Reestablishment of transzonal projections and growth of bovine oocytes in vitro.
Ontology highlight
ABSTRACT:
SUBMITTER: Fushii M
PROVIDER: S-EPMC8568608 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Oocytes that undergo in vitro maturation (IVM) are metabolically abnormal and accumulate excess lipid content. However, the mechanism of lipid accumulation and the role of cumulus cells in this process are unclear. Recently, it was shown that fatty acid binding proteins (FABPs) performed intra- and extracellular fatty acid transport. We postulated that FABP3 might be responsible for fatty acid transport from cumulus cells to the oocytes via transzonal projections (TZPs) during IVM. Transcript and protein levels of FABP3 were analyzed in both in vivo- and in vitro-matured cumulus-oocyte-complexes and were increased in IVM samples. Further analysis showed increased lipid content in oocytes and cumulus cells in IVM samples compared to in vivo-derived. We therefore speculated that altered traffic of fatty acids via FABP3 during IVM was the mechanism leading to the excess of lipids accumulated within IVM oocytes. Furthermore, we demonstrated an increase in FABP3 levels and lipid content during the first 9 h of IVM, further strengthening the possibility of fatty acid transport via FABP3 and TZPs. Additionally, disruptions of TZPs during IVM decreased lipid accumulation in oocytes. Our results shed light on a possible mechanism involving FABP3 and TZPs that causes excess lipid accumulation in oocytes during IVM.
| S-EPMC5453981 | biostudies-literature
Project description:PurposeThe present study investigated the effects of docosahexaenoic acid (DHA) on the growth of bovine oocytes.MethodsOocytes and granulosa cell complexes (OGCs) were collected from early antral follicles (0.4-0.7 mm) on the surface of ovaries harvested from a slaughterhouse. The OGCs were cultured with 0, 1, and 10 μmol/L docosahexanoic acid (DHA) for 16 days.ResultsAntrum formation of the OGCs and the number of granulosa cells (GCs) surrounding the oocytes were comparable among groups, whereas supplementation of 0.1 μmol/L of DHA significantly improved oocyte growth. Oocytes grown with DHA had a higher fertilization rate, acetylation levels of H4K12, and ATP contents, as well as a lower lipid content compared with those grown without DHA. In addition, GCs surrounding OGCs grown with DHA had low lipid content compared with vehicle counterparts. Furthermore, when GCs were cultured in vitro, DHA increased ATP production, mitochondrial membrane potential, and reduced lipid content and levels of reactive oxygen species. RNA-seq of GCs revealed that DHA increased CPT1A expression levels and affect genes associated with focal adhesion, oxidative phosphorylation, and PI3K-AKT etc.ConclusionThe results suggest that DHA supplementation affects granulosa cell characteristics and supports oocyte growth in vitro.
| S-EPMC8499585 | biostudies-literature
Project description:Aging processes accelerate dramatically in oocytes that have reached the metaphase-II (M-II) stage. The present work aimed to study the patterns and intracellular pathways of actions of prolactin (PRL) and growth hormone (GH) on age-associated changes in bovine M-II oocytes aging in vitro. To this end, we analyzed spontaneous parthenogenetic activation (cytogenetic assay), apoptosis (TUNEL assay), and the developmental capacity (IVF/IVC) of in vitro-matured oocytes after prolonged culturing. Both PRL and GH reduced the activation rate of aging cumulus-enclosed oocytes (CEOs) and denuded oocytes (DOs), and their respective hormone receptors were revealed in the ova. The inhibitor of Src-family tyrosine kinases PP2 eliminated the effects of PRL and GH on meiotic arrest in DOs, whereas the MEK inhibitor U0126 only abolished the PRL effect. Furthermore, PRL was able to maintain the apoptosis resistance and developmental competence of aging CEOs. The protein kinase C inhibitor calphostin C suppressed both the actions of PRL. Thus, PRL and GH can directly support meiotic arrest in aging M-II oocytes by activating MAP kinases and/or Src-family kinases. The effect of PRL in maintaining the developmental capacity of aging oocytes is cumulus-dependent and related to the pro-survival action of the protein kinase C-mediated signal pathway.
| S-EPMC8308928 | biostudies-literature
Project description:The role of prostaglandin E2 (PGE2) in the successful resumption of oocyte meiosis and cumulus expansion has been well-documented. However, there remains very little information available on the influence of PGE2 on other processes that occur during oocyte maturation. In this study, we supplemented a maturation medium with PGE2 and monitored oocyte quality markers, glucose metabolism, mitochondrial status, oxidative stress, and apoptosis in the cumulus-oocyte complexes (COCs), using a well-established in vitro model of embryo production in cattle. We found that this increased availability of PGE2 during maturation led to an increase in the expression of genes associated with oocyte competence and improved the quality of blastocysts produced. Prostaglandin E2 also appeared to stimulate glucose uptake and lactate production in the COCs, both influencing the expression of enzymes involved in glycolysis and the hexosamine biosynthetic pathway. We found that PGE2 reduced intracellular reactive oxygen species levels, and simultaneously increased glutathione concentration and stimulated antioxidant gene expression in the oocyte. These results indicate that PGE2 has an important role in the protection of oocytes against oxidative stress. Mitochondrial membrane potential was also improved in PGE2-treated oocytes, and there was a reduction in the occurrence of apoptosis in the COCs. Promotion of an anti-apoptotic balance in transcription of genes involved in apoptosis was present in both oocytes and the cumulus cells. In summary, PGE2 could represent a novel autocrine/paracrine player in the mechanisms that can facilitate successful oocyte maturation and oocyte survival in the cow.
