RESUMEN
G-protein-coupled receptor kinase 2 (GRK2) interacts with Gßγ and Gαq, subunits of G-protein alpha, to regulate cell signalling. The second messenger inositol trisphosphate, produced by activated Gαq, promotes calcium release from the endoplasmic reticulum (ER) and regulates maturation-promoting factor (MPF) activity. This study aimed to investigate the role of GRK2 in MPF activity during the meiotic maturation of porcine oocytes. A specific inhibitor of GRK2 (ßi) was used in this study. The present study showed that GRK2 inhibition increased the percentage of oocyte arrest at the metaphase I (MI) stage (control: 13.84 ± 0.95%; ßi: 31.30 ± 4.18%), which resulted in the reduction of the maturation rate (control: 80.36 ± 1.94%; ßi: 65.40 ± 1.14%). The level of phospho-GRK2 decreased in the treated group, suggesting that GRK2 activity was reduced upon GRK2 inhibition. Furthermore, the addition of ßi decreased Ca2+ release from the ER. The protein levels of cyclin B and cyclin-dependent kinase 1 were higher in the treatment group than those in the control group, indicating that GRK2 inhibition prevented a decrease in MPF activity. Collectively, GRK2 inhibition induced meiotic arrest at the MI stage in porcine oocytes by preventing a decrease in MPF activity, suggesting that GRK2 is essential for oocyte meiotic maturation in pigs.
Asunto(s)
Calcio , Quinasa 2 del Receptor Acoplado a Proteína-G , Meiosis , Oocitos , Animales , Oocitos/efectos de los fármacos , Meiosis/efectos de los fármacos , Quinasa 2 del Receptor Acoplado a Proteína-G/metabolismo , Femenino , Calcio/metabolismo , Porcinos , Factor Promotor de Maduración/metabolismo , Técnicas de Maduración In Vitro de los Oocitos/veterinariaRESUMEN
BACKGROUND: Sterigmatocystin (STE) is a mycotoxin widely found in contaminated food and foodstuffs, and excessive long-term exposure to STE is associated with several health issues, including infertility. However, there is little information available regarding the effects of STE toxin on the female reproductive system, particularly concerning oocyte maturation. METHODS: In the present study, we investigated the toxic effects of STE on mouse oocyte maturation. We also used Western blot, immunofluorescence, and image quantification analyses to assess the impact of STE exposure on the oocyte maturation progression, mitochondrial distribution, oxidative stress, DNA damages, oocyte ferroptosis and asymmetric division defects. RESULTS: Our results revealed that STE exposure disrupted mouse oocyte maturation progression. When we examined the cellular changes following 100 µM STE treatment, we found that STE adversely affected polar body extrusion and induced asymmetric division defects in oocytes. RNA-sequencing data showed that STE exposure affects the expression of several pathway-correlated genes during oocyte meiosis in mice, suggesting its toxicity to oocytes. Based on the RNA-seq data, we showed that STE exposure induced oxidative stress and caused DNA damage in oocytes. Besides, ferroptosis and α-tubulin acetylation were also found in STE-exposed oocytes. Moreover, we determined that STE exposure resulted in reduced RAF1 protein expression in mouse oocytes, and inhibition of RAF1 activity also causes defects in asymmetric division of mouse oocytes. CONCLUSIONS: Collectively, our research provides novel insights into the molecular mechanisms whereby STE contributes to abnormal meiosis.
Asunto(s)
Ferroptosis , Oocitos , Esterigmatocistina , Animales , Oocitos/efectos de los fármacos , Oocitos/metabolismo , Ratones , Femenino , Ferroptosis/efectos de los fármacos , Esterigmatocistina/toxicidad , Estrés Oxidativo/efectos de los fármacos , Meiosis/efectos de los fármacos , Daño del ADNRESUMEN
Oogenesis is the central process required to produce viable oocytes in female mammals. It is initiated during embryonic development, and it involves the specification of primordial germ cells (PGCs) and progresses through the activation of the meiotic program, reaching a crucial phase in prophase I before pausing at diplotene around the time of birth. The significance of meiosis, particularly the prophase I stage, cannot be overstated, as it plays a pivotal role in ensuring the formation of healthy gametes, a prerequisite for successful reproduction. While research has explored meiosis across various organisms, understanding how environmental factors, including radiation, drugs, endocrine disruptors, reproductive age, or diet, influence this complex developmental process remains incomplete. In this chapter, we describe an ex vivo culture method to investigate meiotic prophase I and beyond and the disruption of oogenesis by external factors. Using this methodology, it is possible to evaluate the effects of individual xenobiotics by administering chemicals at specific points during oogenesis. This culture technique was optimized to study the effects of two selected endocrine disruptors (vinclozolin and MEHP), demonstrating that vinclozolin exposure delayed meiotic differentiation and MEHP exposure reduced follicle size. This approach also opens avenues for future applications, involving the exploration of established or novel pharmaceutical substances and their influence on essential events during prophase I, such as homologous recombination and chromosome segregation. These processes collectively dictate the ultimate fitness of oocytes, with potential implications for factors relevant to the reproductive age and fertility.
