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1.
PLoS One ; 15(10): e0239779, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-33044971

RESUMO

BACKGROUND: The conditions of diminished ovarian reserve and primary ovarian insufficiency, characterized by poor fertility outcomes, currently comprise a major challenge in reproductive medicine, particularly in vitro fertilization. Currently in the IVF industry, blastocyst developmental success rate per treatment is routinely overlooked when a live birth results from treatment. Limited data are available on this significant and actionable variable of blastocyst development optimization, which contributes to improvement of treatment success Women with elevated basal FSH concentration are reported to still achieve reasonable pregnancy rates, although only a few studies report correlations with blastocysts development. Diagnostic values of AMH/basal FSH concentrations can be useful for determining the optimal stimulation protocol as well as identification of individuals who will not benefit from IVF due to poor prognosis. The objective of this study is to identify actionable clinical and culture characteristics of IVF treatment that influence blastocyst developmental rate, with the goal of acquiring optimal success. METHODS AND FINDINGS: A retrospective observational study was performed, based on 106 women undergoing IVF, regardless of prognosis, over a six-month period from January 1, 2015 to June 31, 2015. Rate of high-quality blastocyst production, which can be used for embryo transfer or vitrification, per normally fertilized oocyte, was evaluated. Treatment was determined successful when outcome was ≥ 40% high-quality blastocysts. The data were initially evaluated with the Evtree algorithm, a statistical computational analysis which is inspired by natural Darwinian evolution incorporating concepts such as mutation and natural selection (see Supplementary Material). The analysis processes all variables simultaneously against the outcome, aiming to maximize discrimination of each variable to then create a "branch" of the tree which can be used as a decision in treatment. The final model results in only those variables which are significant to outcomes. Generalized linear model (GLM) employing logistic regression and survival analysis with R software was used and the final fitting of the model was determined through the use of random forest and evolutionary tree algorithms. Individuals presenting with an [AMH] of >3.15 ng/ml and a good prognosis had a lower success per treatment (n = 11, 0% success) when total gonadotropin doses were greater than 3325 IU. Individuals that presented with an [AMH] of <1.78 ng/ml and a poor prognosis exhibited a greater success per treatment (n = 11, 80% success). AMH emerged as a superior indicator of blastocyst development compared to basal FSH. The accuracy of the prediction model, our statistical analysis using decision tree, evtree methodology is 86.5% in correctly predicting outcome based on the significant variables. The likelihood that the outcome with be incorrect of the model, or the error rate is 13.5%. CONCLUSIONS: [AMH] is a superior indicator of ovarian stimulation response and an actionable variable for stimulation dose management for optimizing blastocyst development in culture. Women whose [AMH] is ≥3.2 mg/ml, having a good prognosis, and developing >12 mature follicles result in <40% blastocysts when gonadotropin doses exceed 3325 IU per treatment. IVF treatments for poor responders that present with infertility due to diminished ovarian reserve, if managed appropriately, can produce more usable blastocyst per IVF treatment, thus increasing rate of blastocyst developmental success and ultimately increasing live birth rates. Future studies are needed to investigate the intra-follicular and the intra-cellular mechanisms that lead to the inverse relationship of blastocysts development and total gonadotropin doses in good responders in contrast to poor responders.


Assuntos
Hormônio Antimülleriano/sangue , Blastocisto/metabolismo , Blastocisto/fisiologia , Desenvolvimento Embrionário/fisiologia , Hormônio Foliculoestimulante/sangue , Adulto , Transferência Embrionária/métodos , Feminino , Fertilização In Vitro/métodos , Humanos , Infertilidade/sangue , Infertilidade/terapia , Nascimento Vivo , Masculino , Folículo Ovariano/metabolismo , Reserva Ovariana/fisiologia , Ovário/metabolismo , Ovário/fisiologia , Indução da Ovulação/métodos , Gravidez , Taxa de Gravidez , Estudos Retrospectivos
2.
Proc Natl Acad Sci U S A ; 117(33): 20015-20026, 2020 08 18.
Artigo em Inglês | MEDLINE | ID: mdl-32759216

RESUMO

We sequenced more than 52,500 single cells from embryonic day 11.5 (E11.5) postembryonic day 5 (P5) gonads and performed lineage tracing to analyze primordial follicles and wave 1 medullar follicles during mouse fetal and perinatal oogenesis. Germ cells clustered into six meiotic substages, as well as dying/nurse cells. Wnt-expressing bipotential precursors already present at E11.5 are followed at each developmental stage by two groups of ovarian pregranulosa (PG) cells. One PG group, bipotential pregranulosa (BPG) cells, derives directly from bipotential precursors, expresses Foxl2 early, and associates with cysts throughout the ovary by E12.5. A second PG group, epithelial pregranulosa (EPG) cells, arises in the ovarian surface epithelium, ingresses cortically by E12.5 or earlier, expresses Lgr5, but delays robust Foxl2 expression until after birth. By E19.5, EPG cells predominate in the cortex and differentiate into granulosa cells of quiescent primordial follicles. In contrast, medullar BPG cells differentiate along a distinct pathway to become wave 1 granulosa cells. Reflecting their separate somatic cellular lineages, second wave follicles were ablated by diptheria toxin treatment of Lgr5-DTR-EGFP mice at E16.5 while first wave follicles developed normally and supported fertility. These studies provide insights into ovarian somatic cells and a resource to study the development, physiology, and evolutionary conservation of mammalian ovarian follicles.


