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1.
Gen Comp Endocrinol ; 285: 113249, 2020 01 01.
Artigo em Inglês | MEDLINE | ID: mdl-31445010

RESUMO

The objective of this study was to document the expression and functional role of BMPs in the placental (caruncle; CAR, cotyledon; COT) during different stages of pregnancy in water buffalo. Samples collected from Early pregnancy 1 (EP1); Early pregnancy 2 (EP2), Mid pregnancy (MP), Late pregnancy (LP) while the third stage of oestrus cycle (NP) was taken as control. Also, the synergistic role of BMP4/BMP7 or combination on mRNA expression of vWF, PCNA, StAR, CYP11A1, 3ßHSD, and BAX were studied in trophoblast cells cultured (TCC) during an early stage. The qPCR and immunoblotting studies revealed that BMP2, BMPR1A, BMPR1B, and BMPR2 mRNA level was significantly (p < 0.05) upregulated during early pregnancy in COTs while in CARs it was significantly upregulated (p < 0.05) during all the stages of pregnancy.BMP4 mRNA level was significantly upregulated (p < 0.05) during early pregnancy in COTs as well as in CARs. BMP6 expression was significantly upregulated (p < 0.05) during early and late stages of pregnancy. BMP7 mRNA level was upregulated (p < 0.05) during the late stage of pregnancy in COTs. At 100 ng/ml, the BMP4 maximally stimulated the transcripts of StAR, CYP11A1, and 3ßHSD while BMP7 maximally stimulated the transcripts of 3ßHSD that paralleled with P4 accretion in the media (P < 0.05). BMP4 as well as BMP7 upregulated the transcripts of PCNA, vWF, and downregulated BAX in the TCC (P < 0.05). In conclusion, BMPs are expressed in a regulated manner with stage-specific differences in the placenta and promotes the angiogenesis, proliferation, cell survivability, and steroidogenesis thereby regulating placental function in an autocrine/paracrine manner in water buffalo.


Assuntos
Proteínas Morfogenéticas Ósseas/genética , Búfalos/genética , Regulação da Expressão Gênica no Desenvolvimento , Placenta/metabolismo , Animais , Proteínas Morfogenéticas Ósseas/metabolismo , Sobrevivência Celular , Feminino , Neovascularização Fisiológica/genética , Placenta/irrigação sanguínea , Gravidez , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Fatores de Tempo , Trofoblastos/citologia
2.
Nat Commun ; 10(1): 4749, 2019 10 18.
Artigo em Inglês | MEDLINE | ID: mdl-31628347

RESUMO

Trophectoderm (TE) lineage development is pivotal for proper implantation, placentation, and healthy pregnancy. However, only a few TE-specific transcription factors (TFs) have been systematically characterized, hindering our understanding of the process. To elucidate regulatory mechanisms underlying TE development, here we map super-enhancers (SEs) in trophoblast stem cells (TSCs) as a model. We find both prominent TE-specific master TFs (Cdx2, Gata3, and Tead4), and >150 TFs that had not been previously implicated in TE lineage, that are SE-associated. Mapping targets of 27 SE-predicted TFs reveals a highly intertwined transcriptional regulatory circuitry. Intriguingly, SE-predicted TFs show 4 distinct expression patterns with dynamic alterations of their targets during TSC differentiation. Furthermore, depletion of a subset of TFs results in dysregulation of the markers for specialized cell types in placenta, suggesting a role during TE differentiation. Collectively, we characterize an expanded TE-specific regulatory network, providing a framework for understanding TE lineage development and placentation.


Assuntos
Células-Tronco Embrionárias/metabolismo , Elementos Facilitadores Genéticos/genética , Regulação da Expressão Gênica no Desenvolvimento , Redes Reguladoras de Genes , Trofoblastos/metabolismo , Animais , Diferenciação Celular/genética , Linhagem da Célula/genética , Embrião de Mamíferos/citologia , Embrião de Mamíferos/embriologia , Embrião de Mamíferos/metabolismo , Feminino , Perfilação da Expressão Gênica/métodos , Camundongos , Placentação/genética , Gravidez , Fatores de Transcrição/genética , Trofoblastos/citologia
3.
PLoS Biol ; 17(10): e3000187, 2019 10.
Artigo em Inglês | MEDLINE | ID: mdl-31596842

RESUMO

Multipotent trophoblasts undergo dynamic morphological movement and cellular differentiation after conceptus implantation to generate placenta. However, the mechanism controlling trophoblast development and differentiation during peri-implantation development in human remains elusive. In this study, we modeled human conceptus peri-implantation development from blastocyst to early postimplantation stages by using an in vitro coculture system and profiled the transcriptome of 476 individual trophoblast cells from these conceptuses. We revealed the genetic networks regulating peri-implantation trophoblast development. While determining when trophoblast differentiation happens, our bioinformatic analysis identified T-box transcription factor 3 (TBX3) as a key regulator for the differentiation of cytotrophoblast (CT) into syncytiotrophoblast (ST). The function of TBX3 in trophoblast differentiation is then validated by a loss-of-function experiment. In conclusion, our results provided a valuable resource to study the regulation of trophoblasts development and differentiation during human peri-implantation development.


