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1.
J Cell Physiol ; 233(1): 530-548, 2018 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-28300292

RESUMO

Although histone acetylation is one of the most widely studied epigenetic modifications, there is still a lack of information regarding how the acetylome is regulated during brain development and pathophysiological processes. We demonstrate that the embryonic brain (E15) is characterized by an increase in H3K9 acetylation as well as decreases in the levels of HDAC1 and HDAC3. Moreover, experimental induction of H3K9 hyperacetylation led to the overexpression of NCAM in the embryonic cortex and depletion of Sox2 in the subventricular ependyma, which mimicked the differentiation processes. Inducing differentiation in HDAC1-deficient mouse ESCs resulted in early H3K9 deacetylation, Sox2 downregulation, and enhanced astrogliogenesis, whereas neuro-differentiation was almost suppressed. Neuro-differentiation of (wt) ESCs was characterized by H3K9 hyperacetylation that was associated with HDAC1 and HDAC3 depletion. Conversely, the hippocampi of schizophrenia-like animals showed H3K9 deacetylation that was regulated by an increase in both HDAC1 and HDAC3. The hippocampi of schizophrenia-like brains that were treated with the cannabinoid receptor-1 inverse antagonist AM251 expressed H3K9ac at the level observed in normal brains. Together, the results indicate that co-regulation of H3K9ac by HDAC1 and HDAC3 is important to both embryonic brain development and neuro-differentiation as well as the pathophysiology of a schizophrenia-like phenotype.


Assuntos
Encéfalo/enzimologia , Histona Desacetilase 1/metabolismo , Histona Desacetilases/metabolismo , Histonas/metabolismo , Neurogênese , Neurônios/enzimologia , Esquizofrenia/enzimologia , Acetilação , Animais , Antipsicóticos/farmacologia , Encéfalo/efeitos dos fármacos , Encéfalo/embriologia , Encéfalo/patologia , Antagonistas de Receptores de Canabinoides/farmacologia , Modelos Animais de Doenças , Epigênese Genética , Regulação da Expressão Gênica no Desenvolvimento , Idade Gestacional , Histona Desacetilase 1/antagonistas & inibidores , Histona Desacetilase 1/genética , Inibidores de Histona Desacetilases/farmacologia , Histona Desacetilases/genética , Acetato de Metilazoximetanol , Camundongos Endogâmicos C57BL , Moléculas de Adesão de Célula Nervosa/genética , Moléculas de Adesão de Célula Nervosa/metabolismo , Neurogênese/efeitos dos fármacos , Neurônios/efeitos dos fármacos , Neurônios/patologia , Processamento de Proteína Pós-Traducional , Ratos Sprague-Dawley , Receptor CB1 de Canabinoide/antagonistas & inibidores , Receptor CB1 de Canabinoide/metabolismo , Fatores de Transcrição SOXB1/genética , Fatores de Transcrição SOXB1/metabolismo , Esquizofrenia/induzido quimicamente , Esquizofrenia/tratamento farmacológico , Esquizofrenia/genética , Transdução de Sinais , Fatores de Tempo
2.
Biol Open ; 5(2): 140-53, 2016 Jan 19.
Artigo em Inglês | MEDLINE | ID: mdl-26787680

RESUMO

The transcription factor Nkx2.5 and the intermediate filament protein desmin are simultaneously expressed in cardiac progenitor cells during commitment of primitive mesoderm to the cardiomyogenic lineage. Up-regulation of Nkx2.5 expression by desmin suggests that desmin may contribute to cardiogenic commitment and myocardial differentiation by directly influencing the transcription of the nkx2.5 gene in cardiac progenitor cells. Here, we demonstrate that desmin activates transcription of nkx2.5 reporter genes, rescues nkx2.5 haploinsufficiency in cardiac progenitor cells, and is responsible for the proper expression of Nkx2.5 in adult cardiac side population stem cells. These effects are consistent with the temporary presence of desmin in the nuclei of differentiating cardiac progenitor cells and its physical interaction with transcription factor complexes bound to the enhancer and promoter elements of the nkx2.5 gene. These findings introduce desmin as a newly discovered and unexpected player in the regulatory network guiding cardiomyogenesis in cardiac stem cells.

