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1.
Signal Transduct Target Ther ; 8(1): 16, 2023 01 11.
Artigo em Inglês | MEDLINE | ID: mdl-36627278

RESUMO

Lkb1 deficiency confers the Kras-mutant lung cancer with strong plasticity and the potential for adeno-to-squamous transdifferentiation (AST). However, it remains largely unknown how Lkb1 deficiency dynamically regulates AST. Using the classical AST mouse model (Kras LSL-G12D/+;Lkb1flox/flox, KL), we here comprehensively analyze the temporal transcriptomic dynamics of lung tumors at different stages by dynamic network biomarker (DNB) and identify the tipping point at which the Wnt signaling is abruptly suppressed by the excessive accumulation of reactive oxygen species (ROS) through its downstream effector FOXO3A. Bidirectional genetic perturbation of the Wnt pathway using two different Ctnnb1 conditional knockout mouse strains confirms its essential role in the negative regulation of AST. Importantly, pharmacological activation of the Wnt pathway before but not after the tipping point inhibits squamous transdifferentiation, highlighting the irreversibility of AST after crossing the tipping point. Through comparative transcriptomic analyses of mouse and human tumors, we find that the lineage-specific transcription factors (TFs) of adenocarcinoma and squamous cell carcinoma form a "Yin-Yang" counteracting network. Interestingly, inactivation of the Wnt pathway preferentially suppresses the adenomatous lineage TF network and thus disrupts the "Yin-Yang" homeostasis to lean towards the squamous lineage, whereas ectopic expression of NKX2-1, an adenomatous lineage TF, significantly dampens such phenotypic transition accelerated by the Wnt pathway inactivation. The negative correlation between the Wnt pathway and AST is further observed in a large cohort of human lung adenosquamous carcinoma. Collectively, our study identifies the tipping point of AST and highlights an essential role of the ROS-Wnt axis in dynamically orchestrating the homeostasis between adeno- and squamous-specific TF networks at the AST tipping point.


Assuntos
Carcinoma de Células Escamosas , Neoplasias Pulmonares , Animais , Camundongos , Humanos , Via de Sinalização Wnt/genética , Proteínas Proto-Oncogênicas p21(ras)/metabolismo , Transdiferenciação Celular/genética , Espécies Reativas de Oxigênio/metabolismo , Neoplasias Pulmonares/patologia , Pulmão/patologia , Proteínas Serina-Treonina Quinases/genética , Carcinoma de Células Escamosas/genética , Carcinoma de Células Escamosas/patologia , Camundongos Knockout , Estresse Oxidativo/genética
2.
Ren Fail ; 44(1): 752-766, 2022 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-35509185

RESUMO

Chronic kidney disease (CKD) has recently become a serious health and social concern. Vascular calcification, a common complication of CKD, is a risk factor that increases the incidence and mortality of cardiovascular events in patients with CKD. However, there are currently no effective therapeutic targets that can facilitate treatment with fewer side effects for vascular calcification in CKD. To identify potential therapeutic targets, we performed label-free quantification (LFQ) analyses of protein samples from rat aortic vascular smooth muscle cells (RASMCs) after high-phosphorus treatment by nano-UPLC-MS/MS. We determined that ubiquitin-specific protease 47 (USP47) may be associated with CKD vascular calcification by regulating the osteogenic transdifferentiation of the vascular smooth muscle cell (VSMC) phenotype, thus suggesting a novel and potentially effective therapeutic target for CKD vascular calcification. USP47 knockdown significantly reduced the expression of ß-transducin repeat-containing protein (BTRC), serine/threonine-protein kinase akt-1 (AKT1), Klotho, fibroblast growth factor (FGF23), and matrix Gla protein (MGP) in RASMCs after high-phosphorus treatment. Consistent with the results of protein-protein interaction (PPI) analyses, USP47 may be involved in regulating osteogenic transdifferentiation markers, such as runt-related transcription factor 2 (RUNX2), Klotho, FGF23, and MGP through the BTRC/AKT1 pathway upon CKD vascular calcification. These data indicate that USP47 may be associated with vascular calcification in CKD by regulating osteogenic differentiation of VSMCs. USP47 may regulate osteogenic transdifferentiation in VSMCs upon CKD vascular calcification through a process involving the BTRC/AKT1 pathway. This study identified a novel potential therapeutic target for the treatment of vascular calcification in CKD.


