RESUMEN
The purpose of this study is to investigate whether chrysin reduces cerebral ischemia-reperfusion injury(CIRI) by inhi-biting ferroptosis in rats. Male SD rats were randomly divided into a sham group, a model group, high-, medium-, and low-dose chrysin groups(200, 100, and 50 mg·kg~(-1)), and a positive drug group(Ginaton, 21.6 mg·kg~(-1)). The CIRI model was induced in rats by transient middle cerebral artery occlusion(tMCAO). The indexes were evaluated and the samples were taken 24 h after the operation. The neurological deficit score was used to detect neurological function. The 2,3,5-triphenyl tetrazolium chloride(TTC) staining was used to detect the cerebral infarction area. Hematoxylin-eosin(HE) staining and Nissl staining were used to observe the morphological structure of brain tissues. Prussian blue staining was used to observe the iron accumulation in the brain. Total iron, lipid pero-xide, and malondialdehyde in serum and brain tissues were detected by biochemical reagents. Real-time quantitative polymerase chain reaction(RT-qPCR), immunohistochemistry, and Western blot were used to detect mRNA and protein expression of solute carrier fa-mily 7 member 11(SLC7A11), transferrin receptor 1(TFR1), glutathione peroxidase 4(GPX4), acyl-CoA synthetase long chain family member 4(ACSL4), and prostaglandin-endoperoxide synthase 2(PTGS2) in brain tissues. Compared with the model group, the groups with drug intervention showed restored neurological function, decreased cerebral infarction rate, and alleviated pathological changes. The low-dose chrysin group was selected as the optimal dosing group. Compared with the model group, the chrysin groups showed reduced content of total iron, lipid peroxide, and malondialdehyde in brain tissues and serum, increased mRNA and protein expression levels of SLC7A11 and GPX4, and decreased mRNA and protein expression levels of TFR1, PTGS2, and ACSL4. Chrysin may regulate iron metabolism via regulating the related targets of ferroptosis and inhibit neuronal ferroptosis induced by CIRI.
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Isquemia Encefálica , Ferroptosis , Daño por Reperfusión , Ratas , Masculino , Animales , Ratas Sprague-Dawley , Transducción de Señal , Isquemia Encefálica/tratamiento farmacológico , Isquemia Encefálica/genética , Isquemia Encefálica/metabolismo , Ciclooxigenasa 2/metabolismo , ARN Mensajero , Infarto Cerebral , Daño por Reperfusión/tratamiento farmacológico , Daño por Reperfusión/genética , Daño por Reperfusión/metabolismo , Malondialdehído , Infarto de la Arteria Cerebral MediaRESUMEN
This study aims to explore the mechanism of "simultaneous treatment of the brain and the heart" of Naoxintong Capsules(NXT) under cerebral ischemia based on Toll-like receptor(TLR) signaling pathway.Male SD rats were randomized into sham operation group, model group, NXT group, and positive drug group.Middle cerebral artery occlusion(MCAO) model rats were used in model group, NXT group, and positive drug group, respectively.Neurological function was scored with the Bederson scale, and brain infarct rate was measured by 2,3,5-triphenyltetrazolium chloride(TTC) staining.Brain edema was detected with wet-dry weight method.Hematoxylin-eosin(HE) staining and TdT-mediated dUTP nick-end labeling(TUNEL) staining were used to observe and count apoptotic cardiocytes.In addition, serum myocardial enzymes were measured.The expression of 8 TLR signaling pathway-related proteins interferon-α(IFN-α), interferon regulatory factor-3(IRF3), interferon regulatory factor-7(IRF7), TLR2, TLR4, TLR7, TLR9, and tumor necrosis factor-α(TNF-α) in the cerebral cortex and heart of rats was detected by Western blot. Brain infarct rate, neurological function score, and brain water content in NXT group decreased significantly compared with those in the model group. At the same time, the reduction in apoptosis rate of cardiocytes and the content of serum aspartate aminotransferase(AST), alanine aminotransferase(ALT), creatine kinase(CK), and lactate dehydrogenase(LDH) were decreased in the NXT group.Systems pharmacological results and previous research showed that TLR signaling pathway played an important role in immune inflammatory response.The study of TLR signaling pathway and related proteins is helpful to elucidate the mechanism of "simultaneous treatment of the brain and the heart". Western blot results showed that NXT significantly inhibited the expression of IRF3, IRF7, TLR2, TLR7, and TNF-α in cerebral cortex and heart under cerebral ischemia.Cerebral ischemia influences cardiac functions, and TLR signaling pathway is one of the pathways for "simultaneous treatment of the brain and the heart" of NXT.
