Effects of age on follicular fluid exosomal microRNAs and granulosa cell transforming growth factor-β signalling during follicle development in the mare.

Age-related decline in fertility is a consequence of low oocyte number and/or low oocyte competence resulting in pregnancy failure. Transforming growth factor (TGF)-β signalling is a well-studied pathway involved in follicular development and ovulation. Recently, small non-coding RNAs, namely microRNAs (miRNAs), have been demonstrated to regulate several members of this pathway; miRNAs are secreted inside small cell-secreted vesicles called exosomes. The overall goal of the present study was to determine whether altered exosome miRNA content in follicular fluid from old mares is associated with changes in TGF-β signalling in granulosa cells during follicle development. Follicular fluid was collected at deviation (n=6), mid-oestrus (n=6) and preovulation (n=6) for identification of exosomal miRNAs from young (3-12 years) and old (20-26 years) mares. Analysis of selected TGF-β signalling members revealed significantly increased levels of interleukin 6 (IL6) in granulosa cells from mid-oestrus compared with preovulatory follicles, and collagen alpha-2(I) chain (COL1A2) in granulosa cells from deviation compared with preovulatory follicles in young mares. In addition, granulosa cells from old mares had significantly altered levels of DNA-binding protein inhibitor ID-2 (ID2), signal transducer and activator of transcription 1 (STAT1) and cell division cycle 25A (CDC25A). Finally, changes in exosomal miRNA predicted to target selected TGF-β members were identified.