9-cis Retinoic acid inhibits cumulus cell apoptosis during the maturation of bovine cumulus-oocyte-complexes1

Cumulus cell (CC) apoptosis is inversely correlated with embryonic development in vitro. Therefore, inhibition of CC apoptosis is important for proper embryonic development and quality. Retinoic acids (all-transRA and 9-cisRA) are natural components of retinoids, and 9-cisRA is the physiologically active metabolite of retinoic acid in vitro. During in vitro maturation, 9-cisRA enhances oocyte competence through multiple mechanisms affecting the oocyte and preimplantation embryo; however, the effect of 9-cisRA on CC apoptosis has yet to be elucidated. The aim of the present study was to evaluate the effect of 9-cisRA on CC apoptosis and to identify the molecular mechanism underlying that effect. Bovine slaughterhouse cumulus-oocyte complexes (COC) were matured in vitro in the absence or presence of 5 nM 9-cisRA. Cumulus cells were collected from immature and matured COC for the detection of apoptosis and gene expression analysis. Results showed that 9-cisRA reduced the number of apoptotic CC by about 2.7 fold (P < 0.023), compared with control. However, apoptosis is rare in CC of immature COC (0.01% ± 0.001). Transcripts involved in the caspase cascade were down-regulated upon exposure to 9-cisRA, including tumor necrosis factor alpha (TNF-α, 11.1 fold, P < 0.001), tumor necrosis factor alpha receptor 1 (TNFRJ, 2.3 fold, P < 0.01), caspase 9 (CASP9, 2.0 fold, P < 0.03 1), caspase 8 (CASP8, 2.2 fold, P < 0.012), and caspase 3 (CASP3, 2.1 fold, P < 0.006), while antiapoptotic B-cell lymphoma 2 (BCL2) transcript was increased (3.1 fold, P < 0.004), compared with control. In addition, 9-cisRA inhibited mitogen actiratc'd protein kinase mRNA expression in CC, including extracellular signal-regulated kinase 1/2 (ERK1, 2.7 fold, P < 0.02; ERK2, 2.7 fold, P < 0.03), and c-Jun N-terminal kinase (JNK, 1.6 fold, P