Assuntos
Insuficiência Renal Crônica , Proteases Específicas de Ubiquitina , Calcificação Vascular , Animais , Transdiferenciação Celular/genética , Células Cultivadas , Feminino , Humanos , Masculino , Músculo Liso Vascular , Miócitos de Músculo Liso/metabolismo , Osteogênese/genética , Fósforo/metabolismo , Ratos , Insuficiência Renal Crônica/complicações , Insuficiência Renal Crônica/metabolismo , Espectrometria de Massas em Tandem , Proteases Específicas de Ubiquitina/metabolismo , Proteases Específicas de Ubiquitina/farmacologia , Calcificação Vascular/metabolismo
3.
PLoS One ; 16(9): e0249438, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34473703

RESUMO

Muscle derived stem cells (MDSCs) and myoblast play an important role in myotube regeneration when muscle tissue is injured. However, these cells can be induced to differentiate into adipocytes once exposed to PPARγ activator like EPA and DHA that are highly suggested during pregnancy. The objective of this study aims at determining the identity of trans-differentiated cells by exploring the effect of EPA and DHA on C2C12 undergoing differentiation into brown and white adipocytes. DHA but not EPA committed C2C12 cells reprograming into white like adipocyte phenotype. Also, DHA promoted the expression of lipolysis regulating genes but had no effect on genes regulating ß-oxidation referring to its implication in lipid re-esterification. Furthermore, DHA impaired C2C12 cells differentiation into brown adipocytes through reducing the thermogenic capacity and mitochondrial biogenesis of derived cells independent of UCP1. Accordingly, DHA treated groups showed an increased accumulation of lipid droplets and suppressed mitochondrial maximal respiration and spare respiratory capacity. EPA, on the other hand, reduced myogenesis regulating genes, but no significant differences were observed in the expression of adipogenesis key genes. Likewise, EPA suppressed the expression of WAT signature genes indicating that EPA and DHA have an independent role on white adipogensis. Unlike DHA treatment, EPA supplementation had no effect on the differential of C2C12 cells into brown adipocytes. In conclusion, DHA is a potent adipogenic and lipogenic factor that can change the metabolic profile of muscle cells by increasing myocellular fat.


Assuntos
Adipócitos Brancos/efeitos dos fármacos , Ácidos Docosa-Hexaenoicos/farmacologia , Ácido Eicosapentaenoico/farmacologia , Adipócitos Marrons/efeitos dos fármacos , Adipócitos Brancos/citologia , Adipogenia/efeitos dos fármacos , Adipogenia/genética , Tecido Adiposo Marrom/citologia , Tecido Adiposo Marrom/efeitos dos fármacos , Animais , Linhagem Celular , Transdiferenciação Celular/efeitos dos fármacos , Transdiferenciação Celular/genética , DNA Mitocondrial , Regulação da Expressão Gênica/efeitos dos fármacos , Metabolismo dos Lipídeos/efeitos dos fármacos , Metabolismo dos Lipídeos/genética , Lipólise/efeitos dos fármacos , Camundongos , Mioblastos/citologia , Mioblastos/efeitos dos fármacos
4.
Development ; 146(14)2019 07 24.
Artigo em Inglês | MEDLINE | ID: mdl-31142539

RESUMO

An early step in pancreas development is marked by the expression of the transcription factor Pdx1 within the pancreatic endoderm, where it is required for the specification of all endocrine cell types. Subsequently, Pdx1 expression becomes restricted to the ß-cell lineage, where it plays a central role in ß-cell function. This pivotal role of Pdx1 at various stages of pancreas development makes it an attractive target to enhance pancreatic ß-cell differentiation and increase ß-cell function. In this study, we used a newly generated zebrafish reporter to screen over 8000 small molecules for modulators of pdx1 expression. We found four hit compounds and validated their efficacy at different stages of pancreas development. Notably, valproic acid treatment increased pancreatic endoderm formation, while inhibition of TGFß signaling led to α-cell to ß-cell transdifferentiation. HC toxin, another HDAC inhibitor, enhances ß-cell function in primary mouse and human islets. Thus, using a whole organism screening strategy, this study identified new pdx1 expression modulators that can be used to influence different steps in pancreas and ß-cell development.


