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1.
PLoS One ; 15(9): e0238948, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32915925

RESUMO

Several equids have gone extinct and many extant equids are currently considered vulnerable to critically endangered. This work aimed to evaluate whether domestic horse oocytes support preimplantation development of zebra embryos obtained by intracytoplasmic sperm injection (ICSI, zebroid) and cloning, and to study the Hippo signaling pathway during the lineage specification of trophectoderm cells and inner cell mass cells. We first showed that zebra and horse sperm cells induce porcine oocyte activation and recruit maternal SMARCA4 during pronuclear formation. SMARCA4 recruitment showed to be independent of the genetic background of the injected sperm. No differences were found in blastocyst rate of ICSI hybrid (zebra spermatozoon into horse egg) embryos relative to the homospecific horse control group. Interestingly, zebra cloned blastocyst rate was significantly higher at day 8. Moreover, most ICSI and cloned horse and zebra blastocysts showed a similar expression pattern of SOX2 and nuclear YAP1 with the majority of the nuclei positive for YAP1, and most SOX2+ nuclei negative for YAP1. Here we demonstrated that horse oocytes support zebra preimplantation development of both, ICSI and cloned embryos, without compromising development to blastocyst, blastocyst cell number neither the expression of SOX2 and YAP1. Our results support the use of domestic horse oocytes as a model to study in vitro zebra embryos on behalf of preservation of valuable genetic.


Assuntos
Desenvolvimento Embrionário , Equidae/embriologia , Equidae/genética , Cavalos/fisiologia , Oócitos/fisiologia , Proteínas Adaptadoras de Transdução de Sinal/genética , Animais , Núcleo Celular/fisiologia , Clonagem de Organismos/veterinária , Citoplasma/fisiologia , Técnicas de Cultura Embrionária/veterinária , Desenvolvimento Embrionário/genética , Desenvolvimento Embrionário/fisiologia , Espécies em Perigo de Extinção , Equidae/metabolismo , Feminino , Perfilação da Expressão Gênica , Cavalos/genética , Técnicas In Vitro , Masculino , Técnicas de Transferência Nuclear/veterinária , Fatores de Transcrição SOXB1/genética , Injeções de Esperma Intracitoplásmicas/veterinária , Sus scrofa
2.
PLoS Genet ; 16(8): e1008644, 2020 08.
Artigo em Inglês | MEDLINE | ID: mdl-32776941

RESUMO

Correct regulation of cell contractility is critical for the function of many biological systems. The reproductive system of the hermaphroditic nematode C. elegans contains a contractile tube of myoepithelial cells known as the spermatheca, which stores sperm and is the site of oocyte fertilization. Regulated contraction of the spermatheca pushes the embryo into the uterus. Cell contractility in the spermatheca is dependent on actin and myosin and is regulated, in part, by Ca2+ signaling through the phospholipase PLC-1, which mediates Ca2+ release from the endoplasmic reticulum. Here, we describe a novel role for GSA-1/Gαs, and protein kinase A, composed of the catalytic subunit KIN-1/PKA-C and the regulatory subunit KIN-2/PKA-R, in the regulation of Ca2+ release and contractility in the C. elegans spermatheca. Without GSA-1/Gαs or KIN-1/PKA-C, Ca2+ is not released, and oocytes become trapped in the spermatheca. Conversely, when PKA is activated through either a gain of function allele in GSA-1 (GSA-1(GF)) or by depletion of KIN-2/PKA-R, the transit times and total numbers, although not frequencies, of Ca2+ pulses are increased, and Ca2+ propagates across the spermatheca even in the absence of oocyte entry. In the spermathecal-uterine valve, loss of GSA-1/Gαs or KIN-1/PKA-C results in sustained, high levels of Ca2+ and a loss of coordination between the spermathecal bag and sp-ut valve. Additionally, we show that depleting phosphodiesterase PDE-6 levels alters contractility and Ca2+ dynamics in the spermatheca, and that the GPB-1 and GPB-2 Gß subunits play a central role in regulating spermathecal contractility and Ca2+ signaling. This work identifies a signaling network in which Ca2+ and cAMP pathways work together to coordinate spermathecal contractions for successful ovulations.


