Protective effect and mechanism of Cistanche tubulosa extract on titanium dioxide nanoparticle-induced spermatogenic disorder

Titanium dioxide nanoparticles (TiO₂ NPs) can induce spermatogenic disorder, yet effective interventions remain limited. Cistanche tubulosa, a traditional Chinese medicinal herb, has long been used to improve reproductive dysfunction. This study aimed to identify the active components of Cistanche tubulosa extract (CTE), and explore its effects on TiO₂ NPs-induced spermatogenic disorder and the underlying molecular mechanism. Sixty-nine components of CTE were profiled by UPLC-Q-TOF-MS/MS; network pharmacology, molecular docking, and molecular dynamics analyses predicted AKT1 as a main key target and PI3K/AKT signaling pathway as the core pathway, with representative components (quercetin, arachidonic acid, suchilactone, genistein, and boschnaloside) forming stable AKT1 complexes. Fifty male Sprague-Dawley rats were assigned to control, TiO₂ NPs (100 mg/kg BW, body weight), and TiO₂ NPs + CTE (200, 400, 800 mg/kg BW) groups, administered by gavage for 8 weeks. CTE began to exert effects at 400 mg/kg BW, significantly restoring serum testosterone (T)/estradiol (E2) balance, enhancing sperm motility, and reducing sperm abnormality rate. At 800 mg/kg BW, it further improved hormone balance, alleviated testicular injury, reduced testicular apoptosis, and reversed TiO₂ NP-induced suppression of the PI3K/AKT pathway by increasing the levels of phosphorylated PI3K (p-PI3K) and phosphorylated AKT (p-AKT). Collectively, CTE alleviates TiO₂ NPs-induced spermatogenic disorder, possibly by multi-component modulation of AKT1 and re-activation of PI3K/AKT signaling pathway.