Heat stress has been recognized as a major environmental factor affecting reproductive performance in livestock. However, the underlying mechanisms through which high temperature impairs testicular function remain elusive. This study aimed to investigate the effects of high temperature on morphology, oxidative stress levels, and testosterone synthesis in porcine testicular tissue in vitro. Testicular tissue samples from boars were subjected to different temperature conditions: control (37 °C) and heat stress (39 °C) for 4 h. The morphology of the testicular tissue was assessed using histological analysis, while oxidative stress levels were evaluated by measuring reactive oxygen species (ROS) production and antioxidant enzyme activities. Additionally, the expression of key enzymes involved in testosterone synthesis was examined using quantitative real-time polymerase chain reaction (qRT-PCR). Our results revealed that exposure to high temperatures significantly altered testicular tissue morphology. Histological analysis demonstrated degeneration and disorganization of seminiferous tubules, reduction in germ cell populations, and disruption of the blood-testis barrier. Moreover, high-temperature exposure significantly increased ROS production and decreased the activities of antioxidant enzymes (p < 0.05), indicating elevated oxidative stress levels in the testicular tissue. Furthermore, qRT-PCR analysis showed that high-temperature exposure suppressed the expression of key enzymes involved in testosterone synthesis, including steroidogenic acute regulatory protein and cytochrome P450 family 11 subfamilies A member 1. These findings suggest high temperature impairs testicular function by disrupting testicular morphology, inducing oxidative stress, and inhibiting testosterone synthesis. In conclusion, our study demonstrates that high-temperature exposure adversely affects morphology, oxidative stress levels, and testosterone synthesis in porcine testicular tissue. These findings provide insights into the potential mechanisms underlying heat-induced reproductive dysfunction in male pigs and highlight the importance of heat stress management in swine production to maintain optimal reproductive performance. Further investigations are warranted to elucidate the precise molecular pathways involved in the heat-induced testicular impairments observed in this study.
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