miR-223对银屑病细胞增殖、周期、凋亡和炎症反应的影响及机制

Effect and mechanism of miR-223 on proliferation,cell cycle,apoptosis,and inflammatory response of psoriasis cells

目的探讨微小RNA-223(miR-223)表达对银屑病细胞增殖、周期、凋亡和炎症反应的影响及机制。方法体外培养人永生化角质形成细胞HaCaT,将细胞分为对照组、模型组、模型+miR-NC组、模型+miR-223 inhibitor组,除对照组外,其余各组均采用M5法诱导HaCaT细胞建立银屑病细胞模型。采用RT-qPCR法检测各组细胞miR-223表达,CCK-8法、流式细胞术分别检测各组细胞增殖活性、细胞周期及凋亡情况,ELISA法检测上清液中IL-6、IL-1β和TGF-β1,Western blotting法检测第10号染色体同源丢失性磷酸酶张力蛋白基因(PTEN)蛋白表达。结果与对照组比较,模型组细胞miR-223表达升高(P<0.05),48、72、96 h时OD值、细胞周期S期占比增高(P均<0.05),细胞周期G0/G1占比和凋亡率降低(P均<0.05),IL-6、IL-1β和TGF-β1水平明显增高(P均<0.05),PTEN蛋白表达降低(P<0.05)。与模型+miR-NC组比较,模型+miR-223 inhibitor组miR-223表达明显下降(P<0.05),48、72、96 h时OD值、细胞周期S期占比明显降低(P均<0.05),细胞周期G0/G1占比和凋亡率明显升高(P均<0.05),IL-6、IL-1β和TGF-β1水平明显降低(P均<0.05),PTEN蛋白表达升高(P<0.05)。模型组与模型+miR-NC组比较,上述各指标差异无统计学意义(P均>0.05)。结论miR-223在银屑病细胞中表达增高,下调miR-223水平能抑制银屑病细胞增殖及炎症反应,将细胞周期阻滞于G0/G1期,促进细胞凋亡;其机制可能与其调控PTEN有关。

Objective To investigate the effects of expression of microRNA-223(miR-223)on cell proliferation,cell cycle,apoptosis and inflammatory reaction of psoriasis and their mechanism.Methods Human immortalized kerati⁃nocytes HaCaT were cultured in vitro.The cells were divided into the control group,model group,model+miR-NC group,model+miR-223 inhibitor group.Except for the control group,cells in the other groups were induced to establish psoriasis cell models by M5 method.Real-time quantitative PCR(RT-qPCR)was used to detect the expression of miR-223 in eachgroup.CCK-8 method and flow cytometry were used to detect cell proliferation activity,cell cycle,and apoptosis in each group.ELISA was used to detect the levels of inflammatory factors such as IL-6,IL-1βand TGF-β1 in supernatant.West⁃ern blotting was used to detect the expression of phosphatase and tensin homolog deleted on chromosome ten(PTEN)pro⁃tein.Results Compared with the control group,the expression level of miR-223 significantly increased(P<0.05),OD values at 48,72 and 96 h and the proportion of cells in the cell cycle S phase significantly increased(all P<0.05),the proportion of cells in the cell cycle G0/G1 and apoptosis rate significantly decreased(all P<0.05),the levels of IL-6,IL-1βand TGF-β1 significantly increased(all P<0.05),while the expression of PTEN protein decreased in the model group(P<0.05).Compared with the model+miR-NC group,the expression level of miR-223 inhibitor group significant⁃ly decreased(P<0.05),OD values at 48,72 and 96 h and the proportion of cells in the S phase decreased(P<0.05),the proportion of cells in the cell cycle G0/G1 and apoptosis rate significantly increased(P<0.05),the levels of IL-6,IL-1βand TGF-β1 significantly decreased(P<0.05),and PTEN protein expression increased in the model+miR-223 inhibitor group(P<0.05).There were no significant differences in the above indexes between the model group and the model+miRNC group(all P>0.05).Conclusions The expression level of miR-223 increases in psoriasis cells.Down-regulating the level of miR-223 can inhibit the proliferation and inflammatory response of psoriasis cells,and promote their apoptosis and block the cell cycle in G0/G1 phase,which may be related to its regulation of PTEN.