文摘目的:探讨长链非编码RNA 8975-1(Long non-coding RNA 8975-1,LNCRNA8975-1)通过微小RNA-203(microRNA-203,miR-203)/血管内皮生长因子-A(Vascular endothelial growth factor-A,VEGF-A)信号轴对人瘢痕疙瘩成纤维细胞(Human keloid fibroblast,HKF)的细胞增殖周期和迁移的影响及其机制。方法:培养人皮肤成纤维细胞(Human skin fibroblasts,HSF)和HKF,采用定量PCR(quantitative polymerase chain reaction,qPCR)及western blot法检测LNCRNA8975-1、miR-203和I型胶原蛋白(Type I collagen,Col-I)的表达量。将HKF细胞分为Control组、小干扰RNA载体的阴性对照(the negative control for small interfering RNA vector,siNC)组、沉默LNCRNA8975-1的小干扰RNA载体(the small interfering RNA vector for silence of LNCRNA8975-1,siLNCRNA8975-1)组、pcDNA3.1空载体(the empty pcDNA3.1 vector,pcDNA)组、过表达LNCRNA8975-1的pcDNA3.1载体(overexpression of LNCRNA8975-1 in the pcDNA3.1 vector,pcDNA-LNCRNA8975-1)组、miR-203拟似物的阴性对照(the negative control of miR-203 mimic,NC mimic)组、miR-203的拟似物(miR-203 mimic)组、miR-203抑制剂的阴性对照(negative control of miR-203 inhibitor,NC inhibitor)组、miR-203的抑制剂(miR-203 inhibitor)组、siLNCRNA8975-1+miR-203 inhibitor组、VEGF-A+miR-203 mimic组。CCK-8法检测细胞增殖活性,免疫荧光检测细胞周期标志物细胞周期蛋白D1(cyclin D1),伤口愈合实验和Transwell实验检测细胞迁移功能。荧光素酶基因报告实验检测HKF细胞中LNCRNA8975-1和miR-203、miR-203和VEGF-A的直接作用。结果:与HSF相比,HKF中LNCRNA8975-1和Col-I表达量上调(P<0.05),而miR-203降低(P<0.05)。与siNC相比,siLNCRNA8975-1抑制了HKF细胞周期蛋白cyclin D1表达和细胞的迁移(P<0.05)。与NC mimic相比,miR-203 mimic抑制了HKF中cyclin D1蛋白表达和细胞的迁移(P<0.05)。LNCRNA8975-1充当miR-203的“分子海绵”靶向抑制miR-203,且VEGF-A是miR-203的靶基因。VEGF-A表达被miR-203 mimic抑制(P<0.05)。miR-203 inhibitor充分逆转了siLNCRNA8975-1对HKF中cyclin D1表达和迁移的抑制作用(P<0.05),而且VEGF-A则逆转了miR-203 mimic对HKF中cyclin D1表达和迁移的抑制功能(P<0.05)。结论:LNCRNA8975-1通过抑制miR-203上调VEGF-A调控HKF的周期和迁移。
