Quercetin mitigates H2O2-induced oxidative stress in bovine parthenogenetic embryos: in vitro cleavage assessment and reactive oxygen species quantification

Abstract

Antioxidants can diminish oxidative stress, improving embryonic development. Here, we evaluated the effects of an oxidative stress inducer, hydrogen peroxide (H2O2), and the antioxidant quercetin (QUE; plant-derived flavonoid) supplementation, by assessing in vitro cleavage development and reactive oxygen species (ROS) levels in bovine parthenotes. The parthenotes were either untreated, exposed to H2O2 (85 μM), QUE (2 μM), or doubleexposed. Development was assessed at 96 hours post-parthenogenetic activation. The rates were estimated for: Cleavage, 2-cell (2C), 4-cell (4C), early 8-cell (e8C: 5-8 cells), late 8-cell (L8C: 10-16 cells), and 5-16 cell (5-16C; e8C+L8C) stages. All cleaved parthenotes were stained with 2’,7’-dichlorofluorescin diacetate to quantify ROS. The possible association between development and ROS levels was evaluated by correlation analysis. The H2O2-exposed parthenotes showed a significantly lower e8C-developmental rate than untreated (P<0.05). Similarly, H2O2-exposed parthenotes revealed less L8Cdevelopmental rate than QUE-treated (P<0.05). Also, in H2O2-treated parthenotes, the 5- 16C-rate was less than untreated (P=0.0557) and QUE-treated (P=0.0502). Concerning ROS, the H2O2-treated parthenotes exhibited higher quantities than QUE-treated 2Cparthenotes (P<0.05). Furthermore, ROS levels were higher in H2O2-exposed than in untreated (P<0.001), QUE-treated, and double-exposed in e8C-parthenotes (P<0.0001). Moreover, The H2O2-exposed parthenotes exhibited higher ROS levels, as compared with untreated (P<0.001), QUE-treated, and double-exposed (P<0.0001) at the 5-16C stage. Finally, the trends in the correlation analysis suggested inverse association between development and ROS levels. In conclusion, parthenogenetic cleavage and ROS levels appear inversely correlated. Quercetin inhibits H2O2-inflicted oxidative stress by mitigating ROS levels and increases in vitro cleavage in bovine parthenotes of five to 16 cells.