Genetical and biotechnological methods of utilization of female reproductive potential in mammals

The investigations included: 1/ Establishment of culture systems that would maintain the three-dimensional structure of bovine intact early antral follicles (EAF) or isolated cumulus-oocyte-granulosa complexes (COCGs) and increase the resulting portion of cumulus-oocyte complexes (COCs) with normal morphology for subsequent in vitro maturation (IVM) and in vitro fertilization (IVF), 2/ Quality assessment of IVM bovine oocytes and resulting day-8 blastocysts produced in TCM199 medium supplemented with fetal bovine serum (FBS), fatty acid free bovine serum albumin (fafBSA) or polyvinylpyrrolidone (PVP40), 3/ Testing the polymorphism of the genes: retinol binding protein (RBP4), epidermal growth factor (EGF), prostaglandin-endoperoxide synthase 2 (PTGS2), insulin-like growth factor 2 (IGF2), amphiregulin (AREG) and prolactin (PRL), and their effects on reproductive traits in swine. Isolated COCGs created in culture follicle-like structures and their oocytes achieved meiotic competence and matured to metaphase II at a higher rate than did oocytes from smaller diameter follicles which were cultured intact. The proportion of COCs with normal morphology significantly increased when isolated COCGs were embedded in microdrops of collagen gel or cultured on inserts covered with gel rather than in large gel volumes. No significant effect of maturation media composition on meiotic spindle morphology and the rate of apoptotic bovine oocytes was observed. Among day-8 embryos derived from oocytes matured with PVP40 a reduced blastocyst rate and elevated apoptotic index were found, whereas total cell count was not affected. Gene expression study also revealed a decrease in relative abundance for IGF2 and its receptor (IGF2R), and heat shock protein 70 (Hsp70) genes in PVP40 group and a significant elevation in fafBSA derived embryos. The significant effect of reproduction traits of swine (litter size and litter weight) was found for RBP4, EGF, IGF2 and AREG genes. A new polymorphism was also revealed within a promoter region of PRL gene.