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Isquemia Encefálica , Factor de Necrosis Tumoral alfa , Animales , Encéfalo/metabolismo , Isquemia Encefálica/metabolismo , Cápsulas , Medicamentos Herbarios Chinos , Infarto de la Arteria Cerebral Media/tratamiento farmacológico , Masculino , Miocitos Cardíacos , Ratas , Ratas Sprague-Dawley , Transducción de Señal , Receptor Toll-Like 2/genética , Receptor Toll-Like 2/metabolismo , Receptor Toll-Like 7/metabolismo , Factor de Necrosis Tumoral alfa/metabolismoRESUMEN
Oocyte activation deficiency leads to female infertility. [Ca2+ ]i oscillations are required for mitochondrial energy supplement transition from the resting to the excited state, but the underlying mechanisms are still very little known. Three mitochondrial Ca2+ channels, Mitochondria Calcium Uniporter (MCU), Na+ /Ca2+ Exchanger (NCLX) and Voltage-dependent Ca2+ Channel (VDAC), were deactivated by inhibitors RU360, CGP37157 and Erastin, respectively. Both Erastin and CGP37157 inhibited mitochondrial activity significantly while attenuating [Ca2+ ]i and [Ca2+ ]m oscillations, which caused developmental block of pronuclear formation. Thus, NCLX and VDAC are two mitochondria-associated Ca2+ transporter proteins regulating oocyte activation, which may be used as potential targets to treat female infertility. SIGNIFICANCE OF THE STUDY: NCLX and VDAC are two mitochondria-associated Ca2+ transporter proteins regulating oocyte activation.
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Canales de Calcio/metabolismo , Calcio/metabolismo , Oocitos/metabolismo , Animales , Canales de Calcio/química , Femenino , Potencial de la Membrana Mitocondrial/efectos de los fármacos , Ratones , Ratones Endogámicos ICR , Mitocondrias/metabolismo , Oocitos/citología , Oocitos/efectos de los fármacos , Compuestos de Rutenio/farmacología , Rojo de Rutenio/farmacología , Intercambiador de Sodio-Calcio/antagonistas & inhibidores , Intercambiador de Sodio-Calcio/metabolismo , Tiazepinas/farmacología , Canales Aniónicos Dependientes del Voltaje/antagonistas & inhibidores , Canales Aniónicos Dependientes del Voltaje/metabolismoRESUMEN
Insufficiency of oocyte activation impairs the subsequent embryo development in assisted reproductive technology (ART). Intracellular Ca2+ concentration ([Ca2+]i) oscillations switch the oocytes to resume the second meiosis and initiate embryonic development. However, the [Ca2+]i oscillation patterns in oocytes are poorly characterized. In this study, we investigated the effects of various factors, such as the oocytes age, pH, cumulus cells, in vitro or in vivo maturation, and ER stress on [Ca2+]i oscillation patterns and pronuclear formation after parthenogenetic activation of mouse oocytes. Our results showed that the oocytes released to the oviduct at 17 h post-human chorionic gonadotrophin (hCG) displayed a significantly stronger [Ca2+]i oscillation, including higher frequency, shorter cycle, and higher peak, compared with oocytes collected at earlier or later time points. [Ca2+]i oscillations in acidic conditions (pH 6.4 and 6.6) were significantly weaker than those in neutral and mildly alkaline conditions (pH from 6.8 to 7.6). In vitro-matured oocytes showed reduced frequency and peak of [Ca2+]i oscillations compared with those matured in vivo. In vitro-matured oocytes from the cumulus-oocyte complexes (COCs) showed a significantly higher frequency, shorter cycle, and higher peak compared with the denuded oocytes (DOs). Finally, endoplasmic reticulum stress (ER stress) severely affected the parameters of [Ca2+]i oscillations, including elongated cycles and lower frequency. The pronuclear (PN) rate of oocytes after parthenogenetic activation was correlated with [Ca2+]i oscillation pattern, decreasing with oocyte aging, cumulus removal, acidic pH, and increasing ER stress. These results provide fundamental but critical information for the mechanism of how these factors affect oocyte activation.