3.
Cell Physiol Biochem ; 33(1): 205-21, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-24481283

RESUMO

BACKGROUND/AIMS: Cell transplantation into the heart is a new therapy after myocardial infarction. Its success, however, is impeded by poor donor cell survival and by limited transdifferentiation of the transplanted cells into functional cardiomyocytes. A promising strategy to overcome these problems is the induction of cardiomyogenic properties in donor cells by small molecules. METHODS: Here we studied cardiomyogenic effects of the small molecule compound cardiogenol C (CgC), and structural derivatives thereof, on lineage-committed progenitor cells by various molecular biological, biochemical, and functional assays. RESULTS: Treatment with CgC up-regulated cardiac marker expression in skeletal myoblasts. Importantly, the compound also induced cardiac functional properties: first, cardiac-like sodium currents in skeletal myoblasts, and secondly, spontaneous contractions in cardiovascular progenitor cell-derived cardiac bodies. CONCLUSION: CgC induces cardiomyogenic function in lineage-committed progenitor cells, and can thus be considered a promising tool to improve cardiac repair by cell therapy.


Assuntos
Compostos de Anilina/farmacologia , Biomarcadores/metabolismo , Linhagem da Célula/efeitos dos fármacos , Miocárdio/metabolismo , Pirimidinas/farmacologia , Bibliotecas de Moléculas Pequenas/farmacologia , Regulação para Cima/efeitos dos fármacos , Compostos de Anilina/química , Animais , Fator Natriurético Atrial/metabolismo , Diferenciação Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Proteína Homeobox Nkx-2.5 , Proteínas de Homeodomínio/metabolismo , Humanos , Ativação do Canal Iônico/efeitos dos fármacos , Camundongos , Mioblastos Esqueléticos/efeitos dos fármacos , Mioblastos Esqueléticos/metabolismo , Miócitos Cardíacos/citologia , Miócitos Cardíacos/efeitos dos fármacos , Miócitos Cardíacos/metabolismo , Pirimidinas/química , Ratos , Bibliotecas de Moléculas Pequenas/química , Células-Tronco , Fatores de Transcrição/metabolismo
4.
Stem Cell Res ; 11(1): 552-62, 2013 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-23611953

RESUMO

The induction of teratoma in mice by the transplantation of stem cells into extra-uterine sites has been used as a read-out for cellular pluripotency since the initial description of this phenomenon in 1954. Since then, the teratoma assay has remained the assay of choice to demonstrate pluripotency, gaining prominence during the recent hype surrounding human stem cell research. However, the scientific significance of the teratoma assay has been debated due to the fact that transplanted cells are exposed to a non-physiological environment. Since many mice are used for a result that is heavily questioned, it is time to reconsider the teratoma assay from an ethical point of view. Candidate alternatives to the teratoma assay comprise the directed differentiation of pluripotent stem cells into organotypic cells, differentiation of cells in embryoid bodies, the analysis of pluripotency-associated biomarkers with high correlation to the teratoma forming potential of stem cells, predictive epigenetic footprints, or a combination of these technologies. Each of these assays is capable of addressing one or more aspects of pluripotency, however it is essential that these assays are validated to provide an accepted robust, reproducible alternative. In particular, the rapidly expanding number of human induced pluripotent stem cell lines, requires the development of simple, affordable standardized in vitro and in silico assays to reduce the number of animal experiments performed.


Assuntos
Bioensaio/métodos , Células-Tronco Pluripotentes/citologia , Células-Tronco Pluripotentes/transplante , Teratoma/patologia , Animais , Diferenciação Celular/fisiologia , Humanos , Camundongos
5.
Cells Tissues Organs ; 197(4): 249-68, 2013.
Artigo em Inglês | MEDLINE | ID: mdl-23343517