Assuntos
Proteínas de Caenorhabditis elegans/metabolismo , Sinalização do Cálcio , Subunidades Catalíticas da Proteína Quinase Dependente de AMP Cíclico/metabolismo , Contração Muscular , 3',5'-AMP Cíclico Fosfodiesterases/metabolismo , Animais , Caenorhabditis elegans , Proteínas de Caenorhabditis elegans/genética , Subunidades Catalíticas da Proteína Quinase Dependente de AMP Cíclico/genética , Células Epiteliais/metabolismo , Células Epiteliais/fisiologia , Subunidades beta da Proteína de Ligação ao GTP/metabolismo , Mutação com Ganho de Função , Células Musculares/metabolismo , Células Musculares/fisiologia , Oócitos/fisiologia
3.
Mutat Res ; 785: 108320, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32800274

RESUMO

It is well established that maternal age is associated with a rapid decline in the production of healthy and high-quality oocytes resulting in reduced fertility in women older than 35 years of age. In particular, chromosome segregation errors during meiotic divisions are increasingly common and lead to the production of oocytes with an incorrect number of chromosomes, a condition known as aneuploidy. When an aneuploid oocyte is fertilized by a sperm it gives rise to an aneuploid embryo that, except in rare situations, will result in a spontaneous abortion. As females advance in age, they are at higher risk of infertility, miscarriage, or having a pregnancy affected by congenital birth defects such as Down syndrome (trisomy 21), Edwards syndrome (trisomy 18), and Turner syndrome (monosomy X). Here, we review the potential molecular mechanisms associated with increased chromosome segregation errors during meiosis as a function of maternal age. Our review shows that multiple exogenous and endogenous factors contribute to the age-related increase in oocyte aneuploidy. Specifically, the weight of evidence indicates that recombination failure, cohesin deterioration, spindle assembly checkpoint (SAC) disregulation, abnormalities in post-translational modification of histones and tubulin, and mitochondrial dysfunction are the leading causes of oocyte aneuploidy associated with maternal aging. There is also growing evidence that dietary and other bioactive interventions may mitigate the effect of maternal aging on oocyte quality and oocyte aneuploidy, thereby improving fertility outcomes. Maternal age is a major concern for aneuploidy and genetic disorders in the offspring in the context of an increasing proportion of mothers having children at increasingly older ages. A better understanding of the mechanisms associated with maternal aging leading to aneuploidy and of intervention strategies that may mitigate these detrimental effects and reduce its occurrence are essential for preventing abnormal reproductive outcomes in the human population.


Assuntos
Aneuploidia , Proteínas de Ciclo Celular/genética , Proteínas Cromossômicas não Histona/genética , Segregação de Cromossomos/genética , Anormalidades Congênitas/genética , Idade Materna , Anormalidades Congênitas/prevenção & controle , Feminino , Humanos , Pontos de Checagem da Fase M do Ciclo Celular/genética , Meiose/genética , Mitocôndrias/fisiologia , Oócitos/fisiologia
4.
J Vis Exp ; (161)2020 07 08.
Artigo em Inglês | MEDLINE | ID: mdl-32716390

RESUMO

The limited reserve of mature, fertilizable oocytes represents a major barrier for the success of assisted reproduction in mammals. Considering that during the reproductive life span only about 1% of the oocytes in an ovary mature and ovulate, several techniques have been developed to increase the exploitation of the ovarian reserve to the growing population of non-ovulatory follicles. Such technologies have allowed interventions of fertility preservation, selection programs in livestock, and conservation of endangered species. However, the vast potential of the ovarian reserve is still largely unexploited. In cows, for instance, some attempts have been made to support in vitro culture of oocytes at specific developmental stages, but efficient and reliable protocols have not yet been developed. Here we describe a culture system that reproduce the physiological conditions of the corresponding follicular stage, defined to develop in vitro growing oocytes collected from bovine early antral follicles to the fully-grown stage, corresponding to the medium antral follicle in vivo. A combination of hormones and a phosphodiesterase 3 inhibitor was used to prevent untimely meiotic resumption and to guide oocyte's differentiation.


Assuntos
Técnicas de Cultura de Células/métodos , Oócitos/fisiologia , Reserva Ovariana/fisiologia , Reprodução/fisiologia , Animais , Bovinos , Ciclo Celular/fisiologia , Diferenciação Celular/fisiologia , Feminino , Oogênese/fisiologia , Folículo Ovariano/fisiologia , Ovário/citologia , Ovário/fisiologia
5.
J Vis Exp ; (160)2020 06 24.
Artigo em Inglês | MEDLINE | ID: mdl-32658178

RESUMO

In wild animals' conservation programs, gamete banking is crucial to safeguard genetic resources of valuable individuals and rare species and to promote biodiversity preservation. In felids, most species are threatened with extinction, and domestic breeds are used as a model to increase the efficiency of protocols for germplasm banking. Among oocyte cryopreservation techniques, vitrification is more and more popular in human and veterinary assisted reproduction. Cryotop vitrification, which was at first developed for human oocytes and embryos, has demonstrated to be well-suited for cat oocytes. This method offers several advantages, such as the feasibility in field conditions and the speed of the procedure, which can be helpful when several samples need to be processed. However, the efficiency is strongly dependent on the operator's skills, and intra- and inter-laboratory standardization are needed, as well as personnel training. This protocol describes minimum volume vitrification of immature feline oocytes on a commercial support in a step by step field-friendly protocol, from oocyte collection to warming. Following the protocol, preservation of oocyte integrity and viability at warming (as high as 90%) can be expected, although there is still room for improvement in post-warming maturation and embryonic development outcomes.


