过表达或沉默lncRNA SNHG8的脂肪干细胞对血管内皮细胞功能障碍的影响  

Effect of adipose-derived stem cells with overexpression or silencing of lncRNA SNHG8 on vascular endothelial cell dysfunction

作　　者：陈自强 胡晓咏 杨朝颖 邹婷[1] 吕忠英[1] 张颖[1] 王欢[1] 李红建[1] CHEN Ziqiang;HU Xiaoyong;YANG Zhaoying;ZOU Ting;LYU Zhongying;ZHANG Ying;WANG Huan;LI Hongjian(Department of Hypertension,Fifth Clinical Medical College,Xinjiang Medical University,Urumqi 830011,China)

机构地区：[1]新疆医科大学第五临床医学院高血压科,新疆乌鲁木齐830011

出　　处：《中国病理生理杂志》2024年第1期18-27,共10页Chinese Journal of Pathophysiology

基　　金：国家自然科学基金资助项目(No.82260089)。

摘　　要：目的:研究过表达或沉默长链非编码RNA(lncRNA)SNHG8的脂肪干细胞(ADSCs)对人脐静脉内皮细胞(HUVECs)活力、迁移、成管及表达血管活性因子的影响。方法:(1)流式细胞术及成脂、成骨诱导实验鉴定病态肥胖患者来源的脂肪干细胞(O-ADSCs);RT-qPCR检测健康人群来源的脂肪干细胞(H-ADSCs)及O-ADSCs内lncRNA SNHG8的表达。(2)Transwell建立ADSCs与HUVECs间接共培养48 h体系,设置O-ADSCs+HUVECs组、HADSCs+HUVECs组和HUVECs组,通过RT-qPCR和Western blot检测HUVECs内血管紧张素Ⅱ(AngⅡ)、内皮素1(ET-1)和内皮型一氧化氮合酶(eNOS)的mRNA及蛋白表达水平。(3)进一步构建lncRNA SNHG8过表达或沉默慢病毒并感染O-ADSCs,设置O-ADSCs-OE-SNHG8+HUVECs组、O-ADSCs-OE-NC+HUVECs组、O-ADSCs-sh-SNHG8+HUVECs组和O-ADSCs-sh-NC+HUVECs组,共培养48 h后,通过CCK-8实验、划痕实验和小管形成实验分别检测HUVECs活力、迁移能力和成管能力;RT-qPCR和Western blot检测HUVECs内AngⅡ、ET-1和eNOS的mRNA及蛋白表达水平;硝酸还原酶法检测HUVECs内NO含量。结果:(1)所培养细胞经鉴定符合ADSCs特征;(2)与H-ADSCs比较,lncRNA SNHG8在O-ADSCs中显著高表达(P<0.01);(3)与H-ADSCs+HUVECs和HUVECs组相比,O-ADSCs+HUVECs组中HUVECs内AngⅡ和ET-1的mRNA及蛋白表达水平上调(P<0.01);(4)过表达lncRNA SNHG8的OADSCs可增强HUVECs的活力、迁移能力及成管能力,上调HUVECs中AngⅡ和ET-1的mRNA及蛋白表达水平,下调eNOS的mRNA及蛋白表达水平,减少HUVECs内NO含量(P<0.05);而沉默O-ADSCs中lncRNA SNHG8的表达则呈现相反的结果(P<0.05)。结论:(1)O-ADSCs可通过旁分泌作用增强内皮细胞活力、迁移能力及成管能力;(2)过表达lncRNA SNHG8的O-ADSCs促使内皮细胞分泌舒张-收缩因子失衡,导致血管内皮细胞功能障碍。AIM:To investigate the effects of adipose-derived stem cells(ADSCs)with overexpression or silencing of long noncoding RNA(lncRNA)SNHG8 on the viability,migration,angiogenesis,and the expression of vasoactive factors in human umbilical vein endothelial cells(HUVECs).METHODS:Identification of ADSCs derived from morbidly obese patients(O-ADSCs)was conducted using flow cytometry and induction of lipogenesis and osteogenesis.The expression of lncRNA SNHG8 in healthy human ADSCs(H-ADSCs)and O-ADSCs was detected by RT-qPCR.Transwell method was used to establish the indirect co-culture system of ADSCs and HUVECs for 48 h,and the cells were divided into O-ADSCs+HUVECs group,H-ADSCs+HUVECs group,and HUVECs alone group.The mRNA and protein expression levels of angiotensinⅡ(AngⅡ),endothelin-1(ET-1)and endothelial nitric oxide synthase(eNOS)in HUVECs were detected by RT-qPCR and Western blot.The lncRNA SNHG8 overexpression and silencing lentiviruses were constructed and used to infect O-ADSCs.The indirect co-cultured ADSCs and HUVECs were divided into O-ADSCs-OE-SNHG8+HUVECs group,O-ADSCs-OE-NC+HUVECs group,O-ADSCs-sh-SNHG8+HUVECs group,and O-ADSCs-sh-NC+HUVECs group.After co-culture for 48 h,the viability,migration and tubule formation of HUVECs were detected by CCK-8,scratch and angiogenesis assays,respectively.The mRNA and protein expression levels of AngⅡ,ET-1 and eNOS in HUVECs were detected by RT-qPCR and Western blot,respectively.The nitrate reductase method was used to detect the content of NO in HUVECs.RESULTS:(1)The cultured cells were identified as ADSCs.(2)Compared with H-ADSCs,lncRNA SNHG8 expression was significantly up-regulated in O-ADSCs(P<0.01).(3)Compared with H-ADSCs+HUVECs group and HUVECs group,the mRNA and protein expression levels of AngⅡand ET-1 in HUVECs in O-ADSCs+HUVECs group were up-regulated(P<0.01).(4)Overexpression of lncRNA SNHG8 in O-ADSCs enhanced the viability,migration and tube formation ability of HUVECs,up-regulated the mRNA and protein expression levels of AngⅡand ET-1,down-re

关 键 词：肥胖相关性高血压 长链非编码RNA SNHG8 脂肪干细胞 人脐静脉内皮细胞 

分 类 号：R544.1[医药卫生—心血管疾病] R363.2[医药卫生—内科学] R34[医药卫生—临床医学]