miR-193b-3p过表达的HCT116外泌体对HUVEC增殖凋亡、迁移、血管形成影响及其与PAK4靶向关系  

Effects of exosomes from HCT116 cells overexpressing miR-193b-3p on proliferation,apoptosis,migration and angiogenesis of HUVEC and their targeted relationship with PAK4

作　　者：孙玮螺 刘素英 李思锦 周龙妹 何培元[1] SUN Weiuo;LIU Suying;LI Sijin;ZHOU Longmei;HE Peiyuan(Department of Gastroenterology,The Affiliated Hospital of Chengde Medical University,Chengde 067000,China)

机构地区：[1]承德医学院附属医院消化内科,河北承德067000

出　　处：《山东医药》2024年第24期13-18,共6页Shandong Medical Journal

基　　金：2023年度河北省医学科学研究课题计划项目(20231365)。

摘　　要：目的观察微小核糖核酸193b-3p(miR-193b-3p)过表达的人结直肠癌(CRC)细胞(HCT116)外泌体对人脐静脉内皮细胞(HUVEC)增殖凋亡、迁移、血管形成影响,并分析miR-193b-3p与p21活化蛋白激酶4(PAK4)的靶向关系。方法收集CRC患者血清和HCT116细胞上清外泌体,透射电镜实验和纳米颗粒示踪分析观察和鉴定外泌体,RT-q PCR实验检测外泌体miR-193b-3p和PAK4 m RNA。取人脐静脉内皮细胞(HUVEC),并分组,PBS组加入100μL的PBS,HCT116-Exo组加入100μL的HCT116细胞外泌体,HCT116-Exo+GW4869组先用GW4869(10μmol/L)处理30 min,然后加入100μL的HCT116细胞外泌体,采用Ed U实验、Transwell实验和成管实验检测各组HUVEC细胞增殖、迁移和血管形成能力。取对数生长期的HUVEC细胞,并分为miR-NC组、miR-193b-3p组,miR-NC组与转染miRNA阴性对照(miR-NC)的HCT116细胞外泌体共培养,miR-193b-3p组与转染miR-193b-3p模拟物(miR-193b-3p mimics)的HCT116细胞外泌体共培养,采用CCK8实验、流式细胞术实验和成管实验检测各组HUVEC细胞活性、细胞周期、细胞凋亡和血管形成能力。双荧光素酶报告基因系统检测miR-193b-3p和PAK4的靶向关系。结果CRC患者血清和细胞外泌体miR-193b-3p表达升高,PAK4 m RNA表达降低(P均<0.05);CRC细胞外泌体可促进HUVEC细胞的增殖、迁移和血管形成(P均<0.05);过表达CRC细胞外泌体miR-193b-3p增加HUVEC细胞活力和血管形成、加速细胞周期进程、抑制细胞凋亡(P均<0.05);miR-193b-3p直接靶向PAK4抑制其荧光素酶活性(P<0.05)。结论CRC血清、癌细胞外泌体miR-193b-3p表达升高,加入HCT116细胞外泌体的HUVEC细胞增殖、迁移和血管形成能力增强,加入转染miR-193b-3p mimics HCT116细胞外泌体的HUVEC细胞增殖、S期占比、血管形成能力升高及G_(0)/G_(1)期占比、凋亡率下降,miR-193b-3p与PAK4存在靶向关系。Objective To observe the effects of exosomes from human colorectal cancer(CRC)cells(HCT116)with overexpression of microRNA 193b-3p(miR-193b-3p)on the proliferation,apoptosis,migration and angiogenesis of human umbilical vein endothelial cells(HUVECs),and to analyze the targeted relationship between miR-193b-3p and p21-activated kinase 4(PAK4).Methods Serum from CRC patients and exosomes in the supernatant of HCT116 cells were collected.Transmission electron microscopy and nanoparticle tracking analysis were used to observe and identify exo-somes.RT-qPCR was performed to detect exosomal miR-193b-3p and PAK4 mRNA.HUVECs were taken and grouped.HUVECs in the PBS group were added with 100µL of PBS,HUVECs in the HCT116-Exo group were added with 100µL of HCT116 cell exosomes,and HUVECs in the HCT116-Exo+GW4869 group were first treated with GW4869(10µmol/L)for 30 min and then were added with 100µL of HCT116 cell exosomes.The proliferation,migration and angiogenesis abili-ties of HUVECs in each group were detected by EdU assay,Transwell assay and tube formation assay.HUVECs in the log-arithmic growth phase were taken and divided into the miR-NC group and the miR-193b-3p group.HUVECs in the miR-NC group were co-cultured with HCT116 cell exosomes transfected with miRNA negative control(miR-NC),and the miR-193b-3p group with HCT116 cell exosomes transfected with miR-193b-3p mimics.The activity,cell cycle,apoptosis and angiogenesis ability of HUVECs in each group were detected by CCK-8 assay,flow cytometry and tube formation as-say.The targeted relationship between miR-193b-3p and PAK4 was detected by the dual-luciferase reporter gene sys-tem.Results The expression of miR-193b-3p in serum and exosomes of CRC patients increased,while the expression of PAK4 mRNA decreased(both P<0.05);exosomes could promote the proliferation,migration,and angiogenesis of HU-VECs(all P<0.05).Overexpression of miR-193b-3p increased the cell viability and angiogenesis,accelerated cell cycle progression,and inhibited the apoptosis of HUVECs(all

关 键 词：结直肠癌 外泌体 微小核糖核酸193b-3p P21活化蛋白激酶4 细胞增殖 细胞迁移 细胞凋亡 细胞周期 血管生成 

分 类 号：R735.35[医药卫生—肿瘤]