119 VITRIFICATION OF IMMATURE FELINE OOCYTES: EFFECTS ON ZONA PELLUCIDA ULTRASTRUCTURE AND DEVELOPMENTAL COMPETENCE

Abstract

Oocyte cryopreservation has been used to restore fertility problems in women undergoing oocyte-toxic chemotherapy and also to salvage the genetic potential of endangered/valuable species. In domestic cats, knowledge regarding the cryopreservation technique of feline oocytes is rather limited. We examined the effect of OPS vitrification on zona pellucida ultrastructure and also the developmental competence of immature cat oocytes. Immature cat oocytes were vitrified using different CPA and CPA exposure techniques: (1) 2-step DMSO, n = 71; (2) 4-step DMSO, n = 78; (3) 2-step EG, n = 73; (4) 4-step EG, n = 117; (5) 2-step EG + DMSO, n = 67 and (6) 4-step EG + DMSO, n = 88. A combination of EG and DMSO or EG alone using 4-step equilibration technique yielded the highest maturation rates (28/88: 31.8% and 44/117: 37.6%, respectively). This was, however, significantly lower than nonvitrified controls (36/59, 61% maturation rate). We further examined the effect of OPS vitrification on ultrastructure of the zona pellucida using scanning electron microscopy and also to test the quality of the zona pellucida using sperm binding assay. Our modified OPS protocols did not significantly affect the appearance of zona pellucida of vitrified oocytes and also the number of tightly bound sperm as compared with controls (80.8 ± 63.8 v. 81.6 ± 70.4, respectively). Vitrification of immature oocytes significantly reduced the number of oocytes reaching cleavage and blastocyst stages compared with nonvitrified controls (24.8 and 7.5% v. 62.5 and 30.8% for cleavage and blastocyst formation rates, respectively). It is concluded that modification of OPS vitrification can be successfully applied for immature cat oocytes, in terms of zona pellucida ultrastructure, ability to bind sperm, and also their developmental competence. This study was financially supported by MRG4980108, CHE-TRF Senior Research Scholars RTA-5080010, The Thailand Research Fund, and The Research Unit of Reproductive and Biological Science, Chulalongkorn University.