LncRNA NEAT1调节微小RNA-144-3p/NOVA1轴对高糖诱导的人视网膜微血管内皮细胞损伤的影响  

Influence of LncRNA NEAT1 on high glucose-induced injury of human retinal microvascular endothelial cells by regulating micro RNA-144-3p/NOVA1 axis

作　　者：周云云 邓展锋 刘萍[2] 吴年周 陈琳瑶 ZHOU Yunyun;DENG Zhanfeng;LIU Ping;WU Nianzhou;CHEN Linyao(Department of Ophthalmology,Second Affiliated Hospital of Shaoyang University,Shaoyang,Hunan 422000,China;Department of Ophthalmology,First Affiliated Hospital of South China University,Hengyang,Hunan 421200,China)

机构地区：[1]邵阳学院附属第二医院眼耳鼻喉科,湖南邵阳422000 [2]南华大学附属第一医院眼科,湖南衡阳421200

出　　处：《岭南心血管病杂志》2023年第5期551-557,562,共8页South China Journal of Cardiovascular Diseases

摘　　要：目的探究长链非编码RNA(long non-coding RNA,LncRNA)核富含丰富的转录本1(nuclear-enriched abundant transcript 1,NEAT1)通过微小RNA(micro RNA,miR)-144-3p/神经肿瘤腹侧抗原1(neural tumor ventral antigen 1,NOVA1)对高糖诱导的人视网膜微血管内皮细胞(human retinal microvascular endothelial cells,hRMECs)损伤的影响。方法将h RMECs分为高糖组(不转染)、对照组(不转染)、高糖+si-NEAT1-阴性对照(negative control,NC)组(转染siRNA-NC)、高糖+si-NEAT1组(转染NEAT1 siRNA)、高糖+si-NEAT1+抑制剂-NC组(共转染NEAT1 siRNA+抑制剂-NC)、高糖+si-NEAT1+miR-144-3p抑制剂组(共转染NEAT1 siRNA+miR-144-3p抑制剂)、高糖+si-NOVA1-NC组(转染NOVA1 siRNA-NC)、高糖+si-NOVA1组(转染NOVA1 siRNA)。实时荧光定量聚合酶链反应技术(real-time fluorescent quantitative polymerase chain reaction,qRT-PCR)检测hRMECs中NEAT1、miR-144-3p表达量;CCK-8、Transwell和血管形成实验分别分析hRMECs增殖、迁移和血管形成能力;双荧光素酶报告实验分析NEAT1与miR-144-3p、miR-144-3p与NOVA1的结合情况;Western blot检测NOVA1、VEGF蛋白表达。结果与对照组比较,高糖组hRMECs中NEAT1表达(2.67±0.13 vs.1.02±0.08)、相对活力、迁移数量[(271.50±20.35)个vs.(144.30±16.41)个]、成管节点数[(426.50±22.75)个vs.(181.70±12.43)个]、管长度[(12725.46±136.57)μm vs.(8009.32±107.32)μm]均较高,miR-144-3p表达(0.46±0.06 vs.1.00±0.05)较低,差异有统计学意义(P<0.05)。敲低NEAT1可明显下调h RMECs中NEAT1的表达,上调miR-144-3p的表达,同时降低NOVA1(0.80±0.09 vs.1.35±0.07)、血管内皮生长因子(vascular endothelial growth factor,VEGF)蛋白表达及hRMECs的相对活力、迁移数量[(231.60±14.25)个vs.(271.50±20.35)个]、成管节点数[(234.30±13.18)个vs.(426.50±22.75)个]和管长度[(9208.74±130.85)μm vs.(12725.46±136.57)μm],差异有统计学意义(P<0.05),上述作用可被miR-144-3p抑制剂减弱。NEAT1可结合并下调miR-144-3p表达,且NOVA1为miObjectives To investigate the influence of long non-coding RNA(LncRNA)nuclear-enriched abundant transcript 1(NEAT1)on high glucose-induced injury of human retinal microvascular endothelial cells(hRMECs)through micro RNA(miR)-144-3p/neural tumor ventral antigen 1(NOVA1).Methods HRMECs were separated into high glucose group(no transfection),control group(no transfection),high glucose+si-NEAT1-negative control(NC)group(transfected with siRNA-NC),and high glucose+si-NEAT1 group(transfected with NEAT1 siRNA-NC),high glucose+si-NEAT1+inhibitor-NC group(co-transfected NEAT1 siRNA+inhibitor-NC),high glucose+si-NEAT1+miR-144-3p inhibitor group(co-transfected with NEAT1 siRNA+miR-144-3p inhibitor),high glucose+si-NOVA1-NC group(transfected with NOVA1 siRNA-NC),and high glucose+si-NOVA1 group(transfected with NOVA1 siRNA).The expression levels of NEAT1 and miR-144-3p in hRMECs were detected by real-time fluorescent quantitative polymerase chain reaction(qRT-PCR);CCK-8,Transwell and angiogenesis assays were used to analyze the proliferation,migration and angiogenesis ability of hRMECs;the binding of NEAT1 to miR-144-3p,and the binding of miR-144-3p to NOVA1 were analyzed by dual luciferase reporter assay;Western blot was used to detect the expressions of NOVA1 and VEGF proteins.Results Compared with control group,the expression of NEAT1(2.67±0.13 vs.1.02±0.08),relative viability,migration numbers[(271.50±20.35)individual vs.(144.30±16.41)individual],number of tube-forming nodes[(426.50±22.75)individual vs.(181.70±12.43)individual]and tube length[(12725.46±136.57)μm vs.(8009.32±107.32)μm]were higher in hRMECs in high glucose group,while the expression of miR-144-3p(0.46±0.06 vs.1.00±0.05)was lower(P<0.05).Knockdown of NEAT1 could significantly down-regulate the expression of NEAT1 in hRMECs,up-regulate the expression of miR-144-3p,and at the same time decrease the expression of NOVA1(0.80±0.09 vs.1.35±0.07)and vascular endothelial growth factor(VEGF)proteins,the relative viability,migration numbers[(231.60±14.2

关 键 词：长链非编码RNA核富含丰富的转录本1 微小RNA-144-3p 人视网膜微血管内皮细胞 神经肿瘤腹侧抗原1 增殖 血管形成- 

分 类 号：R33[医药卫生—人体生理学]