231. Mouse oocyte paracrine signalling to cumulus cells by TGF-β superfamily molecules is indispensable for cumulus expansion

Abstract

Oocyte-secreted factors are required for expansion of the mouse cumulus-oocyte complex (COC), which is necessary for ovulation. Members of the transforming growth factor-β (TGF-β) superfamily are prime candidates for the mouse cumulus expansion-enabling factor (CEEF), and we have recently determined that growth differentiation factor 9 (GDF9) alone is not the CEEF. This study was conducted to examine TGF-β superfamily processes regulating cumulus expansion. COCs were collected from eCG-primed mice and the oocyte microsurgically removed to generate oocytectomised (OOX) complexes. An established scoring system was used to measure FSH-induced cumulus expansion; 0 (no expansion) to +4 (maximum expansion). OOX complexes treated with FSH alone failed to expand (score: 0), whereas expansion was significantly (P < 0.05) induced by either GDF9 (score: mean ± SEM, 3.7 ± 0.1), activin A (2.6 ± 0.1), or co-culture with oocytes (3.2 ± 0.2). The type-I receptors for GDF9 and activin are activin receptor-like kinase 5 (ALK5) and ALK4, respectively. We tested the ability of the ALK4/5/7 kinase inhibitor, SB431542, to neutralise cumulus expansion. SB431542 completely neutralised (P < 0.05) the response of OOX complexes to GDF9, activin and oocyte-induced cumulus expansion. SB431542 also neutralised (P < 0.05) COC expansion in a dose dependent manner. Follistatin, an activin antagonist was effective at neutralising the response of OOX complexes to activin (score: 0), but had no significant effect (P > 0.05) on the expansion of OOX complexes co-cultured with oocytes (score: 2.7 ± 0.2). This study provides evidence that activin is not the sole CEEF, but signalling through the ALK4/5/7 pathway is indispensable for mouse cumulus expansion.