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Desarrollo Embrionario/genética , Estrés del Retículo Endoplásmico/genética , Técnicas de Maduración In Vitro de los Oocitos , Oocitos/crecimiento & desarrollo , Animales , Gonadotropina Coriónica/genética , Células del Cúmulo/metabolismo , Femenino , Meiosis/genética , Ratones , Partenogénesis/genética , EmbarazoRESUMEN
In vitro maturation of oocytes is a promising assisted reproductive technology (ART) for infertility treatment, although it is still not a routine technique for human ART due to reduced embryonic development. The aim of the present study was to clarify the possible reasons for reduced capacity of in vitro matured oocytes. Our results showed that the oocytes matured in vitro displayed increased abnormal mitochondrial distribution, reduced mitochondrial membrane potential, and increased reactive oxygen species levels when compared to in vivo matured oocytes. These results were not different in oocytes matured in vitro with or without cumulus cells. Notably, in vitro matured oocytes displayed increased mitochondrial DNA numbers probably due to functional compensation. In vitro matured oocytes showed significantly lower activation and embryonic development rates, and their ability to produce Ca2+ oscillations was much lower in response to parthenogenetic activation, especially in oocytes matured in vitro without cumulus cells with nearly half of them failing to produce calcium waves upon strontium chloride stimulation. These data are important for understanding the reasons for reduced developmental potential of in vitro matured oocytes and the importance of cumulus cells for oocyte quality.
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ADN Mitocondrial/genética , Técnicas de Maduración In Vitro de los Oocitos/métodos , Mitocondrias/genética , Oocitos/crecimiento & desarrollo , Animales , Células del Cúmulo/metabolismo , Desarrollo Embrionario/genética , Femenino , Humanos , Ratones , Mitocondrias/metabolismo , Recuperación del Oocito/métodos , Oocitos/metabolismo , Embarazo , Especies Reactivas de Oxígeno/metabolismo , Técnicas Reproductivas AsistidasRESUMEN
During follicle growth, DNA methylation is gradually established, which is important for oocyte developmental competence. Due to the facts that oocytes from prepubertal individuals show reduced developmental outcomes when compared to those from sexually mature individuals, and the fact that oocytes derived from in vitro follicle culture have much lower developmental competence, it is worth exploring whether prepubertal superovulation and in vitro follicle culture will cause changes in DNA methylation imprinting status in oocytes. In this study, we found that the CpG island in maternally imprinted GNAS clusters was hypermethylated in the MII-stage oocytes from sexually mature mice, but was hypomethylated in oocytes from prepuberty individuals. The GNAS clusters in the MII-stage oocytes obtained by in vitro follicle culture showed heterogeneous methylation levels, indicating different qualities of oocytes, however, three other maternally imprinted genes, Peg1, Lot1 and Impact, were all hypermethylated in the MII-stage oocytes derived from both prepubertal superovulation and in vitro follicle culture. Taken together, the findings suggest that the methylation status in GNAS clusters may potentially represent a novel epigenetic marker for oocyte quality detection.
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Islas de CpG , Metilación de ADN , Impresión Genómica , Oocitos/metabolismo , Factores de Edad , Animales , Biomarcadores , Células Cultivadas , Femenino , Ratones , Folículo Ovárico/citologíaRESUMEN
The genome methylation is globally erased in early fetal germ cells, and it is gradually re-established during gametogenesis. The expression of some imprinted genes is regulated by the methylation status of CpG islands, while the exact time of DNA methylation establishment near maternal imprinted genes during oocyte growth is not well known. Here, growing oocytes were divided into three groups based on follicle diameters including the S-group (60-100 µm), M-group (100-140 µm), and L-group (140-180 µm). The fully grown germinal vesicle (GV)-stage and metaphase II (M2)-stage mature oocytes were also collected. These oocytes were used for single-cell bisulfite sequencing to detect the methylation status of CpG islands near imprinted genes on chromosome 7. The results showed that the CpG islands near Ndn, Magel2, Mkrn3, Peg12, and Igf2 were completely unmethylated, but those of Peg3, Snrpn, and Kcnq1ot1 were hypermethylated in MII-stage oocytes. The methylation of CpG islands near different maternal imprinted genes occurred asynchronously, being completed in later-stage growing oocytes, fully grown GV oocytes, and mature MII-stage oocytes, respectively. These results show that CpG islands near some maternally imprinted genes are not necessarily methylated, and that the establishment of methylation of other maternally imprinted genes is completed at different stages of oocyte growth, providing a novel understanding of the establishment of maternally imprinted genes in oocytes.
