P–596 Association of oleic acid production in cumulus-granulosa cells with glutathione of in vitro matured oocytes

Abstract

Abstract Study question Does oleic acid production in cumulus-granulosa cells affect glutathione levels of in vitro matured oocytes? Summary answer Oleic acid availability in cumulus-granulosa cells is associated with a higher glutathione level in in vitro matured oocytes. What is known already The monounsaturated fatty acid oleic acid is de novo synthesized by desaturation of stearic acid and can promote steroidogenesis and oocyte development in vitro. The endogenous antioxidant glutathione content in metaphase II oocyte is significantly higher than immature stages and is related to the normal oocyte maturation. Study design, size, duration Mouse germinal vesicles were co-cultured for 24 hours, during in vitro maturation, with granulosa cells treated with a specific inhibitor of oleic acid synthesis alone or in combination with oleic acid. Participants/materials, setting, methods Fluorescence staining was used to assess the glutathione content of mouse metaphase II oocytes following in vitro maturation as an indicator of cytoplasmic maturation. Glutathione was stained using Cell Tracker Blue –CMAC for 30 min at 37 °C. After being washed in fresh media, stained oocytes were photographed by a fluorescence microscope. Cell area and associated fluorescence were quantified in 20 metaphase II mouse oocytes randomly chosen from in vitro matured oocytes for each condition. Main results and the role of chance The intracellular glutathione content was profoundly lower in metaphase II oocytes obtained from co-cultures with inhibitor-treated cumulus-granulosa cells than with the control cumulus cells (–50%, p < 0.01). Oleic acid effectively recovered the negative effect of inhibitor on glutathione level nearly up to the level of the mock-treated cells. Limitations, reasons for caution The findings are limited to metaphase II. Measurement at more advanced stages of oocyte development is of interest. Inhibition of cellular fatty acid synthesis was performed solely with a specific chemical. Wider implications of the findings: Involvement of the oleic acid availability for cumulus-granulosa cells in normal oocyte maturation may be of relevance in reproductive disorders, particularly in the pathological mechanism of impaired oogenesis. Trial registration number 400/3226