Assuntos
Células da Granulosa/citologia , Camundongos/embriologia , Folículo Ovariano/embriologia , Animais , Diferenciação Celular , Linhagem da Célula , Feminino , Proteína Forkhead Box L2/genética , Proteína Forkhead Box L2/metabolismo , Células da Granulosa/metabolismo , Camundongos/metabolismo , Folículo Ovariano/citologia , Folículo Ovariano/metabolismo , Gravidez , Receptores Acoplados a Proteínas-G/genética , Receptores Acoplados a Proteínas-G/metabolismo
3.
PLoS One ; 15(7): e0234795, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32645018

RESUMO

Forkhead box L2 (FOXL2) is a single-exon gene encoding a forkhead transcription factor, which is mainly expressed in the ovary, eyelids and the pituitary gland. FOXL2 plays an essential role in ovarian development. To reveal the effects of FOXL2 on the biological process and gene expression of ovarian granulosa cells (GCs), we established stable FOXL2-knockdown GCs and then analysed them using transcriptome sequencing. It was observed that knocking down FOXL2 affected the biological processes of cell proliferation, DNA replication, and apoptosis and affected cell cycle progression. FOXL2 knockdown promoted cell proliferation and DNA replication, decreased cell apoptosis, and promoted mitosis. In addition, by comparing the transcriptome after FOXL2 knockdown, we found a series of DEGs (differentially expressed genes) and related pathways. These results indicated that, through mediating these genes and pathways, the FOXL2 might induce the cell proliferation, cycle, and DNA replication, and play a key role during ovarian development and maintenance.


Assuntos
Proteína Forkhead Box L2/genética , Proteína Forkhead Box L2/metabolismo , Ovário/metabolismo , Animais , Ciclo Celular/genética , Divisão Celular/genética , Proliferação de Células/genética , Galinhas/genética , Replicação do DNA/genética , Feminino , Fatores de Transcrição Forkhead/metabolismo , Regulação da Expressão Gênica/genética , Células da Granulosa/metabolismo , Folículo Ovariano/metabolismo , RNA Mensageiro/genética , Transcriptoma , Sequenciamento Completo do Exoma
4.
Metabolism ; 107: 154241, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32304754

RESUMO

BACKGROUND: Hyperandrogenism is one of the major characteristics of polycystic ovary syndrome (PCOS). Abnormal miR-125b-5p expression has been documented in multiple diseases, but whether miR-125b-5p is associated with aberrant steroidogenesis in preantral follicles remains unknown. METHODS: Steriod hormone concentrations and miR-125b-5p expression were measured in clinical serum samples from PCOS patients. Using a mouse preantral follicle culture model and a letrozole-induced PCOS mouse model, we investigated the mechanism underlying miR-125b-5p regulation of androgen and oestrogen secretion. RESULTS: The decreased miR-125b-5p expression was observed in the sera from hyperandrogenic PCOS (HA-PCOS) patients. In mouse preantral follicles, inhibiting miR-125b-5p increased the expression of androgen synthesis-related genes and stimulated the secretion of testosterone, while simultaneously downregulating oestrogen synthesis-related genes and decreasing oestradiol release. Ectopically expressed miR-125b-5p reversed the effects on steroidogenesis-related gene expression and hormone release. Mechanistic studies identified Pak3 as a direct target of miR-125b-5p. Furthermore, inhibiting miR-125b-5p facilitated the activation of ERK1/2 in mouse preantral follicles, while inhibiting Pak3 abrogated this activating effect. These results were recapitulated in letrozole-induced PCOS mouse ovaries. Of note, inhibiting PAK3 antagonised the positive effect of miR-125b-5p siRNA on the expressions of androgen synthesis-related enzymes and testosterone secretion. Luteinizing hormone (LH) inhibited miR-125b-5p expression, and stimulated Pak3 expression. CONCLUSION: High serum LH concentrations in PCOS patients repress miR-125b-5p expression, which further increases Pak3 expression, leading to activation of ERK1/2 signalling, thus stimulating the expression of androgen synthesis-related enzymes and testosterone secretion in HA-PCOS.


Assuntos
MicroRNAs/genética , Folículo Ovariano/metabolismo , Esteroides/biossíntese , Androgênios/biossíntese , Androgênios/genética , Animais , Estradiol/metabolismo , Estrogênios/biossíntese , Estrogênios/genética , Feminino , Regulação da Expressão Gênica/genética , Hiperandrogenismo/induzido quimicamente , Hiperandrogenismo/metabolismo , Letrozol , Hormônio Luteinizante/metabolismo , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Sistema de Sinalização das MAP Quinases/genética , Camundongos , Camundongos Endogâmicos C57BL , Síndrome do Ovário Policístico/induzido quimicamente , Síndrome do Ovário Policístico/genética , Síndrome do Ovário Policístico/metabolismo , Quinases Ativadas por p21/genética , Quinases Ativadas por p21/metabolismo
5.
Endocrinology ; 161(4)2020 04 01.
Artigo em Inglês | MEDLINE | ID: mdl-32141511

RESUMO

Over the entire reproductive lifespan in mammals, a fixed number of primordial follicles serve as the source of mature oocytes. Uncontrolled and excessive activation of primordial follicles can lead to depletion of the ovarian reserve. We observed that disruption of estrogen receptor ß (ESR2) signaling results in increased activation of primordial follicles in Esr2-null (Esr2-/-) rats. However, follicle assembly was unaffected, and the total number of follicles remained comparable between neonatal wild-type and Esr2-/- ovaries. While the activated follicle counts were increased in Esr2-/- ovary, the number of primordial follicles were markedly decreased. Excessive recruitment of primordial follicles led to premature ovarian senescence in Esr2-/- rats and was associated with reduced levels of serum AMH and estradiol. Disruption of ESR2 signaling through administration of a selective antagonist (PHTPP) increased the number of activated follicles in wildtype rats, whereas a selective agonist (DPN) decreased follicle activation. In contrast, primordial follicle activation was not increased in the absence of ESR1, indicating that the regulation of primordial follicle activation is ESR2 specific. Follicle activation was also increased in Esr2 mutants lacking the DNA binding domain, suggesting a role for the canonical transcriptional activation function. Both primordial and activated follicles express ESR2, suggesting a direct regulatory role for ESR2 within these follicles. We also detected that loss of ESR2 augmented the activation of AKT, ERK, and mTOR pathways. Our results indicate that the lack of ESR2 upregulated both granulosa and oocyte factors, which can facilitate AKT and mTOR activation in Esr2-/- ovaries leading to increased activation of primordial follicles.