Asunto(s)
Meiosis , Ovario , Animales , Femenino , Ratones , Ovario/citología , Meiosis/efectos de los fármacos , Oogénesis/efectos de los fármacos , Oocitos/citología , Oocitos/efectos de los fármacos , Profase Meiótica I/efectos de los fármacos , Disruptores Endocrinos/farmacología , Oxazoles/farmacología , Embrión de Mamíferos/citología , Embrión de Mamíferos/efectos de los fármacosRESUMEN
Bisphenol M (BPM), an alternative to bisphenol A (BPA), is commonly utilized in various industrial applications. However, BPM does not represent a safe substitute for BPA due to its detrimental effects on living beings. This research aimed to assess the influence of BPM exposure on the in vitro maturation of mouse oocytes. The findings revealed that BPM exposure had a notable impact on the germinal vesicle breakdown (GVBD) rate and polar body extrusion (PBE) rate throughout the meiotic progression of mouse oocytes, ultimately resulting in meiotic arrest. Investigations demonstrated that oocytes exposure to BPM led to continued activation of spindle assembly checkpoint. Further studies revealed that securin and cyclin B1 could not be degraded in BPM-exposed oocytes, and meiosis could not realize the transition from the MI to the AI stage. Mechanistically, BPM exposure resulted in abnormal spindle assembly and disrupted chromosome alignment of oocytes. Additionally, abnormal positioning of microtubule organizing center-associated proteins implied that MTOC may be dysfunctional. Furthermore, an elevation in the acetylation level of α-tubulin in oocytes was observed after BPM treatment, leading to decreased microtubule stability. In addition to its impact on microtubules, BPM exposure led to a reduction in the expression of the actin, signifying the disruption of actin assembly. Further research indicated a heightened incidence of DNA damage in oocytes following BPM exposure. Besides, BPM exposure induced alterations in histone modifications. The outcomes of this experiment demonstrate that BPM exposure impairs oocyte quality and inhibits meiotic maturation of mouse oocytes.
Asunto(s)
Compuestos de Bencidrilo , Citoesqueleto , Meiosis , Oocitos , Fenoles , Animales , Oocitos/efectos de los fármacos , Fenoles/toxicidad , Femenino , Compuestos de Bencidrilo/toxicidad , Citoesqueleto/efectos de los fármacos , Meiosis/efectos de los fármacos , Ratones , Ciclo Celular/efectos de los fármacos , Técnicas de Maduración In Vitro de los Oocitos , Huso Acromático/efectos de los fármacos , Oogénesis/efectos de los fármacos , Tubulina (Proteína)/metabolismo , Ratones Endogámicos ICRRESUMEN
3-methyl-4-nitrophenol (PNMC), a degradation product of organophosphorus insecticides and a byproduct of fuel combustion, exerting endocrine-disrupting effects. However, its impact on the meiotic process of oocytes remains unclear. In the present study, we investigated the effects of PNMC on meiotic maturation of mouse oocytes in vitro and related mechanisms. Morphologically, PNMC-exposure affected germinal vesicle breakdown (GVBD) and polar body extrusion (PBE) in mouse oocytes. Proteomic analysis suggested that PNMC-exposure altered oocyte protein expression that are associated with cytoskeleton, mitochondrial function and oxidative stress. Further studies demonstrated that PNMC-exposure disrupted spindle assembly and chromosome alignment, caused sustained activation of spindle assembly checkpoint (SAC), and arrested meiosis in oocytes. Specifically, PNMC-exposure interfered with the function of microtubule organizing centers (MTOCs) by significantly reducing phosphorylated mitogen activated protein kinase (p-MAPK) expression and disrupting the localization of Pericentrin and p-Aurora A, leading to spindle assembly failure. Besides, PNMC-exposure also increased α-tubulin acetylation, decreased microtubule stability. Moreover, PNMC-exposure impaired mitochondrial function, evidenced by abnormal mitochondrial distribution, decreased mitochondrial membrane potential and ATP levels, release of Cytochrome C into the cytoplasm, and elevated ROS levels. As a result, exposure to PNMC caused DNA damage and early apoptosis in oocytes. Fortunately, melatonin was able to promote oocyte maturation by removing the excessive ROS and enhancing mitochondrial function. These results highlight the adverse effects of PNMC on meiotic maturation, and underscore the protective role of melatonin against PNMC-induced damage.
