过表达miR-515-5p的视网膜母细胞瘤细胞增殖、凋亡、侵袭能力变化及其机制  

Effects of overexpression of miR-515-5p on proliferation,apoptosis,and invasion of retinoblastoma cells

作　　者：鲍慧婧 刘萍[1] 杜允宏 刘文静[1] 马义宾[1] BAO Huijing;LIU Ping;DU Yunhong;LIU Wenjing;MA Yibin(Ophthalmology Department,Taian Central Hospital Affiliated to Qingdao University,Taian 271000,China)

机构地区：[1]青岛大学附属泰安市中心医院眼科,山东泰安271000

出　　处：《山东医药》2023年第11期19-23,共5页Shandong Medical Journal

基　　金：山东省自然科学基金资助项目(ZR2019PH065);山东省泰安市科技发展计划项目(2019NS110)。

摘　　要：目的探讨过表达miR-515-5p对视网膜母细胞瘤细胞增殖、凋亡、侵袭能力的影响及其机制。方法利用StarBase数据库进行生物信息学分析并预测miR-515-5p的靶基因为CBX4,并经双荧光素酶报告基因实验验证。选择视网膜母细胞瘤细胞系SO-RB50、Y79、HXO-RB-44细胞中miR-515-5p表达降低最明显的SO-RB50细胞进行实验,分为miR-515-5p NC组、miR-515-5p mimics组、miR-515-5p mimics+CBX4组、miR-515-5p mimics+CBX4-NC组,分别转染miR-515-5p阴性对照、miR-515-5p mimics、miR-515-5p mimics+CBX4过表达载体、miR-515-5p mimics+CBX4空载体。采用实时荧光定量PCR法检测各组miR-515-5p及CBX4 mRNA表达,Western blotting法检测CBX4蛋白表达,CCK-8法检测转染12、24、48、72 h的细胞增殖能力(OD值),TUNEL法检测细胞凋亡率,Transwell小室实验检测细胞侵袭能力。结果miR-515-5p mimics组、miR-515-5p mimics+CBX4组、miR-515-5p mimics+CBX4-NC组miR-515-5p相对表达量均高于miR-515-5p NC组(P均<0.05)。miR-515-5p mimics组、miR-515-5p mimics+CBX4-NC组、miR-515-5p mimics+CBX4组CBX4 mRNA相对表达量及培养48、72 h的OD值均低于miR-515-5p NC组,且miR-515-5p mimics组、miR-515-5p mimics+CBX4-NC组降低更明显(P均<0.05)。培养12 h,各组细胞OD值比较P>0.05。培养24 h,miR-515-5p mimics组、miR-515-5p mimics+CBX4-NC组OD值均低于miR-515-5p NC组(P均<0.05)。与miR-515-5p NC组、miR-515-5p mimics+CBX4组比较,miR-515-5p mimics组、miR-515-5p mimics+CBX4-NC组细胞凋亡率均升高,CBX4蛋白相对表达量及侵袭细胞数均降低(P均<0.05)。结论过表达miR-515-5p能够抑制视网膜母细胞瘤细胞SO-RB50的增殖、侵袭能力并促进其凋亡,其机制可能与抑制CBX4表达有关。Objective To investigate the effects of overexpression of miR-515-5p on proliferation,apoptosis,and invasion of retinoblastoma cells and their mechanism of action.Methods CBX4 was predicted as the target gene of miR-515-5p by bioinformatics analysis using StarBase database,which was verified by dual luciferase reporter gene assay.SORB50 cells with the most significant reduction in miR-515-5p expression among retinoblastoma cell lines SO-RB50,Y79 and HXO-RB-44 were selected for experiments.They were divided into the miR-515-5p NC group,miR-515-5p mimics group,miR-515-5p mimics+CBX4 group,and miR-515-5p mimics+CBX4-NC group;miR-515-5p NC,miR-515-5p mimics,miR-515-5p mimics+CBX4 overexpression vector,and miR-515-5p mimics+CBX4 empty vector were transfected,respectively.The expression of miR-515-5p and CBX4 mRNA in each group was detected by real-time fluorescent quantitative PCR,and the expression of CBX4 protein was detected by Western blotting.The cell proliferation ability(OD value)was detected by CCK-8 at 12,24,48,and 72 h after transfection.The apoptosis was detected by TUNEL experiment.Transwell chamber assay was used to detect cell invasion ability.Results The relative expression of miR-515-5p in the miR-515-5p mimics group,miR-515-5p mimics+CBX4 group,and miR-515-5p mimics+CBX4-NC group was significantly higher than that in miR-515-5p NC group(all P<0.05).The relative expression levels of CBX4 mRNA and protein in the miR-515-5p mimics group,miR-515-5p mimics+CBX4-NC group,and miR-515-5p mimics+CBX4 group were lower than those in the miR-515-5p NC group at 48 and 72 h;the decreased in the miR-515-5p mimics group and miR-515-5p mim-ics+CBX4-NC group was more significant(all P<0.05).At 12 of culture,no significant difference was found in the OD value of cells among groups(P>0.05).At 24 h of culture,the OD values of the miR-515-5p mimics group and miR-515-5p mimics+CBX4-NC group were lower than those of miR-515-5p NC group(all P<0.05).Compared with the miR-515-5p NC group and miR-515-5p mimics+CBX4 group,the apo

关 键 词：微小RNA-515-5p CBX4蛋白 细胞增殖 细胞凋亡 细胞侵袭 视网膜母细胞瘤 

分 类 号：R775[医药卫生—眼科]