Ganoderic acid D as a potential modulator of steroidogenic enzymes: Protective effects of Cucurbita maxima Duch. seed extract against nonylphenol-induced reproductive toxicity.

Nonylphenol (NP), a widespread environmental endocrine disruptor, adversely affects male reproductive health through oxidative stress and impaired steroidogenesis. Although natural products have shown protective effects against reproductive toxicity, the bioactive compounds responsible for these effects and their molecular mechanisms remain inadequately characterized. This study investigated the protective potential of Cucurbita maxima seed extract (CMSE) against NP-induced reproductive toxicity in adult male albino mice and identified key phytochemicals associated with its activity. Male mice were divided into six groups: Control, NP-treated, CMSE (HD), NP+CMSE (LD), NP+CMSE (HD), and NP + CC, and treated for 35 days. LC-MS analysis identified eleven bioactive compounds in CMSE, which were subsequently evaluated through molecular docking and molecular dynamics simulations against key steroidogenic enzymes. Among the identified constituents, Ganoderic acid D exhibited the highest binding affinity and the most stable interactions with steroidogenic targets, suggesting a potential role in regulating testosterone biosynthesis and reproductive function. In vivo analyses demonstrated that CMSE significantly attenuated NP-induced reproductive toxicity. Treatment improved gonadosomatic index, restored antioxidant defenses (GSH, SOD, and CAT), reduced lipid peroxidation (MDA), normalized testicular protein and cholesterol levels, enhanced 3β-HSD and 17β-HSD activities, and restored serum testosterone concentrations. Histopathological examination further confirmed substantial recovery of testicular architecture following CMSE administration. The integrated in silico and in vivo evidence provides novel mechanistic insights into the reproductive protective effects of Cucurbita maxima and highlights Ganoderic acid D as a promising natural therapeutic candidate against NP-induced male reproductive dysfunction.