Assuntos
Avaliação Pré-Clínica de Medicamentos/métodos , Ilhotas Pancreáticas/embriologia , Modelos Animais , Organogênese/efeitos dos fármacos , Bibliotecas de Moléculas Pequenas/análise , Peixe-Zebra , Animais , Animais Geneticamente Modificados , Células COS , Diferenciação Celular/efeitos dos fármacos , Diferenciação Celular/fisiologia , Transdiferenciação Celular/efeitos dos fármacos , Transdiferenciação Celular/genética , Células Cultivadas , Chlorocebus aethiops , Embrião não Mamífero , Regulação da Expressão Gênica no Desenvolvimento/efeitos dos fármacos , Inibidores de Histona Desacetilases/isolamento & purificação , Inibidores de Histona Desacetilases/farmacologia , Proteínas de Homeodomínio/genética , Proteínas de Homeodomínio/metabolismo , Humanos , Células Secretoras de Insulina/citologia , Células Secretoras de Insulina/efeitos dos fármacos , Células Secretoras de Insulina/fisiologia , Ilhotas Pancreáticas/efeitos dos fármacos , Ilhotas Pancreáticas/crescimento & desenvolvimento , Ilhotas Pancreáticas/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Organogênese/genética , Bibliotecas de Moléculas Pequenas/isolamento & purificação , Transativadores/genética , Transativadores/metabolismo , Ácido Valproico/isolamento & purificação , Ácido Valproico/farmacologia , Peixe-Zebra/embriologia , Peixe-Zebra/genética
5.
PLoS One ; 13(7): e0200210, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-29979748

RESUMO

Hearing loss is the most common sensorineural disorder, affecting over 5% of the population worldwide. Its most frequent cause is the loss of hair cells (HCs), the mechanosensory receptors of the cochlea. HCs transduce incoming sounds into electrical signals that activate auditory neurons, which in turn send this information to the brain. Although some spontaneous HC regeneration has been observed in neonatal mammals, the very small pool of putative progenitor cells that have been identified in the adult mammalian cochlea is not able to replace the damaged HCs, making any hearing impairment permanent. To date, guided differentiation of human cells to HC-like cells has only been achieved using either embryonic stem cells (ESCs) or induced pluripotent stem cells (iPSCs). However, use of such cell types suffers from a number of important disadvantages, such as the risk of tumourigenicity if transplanted into the host´s tissue. We have obtained cells expressing hair cell markers from cultures of human fibroblasts by overexpression of GFI1, Pou4f3 and ATOH1 (GPA), three genes that are known to play a critical role in the development of HCs. Immunocytochemical, qPCR and RNAseq analyses demonstrate the expression of genes typically expressed by HCs in the transdifferentiated cells. Our protocol represents a much faster approach than the methods applied to ESCs and iPSCs and validates the combination of GPA as a set of genes whose activation leads to the direct conversion of human somatic cells towards the hair cell lineage. Our observations are expected to contribute to the development of future therapies aimed at the regeneration of the auditory organ and the restoration of hearing.


Assuntos
Transdiferenciação Celular/fisiologia , Células Ciliadas Auditivas/citologia , Células Ciliadas Auditivas/metabolismo , Fatores de Transcrição/metabolismo , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Biomarcadores/metabolismo , Linhagem da Célula/efeitos dos fármacos , Linhagem da Célula/genética , Linhagem da Célula/fisiologia , Transdiferenciação Celular/efeitos dos fármacos , Transdiferenciação Celular/genética , Células Cultivadas , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Fator de Crescimento Epidérmico/farmacologia , Fibroblastos/citologia , Fibroblastos/efeitos dos fármacos , Fibroblastos/metabolismo , Expressão Gênica/efeitos dos fármacos , Células Ciliadas Auditivas/efeitos dos fármacos , Proteínas de Homeodomínio/genética , Proteínas de Homeodomínio/metabolismo , Humanos , Miosina VIIa , Miosinas/genética , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Fator de Transcrição Brn-3C/genética , Fator de Transcrição Brn-3C/metabolismo , Fatores de Transcrição/genética , Tretinoína/farmacologia
6.
Integr Cancer Ther ; 17(3): 1000-1008, 2018 09.
Artigo em Inglês | MEDLINE | ID: mdl-29896984