| S-EPMC7216604 | biostudies-literature
Project description:Rationale: Polycystic ovary syndrome (PCOS) is closely linked to follicular dysplasia and impaired bidirectional oocyte-granulosa cell (GC) communication. Given that PCOS is a heterogeneous, multifactorial endocrine disorder, it is important to clarify the pathophysiology of this ovarian disease and identify a specific treatment. Methods: We generated PCOS rat models based on neonatal tributyltin (TBT) exposure and studied the therapeutic effect and mechanism of resveratrol (RSV), a natural plant polyphenol. Transcriptome analysis was conducted to screen the significantly changed pathways, and a series of experiments, such as quantitative real-time polymerase chain reaction (PCR), Western blot and phalloidin staining, were performed in rat ovaries. We also observed similar changes in human PCOS samples using Gene Expression Omnibus (GEO) database analysis and quantitative real-time PCR. Results: We first found that injury to transzonal projections (TZPs), which are specialized filopodia that mediate oocyte-GC communication in follicles, may play an important role in the etiology of PCOS. We successfully established PCOS rat models using TBT and found that overexpressed calcium-/calmodulin-dependent protein kinase II beta (CaMKIIβ) inhibited TZP assembly. In addition, TZP disruption and CAMK2B upregulation were also observed in samples from PCOS patients. Moreover, we demonstrated that RSV potently ameliorated ovarian failure and estrus cycle disorder through TZP recovery via increased cytoplasmic calcium levels and excessive phosphorylation of CaMKIIβ. Conclusions: Our data indicated that upregulation of CaMKIIβ may play a critical role in regulating TZP assembly and may be involved in the pathogenesis of PCOS associated with ovarian dysfunction. Investigation of TZPs and RSV as potent CaMKIIβ activators provides new insight and a therapeutic target for PCOS, which is helpful for improving female reproduction.
| S-EPMC8692920 | biostudies-literature
Project description:Each mammalian oocyte is nurtured by its own multi-cellular structure, the ovarian follicle. We used new methods for serial section electron microscopy to examine entire cumulus and mural granulosa cells and their projections in mouse antral ovarian follicles. Transzonal projections (TZPs) are thin cytoplasmic projections that connect cumulus cells to the oocyte and are crucial for normal oocyte development. We studied these projections in detail and found that most TZPs do not reach the oocyte, and that they often branch and make gap junctions with each other. Furthermore, the TZPs that connect to the oocyte are usually contacted on their shaft by oocyte microvilli. Mural granulosa cells were found to possess randomly oriented cytoplasmic projections that are strikingly similar to the free-ended TZPs. We propose that granulosa cells use cytoplasmic projections to search for the oocyte, and cumulus cell differentiation results from a contact-mediated paracrine interaction with the oocyte.
| S-EPMC6362238 | biostudies-literature
Project description:OBJECTIVE:Epigallocatechin-3-gallate (EGCG) is a major ingredient of catechin polyphenols and is considered one of the most promising bioactive compounds in green tea because of its strong antioxidant properties. However, the protective role of EGCG in bovine oocyte in vitro maturation (IVM) has not been investigated. Therefore, we aimed to study the effects of EGCG on IVM of bovine oocytes. METHODS:Bovine oocytes were treated with different concentrations of EGCG (0, 25, 50, 100, and 200 μM), and the nuclear and cytoplasmic maturation, cumulus cell expansion, intracellular reactive oxygen species (ROS) levels, total antioxidant capacity, the early apoptosis and the developmental competence of in vitro fertilized embryos were measured. The mRNA abundances of antioxidant genes (nuclear factor erythriod-2 related factor 2 [NRF2], superoxide dismutase 1 [SOD1], catalase [CAT], and glutathione peroxidase 4 [GPX4]) in matured bovine oocytes were also quantified. RESULTS:Nuclear maturation which is characterized by first polar body extrusion, and cytoplasmic maturation characterized by peripheral and cortical distribution of cortical granules and homogeneous mitochondrial distribution were significantly improved in the 50 μM EGCG-treated group compared with the control group. Adding 50 μM EGCG to the maturation medium significantly increased the cumulus cell expansion index and upregulated the mRNA levels of cumulus cell expansion-related genes (hyaluronan synthase 2, tumor necrosis factor alpha induced protein 6, pentraxin 3, and prostaglandin 2). Both the intracellular ROS level and the early apoptotic rate of matured oocytes were significantly decreased in the 50 μM EGCG group, and the total antioxidant ability was markedly enhanced. Additionally, both the cleavage and blastocyst rates were significantly higher in the 50 μM EGCG-treated oocytes after in vitro fertilization than in the control oocytes. The mRNA abundance of NRF2, SOD1, CAT, and GPX4 were significantly increased in the 50 μM EGCG-treated oocytes. CONCLUSION:In conclusion, 50 μM EGCG can improve the bovine oocyte maturation, and the protective role of EGCG may be correlated with its antioxidative property.
