miR-410-3p通过靶向YAP1调节IL-22诱导的人角质形成细胞增殖和凋亡

miR-410-3p modulates proliferation and apoptosis of IL-22-induced humankeratinocytes via targeting YAP1

目的:探究miR-410-3p在银屑病中的表达及其对角质形成细胞增殖和凋亡的影响及机制。方法:收集12例寻常型银屑病患者和12例健康志愿者空腹静脉血,qRT-PCR检测血清miR-410-3p表达。将常规培养的人角质形成细胞HaCaT分为:对照组、IL-22组、miR-NC组和miR-410-3p组。后3组使用100 ng/ml IL-22刺激24 h,体外建立银屑病模型。miRNC组和miR-410-3p组加入IL-22前48 h分别转染miR-NC和miR-410-3p,对照组不进行任何处理。CCK-8分析细胞活力,BrdU检测细胞增殖,流式细胞术检测细胞凋亡,qRT-PCR检测miR-410-3p和YAP1 mRNA表达,Western blot分析YAP1蛋白表达。Starbase软件和双荧光素酶报告基因分析miR-410-3p和YAP1的靶向关系。转染miR-410-3p的HaCaT银屑病细胞模型中同时过表达YAP1,检测细胞活力和凋亡。结果:miR-410-3p在银屑病患者血清中的表达显著低于健康志愿者(P<0.05)。与对照组相比,IL-22组miR-410-3p表达降低,YAP1 mRNA和蛋白表达增加,细胞活力增强,BrdU阳性细胞增多,凋亡率降低(均P<0.05);与IL-22组相比,miR-410-3p组miR-410-3p表达增加,YAP1 mRNA和蛋白表达减少,细胞活力减弱,BrdU阳性细胞减少,凋亡率升高(均P<0.05)。此外,本实验证实miR-410-3p与YAP1存在靶向结合关系,过表达YAP1可逆转miR-410-3p对IL-22诱导的HaCaT细胞增殖和凋亡的影响。结论:miR-410-3p在银屑病患者中表达减少,上调其表达可通过靶向YAP1抑制IL-22诱导的角质形成细胞增殖,促进细胞凋亡。

Objective:To investigate miR-410-3p expression in psoriasis and its effects and mechanism on proliferation and apoptosis of keratinocytes.Methods:Fasting venous blood was collected from 12 patients with psoriasis vulgaris and 12 healthy sub-jects,and serum miR-410-3p expression was detected by qRT-PCR.Conventionally cultured keratinocytes HaCaT were divided into control,IL-22,miR-NC and miR-410-3p groups.Cells in last three groups were stimulated with 100 ng/ml IL-22 for 24 h to establish psoriatic model in vitro.Cells in miR-NC and miR-410-3p groups were transfected with miR-NC or miR-410-3p 48 h prior to IL-22 stimulation,respectively.While cells in control group did not perform any treatment.Cell viability was determined by CCK-8.BrdU was used to detect cell proliferation.Apoptosis of cells was detected by flow cytometry.Expressions of miR-410-3p and YAP1 mRNA were measured by qRT-PCR.Western blot was used to examine YAP1 protein expression.Starbase software and dual-luciferase reporter assay were used to analyze targeting relationship between miR-410-3p and YAP1.YAP1 was over-expressed in miR-410-3p transfected HaCaT keratinocytes to detect cell viability and apoptosis.Results:Serum miR-410-3p expression was lower in patients with psoriasis than that in healthy subjects(P<0.05).Compared with control group,miR-410-3p expression was decreased,mRNA and protein levels of YAP1 as well as cell viability and BrdU-positive cells were elevated,and apoptotic rate was decreased in IL-22 group(all P<0.05).Compared with IL-22 group,miR-410-3p expression was increased,mRNA and protein levels of YAP1 as well as cell viability and BrdU-positive cells were reduced,and apoptotic rate was increased in miR-410-3p group(all P<0.05).Targeting relationship between miR-410-3p and YAP1 was confirmed in this study.Overexpression of YAP1 significantly reversed effects of miR-410-3p on prolifera-tion and apoptosis of IL-22-stimulated HaCaT cells.Conclusion:miR-410-3p expression is down-regulated in psoriasis.Up-regulation of miR-410-3p can inhibit cell proliferation,and promote cell apoptosis in IL-22-stimulated keratinocytes via targeting YAP1.