Cryopreservation did not affect sperm DNA methylation levels of genes related to fertilization and embryonic development of cynomolgus macaque (Macaca fascicularis).

DNA methylation alters gene expression in numerous biological processes, including embryonic development. It is little known about the effect of cryopreservation on sperm DNA methylation. The present study has investigated whether cryopreservation causes abnormal DNA methylation in cynomolgus macaque sperm for five critical genes that includes the maternally imprinted gene (SNRPN), genes associated with male infertility (HSPA1L, MTHFR) and genes involved in embryonic development (TET3, LZTR1). Our results showed that sperm motility, the percentage of acrosomal integrity, DNA integrity and mitochondrial membrane potential were decreased after cryopreservation either being frozen with penetrating cryoprotectant, glycerol (Gly) or ethylene glycol (EG), compared to fresh sperm (p = 0.000), but the methylation patterns of the five target genes from cynomolgus macaque sperm samples were not affected after cryopreservation as evaluated by the Bisulfite Sequencing PCR (BSP) method. The data indicates that the current protocol for sperm cryopreservation of cynomolgus macaque is safe in terms of DNA methylation levels in these genes related to critical sperm functions.