Assuntos
Hormônio Antimülleriano/sangue , Estradiol/sangue , Receptor beta de Estrogênio/genética , Folículo Ovariano/metabolismo , Reserva Ovariana/fisiologia , Animais , Moduladores de Receptor Estrogênico/farmacologia , Receptor beta de Estrogênio/agonistas , Receptor beta de Estrogênio/antagonistas & inibidores , Receptor beta de Estrogênio/metabolismo , Feminino , Alvo Mecanístico do Complexo 1 de Rapamicina , Nitrilos/farmacologia , Folículo Ovariano/efeitos dos fármacos , Reserva Ovariana/efeitos dos fármacos , Fosfatidilinositol 3-Quinases/metabolismo , Pirazóis/farmacologia , Pirimidinas/farmacologia , Ratos , Ratos Sprague-Dawley , Ratos Transgênicos , Transdução de Sinais/efeitos dos fármacos
6.
Endocrinology ; 161(4)2020 04 01.
Artigo em Inglês | MEDLINE | ID: mdl-32141513

RESUMO

Follicle-stimulating hormone (FSH)-induced growth of ovarian follicles is independent of follicular vascularization. Recent evidence has indicated that follicular vascularization is critical to ovarian follicle development and survival. FSH, a gonadotropin that induces follicular growth and development, also acts as the major survival factor for antral follicles. FSH has been reported to stimulate angiogenesis in the theca layers mediated in part by the vascular endothelial growth factor A (VEGFA) and the transcription factor hypoxia inducible factor 1α (HIF-1α). However, it remains largely undetermined whether FSH-dependent growth and survival of antral follicles relies on FSH-induced vascularization. Here, we first demonstrated that induction of angiogenesis through the FSH-HIF-1α-VEGFA axis is not required for FSH-stimulated follicular growth in mouse ovary. FSH increased the total number of blood vessels in mouse ovarian follicles, which was correlated with elevated expression of VEGFA and HIF-1α in granulosa cells. In contrast, blocking of follicular angiogenesis using inhibitors against the HIF-1α-VEGFA pathway repressed vasculature formation in follicles despite FSH administration. Interestingly, by measuring follicular size and ovarian weight, we found that the suppression of angiogenesis via HIF-1α-VEGFA pathway did not influence FSH-mediated follicular growth. However, inhibition of FSH-induced follicular vascularization by PX-478, a small-molecule inhibitor that suppresses HIF-1α activity, blocked ovulation and triggered atresia in large follicles. On the other hand, PX-478 injection reduced oocyte quality via impairing the meiotic apparatus, showing a prominently defective spindle assembly and actin dynamics. Collectively, our findings unveiled a vascularization-independent effect of FSH on follicular growth, whereas follicular survival, ovulation, and oocyte development relies on FSH-mediated angiogenesis in the follicles.


Assuntos
Hormônio Foliculoestimulante/farmacologia , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Oócitos/crescimento & desenvolvimento , Folículo Ovariano/crescimento & desenvolvimento , Ovulação/metabolismo , Transdução de Sinais/fisiologia , Fator A de Crescimento do Endotélio Vascular/metabolismo , Animais , Feminino , Camundongos , Neovascularização Fisiológica/efeitos dos fármacos , Neovascularização Fisiológica/fisiologia , Oócitos/efeitos dos fármacos , Oócitos/metabolismo , Folículo Ovariano/efeitos dos fármacos , Folículo Ovariano/metabolismo , Ovulação/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos
7.
Life Sci ; 249: 117515, 2020 May 15.
Artigo em Inglês | MEDLINE | ID: mdl-32147428

RESUMO

AIMS: This study investigated the effects of curcumin-loaded super-paramagnetic iron oxide (Fe3O4) nanoparticles (NPs) (SPIONs) on histological parameters and apoptosis-inducing factors (AIFs) in an experimental mouse model of polycystic ovary syndrome (PCOS). MATERIALS AND METHODS: A total number of 40 female prepuberal BALB/c mice were randomly divided into four groups. Group 1 was selected as control and Group 2 was considered as a vehicle taking sesame oil, in the form of a curcumin carrier. Moreover, Group 3 was administered with dehydroepiandrosterone (DHEA) at 6 mg/100 g of the body weight and Group 4 received the DHEA plus the NPs of curcumin (5.4 mg/100 g) for twenty consecutive days. Finally, histology, stereology, and apoptosis of the ovary were evaluated. KEY FINDINGS: The results revealed that the NPs of curcumin had reduced ovarian volume (p < 0.05) and a total number of primary, secondary, antral, and primordial follicles in comparison with the PCOS and vehicle groups (p < 0.05). Furthermore, curcumin treatment following administration of the DHEA resulted in a significant decrease in BAX (p < 0.001) and levels of expression of Caspase3 (CASP3) protein, increased levels of B-cell lymphoma 2 (Bcl2) expression (p < 0.05), and moderated apoptosis in granulosa cells in comparison with the ones seen in the PCOS group. SIGNIFICANCE: Ovarian injuries and DHEA-induced apoptosis were efficiently suppressed by curcumin, indicating the probable protective property of NPs of curcumin against PCOS.


Assuntos
Apoptose/efeitos dos fármacos , Curcumina/administração & dosagem , Desidroepiandrosterona/administração & dosagem , Compostos Férricos/administração & dosagem , Nanopartículas Metálicas/química , Síndrome do Ovário Policístico/induzido quimicamente , Síndrome do Ovário Policístico/tratamento farmacológico , Maturidade Sexual , Animais , Antioxidantes/administração & dosagem , Apoptose/genética , Caspase 3/metabolismo , Modelos Animais de Doenças , Feminino , Compostos Férricos/química , Expressão Gênica/efeitos dos fármacos , Camundongos , Camundongos Endogâmicos BALB C , Folículo Ovariano/metabolismo , Folículo Ovariano/patologia , Estresse Oxidativo
8.
Curr Opin Insect Sci ; 37: 39-48, 2020 02.
Artigo em Inglês | MEDLINE | ID: mdl-32087562

RESUMO

The follicle stem cell (FSC) lineage in the Drosophila ovary is a highly informative model of in vivo epithelial stem cell biology. Studies over the past 30 years have identified roles for every major signaling pathway in the early FSC lineage. These pathways regulate a wide variety of cell behaviors, including self-renewal, proliferation, survival and differentiation. Studies of cell signaling in the follicle epithelium have provided new insights into how these cell behaviors are coordinated within an epithelial stem cell lineage and how signaling pathways interact with each other in the native, in vivo context of a living tissue. Here, we review these studies, with a particular focus on how these pathways specify differences between the FSCs and their daughter cells. We also describe common themes that have emerged from these studies, and highlight new research directions that have been made possible by the detailed understanding of the follicle epithelium.