Asunto(s)
Meiosis , Melatonina , Mitocondrias , Oocitos , Huso Acromático , Animales , Oocitos/efectos de los fármacos , Melatonina/farmacología , Ratones , Mitocondrias/efectos de los fármacos , Femenino , Huso Acromático/efectos de los fármacos , Meiosis/efectos de los fármacos , Estrés Oxidativo/efectos de los fármacos , Disruptores Endocrinos/toxicidad , Especies Reactivas de Oxígeno/metabolismo , Antioxidantes , Insecticidas/toxicidadRESUMEN
The presence of cyclic adenosine monophosphate (cAMP) has been considered to be a fundamental factor in ensuring meiotic arrest prior to ovulation. cAMP is regarded as a key molecule in the regulation of oocyte maturation. However, it has been reported that increased levels of intracellular cAMP can result in abnormal cytokinesis, with some MI oocytes leading to symmetrically cleaved 2-cell MII oocytes. Consequently, we aimed to investigate the effects of elevated intracellular cAMP levels on abnormal cytokinesis and oocyte maturation during the meiosis of mouse oocytes. This study found that a high concentration of isobutylmethylxanthine (IBMX) also caused chromatin/chromosomes aggregation (AC) after the first meiosis. The rates of AC increased the greater the concentration of IBMX. In addition, AC formation was found to be reversible, showing that the re-formation of the spindle chromosome complex was possible after the IBMX was removed. In human oocytes, the chromosomes aggregate after the germinal vesicle breakdown and following the first and second polar body extrusions (the AC phase), while mouse oocytes do not have this AC phase. The results of our current study may indicate that the AC phase in human oocytes could be related to elevated levels of intracytoplasmic cAMP.
Asunto(s)
1-Metil-3-Isobutilxantina , Cromatina , Oocitos , Animales , Oocitos/metabolismo , Femenino , Cromatina/metabolismo , 1-Metil-3-Isobutilxantina/farmacología , Ratones , Humanos , Meiosis/efectos de los fármacos , AMP Cíclico/metabolismo , Inhibidores de Fosfodiesterasa/farmacologíaRESUMEN
During meiosis, defects in cohesin localization within the centromere region can result in various diseases. Accurate cohesin localization depends on the Mis4-Ssl3 loading complex. Although it is known that cohesin completes the loading process with the help of the loading complex, the mechanisms underlying its localization in the centromere region remain unclear. Previous studies suggest cohesin localization in the centromere is mediated by phosphorylation of centromeric proteins. In this study, we focused on the Fta2 protein, a component of the Sim4 centromere protein complex. Using bioinformatics methods, potential phosphorylation sites were identified, and fta2-9A and fta2-9D mutants were constructed in Schizosaccharomyces pombe. The phenotypes of these mutants were characterized through testing thiabendazole (TBZ) sensitivity and fluorescent microscopy localization. Results indicated that Fta2 phosphorylation did not impact mitosis but affected chromosome segregation during meiosis. This study suggests that Fta2 phosphorylation is vital for meiosis and may be related to the specific localization of cohesin during this process.
Asunto(s)
Meiosis , Proteínas de Schizosaccharomyces pombe , Schizosaccharomyces , Proteínas de Ciclo Celular/metabolismo , Proteínas de Ciclo Celular/genética , Centrómero/metabolismo , Proteínas Cromosómicas no Histona/metabolismo , Proteínas Cromosómicas no Histona/genética , Segregación Cromosómica/efectos de los fármacos , Cohesinas , Meiosis/efectos de los fármacos , Fosforilación , Schizosaccharomyces/citología , Schizosaccharomyces/efectos de los fármacos , Schizosaccharomyces/genética , Schizosaccharomyces/metabolismo , Proteínas de Schizosaccharomyces pombe/metabolismo , Proteínas de Schizosaccharomyces pombe/genéticaRESUMEN
Vinorelbine is a commonly used drug to treat various malignancies, such as breast cancer, non-small cell lung cancer, and metastatic pleural mesothelioma. Its side effects include severe neutropenia, local phlebitis, gastrointestinal reactions, and neurotoxicity. In view of the scarcity of research on vinorelbine's reproductive toxicity, this study evaluated the impact of vinorelbine ditartrate, a commonly used form of vinorelbine, on oocyte maturation in vitro. Our investigation revealed that vinorelbine ditartrate had no effect on oocyte meiotic resumption. However, it did reduce the rate of first polar body extrusion, suggesting that it could significantly impede the meiotic maturation of oocytes. Vinorelbine ditartrate exposure was found to disturb the regular spindle assembly and chromosome alignment, leading to the continuous activation of the spindle assembly checkpoint (SAC) and a delayed activation of the anaphase-promoting complex/cyclosome (APC/C), ultimately causing aneuploidy in oocytes. Consequently, the administration of vinorelbine is likely to result in oocyte aneuploidy, which can be helpful in providing a drug reference and fertility guidance in a clinical context.
