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In this study, we reported for the first time the dose-dependent dual effects of Alpha-Ketoglutarate (AKG) on cumulus oocyte complexes (COCs) during in vitro maturation (IVM). AKG at appropriate concentration (30 µM) has beneficial effects on IVM. This includes improved cumulus expansion, oocyte quality, and embryo development. These effects are mediated through multiple underlying mechanisms. AKG reduced the excessive accumulation of reactive oxygen species (ROS) in cumulus cells, reduced the consumption of GSH and NADPH. Cumulus GSH and NADPH were transported to oocytes via gap junctions, thereby reducing the oxidative stress, apoptosis and maintaining the redox balance in oocytes. In addition, AKG improved the mitochondrial function by regulating the mitochondrial complex 1 related gene expression in oocytes to maintain mitochondrial membrane potential and ATP production. On the other hand, oocyte generated GDF9 could also be transported to cumulus cells to promote cumulus expansion. Conversely, a high concentration of AKG (750 µM) exerted adverse effects on IVM and suppressed the cumulus expansion as well as reduced the oocyte quality. The suppression of the cumulus expansion caused by high concentration of AKG could be rescued with GDF9 supplementation in COCs, indicating the critical role of GDF9 in IVM. The results provide valuable information on the variable effects of AKG at different concentrations on reproductive physiology. [ABSTRACT FROM AUTHOR]

Abstract In this study, we reported for the first time the dose-dependent dual effects of Alpha-Ketoglutarate (AKG) on cumulus oocyte complexes (COCs) during in vitro maturation (IVM). AKG at appropriate concentration (30 µM) has beneficial effects on IVM. This includes improved cumulus expansion, oocyte quality, and embryo development. These effects are mediated through multiple underlying mechanisms. AKG reduced the excessive accumulation of reactive oxygen species (ROS) in cumulus cells, reduced the consumption of GSH and NADPH. Cumulus GSH and NADPH were transported to oocytes via gap junctions, thereby reducing the oxidative stress, apoptosis and maintaining the redox balance in oocytes. In addition, AKG improved the mitochondrial function by regulating the mitochondrial complex 1 related gene expression in oocytes to maintain mitochondrial membrane potential and ATP production. On the other hand, oocyte generated GDF9 could also be transported to cumulus cells to promote cumulus expansion. Conversely, a high concentration of AKG (750 µM) exerted adverse effects on IVM and suppressed the cumulus expansion as well as reduced the oocyte quality. The suppression of the cumulus expansion caused by high concentration of AKG could be rescued with GDF9 supplementation in COCs, indicating the critical role of GDF9 in IVM. The results provide valuable information on the variable effects of AKG at different concentrations on reproductive physiology. Graphical abstract

In mammalian ovaries, oocytes are physically coupled to somatic granulosa cells, and this coupling is crucial for the growth and development of competent oocytes as it mediates the transfer of metabolic support molecules. However, aging-mediated dysregulation in communication between the oocytes and granulosa cells affects the oocyte quality. In the present study, we examined the defected germline-soma communication and reduced mRNA levels encoding key structural components of transzonal projections (TZPs) in maternally aged oocytes. Oral administration of melatonin to aged mice substantially increased TZPs and maintained the cumulus cells-oocyte communication, which played a central role in the production of adequate oocyte ATP levels and reducing the accumulation of reactive oxygen species (ROS), apoptosis, DNA damage, endoplasmic reticulum (ER) stress and spindle/chromosomal defects. This beneficial effect of melatonin was inhibited by carbenoxolone (CBX), a gap junctional uncoupler, which disrupts bidirectional communications between oocyte and somatic cells. Simultaneously, melatonin significantly increased the mRNA and protein levels corresponding to genes associated with TZPs and prevented TZP retraction in in vitro-cultured cumulus-oocyte complex (COCs). Furthermore, we infused melatonin and CBX into the COCs in vitro culture system and monitored the levels of nicotinamide adenine dinucleotide phosphate (NADPH) and glutathione (GSH) in cumulus cells and oocytes. Notably, COCs treated with melatonin demonstrated improved NADPH and GSH levels. Of note, CBX was capable of reducing NADPH and GSH levels, aggravated the ROS accumulation and ER stress. Collectively, our data demonstrate the role of melatonin in preventing age-associated germline-soma communication defects, aiding the relay of antioxidant metabolic molecules for the maintenance of oocyte quality from cumulus cells, which have important potential for improving deficient phenotypes of maternally aged oocytes and the treatment of woman infertility.

In mammalian ovaries, oocytes are physically coupled to somatic granulosa cells, and this coupling is crucial for the growth and development of competent oocytes as it mediates the transfer of metabolic support molecules. However, aging-mediated dysregulation in communication between the oocytes and granulosa cells affects the oocyte quality. In the present study, we examined the defected germline-soma communication and reduced mRNA levels encoding key structural components of transzonal projections (TZPs) in maternally aged oocytes. Oral administration of melatonin to aged mice substantially increased TZPs and maintained the cumulus cells-oocyte communication, which played a central role in the production of adequate oocyte ATP levels and reducing the accumulation of reactive oxygen species (ROS), apoptosis, DNA damage, endoplasmic reticulum (ER) stress and spindle/chromosomal defects. This beneficial effect of melatonin was inhibited by carbenoxolone (CBX), a gap junctional uncoupler, which disrupts bidirectional communications between oocyte and somatic cells. Simultaneously, melatonin significantly increased the mRNA and protein levels corresponding to genes associated with TZPs and prevented TZP retraction in in vitro-cultured cumulus-oocyte complex (COCs). Furthermore, we infused melatonin and CBX into the COCs in vitro culture system and monitored the levels of nicotinamide adenine dinucleotide phosphate (NADPH) and glutathione (GSH) in cumulus cells and oocytes. Notably, COCs treated with melatonin demonstrated improved NADPH and GSH levels. Of note, CBX was capable of reducing NADPH and GSH levels, aggravated the ROS accumulation and ER stress. Collectively, our data demonstrate the role of melatonin in preventing age-associated germline-soma communication defects, aiding the relay of antioxidant metabolic molecules for the maintenance of oocyte quality from cumulus cells, which have important potential for improving deficient phenotypes of maternally aged oocytes and the treatment of woman infertility., (Copyright © 2021 The Authors. Published by Elsevier B.V. All rights reserved.)