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1.
Arch Gynecol Obstet ; 307(6): 2001-2010, 2023 06.
Artigo em Inglês | MEDLINE | ID: mdl-37061986

RESUMO

PURPOSE: This study attempted at identifying the main parameters influencing the outcome of frozen embryo transfers. METHODS: This is a single-center retrospective cohort study of 830 frozen-embryo-transfer cycles performed at a German university hospital from January 2012 to December 2016. Main outcome parameters were the clinical pregnancy and live birth rate. Twelve patient- and cycle-dependent factors were analyzed in terms of their influence on the outcome of frozen embryo transfers. Multivariate logistic regression analysis was used for the modelling of the dependency of the different parameters on outcomes. RESULTS: The clinical pregnancy rate in our study was 25.5%, the live birth rate was 16.1% with an average maternal age of 34.2 years at the time of the oocyte retrieval. In the univariate analysis age, number of transferred embryos, blastocyst versus cleavage stage transfer, embryo quality and mode of endometrial preparation affected the birth rate significantly. The birth rate after artificial endometrial preparation was significantly lower than the birth rate after transfers in modified natural cycles (12.8 versus 20.6% with p = 0.031). The multivariate logistic regression analysis showed a significant independent influence of age, number of transferred embryos, culture duration and mode of endometrial preparation on the frozen embryo transfer success rates. Body mass index, nicotine abuse, a history of PCO syndrome or endometriosis and the co-transfer of a second poor-quality embryo to a good-quality embryo appeared to be irrelevant for the outcome in our collective. CONCLUSION: Age, number of transferred embryos, embryo culture duration and the mode of endometrial preparation are independent predictive factors of frozen embryo transfer outcomes.


Assuntos
Coeficiente de Natalidade , Criopreservação , Transferência Embrionária , Feminino , Humanos , Gravidez , Nascido Vivo , Recuperação de Oócitos , Taxa de Gravidez , Estudos Retrospectivos
2.
J Clin Med ; 10(17)2021 Aug 30.
Artigo em Inglês | MEDLINE | ID: mdl-34501340

RESUMO

We aimed to determine whether a functional link with impact on female ovarian reserve exists between FMR1 expression and expression ratios of AKT/mTOR signaling genes in human granulosa cells in vivo, as suggested from prior in vitro data. Three hundred and nine women, who were classified as normal (NOR; n = 225) and poor (POR; n = 84) responders based on their ovarian reserve, were recruited during stimulation for assisted reproductive techniques. Expressions of FMR1 and of key genes of the AKT/mTOR and AKT/FOXO1/3 signaling pathways were comparatively analyzed in their granulosa cells. FMR1 expression in granulosa cells of NOR and POR correlated significantly with AKT1, TSC2, mTOR, and S6K expression. No correlation was found between FMR1 and FOXO1 in all, and FOXO3 expression in POR, patients. AKT1 expression was significantly higher and FOXO1 expression lower in POR samples, whereas AKT1 expression was lower and FOXO1 expression was higher in NOR samples. In human native granulosa cells, FMR1 expression significantly correlated with the expression of key genes of the AKT/mTOR signaling pathway, but not with the FOXO1/3 signaling pathway. Our data point to a functional link between FMR1 expression and expression of the AKT/mTOR signaling pathway genes controlling human follicular maturation.

