Rapid warming increases survival of slow-frozen sibling oocytes: a step towards a single warming procedure irrespective of the freezing protocol?

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MicroRNAs are stored in human MII oocyte and their expression profile changes in reproductive aging

Maternal RNAs are synthesized by the oocyte during its growth; some of them are utilized for oocyte-specific processes and metabolism, others are stored and used during early development before embryonic genome activation. The appropriate expression of complex sets of genes is needed for oocyte maturation and early embryo development. In spite of the basic role of noncoding RNAs in the regulation of gene expression, few studies have analyzed their role in human oocytes. In this study, we identified the microRNAs (miRNAs) expressed in human metaphase II stage oocytes, and found that some of them are able to control pluripotency, chromatin remodeling, and early embryo development. We demonstrated that 12 miRNAs are differentially expressed in women of advanced reproductive age and, by bioinformatics analysis, we identified their mRNA targets, expressed in human oocytes and involved in the regulation of pathways altered in reproductive aging. Finally, we found the upregulation of miR-29a-3p, miR-203a-3p, and miR-494-3p, evolutionarily conserved miRNAs, also in aged mouse oocytes, and demonstrated that their overexpression is antithetically correlated with the downregulation of DNA methyltransferase 3A (Dnmt3a), DNA methyltransferase 3B (Dnmt3b), phosphatase and tensin homolog (Pten), and mitochondrial transcription factor A (Tfam). We propose that oocyte miRNAs perform an important regulatory function in human female germ cells, and their altered regulation could explain the changes occurring in oocyte aging

The natural carotenoid crocetin and the synthetic tellurium compound as101 protect the ovary against cyclophosphamide by modulating sirt1 and mitochondrial markers

Cancer therapies are associated with increased infertility risk due to accelerated reproductive aging. Oxidative stress (OS) is a potential mechanism behind ovarian toxicity by cyclophosphamide (CPM), the most ovotoxic anticancer drug. An important sensor of OS is SIRT1, a NAD+-dependent deacetylase which regulates cellular defence and cell fate. This study investigated whether the natural carotenoid crocetin and the synthetic compound AS101 protect the ovary against CPM by modulating SIRT1 and mitochondrial markers. We found that the number of primordial follicles of female CD1 mice receiving crocetin plus CPM increased when compared with CPM alone and similar to AS101, whose protective effects are known. SIRT1 increased in CPM mouse ovaries revealing the occurrence of OS. Similarly, mitochondrial SIRT3 rose, whilst SOD2 and the mitochondrial biogenesis activator PGC1-α decreased, suggesting the occurrence of mitochondrial damage. Crocetin and AS101 administration prevented SIRT1 burst suggesting that preservation of redox balance can help the ovary to counteract ovarian damage by CPM. Decreased SIRT3 and increased SOD2 and PGC1-α in mice receiving crocetin or AS101 prior to CPM provide evidence for mitochondrial protection. Present results improve the knowledge of ovarian damage by CPM and may help to develop interventions for preserving fertility in cancer patients

Possible involvement of integrin-mediated signalling in oocyte activation: evidence that a cyclic RGD-containing peptide can stimulate protein kinase C and cortical granule exocytosis in mouse oocytes

BACKGROUND: Mammalian sperm-oocyte interaction at fertilization involves several combined interactions between integrins on the oocyte and integrin ligands (disintegrins) on the sperm. Recent research has indicated the ability of peptides containing the RGD sequence that characterized several sperm disintegrins, to induce intracellular Ca2+ transients and to initiate parthenogenetic development in amphibian and bovine oocytes. In the present study, we investigate the hypothesis that an integrin-associated signalling may participate in oocyte activation signalling by determining the ability of a cyclic RGD-containing peptide to stimulate the activation of protein kinase C (PKC) and the exocytosis of cortical granules in mouse oocytes. METHODS: An In-Vitro-Fertilization assay (IVF) was carried in order to test the condition under which a peptide containing the RGD sequence, cyclo(Arg-Gly-Asp-D-Phe-Val), was able to inhibit sperm fusion with zona-free mouse oocytes at metaphase II stage. PKC activity was determined by means of an assay based on the ability of cell lysates to phosphorylate MARKS peptide, a specific PKC substrate. Loss of cortical granules was evaluated by measuring density in the oocyte cortex of cortical granules stained with LCA-biotin/Texas red-streptavidin. In all the experiments, effects of a control peptide containing a non RGD sequence, cyclo(Arg-Ala-Asp-D-Phe-Val), were evaluated. RESULTS: The IVF assay revealed that the fusion rate declined significantly when insemination was carried out in the presence of cyclic RGD peptide at concentrations > or = 250 microM (P < 0.05, Student-Newman-Keuls Method). When the peptide was applied to the oocytes at these concentrations, a dose-dependent increase of PKC activity was observed, in association with a loss of cortical granules ranging from 38+/-2.5 % to 52+/-5.4 %. Evaluation of meiotic status revealed that cyclic RGD peptide was ineffective in inducing meiosis resumption under conditions used in the present study. CONCLUSION: The presents results provide evidence that a cyclic RGD peptide highly effective in inhibiting sperm-oocyte interaction stimulates in mouse oocytes the activation of PKC and the exocytosis of cortical granules. These data support the view that RGD-binding receptors may function as signalling receptors giving rise integrated signalling not sufficient for a full oocyte activation response. This study may contribute to the understanding of possible negative effects of skipping gamete interaction in IVF techniques