Asunto(s)
Aneuploidia , Antineoplásicos Fitogénicos , Meiosis , Oocitos , Vinorelbina , Vinorelbina/toxicidad , Oocitos/efectos de los fármacos , Animales , Femenino , Meiosis/efectos de los fármacos , Antineoplásicos Fitogénicos/toxicidad , Ratones , Ciclosoma-Complejo Promotor de la Anafase , Huso Acromático/efectos de los fármacosRESUMEN
Anti-Müllerian hormone (AMH) is a key paracrine/autocrine factor regulating folliculogenesis in the postnatal ovary. As antral follicles mature to the preovulatory stage, AMH production tends to be limited to cumulus cells. Therefore, the present study investigated the role of cumulus cell-derived AMH in supporting maturation and competence of the enclosed oocyte. Cumulus-oocyte complexes (COCs) were isolated from antral follicles of rhesus macaque ovaries for in vitro maturation with or without AMH depletion. Oocyte meiotic status and embryo cleavage after in vitro fertilization were assessed. In vitro maturation with AMH depletion was also performed using COCs from antral follicles of human ovarian tissue. Oocyte maturation and morphology were evaluated. The direct AMH action on mural granulosa cells of the preovulatory follicle was further assessed using human granulosa cells cultured with or without AMH supplementation. More macaque COCs produced metaphase II oocytes with AMH depletion than those of the control culture. However, preimplantation embryonic development after in vitro fertilization was comparable between oocytes derived from COCs cultured with AMH depletion and controls. Oocytes resumed meiosis in human COCs cultured with AMH depletion and exhibited a typical spindle structure. The confluency and cell number decreased in granulosa cells cultured with AMH supplementation relative to the control culture. AMH treatment did not induce cell death in cultured human granulosa cells. Data suggest that reduced AMH action in COCs could be beneficial for oocyte maturation. Cumulus cell-derived AMH is not essential for supporting oocyte competence or mural granulosa cell viability.
Asunto(s)
Hormona Antimülleriana , Células del Cúmulo , Técnicas de Maduración In Vitro de los Oocitos , Macaca mulatta , Oocitos , Hormona Antimülleriana/metabolismo , Oocitos/metabolismo , Oocitos/citología , Oocitos/efectos de los fármacos , Femenino , Células del Cúmulo/metabolismo , Células del Cúmulo/citología , Células del Cúmulo/efectos de los fármacos , Animales , Humanos , Técnicas de Maduración In Vitro de los Oocitos/métodos , Oogénesis/fisiología , Oogénesis/efectos de los fármacos , Células Cultivadas , Fertilización In Vitro/métodos , Meiosis/fisiología , Meiosis/efectos de los fármacos , Células de la Granulosa/metabolismo , Células de la Granulosa/citología , Folículo Ovárico/metabolismo , Folículo Ovárico/citología , Folículo Ovárico/fisiología , Desarrollo Embrionario/fisiologíaRESUMEN
Fangchinoline (FA) is an alkaloid derived from the traditional Chinese medicine Fangji. Numerous studies have shown that FA has a toxic effect on various cancer cells, but little is known about its toxic effects on germ cells, especially oocytes. In this study, we investigated the effects of FA on mouse oocyte maturation and its potential mechanisms. Our results showed that FA did not affect meiosis resumption but inhibited the first polar body extrusion. This inhibition is not due to abnormalities at the organelle level, such as chromosomes and mitochondrial, which was proved by detection of DNA damage and reactive oxygen species. Further studies revealed that FA arrested the oocyte at the metaphase I stage, and this arrest was not caused by abnormal kinetochore-microtubule attachment or spindle assembly checkpoint activation. Instead, FA inhibits the activity of anaphase-promoting complexes (APC/C), as evidenced by the inhibition of CCNB1 degeneration. The decreased activity of APC/C may be due to a reduction in CDC25B activity as indicated by the high phosphorylation level of CDC25B (Ser323). This may further enhance Maturation-Promoting Factor (MPF) activity, which plays a critical role in meiosis. In conclusion, our study suggests that the metaphase I arrest caused by FA may be due to abnormalities in MPF and APC/C activity.
Asunto(s)
Bencilisoquinolinas , Factor Promotor de Maduración , Meiosis , Mesotelina , Oocitos , Animales , Meiosis/efectos de los fármacos , Oocitos/efectos de los fármacos , Femenino , Bencilisoquinolinas/farmacología , Factor Promotor de Maduración/metabolismo , Ratones , Fosfatasas cdc25/metabolismo , Fosfatasas cdc25/genética , Ciclosoma-Complejo Promotor de la Anafase/metabolismo , Ratones Endogámicos ICR , Especies Reactivas de Oxígeno/metabolismo , Daño del ADN/efectos de los fármacos , Ciclina B1/metabolismo , Ciclina B1/genéticaRESUMEN
Decabromodiphenyl ether (BDE-209) is an organic compound that is widely used in rubber, textile, electronics, plastics and other industries. It has been found that BDE-209 has a destructive effect on the reproductive system of mammals. However, the effect of BDE-209 exposure on oocyte quality and whether there is a viable salvage strategy have not been reported. Here, we report that murine oocytes exposed to BDE-209 produce a series of meiostic defects, including increased fragmentation rates and decreased PBE. Furthermore, exposure of oocytes to BDE-209 hinders mitochondrial function and disrupts mitochondrial integrity. Our observations show that supplementation with NMN successfully alleviated the meiosis impairment caused by BDE-209 and averted oocyte apoptosis by suppressing ROS generation. In conclusion, our findings suggest that NMN supplementation may be able to alleviate the oocyte quality impairment induced by BDE-209 exposure, providing a potential strategy for protecting oocytes from environmental pollutant exposure.
