微小RNA-4463对人瘢痕疙瘩成纤维细胞增殖和侵袭的影响  

Effect of miR-4463 on proliferation and invasion of human keloid fibroblasts

作　　者：高栋梁[1] 张雷[1] 张宏峰[1] 杨喜明[1] GAO Dongliang;ZHANG Lei;ZHANG Hongfeng;YANG Ximing(Burn and Hand Plastic Surgery,Yan'an University Affiliated Hospital,Yan'an,Shaanxi 716000,China)

机构地区：[1]延安大学附属医院烧伤整形手外科,陕西延安716000

出　　处：《安徽医药》2021年第6期1140-1143,I0004,共5页Anhui Medical and Pharmaceutical Journal

摘　　要：目的研究微小RNA-4463(miR-4463)对人瘢痕疙瘩成纤维细胞增殖、迁移、侵袭的影响以及机制。方法收集延安大学附属医院2017年12月至2019年6月整形手术病人40例,每例切除瘢痕疙瘩组织1个以及邻近正常皮肤组织1个,利用定量聚合酶链反应(qPCR)检测人正常成纤维细胞和瘢痕疙瘩成纤维细胞中miR-4463的表达量;将miR-4463模拟物对照质粒和miR-4463模拟物分别在Lipofectamine 2000介导下转染入瘢痕疙瘩成纤维细胞,分为miR-NC组和miR-4463组;常规培养的瘢痕疙瘩成纤维细胞作为对照;qPCR检测转染后miR-4463的表达量;qPCR检测上调miR-4463对瘢痕疙瘩纤维化相关基因Col1 A1、Col 3 A1表达的影响;细胞计数试剂盒(CCK-8)检测上调miR-4463对瘢痕疙瘩成纤维细胞增殖的影响,Transwell实验检测细胞的迁移、侵袭;采用TargetScan软件预测miR-4463与B细胞淋巴瘤-2(Bcl-2)相关的致病基因BAG4的靶向关系,双荧光素酶报告基因进一步进行验证。结果miR-4463在瘢痕疙瘩成纤维细胞中的表达量较人正常成纤维细胞显著降低[(0.218±0.036)比(1.000±0.071),P<0.05];与对照组相比,转染miR-4463模拟物明显增加瘢痕疙瘩成纤维细胞中miR-4463的表达量[(1.000±0.073)比(3.313±0.242),P<0.05];上调miR-4463显著抑制瘢痕疙瘩纤维化相关基因Col 1 A1[(1.000±0.065)比(0.334±0.010)]、Col 3 A1[(1.000±0.070)比(0.301±0.008)]的表达量(P<0.05);上调miR-4463抑制瘢痕疙瘩成纤维细胞增殖[(0.836±0.081)比(0.656±0.072),P<0.05],降低其迁移[(153.269±12.471)个比(82.363±7.254)个]、侵袭细胞数[(73.623±6.451)个比(36.459±3.235)个](P<0.05);BAG4是miR-4463下游靶基因。结论miR-4463可能通过调控BAG4抑制细胞外基质中纤维化相关胶原基因的表达,抑制瘢痕疙瘩成纤维细胞增殖、迁移、侵袭。Objective To study the effects of microRNA-4463(miR-4463)on the proliferation,migration and invasion of human keloid fibroblasts and its mechanism.Methods Forty patients with plastic surgery in Yan’an University Affiliated Hospital from December 2017 to June 2019 were collected.One keloid tissue and one adjacent normal skin tissue were excised from each patient.The expression of mir-4463 in human normal fibroblasts and keloid fibroblasts was detected by quantitative polymerase chain reaction(qPCR).The control plasmid and the mimic of miR-4463 were respectively transfected into keloid fibroblasts by Lipofectamine 2000 and assigned into miR-NC group and miR-4463 group,while the conventional cultured keloid fibroblasts were selected as controls.The expression of miR-4463 after transfection was detected by qPCR.qPCR was used to detect the effect of up-regulated miR-4463 on the expression of keloid fibrosis related genes Col 1 A1 and Col 3 A1.Cell counting kit-8(CCK-8)was used to detect the effect of up-regulation of miR-4463 on the proliferation of keloid fibroblasts.The cell migration and invasion were analyzed by Transwell assay.The targeting relationship between miR-4463 and Bcl-2-associated athanogene(BAG4)was predicted by TargetScan and further validated by double luciferase reporter gene.Results The expression of miR-4463 in keloid fibroblasts was significantly lower than that in human normal fibroblasts[(0.218±0.036)vs.(1.000±0.071),P<0:05].Compared with the control group,the expression of mill-4463 in keloid fibroblasts transfection with miR-4463 mimics was significantly increased[(1.000±0.073)vs.(3.313±0.242),(P<0.05)].Up-regulation of miR-4463 significantly inhibited the expression of Col 1 A1[(1.000±0.065)vs.(0.334±0.010)]and Col 3 A1[(1.000±0.070)vs.(0.301±0.008)]in keloid fibrosis(P<0.05).Up-regulation of miR-4463 inhibited the proliferation of keloid fibroblasts[(0.836±0.081)vs.(0.656±0.072),P<0.05],and decreased the number of migrating[(153.269±12.471)vs.(82.363±7.254)]and invading[(73.62

关 键 词：瘢痕疙瘩 成纤维细胞 miR-4463 Bcl2相关致病基因4(BAG4) 

分 类 号：R622[医药卫生—整形外科]