The Joint Annual Scientific Meetings of the Endocrine Society of Australia and the Society for Reproductive Biology 2018

The potential role of haemoglobin in sequestering menadione-induced ROS during in vitro maturation of mouse cumulus-oocyte complexes (#326)

Megan Lim 1 2 , Hannah M Brown 3 , Jeremy G Thompson 1 2 , Kylie R Dunning 1 2
  1. Australian Research Council Centre for Nanoscale BioPhotonics, The University of Adelaide, Adelaide, South Australia, Australia
  2. Robinson Research Institute, Adelaide Medical School, The University of Adelaide, Adelaide, South Australia, Australia
  3. South Australian Health and Medical Research Institute, Adelaide, South Australia, Australia

Reactive oxygen species (ROS) are known to have detrimental effects across many stages of early development, including oocyte maturation and embryo implantation, with downstream health impacts such as pre-eclampsia and birth defects. This is especially pertinent when gametes and embryos undergo artificial reproductive technologies (ART), as the in vitro environment may lack the endogenous in vivo mechanisms to quench ROS. Various strategies have attempted to quench ROS during in vitro culture through the use of antioxidants, including L-carnitine and glutathione. We have recently shown that haemoglobin (Hb) is present at high levels in oocytes and embryos developed in vivo, but absent from those derived from in vitro culture. Hb is a gas-binding protein commonly found in red blood cells and acts to decrease oxidative stress in other cell types, namely hepatocytes, brain and retina cells, where it binds ROS such as hydrogen peroxide and other superoxides. This raises the possibility that Hb acts as an antioxidant in the oocyte and embryo and that this antioxidant system is lacking during in vitro culture. Whether Hb acts as an effective antioxidant during the in vitro maturation (IVM) of cumulus-oocyte complexes (COCs) has not been determined. We first established a model of increased ROS during IVM through the addition of menadione (100uM), as demonstrated by increased CellROX Deep Red fluorescence compared to control. High concentrations of menadione (400 mM) during IVM was detrimental to oocyte quality with a significant 83% and 60% reduction in fertilisation and blastocyst rate respectively. In the presence of high levels of menadione during IVM, the addition of Hb could not restore the negative impact on oocyte quality. Experiments are now being pursued to evaluate if Hb has an antioxidant mechanism in COCs under a range of menadione concentrations.