RESUMEN
BRG1-associated factor 250a (BAF250a) is a component of the SWI/SNF adenosine triphosphate-dependent chromatin remodeling complex, which has been shown to control chromatin structure and transcription. BAF250a was reported to be a key component of the gene regulatory machinery in embryonic stem cells controlling self-renewal, differentiation, and cell lineage decisions. Here we constructed Baf250aF/F ;Gdf9-cre (Baf250aCKO ) mice to specifically delete BAF250a in oocytes to investigate the role of maternal BAF250a in female germ cells and embryo development. Our results showed that BAF250a deletion did not affect folliculogenesis, ovulation, and fertilization, but it caused late embryonic death. RNA sequencing analysis showed that the expression of genes involved in cell proliferation and differentiation, tissue morphogenesis, histone modification, and nucleosome remodeling were perturbed in Baf250aCKO MII oocytes. We showed that covalent histone modifications such as H3K27me3 and H3K27ac were also significantly affected in oocytes, which may reduce oocyte quality and lead to birth defects. In addition, the DNA methylation level of Igf2r, Snrpn, and Peg3 differentially methylated regions was decreased in Baf250aCKO oocytes. Quantitative real-time polymerase chain reaction analysis showed that the relative messenger RNA (mRNA) expression levels of Igf2r and Snrpn were significantly increased. The mRNA expression level of Dnmt1, Dnmt3a, Dnmt3l, and Uhrf1 was decreased, and the protein expression in these genes was also reduced, which might be the cause for impaired imprinting establishment. In conclusion, our results demonstrate that BAF250a plays an important role in oocyte transcription regulation, epigenetic modifications, and embryo development.
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Proteínas de Unión al ADN/genética , Desarrollo Embrionario/genética , Epigénesis Genética/genética , Oocitos/metabolismo , Factores de Transcripción/genética , Animales , Diferenciación Celular/genética , Linaje de la Célula/genética , Células Cultivadas , Metilación de ADN/genética , Células Madre Embrionarias/metabolismo , Células Madre Embrionarias/fisiología , Femenino , Eliminación de Gen , Impresión Genómica , Técnicas de Maduración In Vitro de los Oocitos , Ratones , Ratones Noqueados , Oocitos/fisiología , EmbarazoRESUMEN
N6-methyladenosine (m6A) is the most prevalent and reversible internal modification of mammalian messenger and noncoding RNAs mediated by specific m6A writer, reader, and eraser proteins. As an m6A writer, the methyltransferase-like 3-methyltransferase-like 14 (METTL14)-Wilms tumor 1-associated protein complex dynamically regulates m6A modification and plays important roles in diverse biologic processes. However, our knowledge about the complete functions of this RNA methyltransferase complex, the contributions of each component to the methylation, and their effects on different biologic pathways are still limited. By using both in vivo and in vitro models, we here report that METTL14 is indispensable for postimplantation embryonic development by facilitating the conversion from naive to primed state of the epiblast. Depletion of Mettl14 leads to conspicuous embryonic growth retardation from embryonic d 6.5, mainly as a result of resistance to differentiation, which further leads to embryonic lethality early in gestation. Our data highlight the critical function of METTL14 as an m6A modification regulator in orchestrating early mouse embryogenesis.-Meng, T.-G., Lu, X., Guo, L., Hou, G.-M., Ma, X.-S., Li, Q.-N., Huang, L., Fan, L.-H., Zhao, Z.-H., Ou, X.-H., OuYang, Y.-C., Schatten, H., Li, L., Wang, Z.-B., Sun, Q.-Y. Mettl14 is required for mouse postimplantation development by facilitating epiblast maturation.