Assuntos
Regulação da Expressão Gênica no Desenvolvimento , Redes Reguladoras de Genes , Modelos Biológicos , Proteínas com Domínio T/genética , Transcriptoma , Trofoblastos/metabolismo , Técnicas de Cultura de Células , Diferenciação Celular , Biologia Computacional/métodos , Implantação do Embrião/genética , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Análise de Célula Única , Proteínas com Domínio T/metabolismo , Trofoblastos/citologia , Zigoto
4.
Nat Commun ; 10(1): 4601, 2019 10 10.
Artigo em Inglês | MEDLINE | ID: mdl-31601798

RESUMO

During pregnancy, trophoblast cells sustain the maternal-fetal tolerance via expressing and secreting various chemokines and cytokines. Our previous study revealed the expression of interleukin-35 (IL-35) in human first-trimester trophoblasts. Here we show that IL-35 is expressed in both human first-trimester primary trophoblast cells and a trophoblast cell line. Trophoblast cells inhibit the proliferation of human naive conventional T cells (Tconv cells) and convert suppressed Tconv cells into iTR35 in an IL-35-dependent manner. Mechanistically, trophoblast cell derived IL-35 mediates its function through phosphorylation of STAT1 and STAT3. In vivo studies confirm that mice with immunologically spontaneous abortion have lower levels of IL-35 and iTR35 cells at the maternal-fetal interface, and neutralizing anti-IL-35 mAb enhances abortion rates. Meanwhile, exogenous IL-35 induces iTR35 and prevents immunological abortion. Our findings thus suggest that trophoblast cells have a critical function in preserving maternal-fetal tolerance via secreting IL-35 during pregnancy.


Assuntos
Interleucinas/metabolismo , Troca Materno-Fetal/fisiologia , Linfócitos T Reguladores/imunologia , Trofoblastos/citologia , Animais , Proliferação de Células , Feminino , Humanos , Tolerância Imunológica , Interleucinas/sangue , Interleucinas/imunologia , Interleucinas/farmacologia , Masculino , Troca Materno-Fetal/imunologia , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos DBA , Placenta/citologia , Placenta/imunologia , Gravidez , Proteínas Recombinantes/genética , Proteínas Recombinantes/imunologia , Proteínas Recombinantes/farmacologia , Linfócitos T/citologia , Linfócitos T/efeitos dos fármacos , Linfócitos T/imunologia , Linfócitos T Reguladores/metabolismo , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Trofoblastos/imunologia , Trofoblastos/metabolismo
5.
Nat Commun ; 10(1): 4155, 2019 09 13.
Artigo em Inglês | MEDLINE | ID: mdl-31519912

RESUMO

Zika virus (ZIKV) infection results in an increased risk of spontaneous abortion and poor intrauterine growth although the underlying mechanisms remain undetermined. Little is known about the impact of ZIKV infection during the earliest stages of pregnancy, at pre- and peri-implantation, because most current ZIKV pregnancy studies have focused on post-implantation stages. Here, we demonstrate that trophectoderm cells of pre-implantation human and mouse embryos can be infected with ZIKV, and propagate virus causing neural progenitor cell death. These findings are corroborated by the dose-dependent nature of ZIKV susceptibility of hESC-derived trophectoderm cells. Single blastocyst RNA-seq reveals key transcriptional changes upon ZIKV infection, including nervous system development, prior to commitment to the neural lineage. The pregnancy rate of mice is >50% lower in pre-implantation infection than infection at E4.5, demonstrating that pre-implantation ZIKV infection leads to miscarriage. Cumulatively, these data elucidate a previously unappreciated association of pre- and peri-implantation ZIKV infection and microcephaly.


Assuntos
Complicações Infecciosas na Gravidez/metabolismo , Infecção por Zika virus/complicações , Infecção por Zika virus/metabolismo , Zika virus/patogenicidade , Aborto Espontâneo/metabolismo , Aborto Espontâneo/fisiopatologia , Animais , Blastocisto/citologia , Blastocisto/metabolismo , Implantação do Embrião/fisiologia , Feminino , Desenvolvimento Fetal/genética , Desenvolvimento Fetal/fisiologia , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Gravidez , RNA Viral/genética , Pesquisa Médica Translacional/métodos , Trofoblastos/citologia , Trofoblastos/metabolismo
6.
Int J Mol Sci ; 20(18)2019 Sep 13.
Artigo em Inglês | MEDLINE | ID: mdl-31540219