RESUMO

Cachexia has been recognized for a long time as an adverse effect of cancer. It is associated with reduced physical function, reduced tolerance to anticancer therapy, and reduced survival. This wasting syndrome is mainly known for an ongoing loss of skeletal muscle leading to progressive functional impairment and is driven by a variable combination of reduced food intake and abnormal metabolism. Cytokines derived from host immune system or the tumor itself is believed to play a role in promoting cancer cachexia. Circulating levels of cytokines, including IL-1α, IL-6, and TNFα have been identified in cancer patients but they probably only represent a small part of a changed and abnormal metabolism. Murine models have shown that browning of white adipose tissue (WAT) takes place early in the progression of cancer cachexia. Thus, browning of white adipose tissue is believed to be a strong contributor to the increased energy expenditure common in cachectic patients. Despite the severe implications of cancer cachexia for the patients and extensive research efforts, a more coherent and mechanistic explanation of the syndrome is lacking, and for many clinicians, cancer cachexia is still a vague concept. From a lung cancer perspective this commentary reviews the current knowledge on cancer cachexia mechanisms and identifies specific ways of clinical management regarding food intake, systemic inflammation, and muscular dysfunction. Much of what we know comes from preclinical studies. More translational research is needed for a future cancer cachexia screening tool to guide clinicians, and here possible variables for a cancer cachexia screening tool are considered.


Assuntos
Caquexia/etiologia , Caquexia/genética , Caquexia/terapia , Neoplasias Pulmonares/complicações , Neoplasias Pulmonares/terapia , Tecido Adiposo/metabolismo , Tecido Adiposo/patologia , Tecido Adiposo Marrom/metabolismo , Tecido Adiposo Marrom/patologia , Animais , Caquexia/diagnóstico , Transdiferenciação Celular/genética , Humanos , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/patologia , Camundongos , Técnicas de Diagnóstico Molecular , Ratos , Transdução de Sinais/genética , Redução de Peso/genética
7.
Curr Protein Pept Sci ; 18(2): 155-166, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-27001065

RESUMO

Utilizing both primary myometrial cells and a myometrial cell line, we show here that myometrial cells undergo transition to a myofibroblast-like phenotype after a biological insult of 72 hours serum starvation and serum add-back (SB: 1% to 10% FBS). We also found that thrombospondin-1 was increased and that the transforming growth factor-beta (TGFB)-SMAD3/4 pathway was activated. This pathway is a key mediator of fibrosis and extracellular matrix (ECM) deposition. Applying the same insult supplemented with TGFB3 (1-10ng/ml) and ascorbic acid (100µg/ml) in the serum add-back treatment, we further demonstrated that cells migrated into nodules containing collagen and fibronectin. The number of cellnodules was inversely related to the percentage serum add-back. Using transmission electron microscopy we demonstrated myofibroblast-like cells and fibril-like structures in the extracellular spaces of the nodules. This study is the first direct evidence of induction of myofibroblast transdifferentiation in cultured myometrial cells which is related to the increase of thrombospondin-1 (THBS1) and the activation of TGFBSMAD 3 / 4 pathways. Combined, these observations provide biochemical and direct morphological evidence that fibrotic responses can occur in cultured myometrial cells. The findings are the first to demonstrate uterine healing mechanisms at a molecular level. Our data support the concept that fibrosis may be an initial event in formation of fibroid which exhibits signaling pathways and molecular features of fibrosis and grow by both cellular proliferation and altered extracellular matrix accumulation. Our data assists in further understanding of myometrium tissue remodeling during gestation and postpartum.


