miR-489在瘢痕疙瘩组织中表达变化及其过表达对瘢痕疙瘩成纤维细胞增殖、迁移、纤维化的影响
Expression of miR-489 in keloid tissues and the effect of its overexpression on proliferation,migration and fibrosis of keloid fibroblasts
刘鹏超 1刘智君 2胡金宏 1梁亚坤 1缪国东 2马志红2
作者信息
- 1. 承德医学院研究生学院,河北 承德 067000
- 2. 承德市中心医院医疗美容科
- 折叠
摘要
目的 观察miR-489在瘢痕疙瘩组织及其成纤维细胞中的表达变化,探讨其过表达对瘢痕疙瘩成纤维细胞增殖、迁移、纤维化的影响.方法 收集瘢痕疙瘩组织、正常皮肤组织各6例份,分别分离原代瘢痕疙瘩成纤维细胞及正常成纤维细胞并进行传代培养,采用实时荧光定量PCR法检测组织及细胞中的miR-489相对表达量.将瘢痕疙瘩成纤维细胞分为对照组、过表达组,分别转染miR-NC及miR-489-mimic,转染48 h后采用CCK-8法检测连续培养5 d的细胞增殖能力(以光密度值表示),采用Transwell小室实验检测细胞迁移能力(以迁移细胞数表示),采用Western blotting法检测细胞Ⅰ型胶原蛋白(COL1)、Ⅲ型胶原蛋白(COL3)抗体、α-平滑肌动蛋白(α-SMA)、Smad3蛋白相对表达量.结果 瘢痕疙瘩组织、正常皮肤组织miR-489相对表达量分别为0.50±0.06、1.00±0.01,二者比较P<0.05;瘢痕疙瘩成纤维细胞、正常成纤维细胞miR-489相对表达量分别为0.25±0.18、1.00±0.01,二者比较P<0.05.过表达组培养第3～5天细胞增殖光密度值低于对照组(P均<0.05),两组培养第1、2天细胞增殖光密度值比较差异均无统计学意义(P均>0.05).过表达组和对照组迁移细胞数分别为(43.00±4.00)、(129.67±9.71)个,两组比较P<0.05.过表达组细胞COL1、COL3、α-SMA、Smad3蛋白相对表达量均低于对照组(P均<0.05).结论 瘢痕疙瘩组织及其成纤维细胞中miR-489表达均降低,过表达miR-489可能通过调节Smad3蛋白表达而抑制瘢痕疙瘩成纤维细胞增殖、迁移及纤维化过程.
Abstract
Objective To observe the expression changes of miR-489 in the keloid tissues and fibroblasts,and to ex-plore the effect of miR-489 overexpression on the proliferation,migration and fibrosis of keloid fibroblasts.Methods Six cases of keloid tissues and six cases of normal skin tissues were collected.Primary keloid fibroblasts and normal fibroblasts were isolated and subcultured.The relative expression of miR-489 in tissues and cells was detected by real-time fluores-cence quantitative PCR.Keloid fibroblasts were divided into the control group and overexpression group,which were trans-fected with miR-NC and miR-489-mimic,respectively.After 48 h of transfection,CCK-8 method was used to detect the cell proliferation ability after continuous culture for 5 days(expressed as OD450),and Transwell chamber experiment was used to detect the cell migration ability(expressed as the number of migration cells).The relative expression levels of type I collagen(COL1),type III collagen(COL3)antibody,α-smooth muscle actin(α-SMA)and Smad3 protein were detect-ed by Western blotting.Results The relative expression levels of miR-489 in the keloid tissues and normal skin tissues were 0.50±0.06 and 1.00±0.00,respectively,with statistically significant difference(both P<0.05).The relative ex-pression levels of miR-489 in keloid fibroblasts and normal fibroblasts were 0.25±0.18 and 1.00±0.00,respectively,with statistically significant difference(both P<0.05).The cell proliferation ability of the overexpression group was lower than that of the control group on the 3rd to 5th days of culture(P<0.05).There was no significant difference in cell prolif-eration ability between the two groups on the 1st and 2nd days of culture(P>0.05).The numbers of migration cells in the overexpression group and the control group were 43.00±4.00 and 129.67±9.71,respectively(P<0.05).The relative expression levels of COL1,COL3,α-SMA and Smad3 proteins in the overexpression group were lower than those in the control group(all P<0.05).Conclusions The expression of miR-489 in keloid tissues and fibroblasts decreased.Over-expression of miR-489 may inhibit the proliferation,migration and fibrosis of keloid fibroblasts by regulating the expression of target gene Smad3 protein.
关键词
微小核糖核酸-489/瘢痕疙瘩/Smad蛋白/纤维化Key words
microRNA-489/keloid/Smad protein/fibrosis引用本文复制引用
出版年
2024
国家科技期刊平台
NSTL
万方数据