Diffused Intra-Oocyte Hydrogen Peroxide Activates Myeloperoxidase and Deteriorates Oocyte Quality

Hydrogen peroxide (H(2)O(2)) is a relatively long-lived signaling molecule that plays an essential role in oocyte maturation, implantation, as well as early embryonic development. Exposure to relatively high levels of H(2)O(2) functions efficiently to accelerate oocyte aging and deteriorate oocyte quality. However, little precise information exists regarding intra-oocyte H(2)O(2) concentrations, and its diffusion to the oocyte milieu. In this work, we utilized an L-shaped amperometric integrated H(2)O(2)-selective probe to directly and quantitatively measure the real-time intra-oocyte H(2)O(2) concentration. This investigation provides an exact measurement of H(2)O(2) in situ by reducing the possible loss of H(2)O(2) caused by diffusion or reactivity with other biological systems. This experiment suggests that the intra-oocyte H(2)O(2) levels of oocytes obtained from young animals are reasonably high and remained constant during the procedure measurements. However, the intra-oocyte H(2)O(2) concentration dropped significantly (40-50% reduction) in response to catalase pre-incubation, suggesting that the measurements are truly H(2)O(2) based. To further confirm the extracellular diffusion of H(2)O(2), oocytes were incubated with myeloperoxidase (MPO), and the diffused H(2)O(2) triggered MPO chlorinating activity. Our results show that the generated hypochlorous acid (HOCl) facilitated the deterioration in oocyte quality, a process that could be prevented by pre-incubating the oocytes with melatonin, which was experimentally proven to be oxidized utilizing HPLC methods. This study is the first to demonstrate direct quantitative measurement of intracellular H(2)O(2), and its extracellular diffusion and activation of MPO as well as its impact on oocyte quality. These results may help in designing more accurate treatment plans in assisted reproduction under inflammatory conditions.