RESUMO

Compelling evidence for the existence of somatic stem cells in the heart of different mammalian species has been provided by numerous groups; however, so far it has not been possible to maintain these cells as self-renewing and phenotypically stable clonal cell lines in vitro. Thus, we sought to identify a surrogate stem cell niche for the isolation and persistent maintenance of stable clonal cardiovascular progenitor cell lines, enabling us to study the mechanism of self-renewal and differentiation in these cells. Using postnatal murine hearts with a selectable marker as the stem cell source and embryonic stem cells and leukemia inhibitory factor (LIF)-secreting fibroblasts as a surrogate niche, we succeeded in the isolation of stable clonal cardiovascular progenitor cell lines. These cell lines self-renew in an LIF-dependent manner. They express both stemness transcription factors Oct4, Sox2, and Nanog and early myocardial transcription factors Nkx2.5, GATA4, and Isl-1 at the same time. Upon LIF deprivation, they exclusively differentiate to functional cardiomyocytes and endothelial and smooth muscle cells, suggesting that these cells are mesodermal intermediates already committed to the cardiogenic lineage. Cardiovascular progenitor cell lines can be maintained for at least 149 passages over 7 years without phenotypic changes, in the presence of LIF-secreting fibroblasts. Isolation of wild-type cardiovascular progenitor cell lines from adolescent and old mice has finally demonstrated the general feasibility of this strategy for the isolation of phenotypically stable somatic stem cell lines.


Assuntos
Células-Tronco Embrionárias/citologia , Fator Inibidor de Leucemia/metabolismo , Miócitos Cardíacos/citologia , Animais , Diferenciação Celular/fisiologia , Linhagem Celular , Técnicas Citológicas/métodos , Embrião de Mamíferos , Células-Tronco Embrionárias/metabolismo , Células Endoteliais/citologia , Células Endoteliais/metabolismo , Feminino , Fibroblastos/citologia , Fibroblastos/metabolismo , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Microscopia Confocal/métodos , Miócitos Cardíacos/metabolismo
6.
Medchemcomm ; 4(8): 1189-1195, 2013 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-25045463

RESUMO

Intra-cardiac cell transplantation is a new therapy after myocardial infarction. Its success, however, is impeded by the limited capacity of donor cells to differentiate into functional cardiomyocytes in the heart. A strategy to overcome this problem is the induction of cardiomyogenic function in cells prior to transplantation. Among other approaches, recently, synthetic small molecules were identified, which promote differentiation of stem cells of various origins into cardiac-like cells or cardiomyocytes. The aim of this study was to develop and characterise new promising cardiomyogenic synthetic low-molecular weight compounds. Therefore, the structure of the known cardiomyogenic molecule cardiogenol C was selectively modified, and the effects of the resulting compounds were tested on various cell types. From this study, VUT-MK142 was identified as the most promising candidate with respect to cardiomyogenic activity. Treatment using this novel agent induced the strongest up-regulation of expression of the cardiac marker ANF in both P19 embryonic carcinoma cells and C2C12 skeletal myoblasts. The activity of VUT-MK142 on this marker superseded CgC; moreover, the novel compound significantly up-regulated the expression of other cardiac markers, and promoted the development of beating cardiomyocytes from cardiovascular progenitor cells. We conclude that VUT-MK142 is a potent new cardiomyogenic synthetic agent promoting the differentiation of pre-cardiac mesoderm into cardiomyocytes, which may be useful to differentiate stem cells into cardiomyocytes for cardiac repair. Additionally, an efficient synthesis of VUT-MK142 is reported taking advantage of continuous flow techniques superior to classical batch reactions both in yield and reaction time.

7.
Int Rev Cell Mol Biol ; 293: 195-267, 2012.
Artigo em Inglês | MEDLINE | ID: mdl-22251563

RESUMO

Self-renewing cells of the vertebrate heart have become a major subject of interest in the past decade. However, many researchers had a hard time to argue against the orthodox textbook view that defines the heart as a postmitotic organ. Once the scientific community agreed on the existence of self-renewing cells in the vertebrate heart, their origin was again put on trial when transdifferentiation, dedifferentiation, and reprogramming could no longer be excluded as potential sources of self-renewal in the adult organ. Additionally, the presence of self-renewing pluripotent cells in the peripheral blood challenges the concept of tissue-specific stem and progenitor cells. Leaving these unsolved problems aside, it seems very desirable to learn about the basic biology of this unique cell type. Thus, we shall here paint a picture of cardiovascular progenitor cells including the current knowledge about their origin, basic nature, and the molecular mechanisms guiding proliferation and differentiation into somatic cells of the heart.