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Desarrollo Embrionario/genética , Estratos Germinativos/citología , Metiltransferasas/fisiología , Adenosina/análogos & derivados , Adenosina/genética , Animales , Sistemas CRISPR-Cas , Femenino , Perfilación de la Expresión Génica , Genes Letales , Metiltransferasas/genética , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Células Madre Embrionarias de Ratones/citología , ARN Mensajero/genéticaRESUMEN
Mitochondrial DNA (mtDNA) is important for oxidative phosphorylation; dysfunctions can play a role in many mitochondrial diseases and can also affect the aging of cells and individuals. DNA methylation is an important epigenetic modification that plays a critical role in regulating gene expression. While recent studies have revealed the existence of mtDNA methylation there are still controversies about mtDNA methylation due to the special structure of mtDNA. Mitochondria and DNA methylation are both essential for regulating oocyte maturation and early embryo development, but whether mtDNA methylation changes during this process is unknown. By employing bisulfite sequencing, we found that in the process of mouse oocyte maturation, postovulatory oocyte aging, and early embryo development, all analyzed mitochondrial genes, including 16S-CpGI, DCR, ND6, 12S, and ATP8, lacked 5'mC. Thus, mtDNA methylation does not occur in the oocyte and early embryo.
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Metilación de ADN , ADN Mitocondrial/metabolismo , Desarrollo Embrionario , Oocitos/citología , Animales , Embrión de Mamíferos , Epigénesis Genética , Ratones , Oocitos/metabolismoRESUMEN
STUDY QUESTION: What is the impact of glucocorticoid (GC) on female reproduction? SUMMARY ANSWER: Corticosterone (CORT) exposure causes little damage to oocyte quality or developmental competence but has an adverse effect on the uterus, which causes decreased implantation, embryo death and subsequent infertility. WHAT IS KNOWN ALREADY: Chronic treatment with high GC doses is effective in controlling most allergic diseases but may lead to metabolic disorders such as obesity that are closely related with reproductive function. STUDY DESIGN, SIZE, DURATION: Hypercortisolism was induced in a female mouse model by supplementing the drinking water with 100 µg/ml of CORT. Controls received vehicle (1% v/v ethanol) only. After 4 weeks treatment mice were either mated or killed in estrus for hormone and organ measurements. In the first experiment, treatment with CORT or control continued during pregnancy but in the second CORT treatment was stopped after mating. To identify the effects of GC exposure on the uterus, blastocysts were generated by IVF of oocytes from CORT and control mice and replaced into recipients receiving the opposite treatment. PARTICIPANTS/MATERIALS, SETTING, METHODS: The effects of hypercortisolism on female mice were first characterized by living body fat content, body weight, food intake, hormone and biochemical measurements, a glucose tolerance test and an insulin resistance test. Fertility was determined with or without CORT-treatment during pregnancy. Oocyte quality was assessed by oocyte maturation, mitochondrial distribution, reactive oxygen species production, mitochondrial DNA mutations and morphology of blastocysts produced in vivo or in vitro. Blastocyst cross-transfer was done to evaluate the causes of embryonic development failure. Fetus development and uterus morphology evaluation as well as culture of oocytes in vitro with gradient concentrations of CORT were also carried out. MAIN RESULTS AND THE ROLE OF CHANCE: In the hypercortisolism female mouse model, body weight and food intake were much higher than in the control, and corticosterone, estradiol, cholesterol (CHO) and triglycerides (TG) in the plasma of CORT-treated mice was significantly increased. The hypercortisolism female mice were infertile when CORT-treatment was sustained during pregnancy but fertile if CORT-treatment was stopped after mating. The rate of successful implantation in hypercortisolism mice with sustained CORT-treatment during pregnancy was significantly lower than in the control, and the implanted embryos could not develop beyond 13.5 dpc. Blastocyst cross-transfer showed that blastocysts from CORT-treated mice could develop to term in the uterus of control mice, but blastocysts from control mice failed to develop to term when they were transferred into CORT-treated mice, providing evidence that the infertility was mainly caused by an altered uterine environment. CORT administration did not affect oocyte maturation, mitochondrial distribution, ROS production and blastocyst morphology, but increased mitochondrial DNA mutations. Culture of oocytes in vitro with gradient concentrations of CORT showed that only very high concentrations of CORT caused damage to oocyte developmental competence. LARGE SCALE DATA: NA. LIMITATIONS, REASONS FOR CAUTION: The mouse model has the advantages of a consistent genetic and physiological background and openness to experimental manipulation over clinical studies but may not represent the human situation. WIDER IMPLICATIONS OF THE FINDINGS: Our findings show that special care should be taken when administering CORT during pregnancy, and provide important information concerning female reproduction when treating patients by subjecting them to chronic GC exposure. STUDY FUNDING/COMPETING INTEREST(S): This study was supported by the National Key R&D Program of China (Nos. 2016YFA0100400 and 2017YFC1000600) and the National Natural Science Foundation of China (31472055). The authors have no conflicts of interest.