Assuntos
Proliferação de Células/efeitos dos fármacos , Colágeno/genética , Fibronectinas/genética , Fibrose/genética , Miométrio/metabolismo , Ácido Ascórbico/farmacologia , Linhagem Celular , Proliferação de Células/genética , Transdiferenciação Celular/efeitos dos fármacos , Transdiferenciação Celular/genética , Feminino , Fibrose/patologia , Humanos , Redes e Vias Metabólicas/efeitos dos fármacos , Miofibroblastos/efeitos dos fármacos , Miofibroblastos/metabolismo , Miométrio/efeitos dos fármacos , Miométrio/patologia , Período Pós-Parto/genética , Período Pós-Parto/metabolismo , Gravidez , Cultura Primária de Células , Proteína Smad3/metabolismo , Trombospondina 1/metabolismo , Fator de Crescimento Transformador beta/genética , Fator de Crescimento Transformador beta3/farmacologia
8.
Immunol Cell Biol ; 93(3): 284-9, 2015 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-25643615

RESUMO

Cellular reprogramming refers to the conversion of one cell type into another by altering its epigenetic marks. This can be achieved by three different methods: somatic cell nuclear transfer, cell fusion and transcription factor (TF)-mediated reprogramming. TF-mediated reprogramming can occur through several means, either reverting backwards to a pluripotent state before redifferentiating to a new cell type (otherwise known as induced pluripotency), by transdifferentiating directly into a new cell type (bypassing the intermediate pluripotent stage), or, by using the induced pluripotency pathway without reaching the pluripotent state. The possibility of reprogramming any cell type of interest not only sheds new insights on cellular plasticity, but also provides a novel use of this technology across several platforms, most notably in cellular replacement therapies, disease modelling and drug screening. This review will focus on the different ways of implementing TF-mediated reprogramming, their associated epigenetic changes and its therapeutic potential.


Assuntos
Transdiferenciação Celular/genética , Reprogramação Celular/genética , Epigênese Genética , Células-Tronco Pluripotentes Induzidas/citologia , Medicina Regenerativa , Fatores de Transcrição/metabolismo , Animais , Terapia Biológica , Humanos
9.
Rev Med Chir Soc Med Nat Iasi ; 118(3): 699-705, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-25341288

RESUMO

A current area of interest is the determination of factors able to promote the transition from muscle to adipose tissue. The current review has highlighted that treatment of myoblasts with fatty acids (especially oleic acid) and thiazolidindiones causes conversion to adipocytes. The molecular mechanisms mediating the adipogenic action of thiazolidinediones and fatty acids in myoblasts could involve peroxisome proliferators-activated receptor-gamma (PPARgamma and CCAAT-enhancer-binding protein C/EBP. The role of 1,25-D3 in adipogenesis is mediated at the molecular level through VDR-dependent inhibition of C/EBP and PPARgamma expression and a decrease in PPARgamma transactivation activity. Vitamin D supplementation increases muscle strength and ultimately reduces the incidence of falls. Additional research is needed to fully clarify the role of nutritional factors in adipogenesis.


Assuntos
Adipócitos/metabolismo , Diferenciação Celular , Transdiferenciação Celular , Células Musculares/metabolismo , Músculo Esquelético/metabolismo , Tecido Adiposo/metabolismo , Fator de Ligação a CCAAT/metabolismo , Calcitriol/metabolismo , Diferenciação Celular/genética , Transdiferenciação Celular/genética , Medicina Baseada em Evidências , Ácidos Graxos/metabolismo , Humanos , Mioblastos/metabolismo , PPAR gama/metabolismo , Tiazolidinedionas/metabolismo , Fatores de Transcrição/metabolismo
10.
Mol Cell Biochem ; 393(1-2): 77-87, 2014 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-24756243

