Stimulatory effect of growth hormone on in vitro maturation of bovine oocytes is exerted through cumulus cells and not mediated by IGF-I.
A previous study reported that the addition of bovine growth hormone (bGH) during in vitro maturation of bovine oocytes accelerates nuclear maturation and stimulates subsequent embryonic development. The aim of this study was to investigate whether bovine cumulus oocyte complexes (COCs) contain growth hormone receptor (GHR), and whether the stimulatory effect of GH on oocyte maturation is cumulus-dependent and mediated by insulin-like-growth factor (IGF-I). The expression of growth hormone receptor mRNA in mural granulosa cells, in cumulus cells, and in the oocyte was studied using reverse transcriptase polymerase chain reaction (RT-PCR). To investigate the importance of cumulus cells for GH-promoted maturation, COCs and denuded oocytes were cultured for 16 hr in M199 with or without bGH s(NIH-GH-B18). To investigate whether GH action is mediated by IGF-I, COCs were cultured in 1) 100 ng/ml bGH, 2) 100 ng/ml bGH plus anti-IGF-I, 1:100 dilution, 3) 100 ng/ml h-IGF-I, 4) 100 ng/ml h-IGF-I plus anti-IGF-I, 1:100 dilution, and 5) anti-IGF-I, 1:100 dilution. Culture was performed at 39 degrees C in a humidified atmosphere with 5% CO2 in air, and the nuclear stage of oocytes was assessed using 4,6-diamino-2-phenyl-indole (DAPI) staining. PCR on cDNA of mural granulosa cells, cumulus cells, and oocytes revealed that mRNA for GHR was present in all cell types. Addition of GH (100 ng/ml) to the culture medium of denuded oocytes did not affect the number of metaphase II oocytes after 16 hr, while a significant (P < 0.001) increase was observed, when COCs were cultured in the presence of GH. Addition of the antibody against IGF-I to the culture medium completely suppressed the stimulatory effect of IGF-I on oocyte maturation and cumulus expansion, while stimulation by GH was not affected by the antibody. It is concluded that bovine cumulus cells, mural granulosa, and oocytes express mRNA for the GH receptor. The stimulatory effect of GH on bovine oocyte maturation is dependent on the cumulus cells and is not mediated by IGF-I.
Izadyar
F
F
Department of Herd Health and Reproduction, Faculty of Veterinary Medicine, Utrecht University, The Netherlands.
Van Tol
H T
HT
Colenbrander
B
B
Bevers
M M
MM
D000818:Animals; D002417:Cattle; D005260:Female; D006107:Granulosa Cells / Q000187:drug effects; D013006:Growth Hormone / Q000494:pharmacology*; D007334:Insulin-Like Growth Factor I / Q000502:physiology*; D008969:Molecular Sequence Data; D009865:Oocytes / Q000166:cytology*; D009866:Oogenesis / Q000187:drug effects*; D006080:Ovarian Follicle / Q000166:cytology / Q000187:drug effects*; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000096:biosynthesis; D011986:Receptors, Somatotropin / Q000096:biosynthesis / Q000235:genetics; D013268:Stimulation, Chemical; D015854:Up-Regulation / Q000187:drug effects
D012333:RNA, Messenger; D011986:Receptors, Somatotropin; D007334:Insulin-Like Growth Factor I; D013006:Growth Hormone