Sperm freezing is an important technique which is widely used for long term storage of sperm. However, this technique induces partially irreversible damages to sperm. These damages can deteriorate anatomical, biochemical and structural organs in sperm such as cellular and epigenetic modification. On the other hand, oxidative damage to sperm resulting from reactive oxygen species generated by the cellular components of semen is one of the main causes for the decline in motility and fertility of sperm during the freeze–thawing process. Some of this reduction is related to epigenetic medication of sperm during freeze-thawing. Therefore, the first aim of sperm-freezing protocols is use of extenders in order to prevent lethal intracellular ice crystal formation and to reduce membrane damage during freezing-thawing. The aim of this study was to evaluate the motility, membrane integrity, viability, DNA stability, mitochondria activity, apoptosis status and epigenetic patterns such as methylation and acetylation in histone and DNA before and post cryopreservation. In this study, we used two extenders for liquid and frozen storage of rooster sperm. Sperm were collected from six matured rooster and then pooled and evaluated at 33 ?C. Finally the pooled sperm was freezed acoording to the standard procedure. The sperm parameters including motility, viability, membrane integrity, mitochondria activity, apoptosis status, DNA methylation, histone acetylation and methylation and fertility response was measured. Result showed that there was not significant difference between quality of sperm in fresh and freeze extenders. However, these parameters were significantly reduced after freezing compared to fresh status. Between freezing extenders, the higher significant of sperm quality such as motility (77.2 ± 1.6), viability (60 ± 1.69), mitochondria activity (32.9 ± 1.89) were obtained in Lake extenders compared to Beltsevill extenders. In addition the epigenetics parameters such as H3k9 acetylation and H3k4 methylation were singnificantly reduced in lake extender. It seems that extenders play a crucial roles in protection of sperm against cryodamages to improve the cellular and epigenetic parameters of rooster sperm during cryopreservation. Keywords : Cryopreservation, Sperm, Epigenetic, Histone, Mitochondria