Assuntos
Coração/embriologia , Miocárdio/citologia , Organogênese/fisiologia , Células-Tronco/fisiologia , Adulto , Animais , Desenvolvimento Embrionário/genética , Desenvolvimento Embrionário/fisiologia , Humanos , Modelos Biológicos , Miocárdio/metabolismo , Organogênese/genética , Transdução de Sinais/genética , Transdução de Sinais/fisiologia , Nicho de Células-Tronco/genética , Nicho de Células-Tronco/fisiologia , Células-Tronco/metabolismo
8.
Cells Tissues Organs ; 195(5): 377-91, 2012.
Artigo em Inglês | MEDLINE | ID: mdl-21860211

RESUMO

Aggregation of embryonic stem cells gives rise to embryoid bodies (EBs) which undergo developmental processes reminiscent of early eutherian embryonic development. Development of the three germ layers suggests that gastrulation takes place. In vivo, gastrulation is a highly ordered process but in EBs only few data support the hypothesis that self-organization of differentiating cells leads to morphology, reminiscent of the early gastrula. Here we demonstrate that a timely implantation-like process is a prerequisite for the breaking of the radial symmetry of suspended EBs. Attached to a surface, EBs develop a bilateral symmetry and presumptive mesodermal cells emerge between the center of the EBs and a horseshoe-shaped ridge of cells. The development of an epithelial sheet of cells on one side of the EBs allows us to define an 'anterior' and a 'posterior' end of the EBs. In the mesodermal area, first cardiomyocytes (CMCs) develop mainly next to this epithelial sheet of cells. Development of twice as many CMCs at the 'left' side of the EBs breaks the bilateral symmetry and suggests that cardiomyogenesis reflects a local or temporal asymmetry in EBs. The asymmetric appearance of CMCs but not the development of mesoderm can be disturbed by ectopic expression of the muscle-specific protein Desmin. Later, the bilateral morphology becomes blurred by an apparently chaotic differentiation of many cell types. The absence of comparable structures in aggregates of cardiovascular progenitor cells isolated from the heart demonstrates that the self-organization of cells during a gastrulation-like process is a unique feature of embryonic stem cells.


Assuntos
Corpos Embrioides/citologia , Células-Tronco Embrionárias/citologia , Miócitos Cardíacos/citologia , Organogênese/fisiologia , Corpos Embrioides/metabolismo , Desenvolvimento Embrionário , Células-Tronco Embrionárias/metabolismo , Humanos , Microscopia Confocal , Miócitos Cardíacos/metabolismo
9.
EMBO J ; 29(23): 3992-4007, 2010 Dec 01.
Artigo em Inglês | MEDLINE | ID: mdl-20967026

RESUMO

Histone deacetylase (HDAC) inhibitors induce cell cycle arrest, differentiation or apoptosis in tumour cells and are, therefore, promising anti-cancer reagents. However, the specific HDAC isoforms that mediate these effects are not yet identified. To explore the role of HDAC1 in tumourigenesis and tumour proliferation, we established an experimental teratoma model using wild-type and HDAC1-deficient embryonic stem cells. HDAC1-deficient teratomas showed no significant difference in size compared with wild-type teratomas. Surprisingly, loss of HDAC1 was not only linked to increased apoptosis, but also to significantly enhanced proliferation. Epithelial structures showed reduced differentiation as monitored by Oct3/4 expression and changed E-cadherin localization and displayed up-regulated expression of SNAIL1, a regulator of epithelial cell plasticity. Increased levels of the transcriptional regulator SNAIL1 are crucial for enhanced proliferation and reduced differentiation of HDAC1-deficient teratoma. Importantly, the analysis of human teratomas revealed a similar link between loss of HDAC1 and enhanced tumour malignancy. These findings reveal a novel role for HDAC1 in the control of tumour proliferation and identify HDAC1 as potential marker for benign teratomas.


