Gene expression and apoptosis in bovine embryos during in vitro culture and in vivo development

Abstract

The first attempts to fertilise in vitro bovine oocytes were done in the late sixties but only in 1982 the first calf was born after transplantation of a complete in vitro produced embryo. Since then the in vitro production system improved a lot but it is still impossible to mimic the in vivo situation completely. Not only the efficiency of the in vitro procedure to produce morulae and blastocysts is lower then in the in vivo situation but also the quality of the in vitro produced morulae and blastocysts is lower compared to in vivo developed counterparts. The pregnancy rate after transfer of in vitro produced embryos is in general lower compared to in vivo developed embryos and a small but significant higher percentage of calves are born with abnormalities known as the "Large Offspring Syndrome". The best parameter to determine embryo quality would be its possibility to develop into a healthy calf. This parameter is very unpractical and therefore several parameters are investigated such as; morphology, cell count of the total embryo and of the inner cell mass, developmental rate, metabolism, chromosomal abnormalities, gene expression and apoptosis. In this thesis differences between completely in vitro produced and entirely in vivo developed embryos are described with special attention for cell number, gene expression and apoptosis. Furthermore it is investigated which step of the in vitro production process is responsible for the observed differences. The common strategy was to let part of the development take place in vivo and after this continuation in vitro. These embryos were then compared with embryos produced completely in vitro or entirely in vivo. Special attention was given to the maturation of the oocyte and the period of activation of the embryonic genome that in bovine embryos takes place during the fourth cell cycle, at the 8- to 16-cell stage. The results of the experiments described in this thesis show that the period after fertilisation is the most important period causing differences in embryo quality, which is in this thesis determined by mRNA expression of develop-mentally important genes, total cell number and level of apoptosis. Both the period before and after the activation of the genome seems to be important for these different aspects of embryo quality.