Mitochondria Biology in Reproductive Function

Mitochondria are multitasking organelles involved in the maintenance of cell homeostasis [...

Dicarbonyl stress and glyoxalases in ovarian function

Tatone C, Eichenlaub-Ritter U, Amicarelli F. Dicarbonyl stress and glyoxalases in ovarian function. Biochemical Society Transactions. 2014;42(2):433-438.The ovary is the main regulator of female fertility. Changes in maternal health and physiology can disrupt intraovarian homoeostasis thereby compromising oocyte competence and fertility. Research has only recently devoted attention to the involvement of dicarbonyl stress in ovarian function. On this basis, the present review focuses on clinical and experimental research supporting the role of dicarbonyl overload and AGES (advanced glycation end-products) as key contributors to perturbations of the ovarian microenvironment leading to lower fertility. Particular emphasis has been given to oocyte susceptibility to methylglyoxal, a powerful glycating agent, whose levels are known to increase during aging and metabolic disorders. According to the literature, the ovary and the oocyte itself can rely on the glyoxalase system to counteract the possible dicarbonyl overload such as that which may occur in reproductive-age women and patients with PCOS (polycystic ovarian syndrome) or diabetes. Overall, although biochemical methods for proper evaluation of dicarbonyl stress in oocytes and the ovarian microenvironment need to be established, AGEs can be proposed as predictive markers and/or therapeutic targets in new strategies for improving reproductive counselling and infertility therapies

FOLLICLE CELL REGULATION OF MAMMALIAN OOCYTE GROWTH

To investigate mechanisms of follicle cell control on mammalian oocyte growth, preantral mouse oocytes free from surrounding follicle cells were individually cocultured with monolayers of different somatic cells competent to form gap junctions, and the rate of in vitro oocyte growth was directly correlated with the level of metabolic coupling on the same cells. The results indicate that 1) at a similar extent of metabolic coupling, mouse oocytes grew on follicle cells but not on 3T3 and Sertoli cell monolayers, and 2) the growth rate of oocytes cultured on follicle cells was dependent on the extent of metabolic coupling. It was concluded that gap-junction-mediated nutrition of ovarian mouse oocytes exerted by somatic cells is necessary but not sufficient to maintain oocyte growth. A specific regulatory role of follicle cells on mammalian oocyte growth is proposed

High Doses of D-Chiro-Inositol Alone Induce a PCO-Like Syndrome and Other Alterations in Mouse Ovaries

Administration of 1000–1500 mg/day D-Chiro-Inositol (DCIns) or a combination of Myo-Inositol (MyoIns) and DCIns in their plasma molar ratio (40:1) for three or more months are among recommended treatments for metabolic syndrome and/or Polycystic Ovary Syndrome (PCOS). We previously confirmed the efficacy of this formulation (8.2 mg/day MyoIns and 0.2 mg/day DCIns for 10 days) in a mouse PCOS model, but also observed negative effects on ovarian histology and function of formulations containing 0.4–1.6 mg/day DCIns. We therefore analyzed effects of higher doses of DCIns, 5, 10 and 20 mg/day, administered to young adult female mice for 21 days, on ovarian histology, serum testosterone levels and expression of the ovarian enzyme aromatase. Five mg/day DCIns (human correspondence: 1200 mg/day) altered ovarian histology, increased serum testosterone levels and reduced the amount of aromatase of negative controls, suggesting the induction of an androgenic PCOS model. In contrast, 10–20 mg/day DCIns (human correspondence: 2400–4800 mg/day) produced ovarian lesions resembling those typical of aged mice, and reduced serum testosterone levels without affecting aromatase amounts, suggesting a failure in steroidogenic gonadal activity. Notwithstanding physiological/biochemical differences between mice and humans, the observed pictures of toxicity for ovarian histology and function recommend caution when administering DCIns to PCOS patients at high doses and/or for periods spanning several ovulatory cycles