Protective effects of melatonin on DEHP‐induced apoptosis and oxidative stress in prepubertal testes via the PI3K/AKT pathway

Abstract Di(2‐ethylhexyl) phthalate (DEHP), an environmental endocrine disruptor, is one of the most common plasticizers and is widely used in various plastic products. DEHP induces apoptosis and oxidative stress and has been shown to have androgenic toxicity. However, the methods to combat DEHP‐induced testicular damage and the mechanisms involved remain to be elucidated. In the present study, we used melatonin, which has strong antioxidant properties, to intervene in prepubertal mice and mouse Leydig cells (TM3) treated with DEHP or its metabolite mono(2‐ethylhexyl) phthalate (MEHP). The results showed that melatonin protected against DEHP‐induced testicular damage in prepubertal mice, mainly by protecting against DEHP‐induced structural destruction of the germinal tubules and by attenuating the DEHP‐induced decrease in testicular organ coefficients and testosterone levels. Transcriptomic analysis found that melatonin may attenuate DEHP‐induced oxidative stress and apoptosis in prepubertal testes. In vitro studies further revealed that MEHP induces oxidative stress injury and increases apoptosis in TM3 cells, while melatonin reversed this damage. In vitro studies also found that MEHP exposure inhibited the expression levels of molecules related to the PI3K/AKT signaling pathway, and melatonin reversed this change. In conclusion, these findings suggest that melatonin protects against DEHP‐induced prepubertal testicular injury via the PI3K/AKT signaling pathway, and provide a theoretical basis and experimental rationale for combating male reproductive dysfunction.