Esculetin inhibits TGF-β2-induced proliferation and epithelial-mesenchymal transition of retinal pigment epithelial cells.

Abstract

Purpose: Proliferative vitreoretinopathy (PVR) is a multifactorial pathological condition resulting from the migration of various cell types, primarily retinal pigment epithelial (RPE) cells, into the vitreous cavity. Currently, the only effective treatment for PVR is surgery; however, its outcomes remain suboptimal. Esculetin, a natural dihydroxycoumarin derivative, exhibits antioxidant, anti-inflammatory and antiproliferative properties. However, its effects on cell transformation and migration induced by transforming growth factor-beta 2 (TGF-β2) have not yet been investigated. This study aimed to investigate the therapeutic potential and underlying molecular mechanisms of esculetin in regulating migration and Epithelial-mesenchymal transition (EMT), key processes in the progression of PVR.

Methods: The effect of the treatment on cell survival was determined by the MTT test. Human retinal pigment epithelial cells (ARPE-19) subjected to serum starvation were treated as follows: cells were incubated for 48 h with either 10 ng/ml TGF-β2, 12.5 μM esculetin, or a combination of both agents applied simultaneously. Cells treated with vehicle alone were defined as the control group.The impact of this treatment on cell migration was evaluated using a wound healing assay, while apoptosis was analyzed by Hoechst staining assay. mRNA and protein expression levels were quantified using quantitative real-time PCR and Western blot assay, respectively.

Results: Esculetin inhibits the proliferation, transformation of RPE cells modified by TGF-β2, reverses cell morphology and inhibits cell migration in a wound healing assay. Treatment with esculetin reduces the expression levels of mesenchymal markers such as MMP-1, -2, -9, fibronectin, α-SMA, vimentin, as well as transcription factors Snail, Slug, ZEB-1 and Twist, which were upregulated by TGF-β2 treatment. Conversely, the epithelial markers E-cadherin, ZO-1 and occludin were up-regulated by esculetin treatment.

Conclusion: These findings suggest that esculetin may inhibit RPE cell migration and EMT processes involved in the development of experimentally induced PVR through various molecular mechanisms. Although these results are limited to in vitro observations, they indicate the potential of esculetin as a non-invasive therapeutic agent for PVR, pending further validation through future in vivo and clinical studies.