Assuntos
Folículo Ovariano/citologia , Transdução de Sinais , Células-Tronco/metabolismo , Animais , Drosophila/crescimento & desenvolvimento , Feminino , Folículo Ovariano/metabolismo , Nicho de Células-Tronco , Células-Tronco/citologia
9.
J Ovarian Res ; 13(1): 15, 2020 Feb 10.
Artigo em Inglês | MEDLINE | ID: mdl-32041647

RESUMO

BACKGROUND: DCN (decorin) is a proteoglycan known to be involved in regulating cell proliferation, collagen fibril organization and migration. In our global transcriptome RNA-sequencing approach to systematically identify new ovulation-associated genes, DCN was identified as one of the highly regulated genes. We therefore hypothesize that DCN may have a role in ovulatory processes such as cell migration and proliferation. AIM: To characterize the expression, regulation and function of the proteoglycan DCN in the human ovarian follicles during the preovulatory period. METHODS: The in-vivo expression of DCN mRNA in mural (MGCs) and cumulus (CGCs) granulosa cells was characterized using quantitative RT-PCR and western blot. A signaling study was performed by treating human MGCs cultures with gonadotropins and different stimulators and inhibitors to determine their effect on DCN expression by qRT- PCR and elucidate the pathways regulating these proteins. In a functional study, KGN granulosa cell line was used to study cell migration with a scratch assay. RESULTS: DCN mRNA expression was significantly higher in MGCs compared to CGCs. DCN mRNA was significantly higher in CGCs surrounding mature metaphase II (MII) oocytes compared to CGCs of germinal vesicle (GV) and metaphase I (MI) oocytes. hCG significantly increased DCN mRNA and protein expression levels in cultured MGCs. Using signal transduction activators and inhibitors, we demonstrated that DCN induction by LH/hCG is carried out via PKA, PKC, ERK/MEK, and PI3K pathways. We showed that DCN expression is also induced in high-density cell cultures, in a dose-dependent pattern. In addition, progesterone induced a significant increase in DCN secretion to the media. MGCs from follicles of endometriosis patients exhibited reduced (about 20% of) mRNA transcriptions levels compared to MGCs follicles of control patients. More significantly, we found that DCN has an inhibiting effect on KGN cell migration. CONCLUSIONS: Our study indicates that DCN is a unique ovulatory gene. Our findings support the hypothesis that DCN plays an important new role during the preovulatory period and ovulation, and stress its involvement in endometriosis infertility. A better understanding of DCN role in ovulation and endometriosis may provide treatment for some types of infertility.


Assuntos
Decorina/metabolismo , Folículo Ovariano/metabolismo , Células Cultivadas , Decorina/genética , Feminino , Humanos , Ovulação/fisiologia , Estudos Prospectivos , RNA Mensageiro/genética , RNA Mensageiro/metabolismo
10.
Transplant Proc ; 52(1): 406-413, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-31911055

RESUMO

BACKGROUND: Subcutaneous ovarian transplantation has recently begun receiving increased attention. Fourteen days after transplantation is used as an important time point for assessing the recovery of ovarian function. The goal of this study is to determine the expression of apoptotic genes in the ovary at this time. METHODS: This study investigated follicle development and the expression of 3 apoptosis genes (Bax, Bcl2, and P53) after mouse ovaries were transplanted. Seven-week-old mouse ovaries were autologously transplanted into back muscle. The ovaries were harvested on day 14, morphology was observed by hematoxylin and eosin staining, and the distribution of 3 proteins was observed by immunohistochemistry. TUNEL staining showed where apoptosis occurred in the ovary. Finally, RT-PCR/Western blotting was used to analyze the differential expression of mRNA/proteins between the transplantation group and the control group. RESULTS: The results revealed follicles at different stages at the edge of the grafts. In immunohistochemical experiments, BAX, BCL2, and P53 were found to be extensively expressed in the transplant group and the control group. P53 was strongly expressed in the medulla of transplanted ovaries. Bax was strongly expressed in the antral follicles of both groups. The results were consistent with the results of the TUNEL experiments. Three genes (Bax, Bcl2, and P53) were downregulated in the transplanted groups. The results showed that significant differences were detected in Bax and P53 mRNA expression levels between the transplanted groups and the control group (P < .01). Bcl2 expression was not significantly different, but the Bax/Bcl2 ratio increased. The results of the protein experiments were the same. CONCLUSION: P53 may downregulate Bax in the early stage of transplantation. Follicle growth and atresia were regulated through modulation of Bcl2- and Bax-mediated apoptotic pathways in heterotopic ovarian transplantation.


