The in vivo nanotoxicity of nanoparticles is drawing increased attention as concerns grow over the biosafety of nanotechnology. TiO₂ nanoparticles are coated to decrease the potential of harmful effects due to their photoactivity. Rutile-type alumina-coated titanium dioxide nanoparticles (Al₂O₃-TiO₂-NPs) are frequently used in cosmetics to improve their dispersion stability. We herein discuss the effects of Al₂O₃-TiO₂-NPs exposure during pregnancy on mouse spermatogenesis. Pregnant mice were injected five times, once each with 0.1 mL of sequentially diluted concentrations of a Al₂O₃-TiO₂-NPs suspension (1, 10, 100 or 1,000 μg/mL) and received doses of 0.5, 5, 50 and 500 μg, respectively. Prior to injection, the size distribution of the Al₂O₃-TiO₂-NPs was analyzed by dynamic light scattering (DLS) measurement. The average diameter was increased in dose-dependent manner from an average of 153.8 nm to 654.6 nm. The offspring testes were examined at 12 weeks postpartum. The agglomerates in the testicular sections were small (< 200 nm). They were confirmed by the characteristic peaks of the Ti and Al elements on field emission-scanning electron microscope/energy dispersive X-ray spectroscopy (FE-SEM/EDS). Low cellular adhesion and degenerated Sertoli cells were observed in the seminiferous epithelium of all of the Al₂O₃-TiO₂-NP recipient groups by a histological analysis. The detrimental function of the Sertoli cells resulted in the formation of abnormal spermatozoa. The results suggested that Al₂O₃-TiO₂-NPs that transferred from the mother’s body affected spermatogenesis in the offspring.