Asunto(s)
Éteres Difenilos Halogenados , Oocitos , Especies Reactivas de Oxígeno , Animales , Éteres Difenilos Halogenados/toxicidad , Oocitos/efectos de los fármacos , Ratones , Especies Reactivas de Oxígeno/metabolismo , Femenino , Apoptosis/efectos de los fármacos , Mitocondrias/efectos de los fármacos , Contaminantes Ambientales/toxicidad , Meiosis/efectos de los fármacos , Retardadores de Llama/toxicidadRESUMEN
Cryopreservation of in vitro matured oocytes is still considered as an experimental alternative to mature oocyte vitrification after ovarian stimulation. Here, we investigated whether rescue-IVM should be performed before or after vitrification. For this, 101 immature oocytes (germinal vesicle stage) from women undergoing ICSI were used. Oocytes were divided into three groups: freshly in vitro matured oocytes (IVM), freshly in vitro matured oocytes subsequently vitrified (IVM + VIT) and vitrified/warmed GV oocytes then in vitro matured (VIT + IVM). Oocyte maturation rates and kinetics were assessed using time-lapse technology. Spindle dimensions and polarity, chromosome alignment and cytoplasmic F-actin filament length and density were determined using confocal microscopy and quantitative image analyses. No differences in IVM rates (fresh IVM: 63.16% and IVM post-VIT: 59.38%, p = 0.72) and timings (17.73 h in fresh IVM, 17.33 h in IVM post-VIT, p = 0.72) were observed whether IVM is performed freshly or after vitrification. Meiotic spindles were shorter in VIT + IVM (10.47 µm vs 11.23 µm in IVM and 11.40 µm in IVM + VIT, p = 0.012 and p = 0.043) and wider in IVM + VIT (9.37 µm vs 8.12 µm in IVM and 8.16 µm VIT + IVM, p = 0.027 and p = 0.026). The length-to-width ratio was lower in vitrified groups (IVM + VIT: 1.19 and VIT + IVM: 1.26) compared to IVM (1.38), p = 0.013 and p = 0.014. No differences in multipolar spindle and chromosome misalignment occurrence and cytoplasmic F-actin filament length and density were observed between groups. Our results suggest vitrification before or after rescue-IVM does not seem to impair maturation rates and kinetics parameters but induces meiotic spindle alterations.
Asunto(s)
Criopreservación , Técnicas de Maduración In Vitro de los Oocitos , Meiosis , Oocitos , Huso Acromático , Vitrificación , Femenino , Humanos , Oocitos/efectos de los fármacos , Huso Acromático/fisiología , Técnicas de Maduración In Vitro de los Oocitos/métodos , Adulto , Criopreservación/métodos , Meiosis/fisiología , Meiosis/efectos de los fármacos , Cinética , Citoesqueleto de Actina/metabolismoRESUMEN
Copper ionophore NSC319726 has attracted researchers' attention in treating diseases, particularly cancers. However, its potential effects on male reproduction during medication are unclear. This study aimed to determine whether NSC319726 exposure affected the male reproductive system. The reproductive toxicity of NSC319726 was evaluated in male mice following a continuous exposure period of 5 weeks. The result showed that NSC319726 exposure caused testis index reduction, spermatogenesis dysfunction, and architectural damage in the testis and epididymis. The exposure interfered with spermatogonia proliferation, meiosis initiation, sperm count, and sperm morphology. The exposure also disturbed androgen synthesis and blood testis barrier integrity. NSC319726 treatment could elevate the copper ions in the testis to induce cuproptosis in the testis. Copper chelator rescued the elevated copper ions in the testis and partly restored the spermatogenesis dysfunction caused by NSC319726. NSC319726 treatment also decreased the level of retinol dehydrogenase 10 (RDH10), thereby inhibiting the conversion of retinol to retinoic acid, causing the inability to initiate meiosis. Retinoic acid treatment could rescue the meiotic initiation and spermatogenesis while not affecting the intracellular copper ion levels. The study provided an insight into the bio-safety of NSC319726. Retinoic acid could be a potential therapy for spermatogenesis impairment in patients undergoing treatment with NSC319726.