3.
Reprod Sci ; 28(7): 1866-1873, 2021 07.
Artigo em Inglês | MEDLINE | ID: mdl-33151525

RESUMO

In order to improve ART outcome, non-invasive embryo assessment is gaining more and more attention. Therefore, the aim of this study is to determine the consecutive implantation potential via the secretome between blastocysts with or without implantation and to analyse possible interactions between these differentially expressed proteins. In this prospective study, 69 spent culture media from blastocysts transferred at day 5 were collected from patients undergoing IVF/ICSI treatment in a single IVF centre between April 2015 and November 2018 after informed consent and analysed individually. Exclusion criteria were the absence of informed consent, PCOS, endometriosis and maternal age > 42 years. Dependent on the treatment outcome, media were subsequently divided into two groups: from embryos who implanted successfully (n = 37) and from embryos without implantation (n = 32). Ninety-two proteins were measured simultaneously using the proximity extension assay (PEA) technology with the Olink® CVD III panel employing oligonucleotide-labelled antibodies. Statistical analysis was performed using the Kolmogorov-Smirnov test, Student's t test, the Mann-Whitney U test and Fisher's exact test. Media from implanted blastocysts showed significantly higher expression of EPHB4, ALCAM, CSTB, BMH, TIMP4, CCL24, SELE, FAS, JAM-A, PON3, PDGF-A, vWF and PECAM-1 compared with media from blastocysts without subsequent implantation. The highest relative expression change could be demonstrated for PECAM-1 and TIMP4. PECAM-1, SELE and vWF were co-expressed. Especially EPHB 4, SELE, ALCAM, MCP-1, CCL24, FAS, JAM-A and PDGF-A have already been described in early embryonic development and metabolism. Therefore, these proteins together with PECAM-1 indicate possible biomarkers for non-invasive embryo assessment in the future. However, due to the innovative methodology, defining a threshold for the use as biomarkers remains to be assessed.


Assuntos
Técnicas de Cultura Embrionária/métodos , Desenvolvimento Embrionário/fisiologia , Fertilização in vitro/métodos , Injeções de Esperma Intracitoplásmicas , Adulto , Meios de Cultura , Transferência Embrionária/métodos , Feminino , Humanos , Gravidez , Estudos Prospectivos
4.
Arch Gynecol Obstet ; 302(4): 1019-1024, 2020 10.
Artigo em Inglês | MEDLINE | ID: mdl-32556511

RESUMO

PURPOSE: Female fertility preservation prior to gonadotoxic therapies can be achieved by the cryopreservation of ovarian cortical tissue. Immature oocytes may be recovered during the preparation, matured in vitro and lead to live births, thereby providing an additional option for fertility preservation. The purpose of this study was to test the feasibility of this approach in a setting with unilateral biopsy of a small piece of ovarian tissue and minimal tissue preparation prior to shipment to an external cryobank. METHODS: A prospective observational clinical study in an academic center was performed from January 2018 through December 2019. Ovarian tissue was obtained laparoscopically. Immature oocytes were recovered by minimal preparation of the tissue before shipment to an external cryobank for cryopreservation. In vitro maturation was performed on recovered immature oocytes. RESULTS: Twelve patients were enrolled. Immature oocytes could be recovered for all. The maturation rate was 38.9% (n = 14/36). Metaphase II (MII) were either directly used for intracytoplasmic sperm injection (ICSI) with a fertilization rate of 66.6% (n = 4/6) or vitrified (n = 8). PNs were cryopreserved (n = 4). Vitrified MII were warmed with a post-warming vitality rate of 75.0% (n = 3/4) and used for ICSI with a fertilization rate of 33.3% (n = 1/3). CONCLUSIONS: Immature oocytes can be successfully retrieved from ovarian tissue through minimal tissue preparation prior to shipment to a cryobank, matured in vitro, fertilized and cryopreserved for potential future fertility treatments. The total number of oocytes available for fertility preservation can be increased even without controlled ovarian stimulation in a situation where only ovarian biopsy for cryopreservation is performed. TRIAL REGISTRATION: German Clinical Trials Register (DRKS), DRKS00013170. Registered 11 December 2017, https://www.drks.de/drks_web/navigate.do?navigationId=trial.HTML&TRIAL_ID=DRKS00013170 .


