Ghrelin relieves the reproductive damage of TiO2 NPs in young male rats via ROS/AMPK/mTOR signaling pathway

Children's daily exposure to titanium dioxide nanoparticles (TiO₂ NPs) poses significant risks to the developing testes, yet few studies have examined their impact on young testicular growth. Given ghrelin's established antioxidant and testis-protective effects in animal models, we hypothesized that it could mitigate TiO₂ NPs–induced damage. In this first systematic investigation of young testicular responses, three-week-old male rats received oral gavage of TiO₂ NPs (100, 200, or 400 mg/kg), followed 30 min later by intraperitoneal ghrelin (50 mg/kg) for four weeks. Exposure to TiO₂ NPs disrupted seminiferous tubule architecture and reduced serum levels of key developmental and sex hormones, including IGF-1, FSH, LH, and testosterone. Mechanistically, TiO₂ NPs activated the ROS/AMPK/mTOR signaling pathway, resulting in a marked reduction of SOD, CAT, and GSH activities and the induction of oxidative imbalance, while simultaneously upregulating the autophagy protein ULK1 and autophagy-related genes to trigger excessive autophagy, ultimately compromising testicular architecture and function. We further assessed ghrelin's protective efficacy against TiO₂ NPs-induced reproductive injury in young rats, demonstrating that ghrelin inhibited the ROS/AMPK/mTOR signaling pathway, attenuated oxidative stress, and suppressed excessive autophagy, thereby mitigating testicular injury. These findings represent the first evidence of the developing testicular heightened vulnerability to TiO₂ NPs toxicity and underscore ghrelin's potential as a novel protective intervention. Our study provides new insights into the reproductive toxicology of engineered nanoparticles and supports targeted strategies to safeguard pediatric reproductive health.