Assuntos
Folículo Ovariano/metabolismo , Folículo Ovariano/patologia , Ovário/metabolismo , Ovário/patologia , Ovário/transplante , Proteína Supressora de Tumor p53/metabolismo , Animais , Apoptose/fisiologia , Feminino , Camundongos , Camundongos Endogâmicos C57BL , Transplante Autólogo , Proteína X Associada a bcl-2/metabolismo
11.
Int J Mol Med ; 45(2): 647-657, 2020 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-31894307

RESUMO

The molecular mechanisms underlying the activation of primordial follicles are poorly understood. The serine/threonine protein kinase phosphoinositide­dependent kinase 1 (PDK1), a pivotal downstream effector of phosphatidyl inositol­3 kinase (PI3K) signaling, plays a vital role in cellular signaling. In order to identify the function of PDK1 in ovarian follicle development, this study used conditional Pdk1 deletion in mouse oocytes by crossing Pdk1loxP/loxP mice with transgenic mice carrying Gdf­9 promoter­mediated Cre recombinase and found that Pdk1flx/flxGdf9Cre mice were subfertile with increased serum follicle­stimulating hormone (FSH) and luteinizing hormone (LH) levels compared with Pdk1flx/flx mice. The deletion of Pdk1 in oocytes induced massive primordial follicle activation, leading to premature ovarian failure (POF). Further investigation revealed that enhanced Yes­associated protein (YAP) expression and an increased pro­inflammatory response also contributed to massive primordial follicle activation. PDK1 formed the complex with the core kinases of Hippo signaling and regulated the expression levels of YAP. On the whole, the findings of the present study demonstrate that PDK1 serves as an indispensable gatekeeper for maintaining the primordial follicle pool and provide a deeper understanding of POF treatment.


Assuntos
Proteínas Quinases Dependentes de 3-Fosfoinositídeo/genética , Proteínas Adaptadoras de Transdução de Sinal/genética , Proteínas de Ciclo Celular/genética , Folículo Ovariano/metabolismo , Insuficiência Ovariana Primária/genética , Animais , Feminino , Deleção de Genes , Inflamação/genética , Inflamação/patologia , Camundongos Endogâmicos ICR , Oócitos/metabolismo , Folículo Ovariano/patologia , Insuficiência Ovariana Primária/patologia , Regulação para Cima
12.
J Ovarian Res ; 13(1): 5, 2020 Jan 08.
Artigo em Inglês | MEDLINE | ID: mdl-31915051

RESUMO

BACKGROUND: The complex regulatory mechanism involved in ovarian follicular development is not completely understood. Neuronal neuropeptide Y (NPY) is involved in the regulation of feeding behavior, energy homeostasis, and reproduction behavior, while its function in ovarian follicular development is not clear. The objective of this study was to investigate if and how NPY regulates follicle development in the ovary. METHODS: All experiments were performed using Sprague Dawley rats. To understand NPY expression pattern at different stages of follicular development, NPY content was assessed using immunohistochemistry in individual follicles. NPY and its receptors expression pattern were evaluated in granulosa cells isolated from preantral (PA), early antral (EA) and late antral follicles (LAF). The influence of NPY on granulosa cell proliferation and apoptosis were further assessed in vitro, using Ki67- and TUNEL-positivity assays. To investigate whether NPY induced-proliferation in EA granulosa cells is mediated through the activation of NPY receptor Y5 (NPY5R) and Mitogen-activated protein kinase (MEK) signal pathway, EA granulosa cells were treated with NPY5R antagonist (CGP71683) and MEK inhibitors (PD98059 and U0126), and Ki67-positive cells were assessed. RESULTS: NPY protein expression was follicular stage-dependent and cell type-specific. NPY signal intensity in EA was higher than those in PA and LAF. Antral granulosa cells showed the highest signal intensity compared to mural granulosa cells, cumulus cells and theca cells. Granulosa cells NPY protein content and mRNA abundance were higher in EA than in LAF. NPY receptor contents in granulosa cells were follicular stage-dependent. While NPY reduced apoptosis of EA granulosa cells, it increased the proliferation through NPY5R and MEK pathway. In contrast, in LAF granulosa cells, NPY reduced proliferation and increased the number of apoptotic cells, with no significant effects on PA granulosa cells. CONCLUSION: This study is the first to evaluate the intraovarian role of NPY in granulosa cells at various stage of follicular development. These results indicate that NPY regulates granulosa cells proliferation and apoptosis in a follicular stage-dependent and autocrine manner. NPY may play a role in pathogenesis of ovarian follicular disorders.


Assuntos
Apoptose/genética , Proliferação de Células/genética , Regulação da Expressão Gênica , Células da Granulosa/metabolismo , Neuropeptídeo Y/genética , Folículo Ovariano/metabolismo , Animais , Western Blotting , Células Cultivadas , Feminino , Células da Granulosa/citologia , Sistema de Sinalização das MAP Quinases , Neuropeptídeo Y/metabolismo , Folículo Ovariano/citologia , Ovário/citologia , Ovário/metabolismo , Ratos Sprague-Dawley , Receptores de Neuropeptídeo Y/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa
13.
J Ovarian Res ; 13(1): 7, 2020 Jan 11.
Artigo em Inglês | MEDLINE | ID: mdl-31926556

RESUMO

BACKGROUND: The timing of the first postpartum ovulation is an important factor affecting the timing of estrous resumption in dairy cows. The first postpartum ovulation is delayed in cows producing large amounts of milk with an intensive negative energy balance. The antral follicle count (AFC) and serum anti-Müllerian hormone concentrations are known to be indicators of the ovarian reserve, which is the number and quality of follicles left in a pair of ovaries and known as an indicator of female fertility. Cows with higher AFC have been proven to show higher pregnancy rate and shorter calving to conception intervals; however, the relationship between the timing of the first postpartum ovulation and ovarian reserve remains unclear. Therefore, this study examined the relationships between postpartum follicular dynamics, the ovarian cycle, nutritional status, and ovarian reserve. METHODS: Transrectal ultrasonography was conducted from calving to 70-120 days in milk (DIM) in 26 cows to monitor AFC, follicular dynamics and the ovarian cycle. Body weight (BW) and milk yield were used as indicators of nutritional status. RESULTS: The first postpartum ovulation was significantly later in cows with low AFC (< 25) than in those with high AFC (≥25), while changes in BW from calving to the nadir and milk production were similar in both groups. The present results also suggested that cows with low AFC and a delayed first postpartum ovulation had a shorter first ovarian cycle after the first postpartum ovulation. The mean DIM of the first postpartum artificial insemination (AI) and days open (days from calving to AI with which pregnancy was achieved) were similar in high and low AFC groups. CONCLUSIONS: The first postpartum ovulation was significantly earlier in cows with high AFC than in those with low AFC. The assumed reason for this result was higher sensitivity to luteinizing hormone and larger androstenedione and estradiol production in follicles in high AFC cows. Therefore, cows with high AFC may be more fertile than those with low AFC while their milk production increase and BW decrease; it means they are in negative energy balance. (340/350 words).