Assuntos
Criopreservação/métodos , Preservação da Fertilidade/métodos , Técnicas de Maturação in Vitro de Oócitos/métodos , Recuperação de Oócitos/métodos , Oócitos/fisiologia , Adulto , Feminino , Humanos , Estudos Prospectivos , Adulto Jovem
5.
Reprod Sci ; 25(8): 1279-1285, 2018 08.
Artigo em Inglês | MEDLINE | ID: mdl-29141509

RESUMO

INTRODUCTION: Time-lapse technology allows almost continuous noninvasive assessment of embryonic development. It was shown previously that relative kinetics defining cleavage synchronicity are better predictors of blastocyst quality than absolute time points. This study aims to compare relative kinetics in embryos from patients with and without endometriosis. METHODS: Time-lapse data were collected retrospectively from 596 patients undergoing infertility treatment for in vitro fertilization from January 2011 to July 2016. Four hundred twenty-eight patients with confounding comorbidities (ie, polycystic ovary syndrome, pathological spermiogram in the included cycle, numerical/structural genetic abnormalities, preimplantation genetic screening performed) or incomplete data sets were excluded. Of the 168 included patients, 72 (42.9%) had endometriosis. Indications for in vitro fertilization of controls were tubal factor, unexplained infertility, or prolonged infertility. Relative kinetics were calculated as defined previously: cleavage synchronicity (CS)2-8=((t3-t2) + (t5-t4))/(t8-t2), CS4-8=(t8-t5)/(t8-t4), CS2-4=(t4-t3)/(t4-t2), DNA replication time ratio (DR)=(t3-t2)/(t5-t3). In women with more than one embryo, the median time was analyzed. RESULTS: Median age, body mass index, smoking status, and AMH levels were similar in both groups. Embryos from patients with endometriosis showed poorer relative kinetics. The relative time CS2-8 was decreased in embryos from patients with endometriosis (0.7 [0.0-0.93] vs 0.8 [0.0-0.94], P < .05) and CS4-8 was increased (0.4 [0.0-1.0] vs 0.3 [0.0-1.0], P < .05). The less powerful diagnostic relative kinetic parameters (CS2-4 and DR) were not significantly different. CONCLUSIONS: Embryos from patients with endometriosis presented with altered relative kinetics suggesting poorer embryo quality. These findings support recently published data demonstrating reduced oocyte quality in patients with endometriosis which is one possible explanation for their poor response to fertility treatment.


Assuntos
Desenvolvimento Embrionário , Endometriose/embriologia , Imagem com Lapso de Tempo/métodos , Adulto , Endometriose/complicações , Feminino , Fertilização in vitro , Humanos , Infertilidade Feminina/complicações , Pessoa de Meia-Idade , Oócitos/fisiologia , Estudos Retrospectivos
6.
Reprod Biol Endocrinol ; 15(1): 37, 2017 May 08.
Artigo em Inglês | MEDLINE | ID: mdl-28482910

RESUMO

BACKGROUND: The aim of this study was to compare pregnancy rates in patients undergoing IVF/ICSI with embryo transfer after 4 and 5 days of culture in a closed incubation system with integrated time-lapse imaging. METHODS: Out of n = 2207 in vitro fertilization (IVF)/ intracytoplasmic sperm injection (ICSI) cycles performed between January 2011 and April 2016 at a tertiary referral university hospital, a total of n = 599 IVF/ICSI cycles with prolonged embryo culture in an integrated time-lapse system (EmbryoScope© (Vitrolife)) until day 4 or 5 were retrospectively analyzed with regard to embryo morphology and pregnancy rates. RESULTS: A transfer on day 5 compared to a transfer on day 4 did not result in higher implantation and clinical pregnancy rates (IR 29.4% on day 4 versus 33.0% on day 5, p = 0.310; CPR 45.2% on day 4 versus 45.7% on day 5, p = 1.0). The percentage of ideal embryos transferred on day 4 was comparable to the rate of ideal embryos transferred on day 5 (41.6% versus 44.1%, p = 0.508). However, on day 4 a significantly higher number of embryos was transferred (1.92 on day 4 versus 1.84 on day 5, p = 0.023), which did not result in higher rates of multiple pregnancies. CONCLUSIONS: Pregnancy rates in IVF/ICSI cycles with integrated time-lapse incubation and transfer on day 4 and 5 are comparable. This finding provides the clinician, IVF laboratory and patient with more flexibility. TRIAL REGISTRATION: This study was retrospectively registered by the local ethics committee of the University of Heidelberg on December 19, 2016 (registration number S-649/2016).