RESUMO

During the peri-implantation period, multinucleated syncytia are formed in the sheep placenta. For over 20 years the scientific consensus has been that during trophoblast syncytialization in sheep, binucleate trophoblast giant cells (BNCs) differentiate from mononuclear trophoblast cells, and individual BNCs fuse with individual luminal epithelial (LE) cells to form trinucleate cells. These trophoblast-LE syncytial plaques then grow through continued BNC migration and fusion. Therefore, LE cells are thought to be incorporated into syncytial plaques. However, these ideas were based on electron microscopy studies, without benefit of molecular markers for BNC and LE cells to support conclusions. The aim of this study was to observe interactions between BNCs and uterine LE cells using immunohistochemical localization for molecular markers for BNCs and uterine LE cells. We performed immunofluorescence staining, laser capture microdissection, and TUNEL staining on the uterine-placental tissues of sheep during early placentation. We observed: (1) syncytial cells containing more than two nuclei within the trophoblast cell layer; (2) depolarized LE cells that express caspase 3 and stain positively for TUNEL; (3) engulfment of caspase 3-positive LE cells by trophoblast giant cells (TGCs) and empty spaces within the LE layer at sites of implantation; (4) rapid enlargement of syncytial plaques; and (5) E-cadherin and TUNEL-positive cells within the uterine stroma underlying degenerating LE was coincident with accumulation of CD45-positive cells at these sites. These data suggest that during early placentation: (1) fusion between trophoblasts is not limited to the formation of BNCs, and the term 'trophoblast giant cell (TGC)' may be appropriate; (2) LE cells undergo apoptosis; (3) apoptotic LE cells are eliminated by TGCs; (4) fusion is not limited to the incorporation of new BNCs but involves the lateral fusion between growing syncytial plaques; and (5) TGCs carry apoptotic LE cells away from the uterine-placental interface for elimination by immune cells within the stroma. These data indicate that uterine LE cells are not incorporated into syncytial plaques, but are engulfed and eliminated, and that early placentation in sheep is more similar to early placentation in humans than is currently understood in that both develop mononucleated cytotrophoblast and multinucleated syncytiotrophoblast layers of entirely placental origin. The elimination of LE cells by sheep TGCs might provide insights into elimination and penetration of LE cells during human embryo implantation.


Assuntos
Biomarcadores/metabolismo , Células Epiteliais/citologia , Células Gigantes/citologia , Placentação , Trofoblastos/citologia , Animais , Caderinas/metabolismo , Caspase 3/metabolismo , Diferenciação Celular , Fusão Celular , Movimento Celular , Células Epiteliais/metabolismo , Feminino , Células Gigantes/metabolismo , Imuno-Histoquímica , Queratinas/metabolismo , Antígenos Comuns de Leucócito/metabolismo , Placenta/citologia , Placenta/metabolismo , Gravidez , Ovinos , Trofoblastos/metabolismo
7.
Int J Mol Sci ; 20(18)2019 Sep 07.
Artigo em Inglês | MEDLINE | ID: mdl-31500240

RESUMO

Omega-3 fatty acids are important to pregnancy and neonatal development and health. One mechanism by which omega-3 fatty acids exert their protective effects is through serving as substrates for the generation of specialized pro-resolving lipid mediators (SPM) that potently limit and resolve inflammatory processes. We recently identified that SPM levels are increased in maternal blood at delivery as compared to umbilical cord blood, suggesting the placenta as a potential site of action for maternal SPM. To explore this hypothesis, we obtained human placental samples and stained for the SPM resolvin D2 (RvD2) receptor GPR18 via immunohistochemistry. In so doing, we identified GPR18 expression in placental vascular smooth muscle and extravillous trophoblasts of the placental tissues. Using in vitro culturing, we confirmed expression of GPR18 in these cell types and further identified that stimulation with RvD2 led to significantly altered responsiveness (cytoskeletal changes and pro-inflammatory cytokine production) to lipopolysaccharide inflammatory stimulation in human umbilical artery smooth muscle cells and placental trophoblasts. Taken together, these findings establish a role for SPM actions in human placental tissue.


Assuntos
Ácidos Docosa-Hexaenoicos/farmacologia , Músculo Liso Vascular/citologia , Receptores Acoplados a Proteínas-G/genética , Trofoblastos/citologia , Adulto , Células Cultivadas , Ácidos Graxos Ômega-3/farmacologia , Feminino , Regulação da Expressão Gênica/efeitos dos fármacos , Células Hep G2 , Humanos , Idade Materna , Músculo Liso Vascular/efeitos dos fármacos , Músculo Liso Vascular/metabolismo , Miócitos de Músculo Liso/citologia , Miócitos de Músculo Liso/efeitos dos fármacos , Miócitos de Músculo Liso/metabolismo , Placenta/citologia , Placenta/efeitos dos fármacos , Placenta/metabolismo , Gravidez , Receptores Acoplados a Proteínas-G/metabolismo , Trofoblastos/efeitos dos fármacos , Trofoblastos/metabolismo , Adulto Jovem
8.
Dis Markers ; 2019: 4976845, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31467616