Assuntos
Células-Tronco Embrionárias/metabolismo , Regulação Neoplásica da Expressão Gênica , Histona Desacetilase 1/genética , Teratoma/enzimologia , Animais , Apoptose , Caderinas/genética , Carcinoma Embrionário/enzimologia , Carcinoma Embrionário/genética , Carcinoma Embrionário/patologia , Linhagem Celular , Linhagem Celular Tumoral , Proliferação de Células , Células-Tronco Embrionárias/citologia , Células-Tronco Embrionárias/patologia , Histona Desacetilase 1/metabolismo , Humanos , Camundongos , Fator 3 de Transcrição de Octâmero/genética , Fenótipo , Fatores de Transcrição da Família Snail , Teratoma/genética , Teratoma/patologia , Fatores de Transcrição/genética
10.
Mol Cell Biol ; 30(5): 1171-81, 2010 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-20028735

RESUMO

Histone deacetylases (HDACs) are chromatin-modifying enzymes that are involved in the regulation of proliferation, differentiation and development. HDAC inhibitors induce cell cycle arrest, differentiation, or apoptosis in tumor cells and are therefore promising antitumor agents. Numerous genes were found to be deregulated upon HDAC inhibitor treatment; however, the relevant target enzymes are still unidentified. HDAC1 is required for mouse development and unrestricted proliferation of embryonic stem cells. We show here that HDAC1 reversibly regulates cellular proliferation and represses the cyclin-dependent kinase inhibitor p21 in embryonic stem cells. Disruption of the p21 gene rescues the proliferation phenotype of HDAC1(-/-) embryonic stem cells but not the embryonic lethality of HDAC1(-/-) mice. In the absence of HDAC1, mouse embryonic fibroblasts scarcely undergo spontaneous immortalization and display increased p21 expression. Chromatin immunoprecipitation assays demonstrate a direct regulation of the p21 gene by HDAC1 in mouse embryonic fibroblasts. Transformation with simian virus 40 large T antigen or ablation of p21 restores normal immortalization of primary HDAC1(-/-) fibroblasts. Our data demonstrate that repression of the p21 gene is crucial for HDAC1-mediated control of proliferation and immortalization. HDAC1 might therefore be one of the relevant targets for HDAC inhibitors as anticancer drugs.


Assuntos
Proliferação de Células , Inibidor de Quinase Dependente de Ciclina p21/metabolismo , Histona Desacetilase 1/metabolismo , Animais , Antígenos Transformantes de Poliomavirus/genética , Transformação Celular Viral , Células Cultivadas , Inibidor de Quinase Dependente de Ciclina p21/deficiência , Inibidor de Quinase Dependente de Ciclina p21/genética , Células-Tronco Embrionárias/citologia , Células-Tronco Embrionárias/metabolismo , Feminino , Expressão Gênica , Histona Desacetilase 1/deficiência , Histona Desacetilase 1/genética , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Proteínas Oncogênicas Virais/genética , Proteínas E7 de Papillomavirus/genética , Fenótipo , Proteínas Repressoras/genética
11.
Differentiation ; 75(7): 616-26, 2007 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-17381546

RESUMO

Desmin contributes to the stability of the myocardium and its amino-terminal domain influences intermediate filament formation and interacts with a variety of proteins and DNAs. Specific serine residues located in this domain are reversibly phosphorylated in a cell cycle and developmental stage-dependent manner as has been demonstrated also for other cytoplasmic type III intermediate filament proteins. Although absence of desmin apparently does not affect cardiomyogenesis, homozygous deletion of the amino-terminal domain of desmin severely inhibited in vitro cardiomyogenesis. To demonstrate the significance of phosphorylation of this domain in cardiomyogenic commitment and differentiation, we inhibited phosphorylation of serine residues 6, 7, and 8 by mutation to alanine, and investigated early cardiomyogenesis in heterozygous embryoid bodies. As control, serine residues 31 and 32, which are not phosphorylated by kinases mutating serine residues 6, 7, and 8, were mutated to alanine in a second set. Desmin(S6,7,8A) interfered with cardiomyogenesis and myofibrillogenesis in a dominant negative fashion, whereas desmin(S31,32A) produced only a mild phenotype. Desmin(S6,7,8A) led to the down-regulation of the transcription factor genes brachyury, goosecoid, nkx2.5, and mef2C and increased apoptosis of presumptive mesoderm and differentiating cardiomyocytes. Surviving cardiomyocytes which were few in number had no myofibrils. Demonstration that some but not any mutant desmin interfered with the very beginning of cardiomyogenesis suggests an important function of temporarily phosphorylated serine residues 6, 7, and 8 in the amino-terminal domain of desmin in cardiomyogenic commitment and differentiation.


