Deciphering the mechanism of sperm cell aging - Functional, structural, and kinematic characteristics in fresh semen of mature and aged dogs

This study aimed to evaluate fresh semen from mature and senile dogs, identify differences in sperm parameters, and determine the correlations between sperm variables and semen quality. Forty dogs were divided into two groups based on age: mature, aged 2-4 years (n = 20); and senile, aged ≥ 9 years (n = 20). Two or three ejaculates, totaling 107 samples, were collected from each dog and evaluated for various parameters including motility and sperm movement kinematics, morphology and viability, membrane lipid disorder, DNA integrity, membrane integrity, mitochondrial potential, seminal plasma total antioxidant capacity, apoptotic-like changes, and proAKAP4 protein levels. The results showed significantly lower values for progressive motility, average path velocity, straight-line velocity, and curvilinear velocity in senile dogs than in mature dogs (P < 0.001). At the same time, the proportions of slow motile and static sperm were higher in senile dogs (P < 0.05). The semen of senile dogs was characterized by a higher percentage of apoptotic sperm, lower mitochondrial potential, higher levels of proAKAP4, and a higher percentage of defective spermatozoa (P < 0.01). The two groups had no significant differences in DNA fragmentation, membrane lipid disorder, and total antioxidant capacity in the seminal plasma. In conclusion, advancing male age in dogs is associated with the expression of early apoptotic-like changes, as evidenced by significantly increased permeability of the sperm membrane to YO-PRO-1 and low mitochondrial potential. These apoptotic markers were correlated with reduced sperm quality parameters, highlighting the impact of aging on sperm functionality.