Assuntos
Ciclo Estral/fisiologia , Folículo Ovariano/fisiologia , Ovulação/fisiologia , Período Pós-Parto/fisiologia , Animais , Hormônio Antimülleriano/sangue , Peso Corporal/fisiologia , Bovinos , Contagem de Células , Feminino , Hormônios/metabolismo , Leite/metabolismo , Folículo Ovariano/citologia , Folículo Ovariano/metabolismo , Reserva Ovariana/fisiologia , Gravidez , Taxa de Gravidez , Fatores de Tempo
14.
Acta Obstet Gynecol Scand ; 99(7): 917-924, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-31945183

RESUMO

INTRODUCTION: Human ovulation is a biologically complex process that involves several biochemical factors, promoting follicular rupture and release of a fertilizable oocyte. Proteins which are present in follicular fluid at high concentrations during ovulation are likely to be active participants in the biochemical pathways of ovulation. The aim of the study was to identify, by use of a modern proteomic technique, proteins of human follicular fluid which are differentially regulated during ovulation of the natural menstrual cycle. MATERIAL AND METHODS: This prospective experimental study over 3 years included women planned for laparoscopic sterilization. During surgery, retrieval of the dominant follicle was performed either at the preovulatory stage or during ovulation. Four women of preovulatory phase and four women of ovulatory phase met the predetermined criteria of hormone levels for respective phases, and samples of these were finally included out of the 15 women operated. Follicular fluid was aspirated from the excised follicle and subjected to mass spectrometry with the isobaric tags for relative and absolute quantification (iTRAQ) technology for isobaric tagging of peptides. This enables simultaneous identification and quantification of proteins. The protein profiles of the follicular fluid of the preovulatory phase and the ovulatory phase were analyzed, and proteins that were present were identified. RESULTS: A total of 502 proteins were identified, several of which previously have not been identified in human follicular fluid. Of the 115 proteins that were found in all samples, 20 proteins were at higher levels during ovulation. These were inflammatory-related proteins, coagulation factors, proteins in lipid metabolism, complement factors and antioxidants. Five proteins were present in lower levels during ovulation, with three being enzymes and the other two proteins of lipid metabolism and iron transport. CONCLUSIONS: Twenty-five follicular fluid proteins, with differential regulation during ovulation, were identified in human follicular fluid of the natural menstrual cycle. These proteins may have essential roles in the ovulatory cascade.


Assuntos
Líquido Folicular/química , Folículo Ovariano/metabolismo , Ovulação/metabolismo , Proteínas/metabolismo , Proteômica , Adulto , Feminino , Fase Folicular/metabolismo , Humanos , Espectrometria de Massas , Estudos Prospectivos , Suécia
15.
BMC Cancer ; 20(1): 67, 2020 Jan 29.
Artigo em Inglês | MEDLINE | ID: mdl-31996163

RESUMO

BACKGROUND: We previously reported that tamoxifen (TAM)-induced ovarian hyperstimulation (OHS) is associated with high serum concentrations of estradiol in premenopausal women with breast cancer. To investigate risk factors for TAM-induced OHS, we performed a retrospective multicenter study. METHODS: Premenopausal patients who received surgical therapy for endocrine-dependent breast cancer (n = 235) were recruited in this study and classified into 4 groups: group A, treated with TAM alone; group B, TAM treatment after 2-year-combined therapy with a gonadotropin-releasing hormone (Gn-RH) agonist; group C, TAM treatment after chemotherapy; group D, 5-year-combined therapy with TAM and a Gn-RH agonist. A serum estradiol value of more than 300 pg/mL or mean follicular diameter of more than 30 mm was defined as OHS. RESULTS: The incidence of OHS in group A (n = 13/26, 50.0%) was significantly higher than those in group B (n = 17/63, 27.0%), group C (n = 20/110, 18.2%), and group D (n = 0/36, 0%). The incidence of OHS was significantly correlated with aging, and the median serum concentration of estradiol in the presence of OHS was 823.0 pg/mL. The incidence of OHS (less than 47 years old) was 62.5% in group A, 48.6% in group B, and 28.2% in group C, respectively. Notably, the incidence rate of OHS following amenorrhea in group C (n = 13/20, 65.0%) was significantly higher than that in group B (n = 1/17, 5.9%). CONCLUSIONS: These findings indicate that the onset of OHS following amenorrhea was common in the post-chemotherapeutic group, while its ratio was low in the group after Gn-RH analog treatment, suggesting that combined treatment-based management involving TAM therapy is necessary for premenopausal patients with breast cancer.


Assuntos
Antineoplásicos Hormonais/efeitos adversos , Folículo Ovariano/efeitos dos fármacos , Folículo Ovariano/metabolismo , Ovário/efeitos dos fármacos , Ovário/metabolismo , Pré-Menopausa , Tamoxifeno/efeitos adversos , Adulto , Fatores Etários , Antineoplásicos Hormonais/administração & dosagem , Antineoplásicos Hormonais/uso terapêutico , Protocolos de Quimioterapia Combinada Antineoplásica/efeitos adversos , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Neoplasias da Mama/sangue , Neoplasias da Mama/diagnóstico , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/fisiopatologia , Esquema de Medicação , Estradiol/sangue , Feminino , Humanos , Sistema Hipotálamo-Hipofisário/efeitos dos fármacos , Sistema Hipotálamo-Hipofisário/metabolismo , Pessoa de Meia-Idade , Modelos Biológicos , Estadiamento de Neoplasias , Folículo Ovariano/crescimento & desenvolvimento , Ovário/crescimento & desenvolvimento , Estudos Retrospectivos , Tamoxifeno/administração & dosagem , Tamoxifeno/uso terapêutico
16.
Cell Mol Life Sci ; 77(6): 1177-1196, 2020 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-31327046

