Gypenoside XLIX ameliorates diabetic retinopathy by downregulating prostaglandin-endoperoxide synthase 2 in retinal pigment epithelium cells to inhibit ferroptosis and preserve tight junction integrity.

Diabetic retinopathy (DR) represents a prevalent and severe eye complication in diabetic patients. With DR progresses, destruction of tight junctions (TJs) in RPE cells leads to irreversible visual impairment. Gypenoside XLIX (Gyp XLIX) is a dammarane-type glycoside, which can suppress inflammation and oxidative stress. This study sought to investigate and verify the mechanism underlying the regulatory effects of Gyp XLIX in the early protection of junctional integrity of DR. We combined bioinformatics and network pharmacology to pinpoint the core therapeutic targets of Gyp XLIX for DR. Mice with diabetes mellitus (DM) and high glucose (HG)-stimulated ARPE-19 cells were treated with Gyp XLIX. Its impact on TJ integrity in RPE cells and ferroptosis was evaluated via Western blotting, immunofluorescence staining, and assays for iron content, lipid peroxidation, and glutathione (GSH) levels. Prostaglandin-endoperoxide synthase 2 (PTGS2) was overexpressed to elucidate the mechanism of action of Gyp XLIX in the early protection of junctional integrity of DR. Among the shared targets between Gyp XLIX and DR, ALB, VEGFA, JUN, ESR1, PTGS2, STAT3, MMP9, HSP90AA1, BCL2L1 and AR were identified. Western blotting and immunofluorescence staining revealed that Gyp XLIX preserved TJ integrity in RPE cells. In addition, iron, lipid peroxidation and GSH assays revealed that Gyp XLIX inhibited ferroptosis in both mice with DM and HG-stimulated ARPE-19 cells. Overexpression of PTGS2 partially reversed the protective impacts induced by Gyp XLIX. This study demonstrated that Gyp XLIX suppressed ferroptosis and preserved TJ integrity in RPE cells, with these effects being closely associated with the downregulation of PTGS2, thereby exerting early protective effects on junctional integrity of DR.