摘要
目的:探究miR-383-5p通过调控DNA错配修复基因MSH6对小儿髓母细胞瘤(medulloblastoma,MB)增殖和侵袭的影响。方法:选取2014年7月至2017年5月于我院肿瘤外科手术切除并经病理确诊为MB的15例患者癌组织及相应的癌旁组织标本,qPCR检测MB组织和细胞系中miR-383-5p的表达水平。将实验分为对照组(NC组)、miR-383-5p过表达组(miR-383-5p组)、MSH6敲降组(si-MSH6组)及MSH6+miR-383-5p同时敲降组(miR-383-5p inhibitor+si-MSH6),采用CCK-8法检测UW473细胞的增殖活性,Transwell法检测UW473细胞侵袭和迁移能力,双荧光素酶报告基因验证miR-383-5p与MSH6的靶向关系,Western blotting(WB)法检测MSH6的表达水平。结果:miR-383-5p在MB组织和细胞系中表达水平显著低于癌旁组织(均P<0.05或P<0.01);过表达miR-383-5p显著抑制UW473细胞增殖、迁移和侵袭(均P<0.05或P<0.01),且下调MSH6在UW473细胞中的表达水平(P<0.01)。双荧光素酶报告基因结果证实miR-383-5p靶向结合MSH6的3'UTR,敲降MSH6可抑制UW473细胞增殖、侵袭和迁移(均P<0.01)。进一步实验证明,同时敲降miR-383-5p和MSH6能够恢复敲降MSH6对UW473细胞增殖、侵袭和迁移的抑制作用(均P<0.01)。结论:miR-383-5p在MB组织和细胞系中低表达,其通过靶向下调MSH6表达水平进而抑制UW473细胞的增殖、迁移及侵袭。
Objective:To investigate the effect of miR-383-5p on the proliferation and invasion of medulloblastoma(MB)by targeting DNA mismatch repair MSH6 gene.Methods:A total of 15 pairs of tumor tissues and corresponding adjacent tissues from MB patients,who were surgically treated and pathologically confirmed in the Department of Oncology of the Second Affiliated Hospital of Hainan Medical College from July 2014 to May 2017,were collected for this study.qPCR was applied to detect the expression of miR-383-5p in MB tissues and cell lines.The experimental cells were divided into control group(NC group),miR-383-3p overexpression group(miR-383-5p group),MSH6 knockdown group(si-MSH6 group)and miR-383-5p inhibitor+si-MSH6 group.CCK-8 assay was used to detect the proliferation of UW473 cells.Transwell assay was used to examine the invasion and migration of UW473 cells,the targeting relationship between miR-383-5p and MSH6 was verified by Dual-luciferase reporter gene assay,and Western blotting(WB)was performed to detect the protein expression of MSH6.Results:The expression level of miR-383-5p was significantly down-regulated in MB tissues and cell lines compared with para-cancer tissues(all P<0.05 or P<0.01).Overexpression of miR-383-5p significantly in-hibited the proliferation,migration and invasion of UW473 cells(all P<0.05 or P<0.01),and down-regulated the expression level of MSH6(all P<0.01).Dual-luciferase reporter gene assay demonstrated that miR-383-5p could targetedly bind to the 3'UTR of MSH6.Knockdown of MSH6 could inhibit the proliferation,invasion and migration of UW473 cells(all P<0.01).Further experiments showed that simultaneous knockdown of miR-383-5p and MSH6 could attenuate the inhibition of MSH6 silence on the proliferation,invasion and migration of UW473 cells.Conclusion:miR-383-5p expression is down-regulated in MB tissues and cell lines,and miR-383-5p suppresses the proliferation,migration and invasion of UW473 cells via targetedly down-regulating MSH6.
作者
陈石娇
陈妮
陈运旺
CHEN Shijiao;CHEN Ni;CHEN Yunwang(Department of Pediatrics,the Second Affiliated Hospital of Hainan Medical College,Haikou 570311,Hainan,China)
出处
《中国肿瘤生物治疗杂志》
CAS
CSCD
北大核心
2020年第9期1024-1029,共6页
Chinese Journal of Cancer Biotherapy