Assuntos
Técnicas de Cultura Embrionária , Transferência Embrionária/métodos , Incubadoras , Infertilidade Feminina/terapia , Taxa de Gravidez , Imagem com Lapso de Tempo , Adulto , Células Cultivadas , Fase de Clivagem do Zigoto/citologia , Fase de Clivagem do Zigoto/fisiologia , Técnicas de Cultura Embrionária/instrumentação , Técnicas de Cultura Embrionária/métodos , Implantação do Embrião , Transferência Embrionária/estatística & dados numéricos , Feminino , Fertilização in vitro , Fetoscópios , Humanos , Infertilidade Feminina/epidemiologia , Gravidez , Estudos Retrospectivos , Injeções de Esperma Intracitoplásmicas , Fatores de Tempo , Imagem com Lapso de Tempo/instrumentação , Imagem com Lapso de Tempo/métodos
7.
Fertil Steril ; 107(3): 606-612.e3, 2017 03.
Artigo em Inglês | MEDLINE | ID: mdl-28139236

RESUMO

OBJECTIVE: To study the impact of in vitro maturation (IVM) on embryonal development with the use of time-lapse imaging. DESIGN: Retrospective case-control study. SETTING: University hospital. PATIENT(S): In total, 294 embryos were cultured after intracytoplasmic sperm injection treatment of three groups: patients with polycystic ovary syndrome (PCOS) and IVM (n = 105; group 1 [G1]), patients after conventional stimulation without PCOS (n = 115; G2) and with PCOS (n = 74; G3). In total, 171 embryos were finally analyzed (57 G1, 65 G2, and 49 G3). INTERVENTION(S): Data of 23 PCOS patients (30 IVM cycles) from January 2012 to July 2015 were matched according to age and number of oocytes to patients after conventional stimulation without PCOS (n = 30; 30 cycles) and with PCOS (n = 16; 19 cycles). Markers of embryo development were analyzed at different time points. Pregnancy rates (PRs) and live birth rates (LBRs) were recorded. MAIN OUTCOME MEASURE(S): Morphokinetic differences in embryo development after IVM compared with conventional stimulation with or without PCOS. RESULT(S): The rate of good-quality embryos was significantly lower in G1. Embryo development in G1 was significantly accelerated to the time of appearance of two pronuclei but slowed down by the time of reaching 6-cell stage and remained slower compared with embryos of G2 and G3. PRs as well as LBRs did not differ significantly among the study groups. CONCLUSION(S): Although growth dynamics of embryos from G1 differ from G2 and G3 and the rate of good-quality embryos was lower in IVM embryos, PRs and LBRs did not differ significantly.


Assuntos
Blastocisto/fisiologia , Técnicas de Maturação in Vitro de Oócitos , Infertilidade/terapia , Meiose , Microscopia de Vídeo , Síndrome do Ovário Policístico/complicações , Injeções de Esperma Intracitoplásmicas , Imagem com Lapso de Tempo/métodos , Adulto , Técnicas de Cultura Embrionária , Transferência Embrionária , Desenvolvimento Embrionário , Feminino , Fertilidade , Hospitais Universitários , Humanos , Infertilidade/diagnóstico , Infertilidade/etiologia , Infertilidade/fisiopatologia , Cinética , Nascido Vivo , Síndrome do Ovário Policístico/diagnóstico , Síndrome do Ovário Policístico/fisiopatologia , Gravidez , Taxa de Gravidez , Estudos Retrospectivos , Injeções de Esperma Intracitoplásmicas/efeitos adversos , Resultado do Tratamento , Adulto Jovem
8.
EMBO Rep ; 16(8): 1005-21, 2015 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-26142281

