Network Pharmacology and Experimental Validation Elucidate the Anti‐Angiogenic Mechanism of Silibinin

ABSTRACT

Retinopathy of prematurity (ROP) is a sight‐threatening vascular disorder driven by pathological neovascularization. Silibinin (SIL), a major flavonolignan from milk thistle, possesses antioxidant and anti‐inflammatory properties. Mechanistically, its therapeutic effect arises from attenuated oxidative stress, which suppresses mitogen‐activated protein kinase (MAPK) signaling upstream. This study employed an integrated network pharmacology and experimental strategy to elucidate SIL's antiangiogenic mechanism. Network analysis identified the MAPK pathway as a key target, and topological analysis highlighted core associated proteins (KDR, ABCB1, and MMP2). Subsequent in vitro experiments using human umbilical vein endothelial cells (HUVECs) showed that SIL (at concentrations of 10 and 20 µM) significantly inhibited hypoxia‐induced MAPK pathway activation (reducing phosphorylation of p38, ERK, and JNK) and downregulated KDR expression. SIL treatment dose‐dependently suppressed endothelial cell proliferation, migration, and tube formation under hypoxic conditions. In an oxygen‐induced retinopathy (OIR) mouse model, in vivo administration of SIL (at a dose of 100 mg/kg) reduced pathological retinal neovascularization (RNV) by ∼57.6% and the avascular area by ∼27.2%. ( p < 0.01 and p < 0.05, respectively). These findings demonstrate that SIL inhibits pathological retinal angiogenesis primarily by modulating the MAPK pathway, providing mechanistic insight and highlighting its potential as a multi‐target therapeutic candidate for ROP.