Assuntos
Diferenciação Celular/fisiologia , Desmina/genética , Miócitos Cardíacos/citologia , Fragmentos de Peptídeos/fisiologia , Serina/genética , Animais , Diferenciação Celular/genética , Linhagem Celular , Proliferação de Células , Desmina/fisiologia , Humanos , Desenvolvimento Muscular/genética , Fragmentos de Peptídeos/genética , Estrutura Terciária de Proteína/genética , Serina/fisiologia , Fatores de Transcrição/biossíntese , Fatores de Transcrição/genética
12.
Differentiation ; 75(7): 605-15, 2007 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-17381547

RESUMO

Desmin contributes to structural integrity and function of the myocardium but its function seems to be redundant in early cardiomyogenesis in the desmin null mouse model. To test the hypothesis that desmin also plays a supportive role in cardiomyogenic commitment and early differentiation of cardiomyocytes we investigated cardiomyogenesis in embryoid bodies expressing different desmin alleles. Constitutive expression of desmin and increased synthesis during mesoderm formation led to the up-regulation of brachyury and nkx2.5 genes, accelerated early cardiomyogenesis and resulted in the development of large, proliferating, highly interconnected, and synchronously beating cardiomyocyte clusters, whereas desmin null cardiomyocytes featured an opposite phenotype. In contrast, constitutive expression of amino-terminally truncated desmin(Delta1-48) interfered with the beginning of cardiomyogenesis, caused down-regulation of mesodermal and myocardial transcription factors, and hampered myofibrillogenesis and survival of cardiomyocytes. These results provide first evidence that a type III intermediate filament protein takes part in regulating the differentiation of mesoderm to cardiomyocytes at the very beginning of cardiomyogenesis.


Assuntos
Diferenciação Celular/fisiologia , Desmina/fisiologia , Proteínas Fetais/biossíntese , Proteínas Fetais/genética , Proteínas de Homeodomínio/biossíntese , Proteínas de Homeodomínio/genética , Miócitos Cardíacos/citologia , Proteínas com Domínio T/biossíntese , Proteínas com Domínio T/genética , Fatores de Transcrição/biossíntese , Fatores de Transcrição/genética , Regulação para Cima/genética , Animais , Agregação Celular/genética , Diferenciação Celular/genética , Linhagem Celular , Desmina/biossíntese , Desmina/deficiência , Desmina/genética , Proteína Homeobox Nkx-2.5 , Mesoderma/citologia , Camundongos , Camundongos Knockout , Miócitos Cardíacos/metabolismo
13.
Biochem Biophys Res Commun ; 343(2): 555-63, 2006 May 05.
Artigo em Inglês | MEDLINE | ID: mdl-16554030

RESUMO

Cardiomyogenesis is influenced by factors secreted by anterior-lateral and extra-embryonic endoderm. Differentiation of embryonic stem cells in embryoid bodies allows to study the influence of growth factors on cardiomyogenesis. By these means SPARC was identified as a new factor enhancing cardiomyogenesis [M. Stary, W. Pasteiner, A. Summer, A. Hrdina, A. Eger, G. Weitzer, Parietal endoderm secreted SPARC promotes early cardiomyogenesis in vitro, Exp. Cell Res. 310 (2005) 331-341]. Here we report a similar and new function for S100A4, a calcium-binding protein of the EF-hand type. S100A4 is secreted by parietal endoderm and promotes early differentiation and proliferation of cardiomyocytes. Oligomeric S100A4 supports cardiomyogenesis in a concentration-dependent manner, whereas inhibition of autocrine S100A4 severely attenuates cardiomyogenesis. S100A4 specifically influences transcription in differentiating cardiomyocytes, as evident from increased expression of cardiac transcription factor genes nkx2.5 and mef2C. These data suggest that S100A4, like SPARC, plays a supportive role in early in vitro cardiomyogenesis.