Assuntos
Oócitos/fisiologia , Manejo de Espécimes/métodos , Animais , Gatos , Criopreservação , Desenvolvimento Embrionário , Feminino , Oócitos/citologia , Gravidez , Técnicas de Reprodução Assistida , Sobrevivência de Tecidos
6.
Gynecol Obstet Invest ; 85(3): 290-294, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32485714

RESUMO

AIM: This study evaluated the competency of oocytes/embryos derived from follicles >15 mm in diameter from obese patients, compared with nonobese patients. PATIENTS AND METHODS: A cohort study was conducted in a single tertiary medical center between July 2018 and May 2019. Before ultrasound-guided follicular aspiration, follicles were measured and those with maximal dimensional size >15 mm were tracked. Microscopic examination of the follicular aspirates was performed by an embryologist. Each follicle aspirated was evaluated for oocyte maturation, oocyte fertilization, and embryo quality. RESULTS: 457 follicles were measured: 380 (83.2%) in nonobese and 77 (16.8%) in obese patients. No in-between group differences were observed in the causes of infertility, patients' demographics, or ovarian stimulation characteristics. Oocytes were achieved during aspiration from 277 (72.8%) and 54 (70.0%) of the nonobese and obese groups, respectively (p = 0.67). No in-between group differences were observed in fertilization (2PN/oocyte), top quality embryo (TQE) per zygote (2PN), and TQE per follicle. CONCLUSION: Oocyte recovery rate from follicles >15 mm is unrelated to patients' BMI. Moreover, the oocytes recovered from obese patients are competent yielding comparable zygote and TQE per follicle/oocyte, compared with nonobese patients. Further investigation is required to strengthen this finding.


Assuntos
Infertilidade/fisiopatologia , Obesidade/fisiopatologia , Recuperação de Oócitos/estatística & dados numéricos , Oócitos/fisiologia , Folículo Ovariano/fisiologia , Adulto , Índice de Massa Corporal , Estudos de Coortes , Embrião de Mamíferos , Feminino , Humanos , Infertilidade/etiologia , Obesidade/complicações , Recuperação de Oócitos/métodos , Indução da Ovulação
7.
PLoS One ; 15(6): e0235140, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32574203

RESUMO

BACKGROUND: Due to improved treatment, there is an increasing focus on the reproductive potential of survivors of childhood cancer. Cytotoxic chemotherapy accelerates the decline in the number of primordial follicles within the mammalian ovary at all ages, but effects on the developmental potential of remaining oocytes following prepubertal cancer treatment are unclear. OBJECTIVES: To investigate whether cyclophosphamide (CY) exposure in the prepubertal period in female mice influences ovarian function and the functional competence of oocytes in adulthood. METHODS: This study used Swiss albino mice as the experimental model. Female mice were treated with 200 mg/kg CY on either postnatal day 14 (CY14), 21 (CY21) or 28 (CY28) i.e at a prepubertal and 2 young postpubertal ages. At 14 weeks of life, ovarian function, functional competence of oocytes, and embryo quality were assessed. RESULTS: The number of primordial follicles decreased significantly in CY14 and CY21 groups compared to control (p < 0.01). The number of oocytes from superovulated was 8.5 ± 1.4, 24.1 ± 2.9 and 26.8 ± 2.1 in CY14, CY21 and CY28 respectively which was significantly lower than control (50.2 ± 3.2; p < 0.001). In vitro culture of CY14 embryos demonstrated only 55.4% blastocyst formation (p < 0.0001) and reduced ability of inner cell mass (ICM) to proliferate in vitro (p < 0.05) at 120 and 216 h post insemination respectively. On the other hand, ICM proliferation was unaltered in 2 young postpubertal ages. CONCLUSION: Our results indicate long-term effects on the developmental competence of oocytes exposed to CY in early but not adult life. These data provide a mechanism whereby long-term fertility can be impaired after chemotherapy exposure, despite the continuing presence of follicles within the ovary, and support the need for fertility preservation in prepubertal girls before alkylating agent exposure.


