ARPE-19 cells were treated with PA to induce EMT, followed by E-cadherin and α-smooth muscle actin (α-SMA) expression and the microRNA expression profile analyses. Subsequently, miR-143-5p mimics/inhibitors, and plasmids expressing its predicted target gene c-JUN-dimerization protein 2 (JDP2), were transfected in ARPE-19 cells using lipofectamine 3000, and followed by PA treatment. Their impacts on EMT were explored using wound healing and Western blot assays. Additionally, miR-143-5p mimics and JDP2-expressing plasmid were co-transfected into ARPE-19 cells and treated with PA to explore whether PA induced EMT of ARPE-19 cells via the miR-143-5p/JDP2 axis.
PA decreased E-cadherin expression and increased those of α-SMA and miR-143-5p. Inhibiting miR-143-5p suppressed the migration of ARPE-19 cells and altered the expressions of E-cadherin and α-SMA. However, additional PA treatment attenuated these alterations. JDP2 was a target of miR-143-5p. Overexpression of JDP2 inhibited the EMT of ARPE-19 cells, resulting in α-SMA downregulation and E-cadherin upregulation, which were reversed by additional PA treatment via inhibiting JDP2 expression. Overexpression of miR-143-5p reversed the effect of JDP2 on the EMT of ARPE-19 cells and additional PA treatment markedly enhanced the effect of miR-143-5p mimics.
PA promotes EMT of ARPE-19 cells via regulating the miR-143-5p/JDP2 axis, and these findings provide significant insights into the potential targeting of this axis to treat proliferative vitreoretinopathy.
方法:用PA处理ARPE-19细胞以诱导EMT,其次是E-钙黏着蛋白和α-平滑肌肌动蛋白(α-SMA)的表达和microRNA表达谱分析。随后,miR-143-5p模拟物/抑制剂,和表达其预测的靶基因c-JUN-二聚化蛋白2(JDP2)的质粒,使用脂质体3000转染ARPE-19细胞,然后进行PA处理。使用伤口愈合和Western印迹测定来探索它们对EMT的影响。此外,将miR-143-5p模拟物和JDP2表达质粒共转染到ARPE-19细胞中并用PA处理以探索PA是否通过miR-143-5p/JDP2轴诱导ARPE-19细胞的EMT。
结果:PA降低了E-cadherin的表达,增加了α-SMA和miR-143-5p的表达。抑制miR-143-5p抑制ARPE-19细胞的迁移并改变E-cadherin和α-SMA的表达。然而,额外的PA治疗减轻了这些改变。JDP2是miR-143-5p的靶标。JDP2过表达抑制ARPE-19细胞的EMT,导致α-SMA下调和E-钙黏着蛋白上调,通过抑制JDP2表达,通过额外的PA处理逆转。miR-143-5p的过表达逆转了JDP2对ARPE-19细胞的EMT的作用,并且额外的PA处理显著增强了miR-143-5p模拟物的作用。
结论:PA通过调节miR-143-5p/JDP2轴促进ARPE-19细胞的EMT,这些发现为该轴治疗增殖性玻璃体视网膜病变的潜在靶向提供了重要见解。