miR-590-5p靶向TGF-β1/Smad3通路调控瘢痕疙瘩成纤维细胞增殖和侵袭的研究

Regulation of proliferation and invasion of keloid fibroblasts by miR-590-5p to TGF-β1/Smad3 pathway

目的探讨miR-590-5p通过TGF-β1/Smad3通路调控瘢痕疙瘩成纤维细胞增殖和侵袭的作用。方法获取瘢痕疙瘩成纤维细胞进行培养采用随机数字表法分组,分为空白对照组、空载体转染组、miR-590-5p mimic组、miR-590-5p inhibitor组、miR-590-5p mimic和siRNA TGF-β1共转染组,其中空白对照组不进行转染,空载体转染组转染空载体、miR-590-5p mimic组转染miR-590-5p mimic、miR-590-5p inhibitor组转染miR-590-5p inhibitor、miR-590-5p mimic和siRNA TGF-β1共转染组转染miR-590-5p mimic和siRNA TGF-β1。采用CCK-8试验检测各组细胞增殖情况,流式细胞仪检测细胞凋亡,Transwell试验检测细胞侵袭能力,蛋白质印迹法检测TGF-β1、Smad3蛋白表达。结果miR-590-5p mimic和siRNA TGF-β1共转染组细胞培养24、48和72 h时细胞增殖活力明显低于空白对照组、空载体转染组、miR-590-5p mimic组、miR-590-5p inhibitor组,差异均有统计学意义(P<0.05)。miR-590-5p mimic和siRNA TGF-β1共转染组细胞凋亡率明显高于空白对照组、空载体转染组、miR-590-5p mimic组、miR-590-5p inhibitor组,差异均有统计学意义(P<0.05)。miR-590-5p mimic和siRNA TGF-β1共转染组细胞侵袭数明显低于空白对照组、空载体转染组、miR-590-5p mimic组、miR-590-5p inhibitor组,差异均有统计学意义(P<0.05)。miR-590-5p mimic和siRNA TGF-β1共转染组细胞TGF-β1、Smad3蛋白相对表达量明显低于空白对照组、空载体转染组、miR-590-5p mimic组、miR-590-5p inhibitor组,差异均有统计学意义(P<0.05)。结论抑制miR-590-5p表达,可促进瘢痕疙瘩成纤维细胞增殖和侵袭以及抑制细胞凋亡,可能与通过TGF-β1/Smad3通路调控有关。

Objective To explore the role of miR-590-5p in regulating the proliferation and invasion of keloid fibroblasts through TGF-β1/Smad3 pathway.Methods Keloid fibroblasts were obtained and cultured,which were divided into blank control group,empty body transfection group,miR-590-5p mimic group,miR-590-5p inhibitor group,miR-590-5p mimic and siRNA TGF-β1 co transfection group according to the random number table method,the blank control group was not transfected,while the empty body transfection group was transfected with empty vectors,miR-590-5p mimic group was transfected with miR-590-5p mimic,miR-590-5p inhibitor group was transfected with miR-590-5p inhibitor,miR-590-5p mimic and siRNA TGF-β1 co transfection group was transfected with miR-590-5p mimic and siRNA TGF-β1.CCK-8 test was used to detect cell proliferation,flow cytometry was used to detect apoptosis,Transwell test was used to detect cell invasion,Western blotting was used to detect TGF-β1,Smad3 protein expression.Results The cell proliferation activity of cells cultured for 24,48 and 72 h in miR-590-5p mimic and siRNA TGF-β1 co transfected group were significantly lower than those of blank control group,empty body transfection group,miR-590-5p mimic group,miR-590-5p inhibitor group,the differences were statistically significant(P<0.05).The apoptosis rate of miR-590-5p mimic and siRNA TGF-β1 co transfection group was significantly higher than that of blank control group,empty body transfection group,miR-590-5p mimic group,miR-590-5p inhibitor group,the difference was statistically significant(P<0.05).The number of cell invasion in miR-590-5p mimic and siRNA TGF-β1 co transfection group was significantly lower than that in blank control group,empty body transfection group,miR-590-5p mimic group,miR-590-5p inhibitor group,the difference was statistically significant(P<0.05).The relative expressions of TGF-β1 and Smad3 protein in miR-590-5p mimic and siRNA TGF-β1 co transfected group were significantly lower than those in blank control group,empty body transfection group,miR-590-5p mimic group,miR-590-5p inhibitor group,the differences were statistically significant(P<0.05).Conclusion Inhibiti on of miR-590-5p expression can promote the proliferation and invasion of keloid fibroblasts and inhibit apoptosis,which may be related to the regulation of TGF-β1/Smad3 pathway.