Oocytes from aged mares show reduced expression of mRNA for key spindle assembly checkpoint components

In man and rodents, advanced maternal age is associated with a marked increase in embryonic aneuploidy, as a result of errors in chromosome segregation during meiosis. This can either lead to sub-fertility, primarily as a result of an increased incidence of pregnancy loss, or to the generation of chromosomally abnormal offspring. During meiosis, the Spindle Assembly Checkpoint (SAC) mechanism ensures correct alignment of chromosomes on the spindle before they are separated during anaphase [Musacchio ANat Rev Mol Cell Biol. 2007May;8(5):379-93]. An age-related reduction in SAC component expression in oocytes from aged females may predispose to mis-segregation events [Jones KT Development. 2013 Sep;140(18):3719-30]. Although little data is available on the incidence of aneuploidy in equine embryos, chromosomal abnormalities could contribute to the increased incidence of early pregnancy loss in older mares. The purpose of this study was to examine whether maternal age affects the expression of SAC components and their regulators in equine oocytes. Cumulus oocytes complexes (COCs) recovered from slaughtered mares were divided into groups depending on the age of the mare (young,< 15 years; old, !15 years) and cumulus morphology (compact or expanded) at recovery, and matured in vitro for 24h. After maturation, oocytes were denuded and sub-divided into those that reached MII and those that didn't (non-MII). RNA was extracted and cDNA synthesized from pools of 10 oocytes (n1⁄44 per group). mRNA expression for SAC com- ponents and regulators (MAD2, BUB1, BUB3, BUB1B, TTK, NDC80, SPC25, AURKB, AURKC) was evaluated using RT-qPCR, with PGK1 and SRP14 as housekeeping genes. The effects of maternal age, cumulus morphology and success of maturation on mRNA expression for SAC components and regulators was analyzed using multiple regression. Gene expression for MAD2, BUB1, BUB3, BUB1B, NDC80, SPC25 and AURKB was similar in oocytes from young and old mares. Expression of TTK and AURKC was significantly reduced in oocytes from old compared to younger mares, irrespective of maturation stage or initial cumulus mor- phology. In mice, TTK1 is required for the proper timing of prometaphase, and is essential for SAC control, chromosome alignment and AURKC localization during meiosis I; its absence severely impairs chromosome segregation [Hached Ket al. Development. 2011 Jun;138(11):2261-71] .We therefore speculate that reduced expression of TTK and AURKC in oocytes from aged mares predisposes to aneuploidy and early embryonic loss.