RESUMO

Mammalian development begins with the segregation of embryonic and extra-embryonic lineages in the blastocyst. Recent studies revealed cell-to-cell gene expression heterogeneity and dynamic cell rearrangements during mouse blastocyst formation. Thus, mechanistic understanding of lineage specification requires quantitative description of gene expression dynamics at a single-cell resolution in living embryos. However, only a few fluorescent gene expression reporter mice are available and quantitative live image analysis is limited so far. Here, we carried out a fluorescence gene-trap screen and established reporter mice expressing Venus specifically in the first lineages. Lineage tracking, quantitative gene expression and cell position analyses allowed us to build a comprehensive lineage map of mouse pre-implantation development. Our systematic analysis revealed that, contrary to the available models, the timing and mechanism of lineage specification may be distinct between the trophectoderm and the inner cell mass. While expression of our trophectoderm-specific lineage marker is upregulated in outside cells upon asymmetric divisions at 8- and 16-cell stages, the inside-specific upregulation of the inner-cell-mass marker only becomes evident at the 64-cell stage. This study thus provides a framework toward systems-level understanding of embryogenesis marked by high dynamicity and stochastic variability.


Assuntos
Blastocisto/fisiologia , Linhagem da Célula , Desenvolvimento Embrionário , Animais , Implantação do Embrião , Regulação da Expressão Gênica no Desenvolvimento , Genes Reporter , Microscopia Intravital , Camundongos
9.
J Biol Chem ; 287(30): 25361-9, 2012 Jul 20.
Artigo em Inglês | MEDLINE | ID: mdl-22605343

RESUMO

The study of chromatin and its regulators is key to understanding and manipulating transcription. We previously exploited the Krüppel-associated box (KRAB) transcriptional repressor domain, present in hundreds of vertebrate-specific zinc finger proteins, to assess the effect of its binding to gene bodies. These experiments revealed that the ectopic and doxycycline (dox)-controlled tet repressor KRAB fusion protein (tTRKRAB) can induce reversible and long-range silencing of cellular promoters. Here, we extend this system to in vivo applications and use tTRKRAB to achieve externally controllable repression of an endogenous mouse locus. We employed lentiviral-mediated transgenesis with promoterless TetO-containing gene traps to engineer a mouse line where the endogenous kinesin family member 2A (Kif2A) promoter drives a YFP reporter gene. When these mice were crossed to animals expressing the TetO-binding tTRKRAB repressor, this regulator was recruited to the Kif2A locus, and YFP expression was reduced. This effect was reversed when dox was given to embryos or adult mice, demonstrating that the cellular Kif2A promoter was only silenced upon repressor binding. Molecular analyses confirmed that tTRKRAB induced transcriptional repression through the spread of H3K9me3-containing heterochromatin, without DNA methylation of the trapped Kif2A promoter. Therefore, we demonstrate that targeting of tTRKRAB to a gene body in vivo results in reversible transcriptional repression through the spreading of facultative heterochromatin. This finding not only sheds light on KRAB-mediated transcriptional processes, but also suggests approaches for the externally controllable and reversible modulation of chromatin and transcription in vivo.


Assuntos
Proteínas de Transporte/metabolismo , Montagem e Desmontagem da Cromatina/fisiologia , Loci Gênicos/fisiologia , Proteínas Nucleares/metabolismo , Regiões Promotoras Genéticas/fisiologia , Proteínas Repressoras/metabolismo , Transcrição Gênica/fisiologia , Animais , Proteínas de Transporte/genética , Heterocromatina/genética , Heterocromatina/metabolismo , Cinesinas/biossíntese , Cinesinas/genética , Camundongos , Camundongos Transgênicos , Proteínas Nucleares/genética , Estrutura Terciária de Proteína , Proteínas Repressoras/biossíntese , Proteínas Repressoras/genética
10.
Dev Biol ; 319(2): 248-57, 2008 Jul 15.
Artigo em Inglês | MEDLINE | ID: mdl-18534570