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Corticosterona/farmacología , Síndrome de Cushing/metabolismo , Glucocorticoides/farmacología , Infertilidad Femenina/metabolismo , Oocitos/efectos de los fármacos , Útero/efectos de los fármacos , Animales , Peso Corporal/efectos de los fármacos , Colesterol/sangre , Síndrome de Cushing/inducido químicamente , Ingestión de Alimentos/efectos de los fármacos , Femenino , Infertilidad Femenina/inducido químicamente , Ratones , Oocitos/metabolismo , Triglicéridos/sangre , Útero/metabolismoRESUMEN
Polycystic ovary syndrome (PCOS), a familial aggregation disease that causes anovulation in women, has well-recognised characteristics, two of which are hyperinsulinaemia and hyperandrogenaemia. To determine whether the DNA methylation status is altered in oocytes by high insulin and androgen levels, we generated a mouse model with hyperinsulinaemia and hyperandrogenaemia by injection of insulin and human chorionic gonadotrophin and investigated DNA methylation changes through single-cell level whole genome bisulphite sequencing. Our results showed that hyperinsulinaemia and hyperandrogenaemia had no significant effects on the global DNA methylation profile and different functional regions of genes, but did alter methylation status of some genes, which were significantly enriched in 17 gene ontology (GO) terms (P<0.05) by GO analysis. Among differently methylated genes, some were related to the occurrence of PCOS. Based on our results, we suggest that hyperinsulinaemia and hyperandrogenaemia may cause changes in some DNA methylation loci in oocytes.
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Metilación de ADN , Hiperandrogenismo/genética , Hiperinsulinismo/genética , Oocitos/metabolismo , Animales , Modelos Animales de Enfermedad , Femenino , Hiperandrogenismo/metabolismo , Hiperinsulinismo/metabolismo , RatonesRESUMEN
Environmental factors such as diet and lifestyle can influence the health of both mothers and offspring. However, its transgenerational transmission and underlying mechanisms remain largely unknown. Here, using a maternal lactation-period low-protein diet (LPD) mouse model, we show that maternal LPD during lactation causes decreased survival and stunted growth, significantly reduces ovulation and litter size, and alters the gut microbiome in the female LPD-F1 offspring. The transcriptome of LPD-F1 metaphase II (MII) oocytes shows that differentially expressed genes are enriched in female pregnancy and multiple metabolic processes. Moreover, maternal LPD causes early stunted growth and impairs metabolic health, which is transmitted over two generations. The methylome alteration of LPD-F1 oocytes can be partly transmitted to the F2 oocytes. Together, our results reveal that LPD during lactation transgenerationally affects offspring health, probably via oocyte epigenetic changes.
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Dieta con Restricción de Proteínas , Lactancia , Animales , Femenino , Lactancia/genética , Dieta con Restricción de Proteínas/efectos adversos , Ratones , Embarazo , Oocitos/metabolismo , Microbioma Gastrointestinal , Epigénesis Genética , Fenómenos Fisiologicos Nutricionales Maternos , Transcriptoma/genética , Masculino , Metilación de ADN , Efectos Tardíos de la Exposición Prenatal/genéticaRESUMEN
Heavy drinking in women is known to adversely affect pregnancy and fertility. However, pregnancy is a complex process, and the adverse effects of ethanol on pregnancy does not mean that ethanol will have adverse effects on all stages from gamete to fetal formation. Similarly, the adverse effects of ethanol before and after adolescence cannot be generalized. To focus on the effects of prepubertal ethanol on female reproductive ability, we established a mouse model of prepubertal ethanol exposure by changing drinking water to 20% v/v ethanol. Some routine detections were performed on the model mice, and details such as mating, fertility, reproductive organ and fetal weights were recorded day by day after discontinuation of ethanol exposure. Prepubertal ethanol exposure resulted in decreased ovarian weight and significantly reduced oocyte maturation and ovulation after sexual maturation, however, normal morphology oocytes with discharged polar body showed normal chromosomes and spindle morphology. Strikingly, oocytes with normal morphology from ethanol exposed mice showed reduced fertilization rate, but once fertilized they had the ability to develop to blastocysts. RNA-seq analysis showed that the gene expression of the ethanol exposed oocytes with normal morphology had been altered. These results show the adverse effects of prepubertal alcohol exposure on adult female reproductive health.