Assuntos
Blastocisto/efeitos dos fármacos , Ciclofosfamida/farmacologia , Desenvolvimento Embrionário/efeitos dos fármacos , Reserva Ovariana/efeitos dos fármacos , Maturidade Sexual/fisiologia , Animais , Hormônio Antimülleriano/sangue , Antineoplásicos Alquilantes/farmacologia , Blastocisto/citologia , Blastocisto/fisiologia , Peso Corporal/efeitos dos fármacos , Peso Corporal/fisiologia , Proliferação de Células/efeitos dos fármacos , Ciclofosfamida/administração & dosagem , Embrião de Mamíferos/efeitos dos fármacos , Embrião de Mamíferos/embriologia , Desenvolvimento Embrionário/fisiologia , Feminino , Fertilidade/efeitos dos fármacos , Fertilidade/fisiologia , Camundongos , Oócitos/efeitos dos fármacos , Oócitos/fisiologia , Tamanho do Órgão/efeitos dos fármacos , Reserva Ovariana/fisiologia , Ovário/anatomia & histologia , Ovário/citologia , Ovário/efeitos dos fármacos , Fatores de Tempo
8.
Anim Sci J ; 91(1): e13385, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32515535

RESUMO

Both oocytes and extracellular vesicles (EV) have emerged as critical regulators of mammalian follicular development; however, the possible interaction between the oocyte-derived paracrine factor (ODPF) and EV signals has never been examined. Therefore, to explore the possibility of an interaction between oocyte and EV signals, the effects of ODPFs on the biogenesis of EVs as well as the expression levels of transcripts related to EV biogenesis in mural granulosa cells (MGCs) were examined using mice. The results showed that, while oocyte coculture has some effects on the expression levels of transcripts related to EV biogenesis, the number of EV particles present in the conditioned medium were not significantly different between ODPF-treated and non-treated MGCs. Therefore, oocytes have no effects on the EV biogenesis by MGCs, at least with respect to the numbers of EV particles.


Assuntos
Vesículas Extracelulares/metabolismo , Vesículas Extracelulares/fisiologia , Células da Granulosa/citologia , Células da Granulosa/metabolismo , Peptídeos e Proteínas de Sinalização Intercelular/fisiologia , Oócitos/fisiologia , Comunicação Parácrina/fisiologia , Animais , Células Cultivadas , Feminino , Camundongos Endogâmicos C57BL , Biogênese de Organelas
9.
J Vet Sci ; 21(3): e48, 2020 May.
Artigo em Inglês | MEDLINE | ID: mdl-32476321

RESUMO

BACKGROUND: Mature oocytes at the metaphase II status (MII-stage oocytes) played an important role in assisted reproductive technology in non-human primates. OBJECTIVES: In order to improve the proportion of MII-stage oocytes retrieval, three different superovulation protocols were performed on 24 female cynomolgus monkeys. METHODS: All the monkeys received once-daily injection of follicle-stimulating hormone (25 international unit [IU]) on day 3 of the menstruation, 3-day intervals, twice daily for 8-12 days until the time of human chorionic gonadotropin (1,500 IU) injection, on the 14-17th day of menstruation collecting oocytes. The difference between protocol I and protocol II was that 0.1 mg the gonadotropin-releasing hormone agonist was injected on day 1 of the menstruation, while the difference between personalized superovulation protocol and protocol II was that oocytes could be collected on the 14-17th day of menstrual cycle according to the length of each monkey. RESULTS: The total number of oocytes harvested using the personalized superovulation protocol was much higher than that using protocol I (p < 0.05), and the proportion of MII-stage oocytes was significantly greater than that from either superovulation protocol I or II (p < 0.001 and p < 0.01 respectively), while the proportion of immature oocytes at the germinal vesicle was less than that from superovulation protocol I (p < 0.05). CONCLUSIONS: The personalized superovulation protocol could increase the rate of MII-stage oocytes acquired, and successfully develop into embryos after intracytoplasmic sperm injection, and eventually generated fetus.


Assuntos
Gonadotropina Coriônica/administração & dosagem , Feto/fisiologia , Macaca fascicularis/fisiologia , Oócitos/fisiologia , Injeções de Esperma Intracitoplásmicas/veterinária , Superovulação/fisiologia , Animais , Gonadotropina Coriônica/farmacologia , Feminino , Feto/embriologia
10.
Proc Natl Acad Sci U S A ; 117(19): 10455-10464, 2020 05 12.
Artigo em Inglês | MEDLINE | ID: mdl-32350135

