Affiliation:
1. Research Centre for Reproductive Health, Department of Obstetrics and Gynaecology, The University of Adelaide, The Queen Elizabeth Hospital, Woodville Road, Woodville, South Australia, 5011, Australia
Abstract
Paracrine factors secreted by the oocyte regulate a broad range of cumulus cell functions. Characteristically, cumulus cells have a low incidence of apoptosis and we proposed that this is due to oocyte-secreted factors acting in an anti-apoptotic manner. Bovine cumulus-oocyte complexes (COC) were aspirated from abattoir-derived ovaries and oocytectomized (OOX) by microsurgical removal of the oocyte. OOX were treated with doses of either denuded oocytes (DO) or various growth factors for 24 hours (± rFSH; 0.1 IU/ml). Proportions of apoptotic cumulus cells were assessed using TUNEL and laser confocal scanning microscopy followed by image analysis. Quantification of Bcl-2 and Bax proteins in OOX was undertaken by western analysis. Oocyte removal led to a significant increase in cumulus cell apoptosis compared with COC controls (35% versus 9% TUNEL positive, respectively; P<0.001). Levels of OOX apoptosis were significantly reversed (P<0.001) in a dose-dependent manner when co-cultured with oocytes. Furthermore, the anti-apoptotic effect of oocyte-secreted factors followed a gradient from the site of the oocyte(s). Growth differentiation factor 9 (GDF9) had no significant effect on cumulus cell apoptosis. By contrast, cumulus cell apoptosis was significantly (P<0.001) reduced by bone morphogenetic proteins (BMP) 15, 6 or 7. Accordingly, levels of anti-apoptotic Bcl-2 were high in OOX+DO and OOX+BMP15 and low with OOX+GDF9 or OOX alone, whereas the reverse was observed for pro-apoptotic Bax. DO, BMP15 and BMP6 were also able to protect cumulus cells from undergoing apoptosis induced by staurosporine. FSH partially prevented apoptosis in all treatment groups (P<0.001). Follistatin and a BMP6 neutralizing antibody, which antagonized the anti-apoptotic effects of BMP15 and BMP6, respectively, whether alone or combined, blocked ∼50% of the anti-apoptotic actions of oocytes. These results are the first to demonstrate that oocyte-secreted factors, and particularly BMP15 and BMP6, maintain the low incidence of cumulus cell apoptosis by establishing a localized gradient of bone morphogenetic proteins.
Publisher
The Company of Biologists
Cited by
280 articles.
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