Tri-Ortho-Cresyl Phosphate Inhibits Proliferation of Mouse Germ Cells by Activating Endoplasmic Reticulum Stress and Suppressing NTE Activity.

Tri-o-cresyl phosphate (TOCP) is widely used as a plasticizer, flame retardant, and lubricant additive, but has been reported to impair spermatogenesis. However, how TOCP affects spermatogenesis remains unclear. Therefore, the objective of this study is to investigate the underlying mechanism by which TOCP disrupts spermatogenesis. In order to achieve this, adult male mice were orally administered TOCP at doses of 0, 200, or 400 mg/kg for two weeks, and we found that TOCP exposure reduced the number of germ cells and decreased sperm density. Moreover, the numbers of PCNA-positive cells and phospho-histone H3 (Ser10)-positive cells in mouse testicular tissues were significantly decreased following TOCP treatment, indicating that germ cell proliferation may be impaired. In addition, TOCP did not affect the protein expression of neuropathy target esterase (NTE) in testicular tissues but markedly inhibited its enzymatic activity (by approximately 30% relative to the control level). In vitro experiments further demonstrate that TOCP suppressed cell proliferation and mitotic progression in mouse GC-1 spg cells and excessively activated endoplasmic reticulum (ER) stress. Treatment with 4-phenylbutyric acid (4-PBA), an ER stress inhibitor, partially reversed the TOCP-induced inhibition of cell proliferation and mitosis. Furthermore, TOCP inhibited NTE activity in GC-1 spg cells, and NTE knockdown produced a phenotype similar to that observed after TOCP exposure, characterized by suppressed cell proliferation and mitotic progression. Surprisingly, ER stress was not activated in GC-1 spg cells following NTE knockdown. Collectively, these findings suggest that TOCP may impair spermatogenesis by inhibiting the proliferation and mitotic progression of mouse germ cells, potentially through mechanisms involving excessive activation of ER stress or suppression of NTE activity.