RESUMO

Aneuploidy is the leading contributor to pregnancy loss, congenital anomalies, and in vitro fertilization (IVF) failure in humans. Although most aneuploid conceptions are thought to originate from meiotic division errors in the female germline, quantitative studies that link the observed phenotypes to underlying error mechanisms are lacking. In this study, we developed a mathematical modeling framework to quantify the contribution of different mechanisms of erroneous chromosome segregation to the production of aneuploid eggs. Our model considers the probabilities of all possible chromosome gain/loss outcomes that arise from meiotic errors, such as nondisjunction (NDJ) in meiosis I and meiosis II, and premature separation of sister chromatids (PSSC) and reverse segregation (RS) in meiosis I. To understand the contributions of different meiotic errors, we fit our model to aneuploidy data from 11,157 blastocyst-stage embryos. Our best-fitting model captures several known features of female meiosis, for instance, the maternal age effect on PSSC. More importantly, our model reveals previously undescribed patterns, including an increased frequency of meiosis II errors among eggs affected by errors in meiosis I. This observation suggests that the occurrence of NDJ in meiosis II is associated with the ploidy status of an egg. We further demonstrate that the model can be used to identify IVF patients who produce an extreme number of aneuploid embryos. The dynamic nature of our mathematical model makes it a powerful tool both for understanding the relative contributions of mechanisms of chromosome missegregation in human female meiosis and for predicting the outcomes of assisted reproduction.


Assuntos
Aneuploidia , Oócitos/metabolismo , Blastocisto , Deleção Cromossômica , Segregação de Cromossomos , Feminino , Fertilização In Vitro , Humanos , Cariótipo , Idade Materna , Meiose/fisiologia , Modelos Teóricos , Não Disjunção Genética/genética , Não Disjunção Genética/fisiologia , Oócitos/fisiologia , Diagnóstico Pré-Implantação
11.
Proc Natl Acad Sci U S A ; 117(21): 11513-11522, 2020 05 26.
Artigo em Inglês | MEDLINE | ID: mdl-32381741

RESUMO

Female fertility and offspring health are critically dependent on an adequate supply of high-quality oocytes, the majority of which are maintained in the ovaries in a unique state of meiotic prophase arrest. While mechanisms of DNA repair during meiotic recombination are well characterized, the same is not true for prophase-arrested oocytes. Here we show that prophase-arrested oocytes rapidly respond to γ-irradiation-induced DNA double-strand breaks by activating Ataxia Telangiectasia Mutated, phosphorylating histone H2AX, and localizing RAD51 to the sites of DNA damage. Despite mobilizing the DNA repair response, even very low levels of DNA damage result in the apoptosis of prophase-arrested oocytes. However, we show that, when apoptosis is inhibited, severe DNA damage is corrected via homologous recombination repair. The repair is sufficient to support fertility and maintain health and genetic fidelity in offspring. Thus, despite the preferential induction of apoptosis following exogenously induced genotoxic stress, prophase-arrested oocytes are highly capable of functionally efficient DNA repair. These data implicate DNA repair as a key quality control mechanism in the female germ line and a critical determinant of fertility and genetic integrity.


Assuntos
Quebras de DNA de Cadeia Dupla , Reparo do DNA/fisiologia , Fertilidade/fisiologia , Oócitos/fisiologia , Animais , Apoptose/fisiologia , Feminino , Masculino , Camundongos Endogâmicos C57BL , Prófase/fisiologia
12.
Nat Struct Mol Biol ; 27(6): 533-539, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32451489

RESUMO

The Na+/I- symporter (NIS), the plasma membrane protein that actively transports I- (stoichiometry 2Na+:1I-) in thyroid physiology and radioiodide-based thyroid cancer treatment, also transports the environmental pollutant perchlorate (stoichiometry 1Na+:1ClO4-), which competes with I- for transport. Until now, the mechanism by which NIS transports different anion substrates with different stoichiometries has remained unelucidated. We carried out transport measurements and analyzed these using a statistical thermodynamics-based equation and electrophysiological experiments to show that the different stoichiometry of ClO4- transport is due to ClO4- binding to a high-affinity non-transport allosteric site that prevents Na+ from binding to one of its two sites. Furthermore, low concentrations of ClO4- inhibit I- transport not only by competition but also, critically, by changing the stoichiometry of I- transport to 1:1, which greatly reduces the driving force. The data reveal that ClO4- pollution in drinking water is more dangerous than previously thought.


Assuntos
Percloratos/metabolismo , Simportadores/química , Simportadores/metabolismo , Regulação Alostérica , Sítio Alostérico , Animais , Ânions/química , Ânions/metabolismo , Sítios de Ligação , Transporte Biológico , Cães , Eletrofisiologia/métodos , Feminino , Humanos , Iodo/metabolismo , Células Madin Darby de Rim Canino , Mutação , Oócitos/metabolismo , Oócitos/fisiologia , Percloratos/química , Ratos , Sódio/metabolismo , Simportadores/genética , Termodinâmica , Xenopus laevis
13.
Gynecol Obstet Invest ; 85(3): 252-258, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32268326

