Abstract
BACKGROUND: Cryopreservation can lead to sperm damage, which is why incorporating extracellular vesicles derived from normal semen can help protect against the damage caused by freezing. MATERIALS AND METHODS: Extracellular vesicles were obtained from normozoospermic semen samples (n = 15), and their effects on the cryopreservation process were studied in three groups: the control group (A, n = 6), the extracellular vesicles treated before freezing group (B, n = 6), and the extracellular vesicles treated after freezing group (C, n = 6). Characteristics of sperm after thawing, including morphology, viability, motility, total antioxidant capacity (TAC), the rate of apoptosis, mitochondrial membrane potential (MMP), and the DNA fragmentation index (DFI), as well as the expression levels of the BAX, BCL(2), CRISP(3), and KRAS genes, were examined. RESULTS: Group B displayed a significant increase in total motility, progressive motility, normal morphology, and MMP compared to groups A, and DNA fragmentation (DFI) showed a significant decrease compared to groups A and C. Also, vitality and TAC were significantly increased in group B compared to groups A and C. Decreased level of apoptosis was evident in group B compared to group A. Group A exhibited a significant increase in the expression of BAX/BCL(2) ratio compared to groups B and C. Significantly higher expression of KRAS and CRISP(3) was evident in group B compared to groups A and C. CONCLUSION: Extracellular vesicles obtained from normozoospermic seminal plasma could protect against the adverse cryopreservation effects through increased MMP and TAC, as well as decreased expression of BAX/BCL(2) ratio and increased KRAS and CRISP(3).