RESUMO

Background: Preeclampsia (PE) is a pregnancy-related syndrome characterized by hypertension and proteinuria after the 20th week of gestation. The long noncoding RNAs (lncRNAs) have been recently discovered for their roles in the pathogenesis of PE. This study is aimed at determining the expression of lncRNA MIR503 host gene (MIR503HG) in PE placental tissues and exploring the molecular mechanism underlying MIR503HG-mediated trophoblast cell proliferation, invasion, and migration. Methods: The expression level of MIR503HG in placental tissues, HTR-8/SVneo, and JEG3 cells was determined by quantitative real-time PCR; western blot detected the relevant protein expression levels in HTR-8/SVneo and JEG3 cells; flow cytometry determined cell apoptosis and cell cycle of HTR-8/SVneo and JEG3 cells; trophoblast cell proliferation, invasion, and migration of HTR-8/SVneo and JEG3 cells were measured by CCK-8, transwell invasion, and wound healing assays, respectively. Results: The highly expressed MIR503HG was detected in PE placental tissues compared to normal placental tissues. MIR503HG overexpression suppressed cell proliferation, invasion, and migration of HTR-8/SVneo and JEG3 cells, while knockdown of MIR503HG increased trophoblast cell proliferation, invasion, and migration. Flow cytometry results showed that MIR503HG overexpression induced apoptosis and caused cell cycle arrest at the G0/G1 phase, while MIR503HG knockdown had the opposite actions in HTR-8/SVneo and JEG3 cells. Western blot assay results showed that MIR503HG overexpression suppressed the matrix metalloproteinase-2/-9 and the snail protein expression and increased the E-cadherin expression in trophoblast cells. In addition, MIR503HG overexpression suppressed the NF-κB signaling pathway by inhibiting the phosphorylation of IκBα and the nuclear translocation of NF-κB signaling subunit p65. On the other hand, MIR503HG knockdown played an opposite role in these protein expression levels. Conclusion: Our results showed that MIR503HG inhibited the proliferation, invasion, and migration of HTR-8/SVneo and JEG3 cells, which may be related to the pathogenesis of PE.


Assuntos
Movimento Celular , Proliferação de Células , Metaloproteinases da Matriz/metabolismo , NF-kappa B/metabolismo , Pré-Eclâmpsia/metabolismo , RNA Longo não Codificante/genética , Trofoblastos/metabolismo , Linhagem Celular Tumoral , Células Cultivadas , Feminino , Humanos , Metaloproteinases da Matriz/genética , Pré-Eclâmpsia/genética , Gravidez , RNA Longo não Codificante/metabolismo , Transdução de Sinais , Fatores de Transcrição da Família Snail/metabolismo , Trofoblastos/citologia
9.
Pestic Biochem Physiol ; 159: 144-153, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31400776

RESUMO

Ivermectin is a pesticide that has been used for over 30 years in livestock. Although there are a number of studies on the therapeutic potential of ivermectin, little is known about the effects of the drug during the early stage of pregnancy. In this study, we investigated the detrimental effects of ivermectin on porcine trophectoderm (pTr) and uterine luminal epithelial (pLE) cells. Ivermectin not only inhibited the proliferation of both cells via the regulation of cell cycle-associated genes, but also induced apoptosis in pTr and pLE cells. We also verified its effect on mitochondrial dysfunction as shown by loss of mitochondrial membrane potential, mitochondrial Ca2+ overload, and reactive oxygen species (ROS) generation in pTr and pLE cells. As a mechanistic approach, we evaluated ivermectin-mediated cell signaling interactions including PI3K, AKT and MAPK pathways. Overall, our results suggest that constant exposure to and accumulation of ivermectin may cause abnormal fetal morphogenesis and placentation during the early stages of pregnancy. Our results may further provide a comprehensive understanding of the detrimental effects of ivermectin during pregnancy and will contribute to the establishment of a complete safety profile for ivermectin and its association with environmental pollution and public health in humans and livestock.


Assuntos
Apoptose/efeitos dos fármacos , Cálcio/metabolismo , Células Epiteliais/citologia , Células Epiteliais/efeitos dos fármacos , Ivermectina/farmacologia , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Espécies Reativas de Oxigênio/metabolismo , Útero/citologia , Animais , Proliferação de Células/efeitos dos fármacos , Feminino , Suínos , Trofoblastos/citologia , Trofoblastos/efeitos dos fármacos
10.
Endocrinology ; 160(9): 2189-2203, 2019 09 01.
Artigo em Inglês | MEDLINE | ID: mdl-31294776

