/Sirt1/PGC-1α/UCP2通路来促进结直肠癌细胞HT-29的凋亡。Objective:To investigate whether miR-150-5p targets GHSR to regulate apoptosis of colorectal cancer cell line HT-29.Methods:TargetScan was used to analyze the correlation between miR-150-5p and GHSR.Secondly,the 3-terminal non-coding region of GHSR was cloned into pMir-Glo miRNA vector,and the dual luciferase reporter system was used to detect whether miR-150-5p was targeted or not.GHSR;GHSR/AMPK/Sirt1/PGC-1α/UCP2 pathway protein expression and expression of apoptosis marker protein were detected by Western Blot in the absence of oxygen and overexpression or knockdown of miR-150-5p.Apoptosis levels of colorectal cancer cell line HT-29 were detected using Annexin-V staining.Results:miR-150-5p targets the 66-72 position of the 3'UTR of GHSR.Under hypoxia,the expression levels of GHSR,Sirt1,UCP2,p-AMPK and PGC-1αincreased after knockdown of miR-150-5p(P<0.05),and the expression of apoptosis-related protein BCL2 increased(P<0.05),BAX and Cleaved caspase3 expression decreased(P<0.05),apoptosis decreased(P<0.05);under hypoxic conditions,overexpressed miR-150-5p,GHSR,Sirt1,UCP2,p-AMPK The expression of PGC-1αdecreased(P<0.05),the expression of apoptosis-related protein BCL2 decreased(P<0.05),the expression of BAX and Cleaved caspase3 increased(P<0.05),and the level of apoptosis increased(P<0.05).Conclusion:miR-150-5p regulates GHSR/AMPK/Sirt1/PGC-1α/UCP2 pathway by targeting GHSR mRNA under hypoxia to promote apoptosis of colorectal cancer cell line HT-29.

关 键 词：miR-150-5p GHSR 结直肠癌细胞HT-29 细胞凋亡 缺氧 

分 类 号：R735.34[医药卫生—肿瘤]