RESUMO

BACKGROUND/OBJECTIVES: Mechanical micro-vibration remains insufficient for improving embryo culture conditions in human immature oocytes. This study compared the clinical outcomes and embryo development between germinal vesicle (GV) oocytes with the micro-vibration culture (MVC) system in in vitro maturation (IVM) cycles and in vivo-matured oocytes in controlled ovarian hyperstimulation (COH) cycles in polycystic ovarian syndrome (PCOS) patients. METHODS: This study investigated 152 PCOS patients who underwent 159 fresh embryo transfer cycles, including IVM cycles with embryos derived from GV oocytes and the COH cycles with embryos derived from in vivo-matured oocytes. The IVM cycles were divided into groups according to the culture system used: static culture (SC) and MVC: In the IVM-S group (n = 47), SC was applied during both IVM and in vitro culture (IVC), whereas in the IVM-MV group (n = 44), MVC was applied during both IVM and IVC. For the COH cycles, in the COH-S group (n = 68), SC was applied during IVC. RESULTS: The number of in vitro-matured oocytes was similar in the IVM-S and IVM-MV groups, but the good-quality embryo (GQE; ≥6-cells) rate was significantly higher in the IVM-MV group (p < 0.01). The GQE rate and clinical outcomes of the COH-S group were significantly better than those of the IVM-S group (p < 0.05) but similar to those of the IVM-MV group. CONCLUSION: Compared with the SC system, the MVC system in IVM cycles improves the embryonic quality of GV oocytes and clinical outcomes, resulting in development of potential equivalent to in vivo-matured oocytes.


Assuntos
Desenvolvimento Embrionário/fisiologia , Técnicas de Maturação in Vitro de Oócitos/métodos , Oócitos/fisiologia , Indução da Ovulação/métodos , Síndrome do Ovário Policístico/fisiopatologia , Adulto , Transferência Embrionária , Feminino , Humanos , Gravidez , Resultado do Tratamento , Vibração
14.
Sci China Life Sci ; 63(6): 849-865, 2020 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-32291558

RESUMO

The number of growth factors involved in female fertility has been extensively studied, but reluctance to add essential growth factors in culture media has limited progress in optimizing embryonic growth and implantation outcomes, a situation that has ultimately led to reduced pregnancy outcomes. Insulin-like growth factor II (IGF-II) is the most intricately regulated of all known reproduction-related growth factors characterized to date, and is perhaps the predominant growth factor in human ovarian follicles. This review aims to concisely summarize what is known about the role of IGF-II in follicular development, oocyte maturation, embryonic development, implantation success, placentation, fetal growth, and in reducing placental cell apoptosis, as well as present strategies that use growth factors in culture systems to improve the developmental potential of oocytes and embryos in different species. Synthesizing the present knowledge about the physiological roles of IGF-II in follicular development, oocyte maturation, and early embryonic development should, on the one hand, deepen our overall understanding of the potential beneficial effects of growth factors in female reproduction and on the other hand support development (optimization) of improved outcomes for assisted reproductive technologies.


Assuntos
Fator de Crescimento Insulin-Like II/genética , Fator de Crescimento Insulin-Like II/fisiologia , Reprodução/fisiologia , Animais , Desenvolvimento Embrionário/fisiologia , Feminino , Desenvolvimento Fetal/fisiologia , Regulação da Expressão Gênica , Humanos , Oócitos/fisiologia , Folículo Ovariano/fisiologia , Gravidez
15.
PLoS One ; 15(4): e0231108, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32251418

RESUMO

Clinical applications of oocytes cryopreservation include preservation of future fertility of young cancer patients, substitution of embryo freezing to avoid associated legal and ethical issues, and delaying childbearing years. While the outcome of oocyte cryopreservation has recently been improved, currently used vitrification method still suffer from increased biosafety risk and handling issues while slow freezing techniques yield overall low success. Understanding better the mechanism of cryopreservation-induced injuries may lead to development of more reliable and safe methods for oocyte cryopreservation. Using the mouse model, a microarray study was conducted on oocyte cryopreservation to identify cryoinjuries to transcriptionally active genome. To this end, metaphase II (MII) oocytes were subjected to standard slow freezing, and then analyzed at the four-cell stage after embryonic genome activation. Non-frozen four-cell embryos served as controls. Differentially expressed genes were identified and validated using RT-PCR. Embryos produced from the cryopreserved oocytes displayed 200 upregulated and 105 downregulated genes, associated with the regulation of mitochondrial function, protein ubiquitination and maintenance, cellular response to stress and oxidative states, fatty acid and lipid regulation/metabolism, and cell cycle maintenance. These findings reveal previously unrecognized effects of standard slow oocyte freezing on embryonic gene expression, which can be used to guide improvement of oocyte cryopreservation methods.