RESUMO

In mammalian ovaries, the theca layers of growing follicles are critical for maintaining their structural integrity and supporting androgen synthesis. Through combining the postnatal monitoring of ovaries by abdominal magnetic resonance imaging, endocrine profiling, hormonal analysis of the follicular fluid of growing follicles, and transcriptomic analysis of follicular theca cells, we provide evidence that the exposure of ovine fetuses to testosterone excess activates postnatal follicular growth and strongly affects the functions of follicular theca in adulthood. Prenatal exposure to testosterone impaired androgen synthesis in the small antral follicles of adults and affected the expression in their theca cells of a wide array of genes encoding extracellular matrix components, their membrane receptors, and signaling pathways. Most expression changes were uncorrelated with the concentrations of gonadotropins, steroids, and anti-Müllerian hormone in the recent hormonal environment of theca cells, suggesting that these changes rather result from the long-term developmental effects of testosterone on theca cell precursors in fetal ovaries. Disruptions of the extracellular matrix structure and signaling in the follicular theca and ovarian cortex can explain the acceleration of follicle growth through altering the stiffness of ovarian tissue. We propose that these mechanisms participate in the etiology of the polycystic ovarian syndrome, a major reproductive pathology in woman.


Assuntos
Síndrome do Ovário Policístico/metabolismo , Efeitos Tardios da Exposição Pré-Natal/metabolismo , Testosterona/metabolismo , Células Tecais/metabolismo , Animais , Células Cultivadas , Feminino , Regulação da Expressão Gênica , Redes Reguladoras de Genes , Humanos , Folículo Ovariano/citologia , Folículo Ovariano/crescimento & desenvolvimento , Folículo Ovariano/metabolismo , Síndrome do Ovário Policístico/genética , Gravidez , Efeitos Tardios da Exposição Pré-Natal/genética , Ovinos , Células Tecais/citologia , Células Tecais/ultraestrutura
17.
Chemosphere ; 244: 125497, 2020 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-31809938

RESUMO

Perfluorobutanesulfonate (PFBS), a short-chain perfluoroalkyl substance, is used in many industrial products. Preliminary evidence suggests that exposure to PFBS may increase the risk of infertility. The aim of this study was to investigate the influence of PFBS on ovarian function. Herein, we show that exposure of adult female mice to PFBS (200 mg/kg/day) (PFBS-mice) caused a decrease in the levels of serum total triiodothyronine and thyroxine, which depended on the activation of peroxisome proliferator-activated receptor α (PPARα). The numbers of secondary, early antral and antral follicles were reduced in PFBS-mice with an increase in the atretic follicles, and these changes were recovered by the replacement of L-thyroxinein or the treatment with PPARα antagonist GW6471. PFBS-induced hypothyroxinemia led to a decrease in the levels of Akt, mTOR and p70S6K phosphorylation in ovarian granular cells and cumulus cells, which suppressed the proliferation of these cells and enhanced autophagic death of granular cells and cumulus cells. The levels of serum estradiol and progesterone were reduced in PFBS-mice with a low expression of the steroidogenic genes Star and P450scc in ovarian tissues, which were sensitive to the replacement of L-thyroxinein or the blockade of PPARα. The results indicate that exposure to PFBS (≥200 mg/kg/day) through reducing thyroid hormones causes down-regulation of Akt-mTOR signaling in granular cells and cumulus cells, leading to the deficits in the development of follicles and the biosynthesis of ovarian hormones.


Assuntos
Fluorcarbonetos/toxicidade , Substâncias Perigosas/toxicidade , Ovário/efeitos dos fármacos , Ácidos Sulfônicos/toxicidade , Tiroxina/metabolismo , Animais , Regulação para Baixo , Feminino , Fluorcarbonetos/metabolismo , Camundongos , Folículo Ovariano/metabolismo , Ovário/metabolismo , Fosforilação , Progesterona/sangue , Proteínas Proto-Oncogênicas c-akt/metabolismo , Proteínas Quinases S6 Ribossômicas 70-kDa , Transdução de Sinais , Serina-Treonina Quinases TOR/metabolismo , Tri-Iodotironina/metabolismo
18.
J Anim Sci ; 98(1)2020 Jan 01.
Artigo em Inglês | MEDLINE | ID: mdl-31832639

RESUMO

The E2F family of transcription factors plays an important role in the control of the cell cycle, cell proliferation, and differentiation, and their role in ovarian function is just emerging. Although some evidence suggests a possible role of E2F1 in ovarian follicular development, what regulates its production in ovarian cells is unknown. Objectives of this study were to determine whether: (i) E2F1 gene expression in granulosa cells (GCs) and theca cells (TCs) change with follicular development and (ii) E2F1 mRNA abundance in TC and GC is hormonally regulated. Using real-time PCR, E2F1 mRNA abundance in GC was 5.5-fold greater (P < 0.05) in small (SM; 1 to 5 mm) than large (LG; >8 mm) follicles, but in TC, E2F1 expression did not differ among follicle sizes. SM-follicle GC had 2.1-fold greater (P < 0.05) E2F1 mRNA than TC. In SM-follicle GC, FGF9 induced a 7.6-fold increase in E2F1 mRNA abundance; however, FGF9 did not affect (P > 0.10) abundance of E2F1 mRNA in LG-follicle TC or GC. Follicle-stimulating hormone (FSH) had no effect (P > 0.10) on E2F1 gene expression in SM- or LG-follicle GC. SM-follicle GC were concomitantly treated with insulin-like growth factor 1 (30 ng/mL), FSH (30 ng/mL), and either 0 or 30 ng/mL of FGF9 with or without 50 µM of an E2F inhibitor (E2Fi; HLM0064741); FGF9 alone increased (P < 0.05) GC numbers, whereas E2Fi alone decreased (P < 0.05) GC numbers, and concomitant treatment of E2Fi with FGF9 blocked (P < 0.05) this stimulatory effect of FGF9. Estradiol production was inhibited (P < 0.05) by FGF9 alone and concomitant treatment of E2Fi with FGF9 attenuated (P < 0.05) this inhibitory effect of FGF9. SM-follicle GC treated with E2Fi decreased (P < 0.05) E2F1 mRNA abundance by 70%. Collectively, our studies show that GC E2F1 mRNA is developmentally and hormonally regulated in cattle. Inhibition of E2F1 reduced FGF9-induced GC proliferation and attenuated FGF9-inhibited estradiol production, indicating that E2F1 may be involved in follicular development in cattle.