RESUMO

Defective placental implantation and vascularization with accompanying hypoxia contribute to preeclampsia (PE), a leading cause of maternal and neonatal morbidity and mortality. Genetic and epigenetic mechanisms underlying differentiation of proliferative cytotrophoblasts (CytTs) to multinucleated syncytiotrophoblast (SynT) are incompletely defined. The SynT performs key functions in nutrient and gas exchange, hormone production, and protection of the fetus from rejection by the maternal immune system. In this study, we used chromatin immunoprecipitation sequencing of midgestation human trophoblasts before CytT and after SynT differentiation in primary culture to analyze changes in binding of RNA polymerase II (Pol II) and of active and repressive histone marks during SynT differentiation. Our findings reveal that increased Pol II binding to promoters of a subset of genes during trophoblast differentiation was closely correlated with active histone marks. This gene set was enriched in those controlling immune response and immune modulation, including interferon-induced tetratricopeptide repeat and placenta-specific glycoprotein gene family members. By contrast, genes downregulated during SynT differentiation included proinflammatory transcription factors ERG1, cFOS, and cJUN, as well as members of the NR4A orphan nuclear receptor subfamily, NUR77, NURR1, and NOR1. Downregulation of proinflammatory transcription factors upon SynT differentiation was associated with decreased promoter enrichment of endogenous H3K27Ac and H3K9Ac and enhanced binding of H3K9me3 and histone deacetylase 1. However, promoter enrichment of H3K27me3 was low in both CytT and SynT and was not altered with changes in gene expression. These findings provide important insight into mechanisms underlying human trophoblast differentiation and may identify therapeutic targets for placental disorders, such as PE.


Assuntos
Epigênese Genética , Regulação da Expressão Gênica , Trofoblastos/citologia , Diferenciação Celular , Fusão Celular , Células Cultivadas , Feminino , Histonas/metabolismo , Humanos , Membro 1 do Grupo A da Subfamília 4 de Receptores Nucleares/genética , Placenta/fisiologia , Gravidez , Regiões Promotoras Genéticas , RNA Polimerase II/metabolismo , Trofoblastos/metabolismo
11.
J Assist Reprod Genet ; 36(8): 1609-1621, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31292818

RESUMO

PURPOSE: This study was to evaluate if spent culture media (SCM) of embryos could be used as a non-invasive tool to achieve aneuploidy screening. Ploidy calls, as well as concordance rates between PGT-A results from trophectoderm (TE) and SCM, were compared. Clinical outcomes of single euploid transfers were also evaluated. METHODS: The study was conducted from March 2017 to June 2018 in a university-based ART center. SCM of day 3 to the day(s) of TE biopsy of all biopsied blastocysts were collected for testing. PGT-A results of SCM were compared with the standard results of TE, with clinical relevance and outcomes examined. RESULTS: NiPGT-A using SCM gave a sensitivity of 81.6%, specificity of 48.3%, positive predictive value of 82.6%, and negative predictive value of 46.7% in ploidy calling. The concordance rates for autosomes and sex determination were 62.1% and 82.4%, respectively. There were 14 single embryo transfer cycles of euploids as determined by TE biopsy. Clinical outcomes not only confirmed 3 false positive results from SCM but also reflected the true ploidy status of the transferred embryo in one case. If ploidy calls were dichotomized without mosaic embryos, the sensitivity and NPV would increase to 91.0% and 66.7% (p = 0.60 and p = 0.25), respectively. CONCLUSIONS: Cell-free DNA found in SCM could provide ploidy information of an embryo as in PGT-A from its TE. Given its potential to reflect the comprehensive chromosomal profile of the whole embryo, more research based on clinical outcomes is required to determine if SCM could be a reliable selection tool in PGT-A.


Assuntos
Aneuploidia , Meios de Cultura/metabolismo , Fertilização In Vitro , Testes Genéticos/métodos , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Diagnóstico Pré-Implantação/métodos , Trofoblastos/metabolismo , Técnicas de Cultura Embrionária , Feminino , Humanos , Recuperação de Oócitos , Indução da Ovulação , Gravidez , Estudos Prospectivos , Trofoblastos/citologia
12.
Bull Exp Biol Med ; 167(1): 169-176, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-31183653

RESUMO

We studied changes in angiogenesis during contact interaction of natural killer cells and endothelial cells in the presence of secretory products of trophoblast cells activated by various cytokines. Activated trophoblast regulates angiogenesis by producing soluble factors that affect endothelial cells either directly or indirectly through activation of proangiogenic activity of natural killer cells. A stimulating effect of the trophoblast supernatants activated by IL-1ß and an inhibitory effect of trophoblast supernatants activated by IL-6 and TGFß for the formation of tube-like structures by endothelial cells were revealed. During contact culturing, natural killer cells increased the length of tube-like structures formed by endothelial cells. The trophoblast activated by IL-1ß affects angiogenesis both directly through the production of proangiogenic factors and indirectly through activation of the proangiogenic potential of natural killer cells. Trophoblast activated by IFNγ affects angiogenesis only by stimulating the proangiogenic potential of natural killer cells. Under conditions of contact interaction of natural killer cells and endothelial cells, soluble factors of trophoblast activated by IL-6 or TGFß attenuated the angiogenesis-stimulating effect of natural killer cells.


