Abstract
Abstract The role of reactive oxygen species (ROS) during oocyte in vitro maturation (IVM) is still controversial. Although an increase in ROS production may cause deleterious effects in cells, these reactive species may also act as signaling molecules influencing different cell functions. The aim of this study was to examine the effect of varying endogenous ROS levels during IVM on the process of bovine oocyte maturation. To do so, different enzymatic antioxidant (catalase, or superoxide dismutase + catalase, or diphenyl iodonium) or pro-oxidant systems (xanthine + xanthine oxidase, or xanthine + xanthine oxidase + catalase) were added to the culture medium. ROS levels were determined by 2′,7′-dichlorodihydrofluorescein diacetate stain, nuclear maturation was evaluated by the presence of the metaphase II chromosome configuration at 22h of IVM and cleavage rate was recorded 48h post- in vitro fertilization. ROS levels were only significantly increased (P<0.05) by the O2 .- generating system (xanthine + xanthine oxidase + catalase), but meiotic maturation rates were significantly lower (P<0.05) in all the evaluated systems compared with the control, except for the diphenyl iodonium group. However, this last group presented a significantly lower (P<0.05) cleavage rate in comparison to the control group. These results indicate that ROS would play an essential role during oocyte maturation, since its increase or decrease beyond a physiological level significantly reduced nuclear or cytoplasmic maturation rates in bovine oocytes.
Highlights
Reduction-oxidation reactions are essential for living systems displaying a dynamic equilibrium, where biologically important oxidations occur in a controlled manner (Aitken, 2020)
No significant differences were observed between treatments, so the fluctuations observed in Figures 1 to 3 can be exclusively attributed to the variations in endogenous reactive oxygen species (ROS) levels detected during in vitro maturation (IVM)
As far as we know, this is the first study which examined the effect of varying endogenous ROS levels during bovine oocyte IVM by adding H2O2 and O2.- enzymatic generating and scavenging systems to the culture medium
Summary
Reduction-oxidation (redox) reactions are essential for living systems displaying a dynamic equilibrium, where biologically important oxidations occur in a controlled manner (Aitken, 2020). ROS can be physiologically produced as the primary function of the NADPH oxidase (NOXs) family These enzymes transfer an electron across the cell membrane from NADPH in the cytosol to O2 in the extracellular space generating O2.which can be rapidly converted to H2O2 (Li et al, 2018). The inclusion of catalase to the xanthine + XOD system would mainly generate O2.- due to the transformation of H2O2 into H2O and O2 mediated by catalase (Von Mengden et al, 2020) Another strategy to modulate endogenous ROS levels could be to regulate NAPDH oxidases, the only enzymes whose primary function seem to be ROS production, using specific inhibitors such as diphenyl iodonium (IDP) (Altenhöfer et al, 2015). The aim of this study was to examine the effect of varying endogenous ROS levels during IVM on the process of bovine oocyte maturation by adding different enzymatic antioxidant (catalase or SOD + catalase or IDP) or pro-oxidant systems (xanthine + XOD or xanthine + XOD + catalase) to the culture medium
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