ABSTRACT
The aims of the present study were to assess the effects of cysteamine as an anti-oxidant on the rate of in vitro maturation [IVM] of buffalo oocytes [experiment 1], and their viability and nuclear status following vitrification [experiment 2]. Immature oocytes with compact cumulus cells obtained from the ovaries of slaughtered animals were harvested and then cultured in the maturation medium with no cysteamine [control] or 50 microM cysteamine [treated]. Oocytes were vitrified in vitrification solution 1 [VS1]: 1.5 M ethylene glycol [EG] + 1.5 M dimethyl sulfoxide [DMSO] for 45 s [step one]. After this initial exposure, oocytes were transferred to VS2: 3 M EG + 3 M DMSO in a holding medium for 25 s [step two]. After warming, oocytes were evaluated morphologically and then cultured for a further 2 h in the cysteamine-supplemented or control maturation media. The oocytes were evaluated morphologically, stained with trypan blue for viability evaluation. The maturation rate of oocytes was higher [P<0.05] for IVM media with cysteamine compared with controls. There was no significant difference in morphology, survivability and maturation rate between the two vitrification groups [cysteamine-treated and untreated groups] but the morphology, survivability and percentages of metaphase-II oocytes in both groups of vitrified oocytes were lower compared with their respective controls. In conclusion, the addition of cysteamine to the maturation medium improved nuclear maturation of buffalo oocytes but had no positive effect on their cryoresistance during vitrification
ABSTRACT
This study was conducted to determine chromatin integrity and DNA damage by DNA electrophoresis and comet assays of buffalo fresh and frozen semen. Semen samples were collected from four buffalo bulls and evaluated after freezing for semen motility, viability, sperm abnormalities, chromatin integrity and DNA damage. A significant variation was found in semen parameters after thawing. Highly significant differences [P<0.001] in chromatin integrity were observed between fresh and frozen semen. For the fresh semen, there was no significant difference between the bulls for chromatin integrity; however, a significant variation [P<0.05] was detected in their frozen semen. No DNA fragmentation was observed by agarose gel electrophoresis. The percentage of sperm with damaged DNA detected by comet assay differed significantly between fresh and frozen semen. A significant negative correlation was recorded between motility and DNA damage [r=-0.68, P<0.05]. Sperm abnormalities and DNA fragmentation were significantly positively correlated [r=0.59, P<0.05]. In conclusion, DNA damage evaluation can provide reassurance about genomic normalcy and guide the development of improved methods of selecting spermatozoa with intact DNA to be used in artificial insemination