Assuntos
Células Endoteliais/citologia , Células Endoteliais/metabolismo , Células Matadoras Naturais/citologia , Células Matadoras Naturais/metabolismo , Trofoblastos/citologia , Trofoblastos/metabolismo , Comunicação Celular/fisiologia , Linhagem Celular , Linhagem Celular Tumoral , Feminino , Humanos , Interleucina-1beta/metabolismo , Interleucina-6/metabolismo , Neovascularização Patológica/patologia , Fator de Crescimento Transformador beta/metabolismo
13.
Zygote ; 27(4): 255-258, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31218974

RESUMO

This study aimed to optimize the derivation of trophectoderm from in vitro-produced camel embryos under feeder-free culture conditions using the basement membrane matrix Matrigel. Trophoblastic vesicles were obtained through mechanical microdissection of in vitro-produced camel (Camelus dromedarius) embryos. Supplementing the culture medium with 10 ng/ml of epidermal growth factor and 10 ng/ml fibroblast growth factor improved the attachment and subsequent outgrowths of cultured trophoblastic vesicles when compared with the control group and the groups supplemented individually with each growth factor. The expression levels of pluripotency genes octamer-binding transcription factor 4 (Oct4), sex determining region Y-box 2 (Sox2), myelocytomatosis proto-oncogene (c-Myc) and anti-apoptotic gene B-cell lymphoma 2 (Bcl2) were increased in trophoblastic vesicles supplemented with both growth factors when compared with the control group. Conversely, both growth factors decreased the expression of apoptotic genes tumour protein p53 (p53) and Bcl-2-associated X protein (Bax). To the best of our knowledge, this may be the first report describing the derivation of trophoblast stem cells from in vitro-produced camel embryos.


Assuntos
Diferenciação Celular/efeitos dos fármacos , Embrião de Mamíferos/efeitos dos fármacos , Células-Tronco Embrionárias/efeitos dos fármacos , Fator de Crescimento Epidérmico/farmacologia , Fator 2 de Crescimento de Fibroblastos/farmacologia , Trofoblastos/efeitos dos fármacos , Animais , Camelus , Diferenciação Celular/genética , Células Cultivadas , Sinergismo Farmacológico , Embrião de Mamíferos/citologia , Embrião de Mamíferos/metabolismo , Células-Tronco Embrionárias/citologia , Células-Tronco Embrionárias/metabolismo , Feminino , Regulação da Expressão Gênica no Desenvolvimento/efeitos dos fármacos , Fator 3 de Transcrição de Octâmero/genética , Proteínas Proto-Oncogênicas c-myb/genética , Fatores de Transcrição SOXB1/genética , Trofoblastos/citologia , Trofoblastos/metabolismo
14.
Prostaglandins Other Lipid Mediat ; 143: 106342, 2019 08.
Artigo em Inglês | MEDLINE | ID: mdl-31176799

RESUMO

Although FPR2 receptor is distributed in the endometrium and placenta, its function in human extravillous trophoblastic (TEV-1) cells still remains enigmatic. In this study, overexpression of FPR2 was performed in TEV-1 cells. Then, CCK8 transwell and wound healing assays were used to assess the cell proliferation, migration and invasion, respectively. The results showed that FPR2 overexpression significantly inhibited proliferation, invasion and migration in TEV-1 cells. In addition, FPR2 overexpression significantly decreased mRNA and protein levels of integrin-linked kinase (ILK), nuclear factor-kappa B (NF--κB), matrix metalloproteinase 9 (MMP9) and vascular endothelial growth factor (VEGF) in TEV-1 cells. These findings indicated that FPR2 overexpression alters proliferation, migration and invasion in human extravillous trophoblastic cellsthrough the ILK/NF-κB signaling pathway; ideal FPR2 levels are important for TEV-1 cells functions.


Assuntos
Movimento Celular , Receptores de Formil Peptídeo/metabolismo , Receptores de Lipoxinas/metabolismo , Trofoblastos/citologia , Linhagem Celular , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Humanos , Lipoxinas/farmacologia , Metaloproteinase 9 da Matriz/metabolismo , NF-kappa B/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , Transdução de Sinais/efeitos dos fármacos , Fator A de Crescimento do Endotélio Vascular/metabolismo
15.
Nat Cell Biol ; 21(6): 687-699, 2019 06.
Artigo em Inglês | MEDLINE | ID: mdl-31160711

RESUMO

We recently derived mouse expanded potential stem cells (EPSCs) from individual blastomeres by inhibiting the critical molecular pathways that predispose their differentiation. EPSCs had enriched molecular signatures of blastomeres and possessed developmental potency for all embryonic and extra-embryonic cell lineages. Here, we report the derivation of porcine EPSCs, which express key pluripotency genes, are genetically stable, permit genome editing, differentiate to derivatives of the three germ layers in chimeras and produce primordial germ cell-like cells in vitro. Under similar conditions, human embryonic stem cells and induced pluripotent stem cells can be converted, or somatic cells directly reprogrammed, to EPSCs that display the molecular and functional attributes reminiscent of porcine EPSCs. Importantly, trophoblast stem-cell-like cells can be generated from both human and porcine EPSCs. Our pathway-inhibition paradigm thus opens an avenue for generating mammalian pluripotent stem cells, and EPSCs present a unique cellular platform for translational research in biotechnology and regenerative medicine.


