Altered expression of GDF9 and FGF16 suggests a role in the low developmental competence of repeat breeder oocytes during summer

The aim of this study was to investigate whether the poor oocyte competence verified for repeat breeder (RB) cows during summer (Ferreira et al., JDS 94:2383-92, 2011) might be associated to altered expression of genes related to oocyte maturation (GDF9, BMP15, FGF8, FGF10, FGF16 and FGF17). These genes encode local factors secreted by the oocyte that are essential to control an adequate maturation of the gamete. The oocytes were collected by OPU from 74 heifers (H), 73 peak lactation (PL) and 71 RB Holstein cows from two commercial dairy farms during the summer and winter. Ten to fifteen percent of the viable oocytes of each animal (total of 100 oocytes) was used to study the expression of selected genes, and the rest was sent to IVP (total of 1,493). Ten reference genes were analyzed with the geNorm application and three of them (RPL15, PPIA and GUSB) were determined as the most stable (Macabelli et al. 2014) and were used herein as references for normalization of target genes. Quantification of gene-specific mRNA transcripts was carried out by real-time reverse transcription (RT)-PCR using TaqMan assays. Data were tested for normality of residuals and homogeneity of variances followed by ANOVA using the GLIMMIX procedure of SAS. Amplification of FGF17 failed in all samples, leading to its exclusion from further analysis. An increased expression of BMP15 (P < 0.0001) and FGF8 (P < 0.0001) was observed during summer, regardless of category, when expression of targets was normalized by the reference genes. On other hand, in the absence of normalization, expression of BMP15 (P = 0.006) and FGF10 (P = 0.004) decreased from winter to summer, regardless of animal category. Furthermore, expression of FGF16 decreased during winter in RB compared to H (P = 0.01) and PL (P = 0.03), as well as it decreased from winter to summer in H (P < 0.0001) and PL (P = 0.0003). Yet, expression of GDF9 increased during summer in RB compared to H (P = 0.04) and PL (P = 0.01), and decreased from winter to summer in H (P < 0.0001) and PL (P < 0.0001). Together, these data provide evidence that GDF9 and FGF16 were differentially expressed in RB oocytes during summer and winter, respectively, suggesting a possible role of them in the low developmental competence of RB oocytes. This is further supported by the finding that GDF9 (P < 0.05) and FGF16 (P < 0.05) correlated positively with the number of total oocytes (r = 0.27 and 0.24, respectively), number of viable oocytes (r = 0.27 and 0.23) and blastocyst rate (r = 0.24 and 0.21). Thus, altered expression of GDF9 and FGF16 suggests that the low development competence of RB breeder oocytes during summer may be related to disrupted maturation of these gametes.