Biosynthesis of bovine oviductal explants : correlation of bovine oestrous-associated glycoprotein (BOGP) to glycosaminoglycan (CAG) and total protein synthesis

Legaspi, Alberto G. (2003). Biosynthesis of bovine oviductal explants : correlation of bovine oestrous-associated glycoprotein (BOGP) to glycosaminoglycan (CAG) and total protein synthesis PhD Thesis, School of Veterinary Science, The University of Queensland.

       
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Author Legaspi, Alberto G.
Thesis Title Biosynthesis of bovine oviductal explants : correlation of bovine oestrous-associated glycoprotein (BOGP) to glycosaminoglycan (CAG) and total protein synthesis
School, Centre or Institute School of Veterinary Science
Institution The University of Queensland
Publication date 2003
Thesis type PhD Thesis
Supervisor Prof. Michael D'Occhio
Total pages 104
Subjects 070799 Veterinary Sciences not elsewhere classified
Formatted abstract In vivo is it known that the oviduct maintains sperm fertility during the pre-ovulatory period. The beneficial effects conferred by the oviduct were attributed to factors secreted into the lumen as well as to physical binding of the sperm to the oviductal epithelium. It was hypothesised that co-encapsulating sperm and oviductal epithelial cells would prolong viability of sperm after insemination. Initial study was undertaken to establish protocol on oviductal cell encapsulation and to determine influence of varied steroidal conditions on the synthesis of sperm viability factor (s). It was found that only high molecular weight poly-1- lysine (a membrane forming polymer) was essential for the viability of encapsulated oviductal epithelial cells. To test the effects on sperm, conditioned medium from cultures of encapsulated oviductal epithelial cells was harvested at different intervals from cultures with either with oestrous cow serum, luteal cow serum, and foetal bovine serum. Capsules with no oviductal cells served as control. Surprisingly, sperm viability was readily compromised in the conditioned media regardless of treatment, although the percentage of sperm motility and viability was higher in cultures with the cow sera. Subsequent assay of the conditioned media revealed relatively high concentration of glycosaminoglycan (GAG) originating from Na alginate, an essential agent for encapsulation. Since GAG is a potent capacitating agent it was concluded that the presently adopted protocol for encapsulation, although optimal for the oviductal cells, was detrimental to sperm. Interestingly, however, assay for bovine oestrous-associated glycoprotein (BOGP, presumed pro-sperm factor) was positive for the entire six days of culture, a finding that was never obtained employing conventional culture technique. Consistent with other studies, the finding in the initial experiment that BOGP correlates with the viability of sperm provided further evidence that it is involved in the maintenance of viability and acrosomal status in the oviduct. The finding provided an opportunity to relate BOGP synthesis with other components secreted by the oviduct such as GAG and albumin, both prevalent in the oviductal fluid and are known to promote capacitation of sperm. Thus, emphasis of the study was directed on gaining insights on the mechanism by which sperm maintains viability in the isthmus oviduct prior to ovulation. The approach was to correlate BOGP synthesis, presumably a pro-sperm factor, to GAG and total protein concentration (both antisperm agents). Thus, subsequent experiments were directed with the above-mentioned objective. In a first of a series of experiments the effects of the ovarian artery-level and the systemic-level of oestradiol and progesterone was evaluated in the context of BOGP, GAG and total protein synthesis. Interestingly, results showed that the ovarian artery-level of steroid hormones was optimal in eliciting synthesis of BOGP. In contrast to BOGP, the GAG synthesis was lower when both steroid hormones were added than if either was present suggesting that in the dominance of progesterone GAG would be elevated in the oviduct. Because the synthesis of BOGP tended to decline on extended incubation, it was hypothesises that its synthesis would require constant exposure to ovarian steroids concentration. Thus, isthmic explants cultures were subjected to different regimens of medium replacements. Results revealed that BOGP synthesis was maintained in cultures with frequent (12-hr) medium replacement, but declined dramatically in cultures with no medium replacement during the 36-hr culture period. The results suggest that the decline in BOGP against an increasing GAG could be the event that triggers capacitation of sperm and the eventual release from the oviductal epithelium. Information in the preceding experiments was subsequently employed to compare the biosynthesis between isthmus and ampulla. Results demonstrated that synthesis of BOGP was higher in the ampulla than in isthmus, but vice versa for total protein synthesis. The findings suggest that with a lower BOGP:total protein ratio as oestradiol declines following ovulation would favour capacitation of sperm in the isthmus but not in the ampulla. The functional significance of higher BOGP:total protein ratio in the ampulla following ovulation would be to prevent premature acrosome reaction in order to maximise the chance of monospermic fertilisation. The study demonstrated for the first time that the ovarian artery-level of steroid hormones is essential to mimic in vivo-like biosynthesis of oviductal explant cultures. Thus, encapsulation technique could be a valuable tool in studying the various physiologic events occurring in the oviduct at least in vitro.
Keyword Cattle -- Artificial insemination
Proteins -- Synthesis

 
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