In Vitro Development of Bovine Embryos after Fertilization using Semen from Different Donors

Contents: The aim of this study was to determine whether the semen donor and/or heparin concentration influences the rate of fertilization of bovine follicular oocytes and their subsequent embryonic development in vitro. Frozen-thawed semen from five highly fertile bulls was treated with one of four concentrations of heparin (0.5,1.0, 2.0 and 5.0 μg/ml) on a 5 x4 factorial basis in an IVM-NF programme. Zygotes/oocytes were cultured in frozen-thawed bovine oviduct cell-conditioned medium for 6 days. The use of semen from different bulls resulted in significantly (P < 0.001) different rates offertilization, as judged by cleavage rates of the oocytes at 72 h post insemination, and subsequent embryonic development through the'8-cell-block'(P < 0.05) in vitro. Development up to the morula/blastocyst stage, however, did not differ significantly (P = 0.06) among groups of oocytes fertilized with spermatozoa from different bulls. Heparin levels ranging from 0.5 to 5.0 μg/ml did not differ in their effect on in vitro fertilization as judged by the rate of normally cleaved oocytes (P = 0.14). The overall parthenogenetic division rate at 72 h post insemination was 12.4% and was not influenced by the heparin concentration. There was a linear relationship (P < 0.001) between fertility estimates based on AI and the estimates basedon the first cleavage following in vitro fertilization.     

Effect of Growth Factors on Bovine Blastocyst Development in a Serum-Free Medium

Shamsuddin, M.: Effect of growth factors on bovine blastocyst development in a serum-free medium. Acta vet. scand. 1994, 35, 141-147. - To investigate the effect of growth factors on pre-implantation development, bovine zygotes, produced by in vitro fertilization (IVF) of in vitro-matured (IVM) oocytes, were cultured in a serum-free medium to which the following growth factors were added one at a time: epidermal growth factor (EGF), acidic fibroblast growth factor (a-FGF), insulin-like growth factor-II (IGF-II), platelet-derived growth factor from human platelets (PDGF), and platelet-derived growth factor-AB, human, recombinant (PDGF-AB). All growth factors were added at a dose of either 10 or 50 ng/ml, except PDGF which was added at a dose of either 5 or 15 ng/ml. The control medium was TCM 199 supplemented with sodium pyruvate (0.25 mmol/1), BSA (10 mg/ml), insulin (5 μg/ml), transferrin (5 μg/ml), and sodium selenite (5 ng/ml). Embryos were cultured for 8 days (day of insemination = Day 0). The mean percentages of first cleavage on Day 2 varied from 67% to 86% and the differences between the 2 doses, or between the control and growth factor- treated groups were not significant (p≥0.13). The effects of the two doses on subsequent development up to the blastocyst stage did not differ either (p≥0.12). There was no stimulatory effect of any of the used exogenous growth factors on embryo development up to the morula or blastocyst stage on Day 7, or blastocyst stage on Day 8. Moreover, medium supplemented with PDGF had fewer blastocysts than the control (p≤0.03). The results indicate that growth factor supplementation may not necessarily increase the yield of blastocysts from bovine IVM-IVF oocytes in a serum-free medium.