RESUMO

The vertebrate pituitary gland is a key endocrine control organ that contains six distinct hormone secreting cell types. In this study, we analyzed the role of direct cell-to-cell Delta-Notch signaling in zebrafish anterior pituitary cell type specification. We demonstrate that initial formation of the anterior pituitary placode is independent of Notch signaling. Later however, loss of Notch signaling in mind bomb (mib) mutant embryos or by DAPT treatment leads to increased numbers of lactotropes and loss of corticotropes in the anterior pars distalis (APD), increased number of thyrotropes and loss of somatotrope cell types in the posterior pars distalis (PPD), and fewer melanotropes in the posterior region of the adenohypophysis, the pars intermedia (PI). Conversely, Notch gain of function leads to the opposite result, loss of lactotrope and thyrotrope cell specification, and an increased number of corticotropes, melanotropes, and gonadotropes in the pituitary. Our results suggest that Notch acts on placodal cells, presumably as a permissive signal, to regulate progenitor cell specification to hormone secreting cell types. We propose that Notch mediated lateral inhibition regulates the relative numbers of specified hormone cell types in the three pituitary subdomains.


Assuntos
Adeno-Hipófise/citologia , Adeno-Hipófise/fisiologia , Receptores Notch/genética , Proteínas de Peixe-Zebra/genética , Animais , Diferenciação Celular/fisiologia , Embrião não Mamífero/fisiologia , Desenvolvimento Embrionário , Regulação da Expressão Gênica no Desenvolvimento , Imuno-Histoquímica , Hibridização In Situ , Adeno-Hipófise/enzimologia , RNA Mensageiro/genética , Transdução de Sinais , Peixe-Zebra/genética
11.
Cells Tissues Organs ; 188(1-2): 46-51, 2008.
Artigo em Inglês | MEDLINE | ID: mdl-18303240

RESUMO

Mammalian preimplantation development serves to form a blastocyst, a structure that is able to implant into the mother's uterus to support further development of the embryo proper. In the developing conceptus, the first differentiation events separate the epithelial trophectoderm from the inner cell mass, which is comprised of the primitive endoderm and embryo-generating epiblast. Although the process of blastocyst formation may appear simple, its morphogenesis and the mechanism(s) of lineage specification are not yet fully understood. Here we discuss findings that suggest the involvement of stochastic processes and the influence of external cues in the patterning process during mouse preimplantation development.


Assuntos
Blastocisto/citologia , Blastocisto/metabolismo , Padronização Corporal , Animais , Massa Celular Interna do Blastocisto/citologia , Divisão Celular , Linhagem da Célula , Desenvolvimento Embrionário , Camundongos , Modelos Biológicos , Processos Estocásticos
12.
Development ; 134(23): 4219-31, 2007 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-17978007

RESUMO

Mouse pre-implantation development gives rise to the blastocyst, which is made up of at least three distinct cell types: the trophectoderm (TE) that surrounds a cavity, and an inner cell mass (ICM) comprising the primitive endoderm (PE) and epiblast (EPI). However, the underlying mechanisms involved in patterning the cleavage-stage embryo are still unresolved. By analyzing the distribution of the transcription factors Oct4 (Pou5f1), Cdx2 and Nanog at precisely defined stages in pre-implantation development, we were able to identify critical events leading to the divergence of TE, EPI and PE lineages. We found that Oct4 is present in all cells until late blastocyst, gradually disappearing from the TE thereafter. The expression patterns of both Cdx2 and Nanog exhibit two specific phases, culminating in their restriction to TE and EPI, respectively. In the first phase, starting after compaction, blastomeres show highly variable Cdx2 and Nanog protein levels. Importantly, the variability in Nanog levels is independent of position within the morula, whereas Cdx2 variability may originate from asymmetric cell divisions at the 8-cell stage in a non-stereotypic way. Furthermore, there is initially no reciprocal relationship between Cdx2 and Oct4 or between Cdx2 and Nanog protein levels. In the second phase, a definite pattern is established, possibly by a sorting process that accommodates intrinsic and extrinsic cues. Based on these results, we propose a model in which early embryonic mouse patterning includes stochastic processes, consistent with the highly regulative capacity of the embryo. This may represent a feature unique to early mammalian development.