Asunto(s)
Cobre , Espermatogénesis , Testículo , Tretinoina , Masculino , Animales , Espermatogénesis/efectos de los fármacos , Tretinoina/farmacología , Cobre/toxicidad , Ratones , Testículo/efectos de los fármacos , Testículo/metabolismo , Testículo/patología , Espermatogonias/efectos de los fármacos , Espermatogonias/metabolismo , Espermatozoides/efectos de los fármacos , Espermatozoides/metabolismo , Meiosis/efectos de los fármacos , Epidídimo/efectos de los fármacos , Epidídimo/metabolismo , Epidídimo/patologíaRESUMEN
Follicular fluid meiosis-activating sterol (FF-MAS) is a small molecule compound found in FF, named for its ability to induce oocyte resumption of meiosis. Granulosa cells (GCs) within the follicle are typically located in a hypoxic environment under physiologic conditions due to limited vascular distribution. Previous research suggests that hypoxia-induced cell cycle arrest and apoptosis in GCs may be crucial triggering factors in porcine follicular atresia. However, the impact of FF-MAS on GCs within follicles has not been explored so far. In this study, we uncovered a novel role of FF-MAS in facilitating GC survival under hypoxic conditions by inhibiting STAT4 expression. We found that STAT4 expression was upregulated in porcine GCs exposed to 1% O2. Both gain and loss of function assays confirmed that STAT4 was required for cell apoptosis under hypoxia conditions, and that the GC apoptosis caused by hypoxia was markedly attenuated following FF-MAS treatment through inhibition of STAT4 expression. Correlation analysis in vivo revealed that GC apoptosis was associated with increased STAT4 expression, while the FF-MAS content in follicular fluid was negatively correlated with STAT4 mRNA levels and cell apoptosis. These findings elucidate a novel role of FF-MAS-mediated protection of GCs by inhibiting STAT4 expression under hypoxia, which might contribute to the mechanistic understanding of follicular development.
Granulosa cells (GCs) influence follicle growth and development, with their proliferation and differentiation promoting follicle development and ovulation, while their programmed cell death and degeneration trigger follicular atresia. In this study, to investigate the effect of FF-MAS on GCs of follicles, we performed gene expression profiling in the domestic pig (Sus scrofa). We discovered STAT4 is required for GC apoptosis under hypoxia conditions both in vitro and in vivo and FF-MAS prevents porcine ovarian granulosa cells from hypoxia-induced apoptosis via inhibiting STAT4 expression.
Asunto(s)
Apoptosis , Líquido Folicular , Células de la Granulosa , Meiosis , Factor de Transcripción STAT4 , Animales , Células de la Granulosa/efectos de los fármacos , Femenino , Apoptosis/efectos de los fármacos , Porcinos , Líquido Folicular/química , Meiosis/efectos de los fármacos , Factor de Transcripción STAT4/metabolismo , Factor de Transcripción STAT4/genética , Esteroles , Hipoxia/veterinariaRESUMEN
The use of bisphenol A (BPA) has been restricted due to its endocrine-disrupting effects. As a widely used alternative to BPA today, environmental levels of bisphenol Z (BPZ) continue to rise and accumulate in humans. Oocyte quality is critical for a successful pregnancy. Nevertheless, the toxic impacts of BPZ on the maturation of mammalian oocytes remain unexplored. Therefore, the impacts of BPZ and BPA on oocyte meiotic maturation were compared in an in vitro mouse oocyte culture model. Exposure to 150⯵M of both BPZ and BPA disrupted the assembly of the meiotic spindle and the alignment of chromosomes, and BPZ exerted stronger toxicological effects than BPA. Furthermore, BPZ resulted in aberrant expression of F-actin, preventing the formation of the actin cap. Mechanistically, BPZ exposure disrupted the mitochondrial localization pattern, reduced mitochondrial membrane potential and ATP content, leading to impaired mitochondrial function. Further studies revealed that BPZ exposure resulted in oxidative stress and altered expression of genes associated with anti-oxidative stress. Moreover, BPZ induced severe DNA damage and triggered early apoptosis in oocytes, accompanied by impaired lysosomal function. Overall, the data in this study suggest that BPZ is not a safe alternative to BPA. BPZ can trigger early apoptosis by affecting mitochondrial function and causing oxidative stress and DNA damage in oocytes. These processes disrupt cytoskeletal assembly, arrest the cell cycle, and ultimately inhibit oocyte meiotic maturation.
