Responsiveness of bovine cumulus-oocyte-complexes (COC) to porcine and recombinant human FSH, and the effect of COC quality on gonadotropin receptor and Cx43 marker gene mRNAs during maturation in vitro.
Substantially less development to the blastocyst stage occurs in vitro than in vivo and this may be due to deficiencies in oocyte competence. Although a large proportion of bovine oocytes undergo spontaneous nuclear maturation, less is known about requirements for proper cytoplasmic maturation. Commonly, supraphysiological concentrations of FSH and LH are added to maturation media to improve cumulus expansion, fertilization and embryonic development. Therefore, various concentrations of porcine FSH (pFSH) and recombinant human FSH (rhFSH) were investigated for their effect on bovine cumulus expansion in vitro. Expression of FSHr, LHr and Cx43 mRNAs was determined in cumulus-oocyte complexes to determine whether they would be useful markers of oocyte competence. In serum-free media, only 1000 ng/ml pFSH induced marked cumulus expansion, but the effect of 100 ng/ml pFSH was amplified in the presence of 10% serum. In contrast, cumulus expansion occurred with 1 ng/ml rhFSH in the absence of serum. FSHr mRNA was highest at 0-6 h of maturation, then abundance decreased. Similarly, Cx43 mRNA expression was highest from 0-6 h but decreased by 24 h of maturation. However, the relative abundance of LHr mRNA did not change from 6-24 h of maturation. Decreased levels of FSHr, LHr and Cx43 mRNAs were detected in COCs of poorer quality. In conclusion, expansion of bovine cumulus occurred at low doses of rhFSH in serum-free media. In summary, FSHr, LHr and Cx43 mRNA abundance reflects COC quality and FSHr and Cx43 mRNA expression changes during in vitro maturation; these genes may be useful markers of oocyte developmental competence.
Calder
Michele D
MD
Department of Obstetrics and Gynaecology, The University of Western Ontario, London, Ontario, Canada. Michele.Calder@fmd.uwo.ca
Caveney
Anita N
AN
Smith
Lawrence C
LC
Watson
Andrew J
AJ
D000818:Animals; D002417:Cattle; D018031:Connexin 43 / Q000235:genetics*; D016895:Culture Media, Serum-Free / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D004958:Estradiol / Q000494:pharmacology; D005260:Female; D005640:Follicle Stimulating Hormone / Q000494:pharmacology*; D006801:Humans; D009865:Oocytes / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D009866:Oogenesis / Q000187:drug effects*; D006080:Ovarian Follicle / Q000166:cytology; D012333:RNA, Messenger / Q000096:biosynthesis*; D011962:Receptors, FSH / Q000096:biosynthesis / Q000235:genetics*; D011974:Receptors, LH / Q000096:biosynthesis / Q000235:genetics*; D011994:Recombinant Proteins / Q000494:pharmacology; D013552:Swine
D018031:Connexin 43; D016895:Culture Media, Serum-Free; D012333:RNA, Messenger; D011962:Receptors, FSH; D011974:Receptors, LH; D011994:Recombinant Proteins; D004958:Estradiol; D005640:Follicle Stimulating Hormone