| Summary: | The aims of this thesis were to further study the involvement of apoptosis in bovine
ovarian follicles and embryos. In the first part of the thesis, studies in vitro and in vivo were
carried out to investigate whether apoptosis is the underlying mechanism of follicular atresia
and to characterize its regulation by hormones. Studies in vitro demonstrated that granulosa
cells (GCs) in follicles undergoing atresia display both morphological and biochemical
characteristics of apoptosis. Apoptosis was observed mainly in GCs and in scattered theca
cells. Apoptosis was not evident in cumulus cells. These results suggest that apoptosis is the
most common pathway of GC deletion which leads to the destruction of the GC layer and
finally triggers follicular atresia. Apoptosis was also detected in GCs of some
morphologically healthy follicles, indicating that apoptosis is detectable before other
morphological and biochemical signs of degeneration appear. A system for culturing GCs in
vitro was developed to study the hormonal regulation of apoptosis. Using this culture
system, it was observed that the rate of DNA fragmentation in cultured GCs from small
follicles was higher than that from medium and large follicles. Follicle stimulating hormone
(FSH) and insulin-like growth factor-I (IGF-I) attenuated spontaneous apoptotic cell death in
cultured GCs, suggesting that they are follicle survival factors.
An in vivo model in which the proestrus dominant follicle (DF) is maintained for 9
days was used to further study the precise regulation of the atresia of the nonovulatory DF
induced by high concentration of progesterone (P4). With the time of P4 administration, DFs
gradually underwent atresia and exhibited different levels of apoptosis in GCs, confirming
our previous findings. P4 did not suppress apoptotic death in cultured GCs, suggesting P4-
induced atresia of non-ovulatory bovine DFs is probably via regulation of luteinizing
hormone (LH). In addition, it was demonstrated that follicular atresia was related to a shift
in the ratio of death repressor (Bcl-2) to death inducer (Bax) protein expression.
Using an in vitro embryo production system, apoptosis was found to be related to
bovine oocyte degeneration and embryo fragmentation, suggesting the existence of a natural
preprogrammed cell death mechanism which can respond to external stimuli and/or internal
defects in bovine oocytes and embryos. The ratio of Bcl-2 and Bax protein expression was
demonstrated to be an important determinant of developmental competence for both oocytes
and embryos.
This thesis provides important insight into the mechanisms of follicular atresia and
early embryonic loss. Evaluation of the nature of these events would increase our
understanding of the limitations with follicular dynamics and early embryonic development.
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