Assuntos
Diferenciação Celular/genética , Reprogramação Celular/genética , Células-Tronco Pluripotentes Induzidas/citologia , Células-Tronco Pluripotentes/citologia , Animais , Blastômeros/citologia , Blastômeros/metabolismo , Linhagem da Célula/genética , Células-Tronco Embrionárias/citologia , Camadas Germinativas/crescimento & desenvolvimento , Camadas Germinativas/metabolismo , Humanos , Camundongos , Medicina Regenerativa , Transdução de Sinais/genética , Suínos , Trofoblastos/citologia , Trofoblastos/metabolismo
16.
Nanoscale ; 11(25): 12285-12295, 2019 Jun 27.
Artigo em Inglês | MEDLINE | ID: mdl-31211316

RESUMO

A complex combination of trafficking and signalling occurs at the surface of the placenta. The system delivers maternal nutrients to the fetus and facilitates gaseous exchange, whilst mediating signal transduction to support and stimulate the growth of the placenta itself. IGF-I is acknowledged as a maternally-derived ligand important in the regulation of placental growth. Here we show that quantum dots bearing IGF can stimulate IGF receptor (IGF1R) phosphorylation in the syncytio- (maternal-facing) and cyto- (fetal-facing) trophoblast bilayer that forms the outer boundary of the placenta, in a distribution similar to the one resulting from exposure to a soluble ligand. The conjugates are internalised by a clathrin-dependent pathway and delivered to a syncytioplasmic compartment that differs from conventional late endosomes and lysosomes. Two discrete downstream responses are evident in different cellular compartments: phosphorylation of P70S6K in the non-proliferative syncytiotrophoblast and of AKT in the cytotrophoblast. Co-conjugation of IGF-quantum dots with an RGD-containing ligand permits penetration beyond the syncytium, into the cytoplasm of the underlying cytotrophoblast. These data reveal the existence of a trans-syncytial pathway that allows maternal mitotic signals to penetrate to the inner progenitor cells, which must proliferate to support placental and consequently fetal growth.


Assuntos
Endocitose , Fator de Crescimento Insulin-Like I/metabolismo , Pontos Quânticos/química , Trofoblastos/metabolismo , Adulto , Feminino , Humanos , Gravidez , Trofoblastos/citologia
17.
Hypertension ; 74(1): 145-153, 2019 07.
Artigo em Inglês | MEDLINE | ID: mdl-31079531

RESUMO

Preeclampsia is a hypertensive pregnancy disease associated with a massive increase in sFlt-1 (soluble form of the vascular endothelial growth factor 1) in the maternal circulation, responsible for angiogenic imbalance and endothelial dysfunction. Pilot studies suggest that extracorporeal apheresis may reduce circulating sFlt-1 and prolong pregnancy. Nonspecific apheresis systems have potential adverse effects because of the capture of many other molecules. Our concept is based on a specific and competitive apheresis approach using VEGF (vascular endothelial growth factor) functionalized magnetic beads to capture sFlt-1 while releasing endogenous PlGF (placental growth factor) to restore a physiological angiogenic balance. Magnetic beads were functionalized with VEGF to capture sFlt-1. Experiments were performed using PBS, conditioned media from human trophoblastic cells, and human plasma. The proof of concept was validated in dynamic conditions in a microfluidic device as an approach mimicking real apheresis. Magnetic beads were functionalized with VEGF and characterized to evaluate their surface ligand density and recognition capabilities. VEGF-coated magnetic beads proved to be an efficient support in capturing sFlt-1 and releasing PlGF. In static conditions, sFlt-1 concentration decreased by 33±13%, whereas PlGF concentration increased by 27±10%. In dynamic conditions, the performances were improved, with 40% reduction of sFlt-1 and up to 2-fold increase of free PlGF. The sFlt-1/PlGF ratio was reduced by 63% in the plasma of preeclamptic patients. Apheresis was also associated with VEGF release. A ligand-based approach using VEGF-coated beads is an effective approach to the capture of sFlt-1 and the release of endogenous PlGF. It offers new perspectives for the treatment of preeclampsia.


Assuntos
Dispositivos Lab-On-A-Chip , Pré-Eclâmpsia/terapia , Fator A de Crescimento do Endotélio Vascular/farmacologia , Receptor 1 de Fatores de Crescimento do Endotélio Vascular/metabolismo , Indutores da Angiogênese , Remoção de Componentes Sanguíneos/métodos , Velocidade do Fluxo Sanguíneo , Células Cultivadas , Feminino , Humanos , Técnicas In Vitro , Magnetismo/métodos , Projetos Piloto , Placenta/citologia , Pré-Eclâmpsia/patologia , Gravidez , Sensibilidade e Especificidade , Trofoblastos/citologia , Trofoblastos/fisiologia
18.
Cell Mol Life Sci ; 76(18): 3479-3496, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31049600

