Abstract
Infertility is becoming a more recurrent issue affecting 15% of couples on average as of 2022, with infertility being defined as the inability to conceive after 12 months of unprotected sexual intercourse. This issue of infertility has driven the development, uptake and improvement of Assisted Reproductive Technologies (ART) such as in vitro fertilization (IVF) and in vitro maturation (IVM). However, success rates with current ART are very low with 23% of ART cycles resulting in a clinical pregnancy, and 19% of cycles resulting in live births.
Many factors can contribute to the diagnosis of infertility, however oocyte quality has been shown to be the key limiting factor. Oocyte quality is an oocyte’s ability to undergo maturation and become competent in order to support embryonic development. Literature has shown that during maturation, mitochondria in mammalian species take on a hooded morphology. It was observed in a previous study that while adult ewe oocytes had an increased prevalence of hooded mitochondria over maturation, prepubertal lamb oocytes displayed a decreased percentage of hooded mitochondria present following maturation. This knowledge paired with the fact that adult oocytes exhibit greater developmental competency compared to their juvenile counterparts, led to the hypothesis that inducing hooded mitochondria formation will improve oocyte quality, while reducing hooded mitochondria formation will decrease oocyte quality. The aim of this study was to assess how mitochondrial inhibitors influence the quality of oocytes and embryos during IVM.
Sheep oocytes from both adults and prepubertal lambs were treated with the mitochondrial inhibitors FCCP and oligomycin during the final two hours of IVM. Oocytes were fertilized and cultured to analyse changes in cleavage, morula and blastocyst rates. Differences in mitochondrial morphology were distinguished through the utilization of transmission electron microscopy, while electron tomography was used to visualise hooded mitochondria three dimensionally.
Oligomycin resulted in decreased cleavage rates in lamb (p=<0.001) and adult oocytes (p=0.06), as well as decreased morula rates in lamb (p=0.006). However, FCCP had no significant effect on cleavage or morula rates in both lamb and adult oocytes, but did show a trend towards an increase in morula rate in lambs (p=0.09). In contrast to what had been previously observed in adult oocytes, both inhibitors had no effect on the percentage of hooded mitochondria present following maturation in lamb oocytes. Three dimensional reconstructions of hooded and spherical (normal) mitochondria proved that there are differences between these two forms that had previously only been observed in two dimensions.
These results demonstrate a relationship between mitochondrial function and oocyte quality as treatment with oligomycin (an inhibitor of the ATP-synthase enzyme) leads to reduced cleavage and morula rates, and overall reduced oocyte competence. Further experiments are required to better understand the function of hooded mitochondria and their impact on the quality of oocytes, with the aim of identifying ways to improve IVM.