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Etanol , Reproducción , Embarazo , Femenino , Ratones , Animales , Etanol/toxicidad , Oocitos , Fertilidad , Células GerminativasRESUMEN
Alternative splicing expands the transcriptome and proteome complexity and plays essential roles in tissue development and human diseases. However, how alternative splicing regulates spermatogenesis remains largely unknown. Here, using a germ cell-specific knockout mouse model, we demonstrated that the splicing factor Srsf10 is essential for spermatogenesis and male fertility. In the absence of SRSF10, spermatogonial stem cells can be formed, but the expansion of Promyelocytic Leukemia Zinc Finger (PLZF)-positive undifferentiated progenitors was impaired, followed by the failure of spermatogonia differentiation (marked by KIT expression) and meiosis initiation. This was further evidenced by the decreased expression of progenitor cell markers in bulk RNA-seq, and much less progenitor and differentiating spermatogonia in single-cell RNA-seq data. Notably, SRSF10 directly binds thousands of genes in isolated THY+ spermatogonia, and Srsf10 depletion disturbed the alternative splicing of genes that are preferentially associated with germ cell development, cell cycle, and chromosome segregation, including Nasp, Bclaf1, Rif1, Dazl, Kit, Ret, and Sycp1. These data suggest that SRSF10 is critical for the expansion of undifferentiated progenitors by regulating alternative splicing, expanding our understanding of the mechanism underlying spermatogenesis.
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Empalme Alternativo , Espermatogonias , Ratones , Animales , Masculino , Humanos , Espermatogénesis/genética , Diferenciación Celular/genética , Meiosis , Ratones Noqueados , Factores de Empalme Serina-Arginina/genética , Factores de Empalme Serina-Arginina/metabolismo , Proteínas Represoras/metabolismo , Proteínas de Ciclo Celular/metabolismoRESUMEN
During oocyte growth, various epigenetic modifications are gradually established, accompanied by accumulation of large amounts of mRNAs and proteins. However, little is known about the relationship between epigenetic modifications and meiotic progression. Here, by using Gdf9-Cre to achieve oocyte-specific ablation of Ehmt2 (Euchromatic-Histone-Lysine-Methyltransferase 2) from the primordial follicle stage, we found that female mutant mice were infertile. Oocyte-specific knockout of Ehmt2 caused failure of homologous chromosome separation independent of persistently activated SAC during the first meiosis. Further studies revealed that lacking maternal Ehmt2 affected the transcriptional level of Ccnb3, while microinjection of exogenous Ccnb3 mRNA could partly rescue the failure of homologous chromosome segregation. Of particular importance was that EHMT2 regulated ccnb3 transcriptions by regulating CTCF binding near ccnb3 gene body in genome in oocytes. In addition, the mRNA level of Ccnb3 significantly decreased in the follicles microinjected with Ctcf siRNA. Therefore, our findings highlight the novel function of maternal EHMT2 on the metaphase I-to-anaphase I transition in mouse oocytes: regulating the transcription of Ccnb3.
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Segregación Cromosómica , Meiosis , Anafase , Animales , Femenino , Meiosis/genética , Ratones , Oocitos/metabolismo , ARN Mensajero/metabolismoRESUMEN
AIMS: Research evidence indicates that epigenetic modifications of gametes in obese or diabetic parents may contribute to metabolic disorders in offspring. In the present study, we sought to address the effect of diabetic uterine environment on the offspring metabolism. METHODS: Type 2 diabetes mouse model was induced by high-fat diet combined with streptozotocin (STZ) administration. We maintained other effect factors constant and changed uterine environment by zygote transfers, and then determined and compared the offspring numbers, symptoms, body weight trajectories, and metabolism indices from different groups. RESULT: We found that maternal type 2 diabetes mice had lower fertility and a higher dystocia rate, accompanying the increased risk of offspring malformations and death. Compared to only a pre-gestational exposure to hyperglycemia, exposure to hyperglycemia both pre- and during pregnancy resulted in offspring growth restriction and impaired metabolism in adulthood. But there was no significant difference between a pre-gestational exposure group and a no exposure group. The deleterious effects, no matter bodyweight or glucose tolerance, could be rescued by transferring the embryos from diabetic mothers into normal uterine environment. CONCLUSION: Our data demonstrate that uterine environment of maternal diabetes makes critical impact on the offspring health.
