Access Type

Open Access Dissertation

Date of Award

January 2016

Degree Type

Dissertation

Degree Name

Ph.D.

Department

Physiology

First Advisor

Husam Abu-Soud

Abstract

Inflammatory reactions mediated by oxidative stress (OS) have been implicated in the deterioration of oocyte quality, which may lead to subfertility. Oxidative stress generated from enhancement of activated macrophages secondary to an inflammatory response are the major source of reactive oxygen species (ROS) such as superoxide (O2•−), hydrogen peroxide (H2O2), hydroxyl radical (•OH), and hypochlorous acid (HOCl), as well as, the pro-inflammatory enzyme myeloperoxidase (MPO). Previously, it has been shown that these ROS have deleterious effect on oocytes; however the link between inflammation through macrophage activity and oocyte quality remains unclear. In this work, we investigated: 1) the mechanism through which direct exposure of ROS and MPO, or through their generation by activated macrophages, deteriorate oocyte quality and whether melatonin (MLT), a potent MPO inhibitor and ROS scavenger, can protect oocyte quality; and 2) the mechanism through which MLT inhibits MPO catalytic activity.

Our results indicated that ROS differentially deteriorate oocyte quality in a dose dependent manner possibly secondary to the overwhelming of the defense antioxidant capacity of the cumulus oocyte complex (COC). Cumulus cells demonstrated protection against H2O2 and •OH insult at low concentrations, but this protection was lost at higher concentrations and all concentrations of HOCl as judged by changes in the organized compact cumulus cell mass into a dispersed mass of cells with decreased cumulus cell number and viability. Therefore, increasing ROS concentration overpowered the antioxidant machinery provided by the oocyte and /or cumulus cells, through loss of cumulus cells, or the lack of scavengers for specific ROS. This mechanism of damage may be associated with infertility related to COC dysfunction and thus deterioration in oocyte quality.

Myeloperoxidase as well as activated macrophages negatively affected oocyte quality in a time dependent fashion. In all circumstances cumulus cells did not offer protection to the oocyte; however significant protection was offered by MLT. Kinetic studies have shown that MLT inhibits the MPO chlorinating (generation of HOCl) activity through its ability to compete with Cl-, the natural substrate of MPO, and serve as a one electron substrate of MPO Compounds I and II. Thus, MLT preserves the MPO peroxidation activity (by consuming H2O2 at slower rates) without the generation of HOCl through a two-step one-electron (1e−) oxidation pathway.

This study is the first to link activated macrophages, a major source of MPO and ROS, and oocyte quality deterioration, highlighting the effects of activated macrophages in infertility caused by inflammation. MLT has beneficial therapeutic effects in preserving oocyte quality, thus improving reproductive outcomes in patients with chronic inflammation.

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