Asunto(s)
Compuestos de Bencidrilo , Daño del ADN , Disruptores Endocrinos , Meiosis , Mitocondrias , Oocitos , Estrés Oxidativo , Fenoles , Animales , Fenoles/toxicidad , Oocitos/efectos de los fármacos , Compuestos de Bencidrilo/toxicidad , Meiosis/efectos de los fármacos , Mitocondrias/efectos de los fármacos , Ratones , Estrés Oxidativo/efectos de los fármacos , Femenino , Disruptores Endocrinos/toxicidad , Apoptosis/efectos de los fármacos , Potencial de la Membrana Mitocondrial/efectos de los fármacos , Actinas/metabolismoRESUMEN
Triphenyltin chloride (TPTCL) is widely used in various industrial and agricultural applications. This study aimed to elucidate the mechanisms underlying the toxicological effects of TPTCL on oocytes. The obtained findings revealed that TPTCL exposure reduced polar body extrusion (PBE) and induced meiotic arrest. Mechanistically, TPTCL disrupted meiotic spindle assembly and chromosome alignment. Further analysis indicated a significant decrease in p-MAPK expression, and disturbances in the localization of Pericentrin and p-Aurora A in TPTCL exposed oocytes, which suggesting impaired microtubule organizing center (MTOC)function. Moreover, TPTCL exposure enhance microtubule acetylation and microtubule instability. Therefore, the spindle assembly checkpoint (SAC) remained activated, and the activity of the anaphase-promoting complex (APC) was inhibited, thereby preventing oocytes from progressing into the entering anaphase I (AI) stage. TPTCL exposure also augmented the actin filaments in the cytoplasm. Notably, mitochondrial function appeared unaffected by TPTCL, as evidenced indicated by stable mitochondrial membrane potential and ATP content. Furthermore, TPTCL treatment altered H3K27me2, H3K27me3 and H3K9me3 levels, suggesting changes in epigenetic modifications in oocytes. Taken together, our results suggest that TPTCL disrupts cytoskeleton assembly, continuously activates SAC, inhibits APC activity, and blocks meiotic progression, ultimately impair oocyte maturation.
Asunto(s)
Citoesqueleto , Meiosis , Oocitos , Compuestos Orgánicos de Estaño , Animales , Oocitos/efectos de los fármacos , Meiosis/efectos de los fármacos , Femenino , Citoesqueleto/efectos de los fármacos , Compuestos Orgánicos de Estaño/toxicidad , Ratones , Ratones Endogámicos ICR , Ciclo Celular/efectos de los fármacosRESUMEN
3-nitropropionic acid (3-NPA), a toxic metabolite produced by mold, is mainly found in moldy sugarcane. 3-NPA inhibits the activity of succinate dehydrogenase that can induce oxidative stress injury in cells, reduce ATP production and induce oxidative stress in mouse ovaries to cause reproductive disorders. Ursolic acid (UA) has a variety of biological activities and is a pentacyclic triterpene compound found in many plants. This experiment aimed to investigate the cytotoxicity of 3-NPA during mouse oocyte in vitro maturation and the protective effects of UA on oocytes challenged with 3-NPA. The results showed that UA could alleviate 3-NPA-induced oocyte meiotic maturation failure. Specifically, 3-NPA induced a decrease in the first polar body extrusion rate of oocytes, abnormal distribution of cortical granules, and an increase in the proportion of spindle abnormalities. In addition, 3-NPA caused mitochondrial dysfunction and induced oxidative stress, including decreases in the GSH, mitochondrial membrane potential and ATP levels, and increases in the ROS levels, and these effects led to apoptosis and autophagy. The addition of UA could significantly improve the adverse effects caused by 3-NPA. In general, our data show that 3-NPA affects the normal development of oocytes during the in vitro culture, and the addition of UA can effectively repair the damage caused by 3-NPA to oocytes.
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Meiosis , Nitrocompuestos , Oocitos , Estrés Oxidativo , Propionatos , Triterpenos , Ácido Ursólico , Animales , Nitrocompuestos/toxicidad , Propionatos/toxicidad , Oocitos/efectos de los fármacos , Oocitos/metabolismo , Femenino , Meiosis/efectos de los fármacos , Ratones , Triterpenos/farmacología , Estrés Oxidativo/efectos de los fármacos , Potencial de la Membrana Mitocondrial/efectos de los fármacos , Especies Reactivas de Oxígeno/metabolismo , Apoptosis/efectos de los fármacos , Mitocondrias/efectos de los fármacos , Mitocondrias/metabolismo , Autofagia/efectos de los fármacos , Adenosina Trifosfato/metabolismo , Ratones Endogámicos ICRRESUMEN
BACKGROUND: Bisphenol-A (BPA) has estrogenic activity and adversely affects humans and animals' reproductive systems and functions. There has been a disagreement with the safety of BPA exposure at Tolerable daily intake (TDI) (0.05 mg/kg/d) value and non-observed adverse effect level (5 mg/kg/d). The current study investigated the effects of BPA exposure at various doses starting from Tolerable daily intake (0.05 mg/kg/d) to the lowest observed adverse effect level (50 mg/kg/d) on the testis development in male mice offspring. The BPA exposure lasted for 63 days from pregnancy day 0 of the dams to post-natal day (PND) 45 of the offspring. RESULTS: The results showed that BPA exposure significantly increased testis (BPA ≥ 20 mg/kg/d) and serum (BPA ≥ 10 mg/kg/d) BPA contents of PND 45 mice. The spermatogenic cells became loose, and the lumen of seminiferous tubules enlarged when BPA exposure at 0.05 mg/kg/d TDI. BPA exposure at a low dose (0.05 mg/kg/d) significantly reduced the expression of Scp3 proteins and elevated sperm abnormality. The significant decrease in Scp3 suggested that BPA inhibits the transformation of spermatogonia into spermatozoa in the testis. The RNA-seq proved that the spliceosome was significantly inhibited in the testes of mice exposed to BPA. According to the RT-qPCR, BPA exposure significantly reduced the expression of Snrpc (BPA ≥ 20 mg/kg/d) and Hnrnpu (BPA ≥ 0.5 mg/kg/d). CONCLUSIONS: This study indicated that long-term BPA exposure at Tolerable daily intake (0.05 mg/kg/d) is not safe because low-dose long-term exposure to BPA inhibits spermatogonial meiosis in mice testis impairs reproductive function in male offspring.
