Black Phosphorus Nanosheets Penetrate the Blood-Testis Barrier and Induce Reproductive Toxicity in Male Mice

Black phosphorus nanosheets (BPNSs), as an emerging subclass of two-dimensional (2D) nanomaterials, have exhibited extensive applicability in electronics, optoelectronics, and medical sciences. However, its expanding application in diverse fields has raised concerns about inadvertent environmental release, which might lead to potential ecological and health risks. Herein, mice were continuously exposed at doses of 0.1 mg/kg/day (possible acceptable daily intake dose, ADI) and 1 mg/kg/day (possible occupational exposure dose) to BPNSs for 28 days. Using hyperspectral imaging and Evans blue fluorescence chromogenic assay, we confirmed that partial BPNSs penetrated the blood-testis barrier (BTB) and distributed throughout the spermatogenic microenvironment of testes. Mice exposed to BPNSs exhibited significant reproductive toxicity, evidenced by abnormal sperm parameters, disordered spermatogenic cell layers, sloughed germ cells, and disrupted sex hormone levels. Further, BPNSs promoted the steroid synthesis pathway and inhibited testosterone aromatization, leading to testicular endocrine homeostasis disorders. BPNSs also damaged germ cells in the testes via the apoptosis pathway. In conclusion, our study not only highlights the male reproductive toxicity of BPNSs but also demonstrates that BPNSs can penetrate the BTB and have bioaccumulation potential in the male reproductive system, which provides new insights into the toxicological implications of 2D nanomaterials for mammalian fertility.