Assuntos
Criopreservação/normas , Embrião de Mamíferos/fisiologia , Congelamento/efeitos adversos , Oócitos/fisiologia , Transcriptoma/genética , Animais , Desenvolvimento Embrionário/genética , Feminino , Fertilização In Vitro/métodos , Regulação da Expressão Gênica no Desenvolvimento , Humanos , Masculino , Metáfase/genética , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos DBA , Mapas de Interação de Proteínas/genética , Reação em Cadeia da Polimerase em Tempo Real
16.
J Ovarian Res ; 13(1): 42, 2020 Apr 21.
Artigo em Inglês | MEDLINE | ID: mdl-32316999

RESUMO

PURPOSE: Variations in many genes may lead to the occurrence of oocyte maturation defects. To investigate the genetic basis of oocyte maturation defects, we performed clinical and genetic analysis of a pedigree. METHODS: The proband with oocyte maturation defect-2 receiving ovulation induction therapy and her parents were selected for clinical detection, whole exome sequencing and Sanger sequencing. One unrelated healthy woman received ovulation induction therapy as control. Mutations were assessed after frequency screening of public exome databases. Then homozygous variants shared by the proband and her parents were selected. RESULTS: Arrest of oocytes maturation was observed. A new missense mutation in TUBB8 (TUBB8: NM_177,987: exon 2: c. C161T: p. A54V) was identified, which was shown to be rare compared with public databases. The variant was highly conserved among primates, and was suggested to be deleterious by online software prediction. CONCLUSIONS: The homozygote of this variant (TUBB8: NM_ 177,987: exon 2:c.C161T: p.A54V) might affect spindle assembly, cause arrest of oocyte maturation and lead to oocyte maturation defect-2.


Assuntos
Grupo com Ancestrais do Continente Asiático/genética , Infertilidade Feminina/genética , Oócitos/fisiologia , Tubulina (Proteína)/genética , Adulto , Consanguinidade , Feminino , Homozigoto , Humanos , Infertilidade Feminina/terapia , Mutação de Sentido Incorreto , Indução da Ovulação
17.
Proc Natl Acad Sci U S A ; 117(17): 9393-9400, 2020 04 28.
Artigo em Inglês | MEDLINE | ID: mdl-32295885

RESUMO

Sperm-oocyte fusion is a critical event in mammalian fertilization, categorized by three indispensable proteins. Sperm membrane protein IZUMO1 and its counterpart oocyte membrane protein JUNO make a protein complex allowing sperm to interact with the oocyte, and subsequent sperm-oocyte fusion. Oocyte tetraspanin protein CD9 also contributes to sperm-oocyte fusion. However, the fusion process cannot be explained solely by these three essential factors. In this study, we focused on analyzing a testis-specific gene 4930451I11Rik and generated mutant mice using the CRISPR/Cas9 system. Although IZUMO1 remained in 4930451I11Rik knockout (KO) spermatozoa, the KO spermatozoa were unable to fuse with oocytes and the KO males were severely subfertile. 4930451I11Rik encodes two isoforms: a transmembrane (TM) form and a secreted form. Both CRISPR/Cas9-mediated TM deletion and transgenic (Tg) rescue with the TM form revealed that only the TM form plays a critical role in sperm-oocyte fusion. Thus, we renamed this TM form Fertilization Influencing Membrane Protein (FIMP). The mCherry-tagged FIMP TM form was localized to the sperm equatorial segment where the sperm-oocyte fusion event occurs. Thus, FIMP is a sperm-specific transmembrane protein that is necessary for the sperm-oocyte fusion process.


Assuntos
Peptídeos e Proteínas de Sinalização Intercelular/metabolismo , Espermatozoides/metabolismo , Testículo/metabolismo , Sequência de Aminoácidos , Animais , Fertilização In Vitro , Humanos , Imunoglobulinas/genética , Imunoglobulinas/metabolismo , Infertilidade Masculina/genética , Peptídeos e Proteínas de Sinalização Intercelular/genética , Masculino , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Camundongos , Camundongos Knockout , Oócitos/fisiologia , Receptores de Superfície Celular/genética , Receptores de Superfície Celular/metabolismo , Interações Espermatozoide-Óvulo/fisiologia
18.
Cell ; 180(6): 1212-1227.e14, 2020 03 19.
Artigo em Inglês | MEDLINE | ID: mdl-32169215

RESUMO

The paternal genome undergoes a massive exchange of histone with protamine for compaction into sperm during spermiogenesis. Upon fertilization, this process is potently reversed, which is essential for parental genome reprogramming and subsequent activation; however, it remains poorly understood how this fundamental process is initiated and regulated. Here, we report that the previously characterized splicing kinase SRPK1 initiates this life-beginning event by catalyzing site-specific phosphorylation of protamine, thereby triggering protamine-to-histone exchange in the fertilized oocyte. Interestingly, protamine undergoes a DNA-dependent phase transition to gel-like condensates and SRPK1-mediated phosphorylation likely helps open up such structures to enhance protamine dismissal by nucleoplasmin (NPM2) and enable the recruitment of HIRA for H3.3 deposition. Remarkably, genome-wide assay for transposase-accessible chromatin sequencing (ATAC-seq) analysis reveals that selective chromatin accessibility in both sperm and MII oocytes is largely erased in early pronuclei in a protamine phosphorylation-dependent manner, suggesting that SRPK1-catalyzed phosphorylation initiates a highly synchronized reorganization program in both parental genomes.


