# Impact of Flavonoid-Enriched Antioxidant Nanoformulation Supplementation on In Vitro Maturation and Gene Expression of Buffalo Oocytes

## TL;DR

This study shows that adding specific antioxidant nanoformulations improves the quality and maturation of buffalo oocytes in lab conditions.

## Contribution

The study introduces novel flavonoid-enriched antioxidant nanoformulations for improving buffalo oocyte maturation in vitro.

## Key findings

- Supplementing IVM medium with 0.5% EMD-300® or EMP3-H200® improved oocyte quality and reduced oxidative stress.
- 0.5% nanoformulations increased total antioxidant capacity and decreased malondialdehyde levels in IVM medium.
- 0.5% treatment enhanced OCT4 gene expression while reducing GPX4, SOD, CAT, and ATF6 gene levels.

## Abstract

Oocytes are exposed to different stressors during in vitro culture, which negatively impacts the developmental competence. Antioxidant supplementation during in vitro maturation (IVM) counteracts the detrimental effects of oxidative stress. We studied the effects of novel flavonoid-enriched antioxidant nanoformulations, namely, EMD-300® and EMP3-H200®, on oocyte IVM and the expression levels of some genes associated with antioxidant activity, apoptosis, and pluripotency in buffalo. We found that supplementation of the IVM medium with either 0.5% EMD-300® or 0.5% EMP3-H200® is associated with a reduction in oxidative stress and improvement in oocyte quality, as confirmed by changes in the gene expression patterns.

Oocytes are exposed to various stressors during in vitro maturation (IVM). Antioxidant supplementation during IVM can mitigate oxidative stress. We investigated the effects of supplementing IVM medium with novel flavonoid-enriched antioxidant nanoformulations, namely, EMD-300® and EMP3-H200®, on oocyte IVM and analyzed the expression of oxidative stress, apoptosis, and pluripotency genes in buffalo. Cumulus oocyte complexes (COCs) obtained from buffalo ovaries were matured in IVM medium supplemented with either EMD-300® or EMP3-H200® at 0.5% and 1.0% for 22 h. Following IVM, nuclear maturation, gene expression, and the levels of total antioxidant capacity (TAC) and malondialdehyde (MDA) were analyzed. Nuclear maturation was lower (p < 0.001) for the 1.0% EMD-300® group than other groups. The expressions of the GPX4, SOD, CAT, and ATF6 genes were lower (p < 0.05) in the 0.5% EMD-300® and EMP3-H200® groups than in the control. OCT4 gene expression was higher (p < 0.05) for the treated groups than control group. The level of TAC in spent IVM medium was higher for the 0.5% EMD-300® and EMP3-H200® groups than for the control. However, the MDA concentrations were lower. In conclusion, supplementing IVM medium with EMD-300® or EMP3-H200® at 0.5% improved nuclear maturation of buffalo oocytes better than 1.0%. Our findings suggest that these compounds had antioxidant effects, which assures their ability in protecting oocytes against oxidative stress.