Proliferative retinopathies are associated with formation of fibrous epiretinal membranes. At present, there is no pharmacological intervention for the treatment of retinopathies. Cytokines such as TGFβ are elevated in the vitreous humor of the patients with proliferative vitro-retinopathy, diabetic retinopathy and age-related macular degeneration. TGFβ isoforms lead to epithelial-mesenchymal transition (EMT) or trans-differentiation of the retinal pigment epithelial (RPE) cells. PI3K/Akt and MAPK/Erk pathways play important roles in the EMT of RPE cells. Therefore, inhibition of EMT by pharmacological agents is an important therapeutic strategy in retinopathy. Dichloroacetate (DCA) is shown to prevent proliferation and EMT of cancer cell lines but its effects are not explored on the prevention of EMT of RPE cells. In the present study, we have investigated the role of DCA in preventing TGFβ2 induced EMT of RPE cell line, ARPE-19. A wound-healing assay was utilized to detect the anti-EMT effect of DCA. The expressions of EMT and cell adhesion markers were carried out by immunofluorescence, western blotting, and quantitative real-time PCR. The expression of MAPK/Erk and PI3K/Akt pathway members was carried out using western blotting. We found that TGFβ2 exposure leads to an increase in the wound healing response, expression of EMT markers (Fibronectin, Collagen I, N-cadherin, MMP9, S100A4, α-SMA, Snai1, Slug) and a decrease in the expression of cell adhesion/epithelial markers (ZO-1, Connexin 43, E-cadherin). These changes were accompanied by the activation of PI3K/Akt and MAPK/Erk pathways. Simultaneous exposure of DCA along with TGFβ2 significantly inhibited wound healing response, expression of EMT markers and cell adhesion/epithelial markers. Furthermore, DCA and TGFβ2 effectively attenuated the activation of MAPK/Erk/JNK and PI3K/Akt/GSK3β pathways. Our results demonstrate that DCA has a strong anti-EMT effect on the ARPE-19 cells and hence can be utilized as a therapeutic agent in the prevention of proliferative retinopathies.

增殖性视网膜病与纤维性视网膜前膜的形成有关。目前，尚无用于治疗视网膜病的药理干预措施。具有增殖性体外视网膜病变，糖尿病性视网膜病变和年龄相关性黄斑变性的患者的玻璃体液中的细胞因子（例如TGFβ）升高。TGFβ同工型会导致视网膜色素上皮细胞（RPE）的上皮-间质转化（EMT）或转分化。PI3K / Akt和MAPK / Erk途径在RPE细胞的EMT中起重要作用。因此，通过药物抑制EMT是视网膜病的重要治疗策略。已显示二氯乙酸盐（DCA）可以阻止癌细胞系的增殖和EMT，但尚未探讨其在预防RPE细胞EMT方面的作用。在目前的研究中，我们研究了DCA在预防TGFβ2诱导的RPE细胞系ARPE-19的EMT中的作用。伤口愈合试验被用来检测DCA的抗EMT作用。通过免疫荧光，蛋白质印迹和定量实时PCR进行EMT和细胞粘附标记物的表达。使用蛋白质印迹法进行MAPK / Erk和PI3K / Akt途径成员的表达。我们发现，TGFβ2暴露导致伤口愈合反应增加，EMT标记物（纤连蛋白，胶原蛋白I，N-钙黏着蛋白，MMP9，S100A4，α-SMA，Snai1，Slug）的表达增加，并且细胞黏附表达的减少/上皮标记物（ZO-1，Connexin 43，E-cadherin）。这些变化伴随着PI3K / Akt和MAPK / Erk途径的激活。同时暴露DCA和TGFβ2会显着抑制伤口愈合反应，EMT标记物的表达和细胞黏附/上皮标记物的表达。此外，DCA和TGFβ2有效减弱了MAPK / Erk / JNK和PI3K / Akt /GSK3β途径的激活。我们的结果证明DCA对ARPE-19细胞具有很强的抗EMT效果，因此可以用作预防增殖性视网膜病的治疗剂。