Gene Expression Alteration of Sperm-Associated Antigens in Human Cryopreserved Sperm

Sperm-associated antigens (SPAGs) are 18 types of proteins, some of which play important roles in various biological functions associated with assisted reproductive technology outcomes, and are consequently important to the success of fertility programs. Despite the favorable outcomes of fecundity rates among male patients with cancer using cryopreserved sperm, the detrimental impact of freezing on cells has been noted in many studies. Cryopreservation has been thought to have adverse effects on sperm quality through disruptions in the expressions of SPAG genes. This study aimed to evaluate the effects of cryopreservation on the expressions of SPAGs genes and their transcriptome alterations in human sperm. A total of 12 normal ejaculations were prepared using the density gradient centrifugation procedure, and the motile sperm fractions were divided into fresh and frozen groups. In the latter, sperm samples were mixed with SpermFreeze^® solution as the cryoprotectant. The cryovial of sperm suspension was first held just over nitrogen vapor and then dipped inside liquid nitrogen. After 3 days, the specimens were thawed in tap water and incubated for 2 hours for recovery. Then, RNA from sperm was extracted for SPAG gene expression analysis, using real-time polymerase chain reaction. Our findings showed a decrease in expression of SPAG5 (p-value = 0.009), SPAG7 (p-value = 0.004), and SPAG12 (SNU13/NHP2L1; p-value = 0.039) genes during cryopreservation. The results indicate that the freezing procedure could negatively affect gene expression and to some extent proteins in human spermatozoa. The alteration of SPAG expression could provide new information on the molecular correlation between cryopreservation and increased failure in intracytoplasmic sperm injection and in vitro fertilization.