STIMULATORY EFFECT OF GROWTH-HORMONE ON IN-VITRO MATURATION OF BOVINEOOCYTES 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 stimula tory effect of GH on oocyte maturation is cumulus-dependent and mediat ed by insulin-like-growth factor (IGF-I), The expression of growth hor mone receptor mRNA in mural granulosa cells, in cumulus cells, and in the oocyte was studied using reverse transcriptase polymerase chain re action (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 (NIH-GH-B18). To investigate whether GH a ction 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 humidifi ed 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 cD NA 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) inc rease was observed, when COCs were cultured in the presence of GH. Add ition of the antibody against IGF-I to the culture medium completely s uppressed the stimulatory effect of IGF-I on oocyte maturation and cum ulus expansion, while stimulation by GH was not affected by the antibo dy. It is concluded that bovine cumulus cells, mural granulosa, and oo cytes express mRNA for the GH receptor, The stimulatory effect of GH o n bovine oocyte maturation is dependent on the cumulus cells and is no t mediated by IGF-I. (C) 1997 Wiley-Liss, Inc.