Assuntos
Blastocisto/fisiologia , Blastômeros/fisiologia , Padronização Corporal/fisiologia , Embrião de Mamíferos/fisiologia , Desenvolvimento Embrionário , Animais , Blastômeros/citologia , Fator de Transcrição CDX2 , Adesão Celular , Diferenciação Celular , Divisão Celular , Embrião de Mamíferos/citologia , Endoderma/crescimento & desenvolvimento , Feminino , Proteínas de Homeodomínio/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C3H , Camundongos Endogâmicos C57BL , Morfogênese , Gravidez , Processos Estocásticos , Fatores de Transcrição/metabolismo
13.
PLoS Biol ; 4(5): e135, 2006 May.
Artigo em Inglês | MEDLINE | ID: mdl-16620153

RESUMO

Knowledge about the mechanism that establishes embryonic polarity is fundamental in understanding mammalian development. In re-addressing several controversial claims, we recently proposed a model in which mouse embryonic polarity is not specified until the blastocyst stage. Before fertilization, the fully differentiated oocyte has been characterized as "polarized," and we indeed observed that the sperm preferentially enters the polar body half. Here we show that preferential sperm entry is not due to an intrinsic polarity of the oocyte, since fertilization takes place uniformly when the zona pellucida is removed. We suggest that the term "asymmetry" denotes morphological differences, whereas "polarity" in addition implies developmental consequences. Thus, the mouse oocyte can be considered "asymmetric" but "non-polarized." The penetration through the zona pellucida is also random, and a significant proportion of sperm binds to the oocyte membrane at a point distant from the zona penetration site. Time-lapse recordings confirmed that sperm swim around the perivitelline space before fertilization. Experimental enlargement of the perivitelline space in the non-polar body half increased the regional probability of fertilization. Based on these experiments, we propose a model in which the space asymmetry exerted by the first polar body and the zona pellucida directs sperm entry preferentially to the polar body half, with no need for oocyte polarity.


Assuntos
Oócitos/citologia , Oócitos/fisiologia , Interações Espermatozoide-Óvulo/fisiologia , Espermatozoides/fisiologia , Animais , Polaridade Celular , Feminino , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Zona Pelúcida
14.
Development ; 132(7): 1579-90, 2005 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-15728669

RESUMO

Hedgehog signaling is required for formation and patterning of the anterior pituitary gland. However, the role of Hedgehog in pituitary precursor cell specification and subsequent placode formation is not well understood. We analyzed pituitary precursor cell lineages and find that pitx3 and distal-less3b (dlx3b) expression domains define lens and pituitary precursor positions. We show that pitx3 is required for pituitary pre-placode formation and cell specification, whereas dlx3b and dlx4b are required to restrict pituitary placode size. In smoothened mutant embryos that cannot transduce Hedgehog signals, median pituitary precursors are mis-specified and form an ectopic lens. Moreover, overexpression of sonic hedgehog (shh) blocks lens formation, and derivatives of lens precursors express genes characteristic of pituitary cells. However, overexpression of shh does not increase median pituitary placode size nor does it upregulate patched (ptc) expression in pituitary precursors during early somitogenesis. Our study suggests that by the end of gastrulation, pitx3-expressing cells constitute an equivalence domain of cells that can form either pituitary or lens, and that a non-Hedgehog signal initially specifies this placodal field. During mid-somitogenesis, Hedgehog then acts on the established median placode as a necessary and sufficient signal to specify pituitary cell types.


Assuntos
Proteínas de Homeodomínio/genética , Cristalino/embriologia , Adeno-Hipófise/embriologia , Proteínas de Peixe-Zebra/genética , Animais , Diferenciação Celular/fisiologia , DNA Complementar/metabolismo , Proteínas de Homeodomínio/biossíntese , Cristalino/citologia , Cristalino/metabolismo , Mutação , Adeno-Hipófise/citologia , Adeno-Hipófise/metabolismo , Receptores Acoplados a Proteínas G/genética , Receptores Acoplados a Proteínas G/metabolismo , Receptor Smoothened , Peixe-Zebra/embriologia , Peixe-Zebra/metabolismo , Proteínas de Peixe-Zebra/biossíntese , Proteínas de Peixe-Zebra/metabolismo
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