Sinensetin inhibits choroidal neovascularization by downregulating the MAPK/ERK pathway

Sinensetin (SIN), a characteristic polymethoxyflavone abundant in Citrus species, is isolated from Citrus peel (Chenpi), a medicinal material widely applied in Traditional Chinese Medicine (TCM) to activate blood circulation and ameliorate blood stasis-associated ocular disorders. Accumulating pharmacological evidence confirms that SIN exerts prominent antioxidant, anti-inflammatory, and anti-angiogenic bioactivities. It is capable of suppressing pathological angiogenesis in zebrafish and endothelial cell models, which renders SIN a promising candidate for angiogenic ocular diseases. Nevertheless, its therapeutic efficacy against choroidal neovascularization (CNV) remains largely uninvestigated.

This study aimed to investigate the inhibitory effects of SIN on CNV and to dissect the underlying mechanism, specifically focusing on its modulation of FGF1-mediated MAPK/ERK signaling.

The anti-CNV effect of SIN was evaluated both in vitro and in vivo. In vivo, a laser-induced CNV model was established in Brown Norway (BN) rats, and SIN was administered via intravitreal injection; CNV severity was assessed by FITC perfusion, OCT, HE staining, and Western blot. In vitro, human umbilical vein endothelial cells (HUVEC) were treated with CNV-mimicking ARPE-19 human retinal pigment epithelial cells (ARPE-19) conditioned medium; the effects of SIN on migration and angiogenesis were evaluated by scratch, transwell, and tube formation assays. Combined transcriptomic and proteomic analyses were employed to screen key targets and pathways, and mechanistic validation was performed using the ERK agonist PMA and inhibitor U0126.

In the laser-induced CNV rat model, intravitreal injection of SIN significantly reduced CNV area and thickness and suppressed the MAPK/ERK signaling pathway, effects that were reversed by the ERK agonist PMA. Consistently, in vitro experiments confirmed that SIN dose-dependently reversed the enhanced migration and tube formation of HUVEC induced by CNV-mimicking conditioned medium. Multi-omics analysis identified FGF1 as a shared candidate downstream mediator of SIN, with SIN downregulating FGF1, VEGF-A, HIF-1α, and p-ERK1/2 expression. Further mechanistic studies revealed that the ERK inhibitor U0126 mimicked the anti-angiogenic effect of SIN, whereas the ERK agonist PMA abolished its inhibitory activity, suggesting that SIN exerts anti-CNV effects, at least in part, through suppression of MAPK/ERK signaling.

Collectively, this study demonstrates that citrus-derived SIN exerts anti-CNV effects, at least in part, through suppression of MAPK/ERK signaling, while additional pathways identified by multi-omics may also contribute. These findings highlight SIN as a promising natural product for the treatment of CNV-related ocular diseases, with FGF1 identified as a potential mediator underlying its anti-angiogenic action.