MiR-96-5p靶向调控MTSS1表达对结肠癌细胞恶性生物学行为的影响  

MiR-96-5p Inhibits the Development of Colon Cancer by Targeting MTSS1

作　　者：许春蕾[1] 李娜[1] 汤旭山[1] XU Chunlei;LI Na;TANG Xushan(Tumor Hospital Affiliated to Xinjiang Medical University,Urumqi,830011)

机构地区：[1]新疆医科大学附属肿瘤医院,830011

出　　处：《实用癌症杂志》2020年第8期1219-1225,共7页The Practical Journal of Cancer

基　　金：新疆维吾尔自治区自然科学基金项目(编号:2018D01C217)。

摘　　要：目的探讨miR-96-5p对人结肠癌细胞生物学行为的影响及其作用机制。方法应用miRNA转染技术,将miR-96-5p抑制物(miR-96-5p inhibitor)及microRNA无关序列(miR-NC)转染入结肠癌HCT116细胞中;采用免疫荧光及RT-PCR验证转染效率后,分别利用MTT、划痕实验、Transwell、凋亡实验检测miR-96-5p对结肠癌细胞的增殖、迁移、侵袭、凋亡能力的影响;采用裸鼠皮下成瘤实验检测miR-96-5p对人结肠癌细胞成瘤能力的影响;miRNA靶基因预测软件分析miR-96-5p潜在靶目标并利用双荧光素酶报告基因和Western blot实验进行验证。结果免疫荧光及RT-PCR结果提示miR-96-5p inhibitor显著下调miR-96-5p在HCT116细胞中的表达。MTT、划痕实验、Transwell及凋亡实验结果提示下调miR-96-5p表达显著抑制结肠癌细胞的增殖、迁移、侵袭能力并且促进细胞发生凋亡。裸鼠皮下成瘤实验提示过表达miR-96-5p能够促进结肠癌细胞成瘤能力,下调miR-96-5p的表达能够明显削弱其成瘤能力。miRNA靶基因预测软件分析提示MTSS1可能为miR-96-5p目标靶基因;双荧光素酶报告基因验证了miR-96-5p对MTSSI的靶向调控,且Western blot实验结果提示下调miR-96-5p可明显促进MTSS1蛋白在结肠癌细胞中的表达。结论下调miR-96-5p的表达可抑制结肠癌细胞的增殖、侵袭、迁移能力并促进细胞发生凋亡,这种作用可能通过对MTSS1的靶向调控来实现。Objective To investigate the effect of miR-96-5p on the biological behavior of human colon cancer cells and to explore its mechanism.Methods MiR-96-5p inhibitor(miR-96-5p inhibitor)and microRNA-independent sequence(miR-NC)were transfected into colon cancer HCT116 cells by microRNA transfection technique.Immunofluorescence and RT-PCR were used to verify the transfection efficiency.MTT,scratch test,Transwell and apoptosis test were used to detect the effects of miR-96-5p on the proliferation,migration,invasion and apoptosis of colon cancer cells.The subcutaneous tumorigenesis of nude mice was used to detect the effect of miR-96-5p on the tumorigenicity of human colon cancer cells.The potential target of miR-96-5p was analyzed by predictive software of microRNA target gene and verified by double luciferase reporter gene and Western blot.Results Immunofluorescence and RT-PCR results indicated that the transfected cells were stable,and the expression of miR-96-5p was significantly down-regulated by miR-96-5p inhibitor.MTT,scratch test,Transwell and apoptotic test indicated that down-regulation of miR-96-5p expression significantly inhibited the proliferation,migration and invasion of colon cancer cells and promoted cell apoptosis.Subcutaneous tumorigenesis in nude mice showed that over-expression of miR-96-5p could promote the tumorigenicity of colon cancer cells,and down-regulation of miR-96-5p could significantly weaken the tumorigenicity of colon cancer cells.Analysis of microRNA target gene prediction software suggested that MTSS1 might be the target gene of miR-96-5p.Dual luciferase reporter gene validates the targeted regulation of MTSSI by miR-96-5p,and Western blot results suggest that down-regulation of miR-96-5p can significantly promote the expression of MTSS1 protein in colon cancer cells.Conclusion Downregulation of the expression of miR-96-5p can inhibit the proliferation,invasion,migration and promote apoptosis of colon cancer cells.This effect may be achieved through targeted regulation of MTSS1.

关 键 词：miR-96-5p 结肠癌 MTSS1 

分 类 号：R735.35[医药卫生—肿瘤]