RESUMO

Abnormal placentation is considered as an underlying cause of various pregnancy complications such as miscarriage, preeclampsia and intrauterine growth restriction, the latter increasing the risk for the development of severe disorders in later life such as cardiovascular disease and type 2 diabetes. Despite their importance, the molecular mechanisms governing human placental formation and trophoblast cell lineage specification and differentiation have been poorly unravelled, mostly due to the lack of appropriate cellular model systems. However, over the past few years major progress has been made by establishing self-renewing human trophoblast stem cells and 3-dimensional organoids from human blastocysts and early placental tissues opening the path for detailed molecular investigations. Herein, we summarize the present knowledge about human placental development, its stem cells, progenitors and differentiated cell types in the trophoblast epithelium and the villous core. Anatomy of the early placenta, current model systems, and critical key regulatory factors and signalling cascades governing placentation will be elucidated. In this context, we will discuss the role of the developmental pathways Wingless and Notch, controlling trophoblast stemness/differentiation and formation of invasive trophoblast progenitors, respectively.


Assuntos
Placenta/metabolismo , Trofoblastos/metabolismo , Doenças Cardiovasculares/metabolismo , Doenças Cardiovasculares/patologia , Diferenciação Celular , Diabetes Mellitus Tipo 2/metabolismo , Diabetes Mellitus Tipo 2/patologia , Feminino , Humanos , Modelos Biológicos , Placenta/anatomia & histologia , Placentação , Gravidez , Transdução de Sinais , Trofoblastos/citologia
19.
Int J Mol Med ; 43(6): 2376-2386, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-30942389

RESUMO

Macrophages can induce Fas ligand (FasL)­mediated apoptosis, and the deregulation of apoptosis is known to be associated with recurrent miscarriage (RM). The aim of the present study was to investigate the possible involvement of FasL in macrophage­mediated trophoblast apoptosis and its potential role in RM. Human decidual and placental villous tissues were collected from 81 women (21 for the RM group, 26 for the spontaneous abortion group and 34 for the control group) at 7­9 weeks of gestation. The distribution changes of macrophages and the expression of FasL on macrophages were evaluated by immunohistochemical, immunofluorescence and western blot analyses. A macrophage and trophoblast co­culture model was used to determine the effects of FasL on the apoptosis of trophoblasts. The results indicated that CD86+ macrophage populations in decidual tissues were significantly increased, accompanied by reduced CD163+ macrophages in the abortion and RM groups. Furthermore, the distribution of CD68+ macrophages was also significantly altered in specimens from the abortion and RM groups, and they were observed to have infiltrated into the trophoblast cells. In addition, elevated expression of FasL on CD68+ and CD86+ macrophages in the decidua was observed in the spontaneous abortion and RM groups of patients, and FasL was demonstrated to mediate the induction of trophoblast apoptosis by macrophages in co­culture. These results indicate that the aberration of macrophage­induced FasL­mediated apoptosis may represent one of the causes of RM.


Assuntos
Aborto Habitual/patologia , Apoptose , Decídua/patologia , Proteína Ligante Fas/análise , Macrófagos/patologia , Trofoblastos/patologia , Aborto Habitual/etiologia , Aborto Habitual/metabolismo , Adulto , Linhagem Celular , Decídua/citologia , Decídua/metabolismo , Proteína Ligante Fas/metabolismo , Feminino , Humanos , Macrófagos/metabolismo , Gravidez , Trofoblastos/citologia , Trofoblastos/metabolismo
20.
PLoS One ; 14(4): e0214110, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30951545

RESUMO

Peg10 (paternally expressed gene 10) is an imprinted gene that is essential for placental development. It is thought to derive from a Ty3-gyspy LTR (long terminal repeat) retrotransposon and retains Gag and Pol-like domains. Here we show that the Gag domain of PEG10 can promote vesicle budding similar to the HIV p24 Gag protein. Expressed in a subset of mouse endocrine organs in addition to the placenta, PEG10 was identified as a substrate of the deubiquitinating enzyme USP9X. Consistent with PEG10 having a critical role in placental development, PEG10-deficient trophoblast stem cells (TSCs) exhibited impaired differentiation into placental lineages. PEG10 expressed in wild-type, differentiating TSCs was bound to many cellular RNAs including Hbegf (Heparin-binding EGF-like growth factor), which is known to play an important role in placentation. Expression of Hbegf was reduced in PEG10-deficient TSCs suggesting that PEG10 might bind to and stabilize RNAs that are critical for normal placental development.


Assuntos
Diferenciação Celular/genética , Fator de Crescimento Semelhante a EGF de Ligação à Heparina/genética , Proteínas Nucleares/genética , Placentação/genética , Fatores de Transcrição/genética , Animais , Linhagem da Célula/genética , Elementos de DNA Transponíveis/genética , Feminino , Regulação da Expressão Gênica no Desenvolvimento , Produtos do Gene gag/genética , Impressão Genômica/genética , Humanos , Camundongos , Placenta/metabolismo , Gravidez , Proteínas de Ligação a RNA/genética , Células-Tronco/citologia , Células-Tronco/metabolismo , Trofoblastos/citologia , Trofoblastos/metabolismo
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