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Compuestos de Bencidrilo/toxicidad , Estrógenos no Esteroides/toxicidad , Meiosis/efectos de los fármacos , Fenoles/toxicidad , Empalmosomas/efectos de los fármacos , Testículo/efectos de los fármacos , Animales , Compuestos de Bencidrilo/metabolismo , Femenino , Humanos , Masculino , Ratones , Embarazo , Semen , Espermatozoides , Testículo/metabolismoRESUMEN
α-Ketoglutarate (α-KG) is a metabolite in the tricarboxylic acid cycle. It has a strong antioxidant function and can effectively prevent oxidative damage. Previous studies have shown that α-KG exists in porcine follicles, and its content gradually increases as the follicles grow and mature. However, the potential mechanism of supplementation of α-KG on porcine oocytes during in vitro maturation (IVM) has not yet been reported. The purpose of this study was to explore the effect of α-KG on the early embryonic development of pigs and the mechanisms underlying these effects. We found that α-KG can enhance the development of early pig embryos. Adding 20 µM α-KG to the in vitro culture medium significantly increased the rate of blastocyst formation and the total cell number. Compared with to that of the control group, apoptosis in blastocysts of the supplement group was significantly reduced. α-KG reduced the production of reactive oxygen species and glutathione levels in cells. α-KG not only improved the activity of mitochondria but also inhibited the occurrence of apoptosis. After supplementation with α-KG, pig embryo pluripotency-related genes (OCT4, NANOG, and SOX2) and antiapoptotic genes (Bcl2) were upregulated. In terms of mechanism, α-KG activates the Nrf2/ARE signaling pathway to regulate the expression of antioxidant-related targets, thus combating oxidative stress during the in vitro culture of oocytes. Activated Nrf2 promotes the transcription of Bcl2 genes and inhibits cell apoptosis. These results indicate that α-KG supplements have a beneficial effect on IVM by regulating oxidative stress during the IVM of porcine oocytes and can be used as a potential antioxidant for IVM of porcine oocytes.
Asunto(s)
Antioxidantes/farmacología , Desarrollo Embrionario/efectos de los fármacos , Ácidos Cetoglutáricos/farmacología , Meiosis/efectos de los fármacos , Factor 2 Relacionado con NF-E2/metabolismo , Oocitos/metabolismo , Oogénesis/efectos de los fármacos , Estrés Oxidativo/efectos de los fármacos , Transducción de Señal/efectos de los fármacos , Animales , Apoptosis/efectos de los fármacos , Blastocisto/metabolismo , Medios de Cultivo/química , Suplementos Dietéticos , Embrión de Mamíferos/efectos de los fármacos , Embrión de Mamíferos/metabolismo , Femenino , Glutatión/metabolismo , Técnicas de Maduración In Vitro de los Oocitos/métodos , Mitocondrias/metabolismo , Oocitos/efectos de los fármacos , Embarazo , Especies Reactivas de Oxígeno/metabolismo , PorcinosRESUMEN
Oocyte maturation is essential for fertilization and early embryo development, and proper organelle functions guarantee this process to maintain high-quality oocytes. The type B trichothecene nivalenol (NIV) is a mycotoxin produced by Fusarium oxysporum and is commonly found in contaminated food. NIV intake affect growth, the immune system, and the female reproductive system. Here, we investigated NIV toxicity on mouse oocyte quality. Transcriptome analysis results showed that NIV exposure altered the expression of multiple genes involved in spindle formation and organelle function in mouse oocytes, indicating its toxicity on mouse oocyte maturation. Further analysis indicated that NIV exposure disrupted spindle structure and chromosome alignment, possibly through tubulin acetylation. NIV exposure induced aberrant mitochondria distribution and reduced mitochondria number, mitochondria membrane potential (MMP), and ATP levels. In addition, NIV caused the abnormal distribution of the Golgi apparatus and altered the expression of the vesicle trafficking protein Rab11. ER distribution was also disturbed under NIV exposure, indicating the effects of NIV on protein modification and transport in oocytes. Thus, our results demonstrated that NIV exposure affected spindle structure and organelles function in mouse oocytes.