RESUMO

Platelet-derived growth factor-BB (PDGF-BB) and transforming growth factor-ß1 (TGF-ß1) are critically involved in idiopathic pulmonary fibrosis by inducing the proliferation and transdifferentiation of lung fibroblasts. In the present study, we examined the impact of diallyl disulfide (DADS), a garlic-derived compound, on such pathological conditions. DADS showed profound inhibitory effects on the PDGF-BB-induced proliferation of human and mouse lung fibroblasts. DADS also abrogated the TGF-ß1-induced expression of α-smooth muscle actin, type I collagen and fibronectin. Following treatment with DADS, the expression of cyclooxygenase-2 (COX-2) and the synthesis of prostaglandin E2 (PGE2) were found to be markedly enhanced, which in turn led to elevated cAMP levels in lung fibroblasts. Notably, the effect of DADS was largely abolished in the presence of either COX inhibitor indomethacin or siRNA-targeting COX-2, or in the absence of the PGE2 receptor EP2, supporting an essential role for the COX-2-PGE2-cAMP autocrine loop. Furthermore, we demonstrated that the upregulated expression of COX-2 was a result of increased level of histone 3 acetylation at COX-2 locus in DADS-treated cells. Together, these results suggest that DADS, by inducing COX-2 expression, may have therapeutic potential in treating lung fibrosis.


Assuntos
Compostos Alílicos/administração & dosagem , Ciclo-Oxigenase 2/biossíntese , Dinoprostona/biossíntese , Dissulfetos/administração & dosagem , Pulmão/efeitos dos fármacos , Animais , Becaplermina , Proliferação de Células/efeitos dos fármacos , Transdiferenciação Celular/efeitos dos fármacos , Transdiferenciação Celular/genética , Fibroblastos/citologia , Fibroblastos/metabolismo , Fibrose/tratamento farmacológico , Fibrose/patologia , Alho/química , Humanos , Pulmão/citologia , Pulmão/patologia , Camundongos , Proteínas Proto-Oncogênicas c-sis/metabolismo , Fator de Crescimento Transformador beta
11.
Ann N Y Acad Sci ; 1302: 30-34, 2013 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-23980536

RESUMO

Two types of fat, white adipose tissue (WAT) and brown adipose tissue (BAT), exist in mammals including adult humans. While WAT stores excess calories and an excessive accumulation of fat causes obesity, BAT dissipates energy to produce heat through nonshivering thermogenesis for protection against cold environments and provides the potential for the development of novel anti-obesity treatments. The hypothalamus plays a central role in the control of energy balance. Specifically, recent observations indicate the importance of the dorsomedial hypothalamus (DMH) in thermoregulation. We have found that the orexigenic neuropeptide Y (NPY) in the DMH has distinct actions in modulating adiposity and BAT thermogenesis. Knockdown of NPY in the DMH elevates the thermogenic activity of classic BAT and promotes the development of brown adipocytes in WAT, leading to increased thermogenesis. These findings identify a novel potential target for combating obesity.


Assuntos
Tecido Adiposo Marrom/fisiologia , Tecido Adiposo Branco/fisiologia , Transdiferenciação Celular/genética , Terapia Genética/métodos , Hipotálamo/fisiologia , Termogênese/genética , Tecido Adiposo Marrom/metabolismo , Tecido Adiposo Branco/metabolismo , Adulto , Animais , Técnicas de Silenciamento de Genes , Humanos , Hipotálamo/metabolismo , Neuropeptídeo Y/genética , Neuropeptídeo Y/metabolismo , Neuropeptídeo Y/fisiologia , Obesidade/genética , Obesidade/terapia , Transdução de Sinais/fisiologia , Termogênese/fisiologia
12.
PLoS One ; 7(3): e33766, 2012.
Artigo em Inglês | MEDLINE | ID: mdl-22438993

RESUMO

MicroRNAs (miRNAs) are non-coding RNAs that regulate gene expression in post-transcriptional fashion, and emerging studies support their importance in regulating many biological processes, including myogenic differentiation and muscle development. miR-29 is a promoting factor during myogenesis but its full spectrum of impact on muscle cells has yet to be explored. Here we describe an analysis of miR-29 affected transcriptome in C2C12 muscle cells using a high throughput RNA-sequencing platform. The results reveal that miR-29 not only functions to promote myogenic differentiation but also suppresses the transdifferentiation of myoblasts into myofibroblasts. miR-29 inhibits the fibrogenic differentiation through down-regulating both extracellular matrix genes and cell adhesion genes. We further demonstrate that miR-29 is under negative regulation by TGF-beta (TGF-ß)-Smad3 signaling via dual mechanisms of both inhibiting MyoD binding and enhancing Yin Yang 1 (YY1)-recruited Polycomb association. Together, these results identify miR-29 as a pleiotropic molecule in both myogenic and fibrogenic differentiation of muscle cells.