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Diabetes affects oocyte nuclear and cytoplasmic quality. In this study, we generated a type 1 diabetes (T1D) mouse model by STZ injection to study the effects of T1D on zona pellucida and genomic DNA methylation of oocytes and granulosa cells. T1D mice showed fewer ovulated oocytes, reduced ovarian reserve, disrupted estrus cycle, and significantly ruptured zona pellucida in 2-cell in vivo embryos compared to controls. Notably, diabetic oocytes displayed thinner zona pellucida and treatment of oocytes with high concentration glucose reduced the zona pellucida thickness. Differential methylation genes in oocytes and granulosa cells were analyzed by methylation sequencing. These genes were significantly enriched in GO terms by GO analysis, and these GO terms were involved in multiple aspects of growth and development. Most notably, the abnormal methylation genes in oocytes may be related to oocyte zona pellucida changes in diabetic mice. These findings provide novel basic data for further understanding and elucidating dysgenesis and epigenetic changes in type 1 diabetes mellitus.
Asunto(s)
Metilación de ADN , Diabetes Mellitus Experimental/genética , Diabetes Mellitus Tipo 1/genética , Células de la Granulosa/química , Oocitos/química , Zona Pelúcida/metabolismo , Animales , Estudios de Casos y Controles , Diabetes Mellitus Experimental/metabolismo , Diabetes Mellitus Tipo 1/inducido químicamente , Diabetes Mellitus Tipo 1/metabolismo , Epigénesis Genética , Ciclo Estral , Femenino , Redes Reguladoras de Genes , Ratones , Análisis de Secuencia de ADN , EstreptozocinaRESUMEN
OBJECTIVE: To observe the effects of core strength training on the body balance of middle-aged men, which could provide evidence for improving the body balance and reducing the risk of falls in the middle-aged men. METHODS: Sixteen 50~60 years old men were randomly divided into experimental group (core strength training, n=8) and control group(n=8). Intermittent core strength trainings were used for dynamic and static training, push-pull training and unarmed lower limb strength training by suspension ropes and yoga mat. The time for training was 50~60 min/day, 4~5 times/week for a total of 10 weeks. The men in control group remained their original living habits. We measured the ability of body static balance and dynamic balance before and after the core strength training. RESULTS: â Compared with before exercise training, both closed single foot standing time and the body dynamic balance were increased significantly in the experimental group after exercise training(P<0.01)(10.63±1.69 s vs 9.00±2.27 s; 77.38±10.94 vs 89.50±5.53). â¡ Compared with before exercise training, star excursion balance test(SEBT) values were significantly increased in the left leg(the right leg support) in eight directions and the right leg(the left leg support)in six directions (P<0.01). CONCLUSIONS: Ten-week core strength training can significantly increase the body static and dynamic balance in the middle-aged men.
Asunto(s)
Equilibrio Postural , Entrenamiento de Fuerza , Accidentes por Caídas/prevención & control , Humanos , Extremidad Inferior , Masculino , Persona de Mediana Edad , Fuerza MuscularRESUMEN
Based on environmental monitoring data, meteorological data and the results of numerical simulation, a typical air pollution process in Nanjing, from 26th May to 1st June, 2014 was deeply analyzed combining aerosol extinction coefficient derived from LiDAR system. Experimental results showed that the entire pollution process was affected by both local pollution and exogenous inputs including dust and smoke. Meteorological factors played a significant role in the generation and elimination of pollutants. Low pressure and temperature inversion also hindered the diffusion of pollutants, while strong rainfall terminated the pollution process. During the pollution, the height of atmospheric boundary layer was lower than normal situation and changed little during the pollution period, which provided a poor diffusion condition for pollutants. LiDAR could accurately detect aerosol vertical structure which was able to capture the temporal and spatial variation of pollutant distributions. Therefore, LiDAR can be of great significance for the atmospheric pollution monitoring.