Effects of scrotal hyperthermia on inducing signaling pathways of Alkaliptosis and Oxeiptosis in testicular tissue of adult male mice

Background: Alkaliptosis and Oxeiptosis are important pathways in cell-death regulation, which can be induced by exposing to high temperature. Alkaliptosis is a pH-dependent form of regulated cell death driven by activation of the IKBKB-NF-B pathway-dependent downregulation of CA9. On the other hand, Oxeiptosis is a novel oxygen radical-induced caspase-independent regulated cell death driven by the activation of the KEAP1-PGAM5-AIFM1 pathway. The objective of this study was to examine the effects of scrotal hyperthermia on the induction of Alkaliptosis and Oxeiptosis molecular pathways, and its influence on testicular tissue and sperm analysis in adult male mice. Method: In the present study, 20 adult male mice weighing 25-30 g were randomly divided into two groups, a hyperthermia group and a control group. Each animal in the hyperthermia group is induced by scrotal hyperthermia at 43°C for 20 minutes. At the end of the experimental period, serum testosterone, sperm analyses including sperm concentration, motility, survival, morphology, and testicular histological studies such as stereoscopic examination and non-apoptotic cell death were performed in both groups. In addition, the expression of genes involved in the alkaliptosis (IKBKB-NF-κB and CA9) and oxeiptosis (KEAP1, PGAM5, and AIFM1) molecular pathways was analyzed using real-time PCR. Results: In the hyperthermia group, the total sperm count and the percentage of sperm exhibiting normal motility, viability, and morphology were significantly lower than those in the control group (P