Assuntos
Embrião de Mamíferos/citologia , Embrião de Mamíferos/fisiologia , Endoderma/metabolismo , Miócitos Cardíacos/citologia , Miócitos Cardíacos/fisiologia , Proteínas S100/administração & dosagem , Proteínas S100/metabolismo , Diferenciação Celular/efeitos dos fármacos , Linhagem Celular , Proliferação de Células , Relação Dose-Resposta a Droga , Embrião de Mamíferos/efeitos dos fármacos , Proteína Homeobox Nkx-2.5 , Proteínas de Homeodomínio/metabolismo , Humanos , Proteínas de Domínio MADS/metabolismo , Fatores de Transcrição MEF2 , Miócitos Cardíacos/efeitos dos fármacos , Fatores de Regulação Miogênica/metabolismo , Proteína A4 de Ligação a Cálcio da Família S100 , Fatores de Transcrição/metabolismo
14.
Exp Cell Res ; 310(2): 331-43, 2005 Nov 01.
Artigo em Inglês | MEDLINE | ID: mdl-16165126

RESUMO

Cardiomyogenesis proceeds in the presence of signals emanating from extra-embryonic lineages emerging before and during early eutherian gastrulation. In embryonic stem cell derived embryoid bodies, primitive endoderm gives rise to visceral and parietal endoderm. Parietal endoderm undergoes an epithelial to mesenchymal transition shortly before first cardiomyocytes start to contract rhythmically. Here, we demonstrate that Secreted Protein, Acidic, Rich in Cysteine, SPARC, predominantly secreted by mesenchymal parietal endoderm specifically promotes early myocardial cell differentiation in embryoid bodies. SPARC enhanced the expression of bmp2 and nkx2.5 in embryoid bodies and fetal cardiomyocytes. Inhibition of either SPARC or Bmp2 attenuated in both cases cardiomyogenesis and downregulated nkx2.5 expression. Thus, SPARC directly affects cardiomyogenesis, modulates Bmp2 signaling, and contributes to a positive autoregulatory loop of Bmp2 and Nkx2.5 in cardiomyocytes.


Assuntos
Proteínas Morfogenéticas Ósseas/metabolismo , Endoderma/metabolismo , Coração/embriologia , Osteonectina/metabolismo , Fator de Crescimento Transformador beta/metabolismo , Animais , Proteína Morfogenética Óssea 2 , Proteínas Morfogenéticas Ósseas/genética , Endoderma/citologia , Regulação da Expressão Gênica no Desenvolvimento , Proteína Homeobox Nkx-2.5 , Proteínas de Homeodomínio/genética , Proteínas de Homeodomínio/metabolismo , Mesoderma/citologia , Mesoderma/fisiologia , Camundongos , Desenvolvimento Muscular/genética , Miócitos Cardíacos/metabolismo , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Fator de Crescimento Transformador beta/genética , Regulação para Cima
15.
Biochem Biophys Res Commun ; 331(4): 1577-86, 2005 Jun 17.
Artigo em Inglês | MEDLINE | ID: mdl-15883053

RESUMO

The unique differentiation potential of inner cell mass derived embryonic stem cells together with their outstanding self-renewal capacity makes them a desirable source for somatic cell therapy of human diseases. Somatic cells are gained by in vitro differentiation of embryonic stem cells, however, the differentiation potential of embryonic stem cells varied even between isogenic cell lines. Variable differentiation potentials may either be a consequence of an inherent inhomogeneity of gene expression in the inner cell mass or may have technical reasons. To understand variations in the differentiation potential, we generated pairs of isogenic, monozygotic twin, and single inner cell mass derived clonal embryonic stem cell lines, and demonstrate that they differentially express the leukaemia inhibitory factor receptor gene. Variations of leukaemia inhibitory factor receptor protein levels are already evident in the inner cell mass and predispose the cardiomyogenic potential of embryonic stem cell lines in a Janus activated kinase dependent manner. Thus, a single inner cell mass may give rise to embryonic stem cell lines with different developmental potentials.


