Gestational prednisone exposure causes subfertility in male offspring mice: Role of testicular E2f1 N6-methyladenosine modification.

Previous studies showed that abnormally high glucocorticoid exposure during pregnancy impaired fetal development and health. However, its impacts on fertility in adult male offspring and the underlying mechanisms remain unknown. Here, pregnant mice were treated intragastrically with prednisone (PDN), a widely used synthetic glucocorticoid and emerging environmental pollutant, from gestational day 0-17. Testicular development was traced in offspring. Sperm quality and male fertility were evaluated at adulthood. Results showed that gestational PDN exposure caused sperm count decline and subfertility in adult male offspring. Microarrays profile and subsequent experiments revealed that gestational PDN exposure inhibited spermatogonial proliferation in offspring. We identified the transcription factor E2f1, known to regulate cell proliferation, as being downregulated in the testes of PDN-exposed offspring. Additionally, testicular E2f1 N6-methyladenosine (m6A) modification levels were also elevated in these offspring. Notably, these molecular changes persisted from the fetal period into adulthood. Moreover, both Mettl3 (the key m6A methyltransferase) knockdown and E2f1 overexpression alleviated the inhibited spermatogonial proliferation induced by prednisolone (the metabolite of PDN) in GC-1 cells. Finally, gestational supplementation with S-adenosylhomocysteine (SAH, a m6A inhibitor) attenuated the PDN-evoked elevation of testicular E2f1 m6A in offspring. Expectedly, PDN-induced subfertility and spermatogonial proliferation suppression in male offspring were also ameliorated by gestational SAH supplementation. These results suggest that gestational PDN exposure causes spermatogonial proliferation inhibition and subfertility via elevating testicular E2f1 m6A in male offspring. Our research provides a warning and basis for further attention regarding the potential environmental health risks associated with gestational PDN exposure in the future.