Assuntos
Bovinos/genética , Fator de Transcrição E2F1/genética , Estradiol/metabolismo , Hormônio Foliculoestimulante/metabolismo , Regulação da Expressão Gênica/genética , Animais , Bovinos/crescimento & desenvolvimento , Bovinos/fisiologia , Proliferação de Células/genética , Feminino , Células da Granulosa/metabolismo , Folículo Ovariano/metabolismo , RNA Mensageiro/genética , Células Tecais/metabolismo
19.
J Sci Food Agric ; 100(1): 92-101, 2020 Jan 15.
Artigo em Inglês | MEDLINE | ID: mdl-31435952

RESUMO

BACKGROUND: Oyster polypeptides have various biofunctions, such as anti-cancer and anti-oxidative stress, but whether it has the protective effects to primary ovarian failure (POF) remains poorly understand. To address this issue, daily gavage of oyster polypeptides was performed to investigate their protective effect, basing on d-galactose-induced POF model in C57BL/6 female mice. RESULTS: Oyster polypeptides restored the irregular estrous cycles and the abnormal serum follicle stimulating hormone (FSH), luteinizing hormone (LH) and progesterone (P) levels as well as the decreased mRNA expression level of Amh that were induced by d-galactose. The follicle development of POF mice was improved by increasing the primordial follicle ratio and decreasing the atretic follicle number after oral administration of oyster polypeptides. Moreover, in the oyster polypeptides treated mice, the total superoxide dismutase (T-SOD) activity was significantly increased, while the malondialdehyde levels were significantly decreased. The mRNA expression levels of stress-related genes (SOD2, SIRT1 and FOXO3a) were remarkably up-regulated after d-galactose induction, but the up-regulation was weakened or disappeared by the gavage of oyster polypeptides. In addition, oyster polypeptides treatment also reduced the apoptosis of the ovarian granulosa cells and down-regulated the mRNA expression levels of apoptosis-related genes (p53 and Bad but not Bcl-2). CONCLUSION: This study reveals that oyster polypeptides may protect ovary against d-galactose-induced POF by their anti-oxidative stress activity to rescue d-galactose-induced ovarian oxidative damage and therefore to prevent ovarian cells apoptosis, thereby tipping the abnormality trigged by POF to get close to the normal levels. © 2019 Society of Chemical Industry.


Assuntos
Ostreidae/química , Peptídeos/administração & dosagem , Insuficiência Ovariana Primária/tratamento farmacológico , Substâncias Protetoras/administração & dosagem , Animais , Apoptose/efeitos dos fármacos , Feminino , Galactose/efeitos adversos , Humanos , Hormônio Luteinizante/metabolismo , Malondialdeído/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Folículo Ovariano/efeitos dos fármacos , Folículo Ovariano/metabolismo , Ovário/efeitos dos fármacos , Ovário/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Insuficiência Ovariana Primária/induzido quimicamente , Insuficiência Ovariana Primária/genética , Insuficiência Ovariana Primária/metabolismo , Progesterona/metabolismo , Sirtuína 1/genética , Sirtuína 1/metabolismo , Superóxido Dismutase/genética , Superóxido Dismutase/metabolismo
20.
Gen Comp Endocrinol ; 285: 113230, 2020 01 01.
Artigo em Inglês | MEDLINE | ID: mdl-31348955

RESUMO

During the ovarian ontogeny in birds, five fundamental events can be recognized: migration and colonization of the primordial germ cells, differentiation and proliferation of oogonies, an organization of germinal nests, beginning of the meiotic process and folliculogenesis. The knowledge of these events is fundamental for the interpretation of the processes involved in the differentiation of female gametes. However, there are only references for some model species such as Gallus gallus domesticus and Coturnix coturnix. In a previous study, the histological structure of embryonic ovaries of Columba livia was revealed. Therefore, the objective of this work is to characterize the processes of meiosis and folliculogenesis C. livia from the analysis of the expression of the GnRH receptor, the 3ßHSD enzyme and the cell proliferation protein PCNA in embryonic and postnatal ovaries. Therefore, the expression of GnRHR, 3ßHSD, and PCNA was revealed in histological testicular and ovarian preparations in embryos (stages 41-43) and neonates (2, 5, 7, 10 and 75 days post-hatching). The present study demonstrates that the fate of germline cells is dictated by their location during gonadal development. Thus, the germline cells located in the cortex of the left gonad enter meiosis, while those in the right gonad and those in the medulla of the left ovary fail to go into meiosis. This indicates that somatic signals, instead of an autonomous cellular mechanism, regulate the entry of the germline cells into meiosis in the C. livia embryo. Future studies will be focused on the analysis of proteins associated with meiotic events and folliculogenesis in embryonic and neonatal ovaries of C. livia, to evaluate the regulation of meiosis in vitro.


Assuntos
3-Hidroxiesteroide Desidrogenases/metabolismo , Columbidae/metabolismo , Meiose , Folículo Ovariano/crescimento & desenvolvimento , Receptores LHRH/metabolismo , Animais , Proliferação de Células , Columbidae/embriologia , Embrião não Mamífero/citologia , Feminino , Células Germinativas/metabolismo , Imuno-Histoquímica , Oócitos/citologia , Oócitos/metabolismo , Folículo Ovariano/metabolismo , Antígeno Nuclear de Célula em Proliferação/metabolismo
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