Demonstration of a suppressive effect of inhibin α-subunit on the developmental competence of in vitro matured bovine oocytes
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Changes in intrafollicular concentrations of different forms of inhibin (free α-subunits and αβ dimers) occur during follicle development and may influence the oocyte maturation process. The aim of this study was to investigate the effects of inhibin A and free α-subunit (pro-αC) isolated from bovine follicular fluid on maturation of bovine cumulus–oocyte complexes, as reflected by their competence for embryo development after in vitro fertilization. Bovine cumulus–oocyte complexes were isolated from ovaries obtained from an abattoir and were cultured for 22–24 h at 38.5°C in TCM-199 medium supplemented with 10% oestrous cow serum, pregnant mares' serum gonadotrophin (2.5 iu ml−1) and either inhibin A (0, 0.2 and 1.0 μg ml−1) or pro-αC (0, 2 and 10 μg ml−1). Neither inhibin A nor free α-subunit affected the cleavage rate of cumulus–oocyte complexes after fertilization (approximately 60%). Inhibin A reduced the proportion of cleaved oocytes reaching the eight-cell stage by 19% (P < 0.05), but did not affect the yield of blastocysts. However, pro-αC decreased the proportion of cleaved oocytes that reached the eight-cell (25%; P < 0.05) and blastocyst (28%; P < 0.05) stages. In addition, a negative correlation (r = −0.55, P < 0.001) was found between concentrations of total immunoreactive (ir) α-inhibin (measured by radioimmunoassay) produced by untreated control cumulus–oocyte complexes and their post-cleavage development to the blastocyst stage. In a second experiment, mouse monoclonal antibodies (20 μg ml−1) against two different regions of the inhibin α-subunit precursor (pro-region and αC fragment) were tested for their ability to neutralize endogenous inhibin α-subunit-related molecules produced by cumulus cells; control cumulus–oocyte complexes were treated with normal mouse IgG (20 μg ml−1). Although the cleavage rate was not affected, the yield of blastocysts was significantly higher in the presence of mouse monoclonal antibodies to both pro-α (77% increase; P < 0.05) and aC (48% increase; P < 0.05). None of the treatments tested affected endogenous production of activin-A or follistatin by cumulus–oocyte complexes. Overall, these results indicate that the inhibin α-subunit (pro-αC) has an inhibitory role in oocyte maturation which is independent of the modulatory effects of activin and follistatin.
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