Maternal age and in vitro culture affect mitochondrial number and function in equine oocytes and embryos

Abstract

Advanced maternal age and in vitro embryo production (IVP) predispose to pregnancy loss in horses. We investigated whether mare age and IVP were associated with alterations in mitochondrial (mt) DNA copy number or function that could compromise oocyte and embryo development. Effects of mare age (,12 vs $12 years) on mtDNA copy number, ATP content and expression of genes involved in mitochondrial replication (mitochondrial transcription factor (TFAM), mtDNA polymerase g subunit B (mtPOLB) and mitochondrial single-stranded DNA-binding protein (SSB)), energy production (ATP synthase-coupling factor 6, mitochondrial-like (ATP-synth_F6)) and oxygen free radical scavenging (glutathione peroxidase 3 (GPX3)) were investigated in oocytes before and after in vitro maturation (IVM), and in early embryos. Expression of TFAM, mtPOLB and ATP-synth-F6 declined after IVM (P,0.05). However, maternal age did not affect oocyte ATP content or expression of genes involved in mitochondrial replication or function. Day 7 embryos from mares $12 years had fewer mtDNA copies (P¼0.01) and lower mtDNA : total DNA ratios (P,0.01) than embryos from younger mares, indicating an effect not simply due to lower cell number. Day 8 IVP embryos had similar mtDNA copy numbers to Day 7 in vivo embryos, but higher mtPOLB (P¼0.013) and a tendency to reduced GPX3 expression (P¼0.09). The lower mtDNA number in embryos from older mares may compromise development, but could be an effect rather than cause of developmental retardation. The general down-regulation of genes involved in mitochondrial replication and function after IVM may compromise resulting embryos.