Assuntos
Cromatina/metabolismo , Protaminas/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , Animais , Proteínas de Ciclo Celular/metabolismo , Núcleo Celular/metabolismo , Cromatina/fisiologia , Montagem e Desmontagem da Cromatina/genética , Montagem e Desmontagem da Cromatina/fisiologia , Fertilização/genética , Histonas/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Camundongos Knockout , Oócitos/metabolismo , Oócitos/fisiologia , Fosforilação , Protamina Quinase/genética , Protamina Quinase/metabolismo , Protaminas/genética , Proteínas Serina-Treonina Quinases/fisiologia , Processamento de RNA/genética , Processamento de RNA/fisiologia , Espermatozoides/metabolismo , Fatores de Transcrição/metabolismo , Zigoto/metabolismo
19.
PLoS Genet ; 16(3): e1008543, 2020 03.
Artigo em Inglês | MEDLINE | ID: mdl-32134927

RESUMO

Following fertilization of a mature oocyte, the formation of a diploid zygote involves a series of coordinated cellular events that ends with the first embryonic mitosis. In animals, this complex developmental transition is almost entirely controlled by maternal gene products. How such a crucial transcriptional program is established during oogenesis remains poorly understood. Here, we have performed an shRNA-based genetic screen in Drosophila to identify genes required to form a diploid zygote. We found that the Lid/KDM5 histone demethylase and its partner, the Sin3A-HDAC1 deacetylase complex, are necessary for sperm nuclear decompaction and karyogamy. Surprisingly, transcriptomic analyses revealed that these histone modifiers are required for the massive transcriptional activation of deadhead (dhd), which encodes a maternal thioredoxin involved in sperm chromatin remodeling. Unexpectedly, while lid knock-down tends to slightly favor the accumulation of its target, H3K4me3, on the genome, this mark was lost at the dhd locus. We propose that Lid/KDM5 and Sin3A cooperate to establish a local chromatin environment facilitating the unusually high expression of dhd, a key effector of the oocyte-to-zygote transition.


Assuntos
Proteínas de Drosophila/genética , Histona Desmetilases/genética , Oócitos/fisiologia , Zigoto/fisiologia , Animais , Núcleo Celular/genética , Cromatina/genética , Montagem e Desmontagem da Cromatina/genética , Drosophila/genética , Feminino , Regulação da Expressão Gênica no Desenvolvimento/genética , Histonas/genética , Masculino , Oogênese/genética , Espermatozoides/fisiologia , Transcrição Genética/genética
20.
Front Biosci (Schol Ed) ; 12: 116-136, 2020 03 01.
Artigo em Inglês | MEDLINE | ID: mdl-32114451

RESUMO

Oocyte quality influences early embryonic survival, establishment and maintenance of pregnancy, fetal development and adult diseases. The developmental competence of oocytes is acquired gradually and increases with follicular development. The ability of an oocyte to develop into an embryo depends on, having enough specific information in the form of mRNA or proteins. If this information is insufficient, defects in nuclear or cytoplasmic maturation, or in both processes, may arise and thus affect the in vitro development of fertilized oocytes. The greater developmental competence of oocytes aspirated from larger follicles is reported as compared with smaller follicles. Oocyte developmental competence is greatly correlated with the morphology of the cumulus oocyte complexes (COCs). Apart from morphological or biochemical markers, molecular markers have also been investigated. Until now, no specific markers of oocyte developmental competence could be described for the oocyte developmental competence. To, utilize female germplasm to its maximum, there is a need to enhance developmental competence of lesser competent oocytes derived from the follicles which are not fully grown. The oocyte pre-maturation and maturation conditions affect gene expression not only in the oocyte but till the blastocyst stages too. Strategies have been discussed in this review would be useful to enhance the developmental competence of oocytes.


Assuntos
Oócitos/fisiologia , Animais , Blastocisto/citologia , Blastocisto/fisiologia , Células do Cúmulo/citologia , Células do Cúmulo/fisiologia , Desenvolvimento Embrionário/fisiologia , Feminino , Humanos , Oócitos/citologia , Oócitos/crescimento & desenvolvimento , Oogênese/fisiologia , Gravidez
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