miR-186-5p通过抑制Twist1诱导人皮肤成纤维细胞衰老

目的探讨miR-186-5p对人皮肤成纤维细胞衰老的影响及其机制。方法采用实时定量PCR方法检测miR-186-5p的表达,应用衰老相关β-半乳糖苷酶(SA-β-gal)染色、CCK-8和EdU等方法研究miR-186-5p对人真皮成纤维细胞(HDFs)衰老的作用。通过生物信息学分析、荧光素酶报告试验、实时定量PCR、Western blot和免疫组化法证实miR-186-5p的下游靶基因。结果首次发现miR-186-5p在传代衰老的HDFs和老年人皮肤真皮组织中表达上调。miR-186-5p的过表达抑制了HDFs细胞的增殖,同时增强了衰老相关指标p16、p21和SA-β-gal的表达。双荧光素酶报告分析显示Twist1是miR-186-5p的下游靶基因。结论miR-186-5p可通过抑制Twist1诱导HDFs衰老,其可能成为干预皮肤老化的潜在靶点。

miR-186-5p Induces Senescence in Human Skin Fibroblasts by Inhibiting Twist1

Objective To investigate the role of miR-186-5p on the aging of human dermal fibroblasts and its mechanism.Methods The expression of miR-186-5p was measured by quantitative real-time PCR(qRT-PCR).Senescence-associatedβ-galactosidase(SA-β-gal)staining,the cell counting Kit-8(CCK-8)and 5-ethynyl-2'-deoxyuridine(EdU)were used to investigate the effects of miR-186-5p on human dermal fibroblast(HDF)senescence.The target gene of miR-186-5p was confirmed by bioinformatics analysis,luciferase reporter assay,qRT-PCR,Western blot and immunohistochemical analysis.Results We first found that miR-186-5p was up-regulated in passage-aged HDFs(pHDFs)and skin dermal tissues from old donors(old skin).Overexpression of miR-186-5p inhibited HDFs proliferation while enhanced the expression of p16,p21 and the SA-β-gal-positive ratio.Dual-luciferase reporter assays showed that Twist1 was the downstream target gene of miR-186-5p.Conclusion miR-186-5p could induce HDF senescence by inhibiting Twist1.It could be a potential therapeutic target for preventing skin aging.