Abstract

Ethnopharmacological relevance Ophiopogonis Radix (Mai-Dong) has been widely used in traditional Chinese medicine for the treatment of cardiovascular diseases, particularly those associated with ischemia and impaired cardiac function. Methylophiopogonanone A (MOA), a bioactive homoisoflavonoid isolated from Ophiopogonis Radix, has demonstrated antioxidant and anti-inflammatory activities; however, its role and molecular mechanisms in myocardial ischemia–reperfusion injury (MIRI) remain unclear. Aim of the study This study aimed to investigate the cardioprotective effects of MOA in MIRI and to determine whether VEGFR2-associated PI3K/Akt signaling, inhibitory phosphorylation of GSK-3β, and preservation of mitochondrial function contribute to these effects. Materials and methods MOA-VEGFR2 target engagement and stabilization were assessed using computational prediction and biochemical target-stability assays. The cardioprotective effects of MOA were evaluated in a rat myocardial ischemia–reperfusion model and in primary cardiomyocytes subjected to oxygen–glucose deprivation/reoxygenation (OGD/R), with emphasis on cardiac function, infarct size, mitochondrial injury, mitochondrial permeability transition pore (mPTP) opening, and VEGFR2/PI3K/Akt/GSK-3β signaling. The involvement of VEGFR2 and PI3K signaling was further examined using VEGFR2 siRNA, SU5416, and LY294002. Results Integrative target screening prioritized VEGFR2/KDR as a functionally relevant candidate target of MOA in MIRI. Molecular docking and molecular dynamics simulations provided structural predictions; DARTS, CETSA, and ITDRF-CETSA supported VEGFR2 target engagement by MOA, whereas CHX-chase analysis showed a prolonged VEGFR2 protein half-life. In vivo, MOA reduced infarct size, improved cardiac function, lowered serum CK-MB, LDH, and cTnI levels, increased myocardial ATP content and SOD activity, enhanced phosphorylation of VEGFR2, PI3K, Akt, and GSK-3β, suppressed mPTP opening, and preserved mitoch