ASSESSMENT AND OPTIMIZATION OF OOCYTE QUALITY TO IMPROVE ASSISTED REPRODUCTIVE TECHNIQUE SUCCESS

WETTEN, P.A.; KLINSKY LAHOZ O.G.; MICHAUT M.A.

Mendoza

Congreso; XL Reunión Científica Anual de la Sociedad de Biología de Cuyo; 2022

Sociedad de Biología de Cuyo

In assisted reproduction technologies (ART), oocytes are inevitably subjected to postovulatory aging. Although quite significant technical progress has been made to improve ART technologies, poor oocyte quality is the crucial factor closely associated with ART failure. Understanding the subjacent mechanisms in the oocyte aging process and finding chemicals that can prevent postovulatory aging are two of the most important research topics today. In previous work, we showed that ROS levels increased during in vitro mouse oocyte aging. In addition, we also observed that: 1) cortical granules density (CGD) decreased, indicating premature exocytosis, and 2) the localization of alpha-SNAP and NSF, two proteins involved in membrane fusion during cortical granule exocytosis, were altered in postovulatory in vitro aged oocytes. Dithiothreitol (DTT) is a dithiol with two end sulfhydryl groups that works as an antioxidant. We hypothesized that DTT treatment might prevent the aforementioned alterations during in vitro mouse oocyte aging. Mature oocytes were obtained from hormonally stimulated CF-1 female mice (8-12 weeks of age). To achieve in vitro aging, oocytes were collected 16 h post hCG (time 0 h, control oocytes) and in vitro cultured by 4 or 8 h (aged oocytes) in the presence or absence of DTT. ROS levels were measured using the fluorescent indicator DCF-DA. When in vitro oocyte aging was performed in presence of DTT, ROS levels diminished significantly to similar levels observed in control oocytes. To determine the DTT effect on the premature cortical granule exocytosis, mouse oocytes were aged in presence of DTT, fixed, and stained with fluorescent LCA to label cortical granules. Results showed that aged oocytes incubated with DTT had a CGD akin to control oocytes. Likewise, indirect immunofluorescence analysis of the of alpha-SNAP and NSF localizations, showed that in aged oocytes treated with DTT both proteins had a similar distribution pattern to control oocytes. Altogether, these results suggest that the addition of antioxidants to the culture medium might be useful to avoid the alterations produced by the postovulatory in vitro oocyte aging and consequently improve the ART rate success.