Assuntos
MicroRNAs/antagonistas & inibidores , MicroRNAs/genética , Mioblastos Esqueléticos/citologia , Mioblastos Esqueléticos/metabolismo , Miofibroblastos/citologia , Miofibroblastos/metabolismo , Proteína Smad3/metabolismo , Fator de Crescimento Transformador beta/metabolismo , Animais , Sequência de Bases , Linhagem Celular , Transdiferenciação Celular/genética , Transdiferenciação Celular/fisiologia , Regulação para Baixo , Camundongos , MicroRNAs/metabolismo , Modelos Biológicos , Proteína MyoD/metabolismo , Proteínas do Grupo Polycomb , Regiões Promotoras Genéticas , RNA Interferente Pequeno/genética , Proteínas Repressoras/metabolismo , Transdução de Sinais/fisiologia , Proteína Smad3/antagonistas & inibidores , Proteína Smad3/genética , Transcriptoma , Fator de Transcrição YY1/metabolismo
13.
Cytotherapy ; 13(9): 1057-65, 2011 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-21846294

RESUMO

BACKGROUND AIMS: GATA-4 is a cardiac transcription factor and plays an important role in cell lineage differentiation during development. We investigated whether overexpression of GATA-4 increases adult mesenchymal stromal cell (MSC) transdifferentiation into a cardiac phenotype in vitro. METHODS: MSC were harvested from rat bone marrow (BM) and transduced with GATA-4 (MSC(GATA-4)) using a murine stem cell virus (pMSCV) retroviral expression system. Gene expression in MSC(GATA-4) was analyzed using quantitative reverse transcription-polymerase chain reaction (RT-PCR) and Western blotting. Native cardiomyocytes (CM) were isolated from ventricles of neonatal rats. Myocardial transdifferentiation of MSC was determined by immunostaining and electrophysiologic recording. The transdifferentiation rate was calculated directly from flow cytometery. RESULTS: The expression of cardiac genes, including brain natriuretic peptide (BNP), Islet-1 and α-sarcomeric actinin (α-SA), was up-regulated in MSC(GATA-4) compared with control cells that were transfected with Green Fluorescent Protein (GFP) only (MSC(Null)). At the same time, insulin-like growth factor-binding protein (IGFBP)-4 was significantly up-regulated in MSC(GATA-4). A synchronous beating of MSC with native CM was detected and an action potential was recorded. Some GFP (+) cells were positive for α-SA staining after MSC were co-cultured with native CM for 7 days. The transdifferentiation rate was significantly higher in MSC(GATA-4). Functional studies indicated that the differentiation potential of MSC(GATA-4) was decreased by knockdown of IGFBP-4. CONCLUSIONS: Overexpression of GATA-4 significantly increases MSC differentiation into a myocardial phenotype, which might be associated with the up-regulation of IGFBP-4.


Assuntos
Células-Tronco Adultas/metabolismo , Medula Óssea/metabolismo , Fator de Transcrição GATA4/metabolismo , Proteína 4 de Ligação a Fator de Crescimento Semelhante à Insulina/metabolismo , Células-Tronco Mesenquimais/metabolismo , Células-Tronco Adultas/patologia , Animais , Medula Óssea/patologia , Linhagem da Célula/genética , Transdiferenciação Celular/genética , Células Cultivadas , Técnicas de Cocultura , Técnicas Eletrofisiológicas Cardíacas , Fator de Transcrição GATA4/genética , Técnicas de Silenciamento de Genes , Ventrículos do Coração/patologia , Proteína 4 de Ligação a Fator de Crescimento Semelhante à Insulina/genética , Células-Tronco Mesenquimais/patologia , Camundongos , Miócitos Cardíacos/patologia , Ratos , Transgenes/genética , Regulação para Cima
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