P-138: Effect of Short Term Storage on Functional Parameters of Post-Thawed Buffalo Spermatozoa

Background: Detection of sperm parameters based on conventional methods, immediately after thawing, have been revealed sperm capacity to in vivo and in vitro fertilization. But this evaluation is not fully reliable because of long journey of sperm during female genital tract to reach the oocyte and fertilize it. Therefore, analyzing postthawed sperm parameters after incubation is more beneficial for estimating sperm fertilizing ability. Because of high precision, repeatability and ability to assist large numbers of cells in a short period of time, recent techniques such as computer assisted sperm analyzer (CASA) as well as flowcytometry have been partially substituted for routine subjective analysis. Therefore, the aim of this study is to evaluate sperm functional and flowcytometric parameters of buffalo frozen-thawed spermatozoa after thawing and following a four-hour incubation Materials and Methods: Twenty ejaculates of 4 buffalo bulls were collected twice a week according to standard artificial insemination procedures. The semen were diluted in Bioxcell extender (IMV France) and frozen in liquid nitrogen. The samples were thawed and sperm motility characteristics, membrane integrity, superoxide anion, and DNA intactness were evaluated with CASA (SCA, Spain), hypo osmotic solution test, dihydroethidium, and sperm chromatin structure assay respectively after thawing and four hours incubation in 37 °C. Mean of data were compared with paired-sample t test by significant level of 0.05. Results: Sperm motility, kinematic parameters, and membrane integrity were reduced after four hours incubation which were not statistically significant. The sperm motility, curvilinear velocity, and straight line velocity were 69.72 %, 73.97 μm/s, and 41.09 μm/s respectively after thawing and 41.20 %, 40.01 μm/s and 20.43 μm/s after 4 hours of incubation. In contrast Reactive oxygen species (52.11 vs. 66.08, p>0.05) and DNA damage (5.32 vs. 7.87, p<0.05) were increased after four hours of incubation. Conclusion: Our results showed that among the sperm functional parameters, DNA integrity was more prone to oxidative damages during 4 hours of incubation.