Assuntos
Embrião de Mamíferos/citologia , Receptores de Citocinas/metabolismo , Células-Tronco/citologia , Animais , Sistema Cardiovascular/embriologia , Linhagem Celular , Feminino , Subunidade alfa de Receptor de Fator Inibidor de Leucemia , Camundongos , Músculos/embriologia , Receptores de OSM-LIF , Reação em Cadeia da Polimerase Via Transcriptase Reversa
16.
FEBS Lett ; 523(1-3): 229-33, 2002 Jul 17.
Artigo em Inglês | MEDLINE | ID: mdl-12123837

RESUMO

Desmin fulfils important functions in maintenance of muscle cells and mutations in the desmin gene have been linked to a variety of myopathies. To ascertain the role of desmin's amino-terminal domain in muscle cells we generated embryonic stem cells constitutively expressing desmin(Delta1-48) in a null background and investigated muscle cell development in vitro. Desmin(Delta1-48) lacking the first 48 amino acid residues promotes fusion of myoblasts, rescues myogenesis and down-regulates vimentin expression in embryoid bodies, but hampers cardiomyogenesis and blocks smooth muscle development. These results demonstrate that desmin's amino-terminus has different roles in skeletal, cardiac, and smooth muscle cell development and function.


Assuntos
Desmina/fisiologia , Desenvolvimento Muscular/fisiologia , Músculo Esquelético/crescimento & desenvolvimento , Músculo Liso/crescimento & desenvolvimento , Diferenciação Celular/fisiologia , Fusão Celular , Linhagem Celular , Desmina/genética , Desmina/metabolismo , Regulação para Baixo , Coração/fisiologia , Humanos , Músculo Esquelético/fisiologia , Músculo Liso/fisiologia , Mutação , Miocárdio , Deleção de Sequência/genética , Células-Tronco/metabolismo , Vimentina/fisiologia
17.
EMBO J ; 21(11): 2672-81, 2002 Jun 03.
Artigo em Inglês | MEDLINE | ID: mdl-12032080

RESUMO

Histone deacetylases (HDACs) modulate chromatin structure and transcription, but little is known about their function in mammalian development. HDAC1 was implicated previously in the repression of genes required for cell proliferation and differentiation. Here we show that targeted disruption of both HDAC1 alleles results in embryonic lethality before E10.5 due to severe proliferation defects and retardation in development. HDAC1-deficient embryonic stem cells show reduced proliferation rates, which correlate with decreased cyclin-associated kinase activities and elevated levels of the cyclin-dependent kinase inhibitors p21(WAF1/CIP1) and p27(KIP1). Similarly, expression of p21 and p27 is up-regulated in HDAC1-null embryos. In addition, loss of HDAC1 leads to significantly reduced overall deacetylase activity, hyperacetylation of a subset of histones H3 and H4 and concomitant changes in other histone modifications. The expression of HDAC2 and HDAC3 is induced in HDAC1-deficient cells, but cannot compensate for loss of the enzyme, suggesting a unique function for HDAC1. Our study provides the first evidence that a histone deacetylase is essential for unrestricted cell proliferation by repressing the expression of selective cell cycle inhibitors.


Assuntos
Quinases Ciclina-Dependentes/metabolismo , Histona Desacetilases/genética , Histona Desacetilases/fisiologia , Alelos , Animais , Southern Blotting , Western Blotting , Proteínas de Ciclo Celular/metabolismo , Divisão Celular , Inibidor de Quinase Dependente de Ciclina p21 , Inibidor de Quinase Dependente de Ciclina p27 , Quinases Ciclina-Dependentes/antagonistas & inibidores , Ciclinas/metabolismo , Éxons , Histona Desacetilase 1 , Histonas/metabolismo , Hibridização In Situ , Marcação In Situ das Extremidades Cortadas , Camundongos , Microscopia de Fluorescência , Modelos Genéticos , Fenótipo , Testes de Precipitina , Ligação Proteica , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Fatores de Tempo , Proteínas Supressoras de Tumor/metabolismo , Regulação para Cima
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