Efeito da adição de colesterol ao meio de congelamento de sêmen de bovino pantaneiro sobre a produção in vitro de embriões

Abstract

The objective of this study was to evaluate the effect of adding cholesterol to the freezing medium of Pantanal cattle semen on in vitro Embryo Production (IVP). Cumulus-Oocyte Complexes (COCs) grades I and II were selected and obtained by aspiration of ovaries from commercial slaughterhouses. Then, the COCs were submitted to in vitro maturation for 24 hours. After maturation, they were distributed in four experimental groups, totaling four drops per group, which were fertilized according to the concentration of cholesterol loaded by cyclodextrin (CCC) used during the process of cryopreservation of semen from a tested Pantaneira bull. The treatments were divided into: T1 - Control (C) - no addition of CCC; T2 - 0.5 mg/mL of CCC; T3- 1 mg/mL of CCC and T4 - 1.5 mg/mL of CCC. After 18 hours of incubation, the structures were "denuded" and transferred to In Vitro Culture (IVC) for eight days. On the second day (D2) the cleavage rate was evaluated and feeding was performed. On the eighth day (D8) the classification of embryos and the rate of blastocyst production were performed. In addition, total and apoptotic embryonic cells were counted using the differential staining method. Finally, the oxidative status was evaluated of the in vitro fertilization (IVF) and IVC media of each treatment by means of the Spontaneous Lipoperoxidation Technique or spontaneous TBARS. Treatment of Pantaneiro bovine semen with increasing concentrations of CCC resulted in a decrease in the number of cleaved structures, percentage of cleaved structures and number of blastocysts produced and total embryonic cell count (P<0.001). No significant differences were found in the other parameters evaluated. Thus, we conclude that the supplementation of cryopreserved Pantanal cattle semen with CCC is deleterious to the IVP process.