| S-EPMC6127565 | biostudies-literature
Project description:This study aimed to evaluate the effect of scriptaid during pre-maturation (PIVM) and/or maturation (IVM) on developmental competence of bovine oocytes. Cumulus-oocyte complexes (COCs) were submitted to PIVM for 6 h in the presence or absence of scriptaid. COCs were distributed into five groups: T1-IVM for 22 h, T2-PIVM for 6 h and IVM for 22 h, T3-PIVM with scriptaid for 6 h and IVM for 22 h, T4-PIVM for 6 h and IVM with scriptaid for 22 h, and T5-PIVM with scriptaid for 6 h and IVM with scriptaid for 22 h. Nuclear maturation, gene expression, cumulus cells (CCs) expansion, and embryo development and quality were evaluated. At the end of maturation, all groups presented the majority of oocytes in MII (P>0.05). Only HAT1 gene was differentially expressed (P<0.01) in oocytes with different treatments. Regarding embryo development at D7, T4 (23%) and T5 (18%) had lower blastocyst rate (P<0.05) than the other treatments (T1 = 35%, T2 = 37% and T3 = 32%). No effect was observed when scriptaid in PIVM was used in less competent oocytes (P>0.05). In conclusion, presence of scriptaid in PIVM and/or IVM did not improve developmental competence or embryo quality.
| S-EPMC7935280 | biostudies-literature
Project description:Glucose or fatty acids (FAs) metabolisms may alter the ovarian follicle environment and thus determine oocyte and the nascent embryo quality. The aim of the experiment was to investigate the effect of selective inhibition of glucose (iodoacetate + DHEA) or FA (etomoxir) metabolism on in vitro maturation (IVM) of bovine COCs (cumulus-oocyte complexes) to investigate oocyte's development, quality, and energy metabolism. After in vitro fertilization, embryos were cultured to the blastocyst stage. Lipid droplets, metabolome, and lipidome were analyzed in oocytes and cumulus cells. mRNA expression of the selected genes was measured in the cumulus cells. ATP and glutathione relative levels were measured in oocytes. Changes in FA content in the maturation medium were evaluated by mass spectrometry. Our results indicate that only glucose metabolism is substantial to the oocyte during IVM since only glucose inhibition decreased embryo culture efficiency. The most noteworthy differences in the reaction to the applied inhibition systems were observed in cumulus cells. The upregulation of ketone body metabolism in the cumulus cells of the glucose inhibition group suggest possibly failed attempts of cells to switch into lipid consumption. On the contrary, etomoxir treatment of the oocytes did not affect embryo development, probably due to undisturbed metabolism in cumulus cells. Therefore, we suggest that the energy pathways analyzed in this experiment are not interchangeable alternatives in bovine COCs.
| S-EPMC8228821 | biostudies-literature
Project description:Increasing the success rate of in vitro maturation (IVM) for human oocytes has a major clinical significance. Previous studies have shown that growth hormone (GH) added into IVM medium could promote IVM of oocytes from non-human beings. However, few studies on systematic IVM for human oocytes with GH have been reported. Human germinal vesicle (GV) oocytes collected for IVM were cultured with different concentrations of GH to optimize the concentration. Metaphase II (MII) stage oocytes obtained from IVM were fertilized by intracytoplasmic sperm injection (ICSI). Maturation rate, fertilization rate, and blastocyst rate were assessed after IVM with or without GH. Furthermore, gene expression profiles were compared in oocytes between the two groups using single-cell RNA-seq. The optimal concentration of GH for IVM was 200 ng/ml, and the maturation rate of this group reached 70% which was double that of the control group (35%, P = 0.004). The fertilization rate (73.1 vs. 60.3%) and blastocyst rate (25.0 vs. 15.5%) both had an increasing trend in the GH group compared to controls. Single-cell RNA-Seq and real-time PCR data showed that GH could significantly enhance the expression of genes associated with meiotic progression and embryo development, such as AURKA (aurora kinase A, P = 0.007), PDIA6 (protein disulfide isomerase family A member 6, P = 0.007), LINGO2 (leucine rich repeat and Ig domain containing 2, P = 0.007), and CENPJ (centromere protein J, P = 0.039). Taken together, GH could promote maturation of human oocytes, probably through accelerating meiotic progression, balancing redox homeostasis of cellular environment, and promoting oocyte developmental competence.
| S-EPMC6667636 | biostudies-literature