免疫分子B7-H3对人脐静脉内皮细胞功能的影响  

The effect of B7-H3 on human umbilical vein endothelial cell

作　　者：孙静[1] 杨洁 吴萍萍[1] 傅丰庆[2] 刘翠平[2] 陈瀚卿 郭云娣[1] SUN Jing;YANG Jie;WU Ping-ping;FU Feng-qing;LIU Cui-ping;CHEN Han-qing;GUO Yundi(The Institute of Medical Biotechnology,Suzhou Vocational Health College,Suzhou 215009,China;Institute of Clinical Immunology,The First Affiliated Hospital of Soochow University,Suzhou215001,China)

机构地区：[1]苏州卫生职业技术学院医学生物技术中心,苏州215009 [2]苏州大学附属第一医院临床免疫研究所,苏州215001

出　　处：《现代免疫学》2020年第3期187-191,共5页Current Immunology

基　　金：国家自然科学基金(31170836);江苏省青年医学人才(2017772);江苏省工程技术研发中心(2018026)。

摘　　要：设计针对B7-H3 1041以及996位点的shRNA,构建相应慢病毒B7-H3载体,研究其表达下调对人脐静脉内皮细胞(human umbilical vein endothelial cell,HUVEC)生物学行为的影响。针对B7-H3mRNA序列设计2对不同位点的shRNA,克隆连接至慢病毒载体中,与穿梭载体LV3共转染至人肾上皮细胞系293T细胞并包装病毒,逐孔稀释法测定病毒滴度。用聚凝胺浸染HUVEC细胞株,real-time PCR、FACS测定靶细胞B7-H3及其蛋白表达抑制情况,CCK-8法检测细胞增殖能力,Transwell实验分析其迁移能力。结果显示,慢病毒载体可抑制B7-H3及其蛋白的表达,抑制率超过84.7%;CCK-8实验显示B7-H3表达降低后HUVEC的增殖和迁移能力明显降低。以上发现可为进一步研究该分子的生物学功能及其在RA中的作用奠定基础。In this study, we constructed a specific lentivirus expression vector of B7-H3 through designing shRNA corresponding to sites 1041 and 996 to study the effect of B7-H3 on the biological behavior of HUVEC(human umbilical vein endothelial cell). Two shRNAs were designed according to B7-H3 mRNA sequence and were cloned into lentivirus vectors. They were co-transfected with shuttle plasmid LV3 into 293 T cells. Viruses were packaged and titers were determined by pore-by-pore dilution method. The virus infected HUVEC line through polybrene and real-time PCR, FACS were performed to measure the inhibition effect. CCK-8 assays and Transwell migration experiments were carried out to analyze the proliferation and migration abilities. The results showed that lentiviruses could inhibit the expression of B7-H3 and its protein, the inhibition rate was over 84.7%. CCK-8 proliferation experiment showed that B7-H3 inhibition decreased the proliferation and migration ability of HUVEC. Our results will lay a foundation for further studying the biological function of B7-H3 and its function in RA.

关 键 词：B7-H3 慢病毒载体 人脐静脉内皮细胞 增殖能力 迁移能力 

分 类 号：R392.11[医药卫生—免疫学]