RESUMO
Ventricular septal defects (VSDs) are recognized as one of the commonest congenital heart diseases (CHD), accounting for up to 40% of all cardiac malformations, and occur as isolated CHDs as well as together with other cardiac and extracardiac congenital malformations in individual patients and families. The genetic etiology of VSD is complex and extraordinarily heterogeneous. Chromosomal abnormalities such as aneuploidy and structural variations as well as rare point mutations in various genes have been reported to be associated with this cardiac defect. This includes both well-defined syndromes with known genetic cause (e.g., DiGeorge syndrome and Holt-Oram syndrome) and so far undefined syndromic forms characterized by unspecific symptoms. Mutations in genes encoding cardiac transcription factors (e.g., NKX2-5 and GATA4) and signaling molecules (e.g., CFC1) have been most frequently found in VSD cases. Moreover, new high-resolution methods such as comparative genomic hybridization enabled the discovery of a high number of different copy number variations, leading to gain or loss of chromosomal regions often containing multiple genes, in patients with VSD. In this chapter, we will describe the broad genetic heterogeneity observed in VSD patients considering recent advances in this field.
Assuntos
Comunicação Interventricular , Humanos , Aberrações Cromossômicas , Variações do Número de Cópias de DNA/genética , Predisposição Genética para Doença/genética , Comunicação Interventricular/genética , Mutação , Fatores de Transcrição/genéticaRESUMO
Chondrocytes in mice developing osteoarthritis (OA) exhibit an aberrant response to the secreted cytokine transforming growth factor (TGF)-ß, consisting in a potentiation of intracellular signaling downstream of the transmembrane type I receptor kinase activin receptor-like kinase (ALK)1 against canonical TGF-ß receptor ALK5-mediated signaling. Unfortunately, the underlying mechanisms remain elusive. In order to identify novel druggable targets for OA, we aimed to investigate novel molecules regulating the ALK1/ALK5 balance in OA chondrocytes. We performed gene expression analysis of TGF-ß signaling modulators in joints from three different mouse models of OA and found an upregulated expression of the TGF-ß co-receptor Cripto (Tdgf1), which was validated in murine and human cartilage OA samples at the protein level. In vitro and ex vivo, elevated expression of Cripto favors the hypertrophic differentiation of chondrocytes, eventually contributing to tissue calcification. Furthermore, we found that Cripto participates in a TGF-ß-ALK1-Cripto receptor complex in the plasma membrane, thereby inducing catabolic SMAD1/5 signaling in chondrocytes. In conclusion, we demonstrate that Cripto is expressed in OA and plays a functional role promoting chondrocyte hypertrophy, thereby becoming a novel potential therapeutic target in OA, for which there is no efficient cure or validated biomarker. © 2021 The Authors. The Journal of Pathology published by John Wiley & Sons, Ltd. on behalf of The Pathological Society of Great Britain and Ireland.
Assuntos
Condrócitos/patologia , Proteínas Ligadas por GPI/metabolismo , Peptídeos e Proteínas de Sinalização Intercelular/metabolismo , Proteínas de Neoplasias/metabolismo , Osteoartrite/patologia , Proteínas Smad/metabolismo , Fator de Crescimento Transformador beta/metabolismo , Animais , Fator de Crescimento Epidérmico/metabolismo , Humanos , Hipertrofia/patologia , Glicoproteínas de Membrana/metabolismo , Camundongos , Transdução de Sinais/fisiologiaRESUMO
The activation of specific gene expression programs depends on the presence of the appropriate signals and the competence of cells to respond to those signals. Although it is well established that cellular competence is regulated in space and time, the molecular mechanisms underlying the loss of competence remain largely unknown. Here, we determine the time window during which zebrafish prospective ectoderm loses its ability to respond to Nodal signals, and show that this coincides with a decrease in the levels of the Nodal co-receptor One-eyed pinhead (Oep). Bypassing Oep using a photoactivatable receptor, or an Oep-independent ligand, allows activation of Nodal target genes for an extended period of time. These results suggest that the reduced expression of Oep causes the loss of responsiveness to Nodal signals in the prospective ectoderm. Indeed, extending the presence of Oep prolongs the window of competence to respond to Nodal signals. Our findings suggest a simple mechanism in which the decreasing level of one component of the Nodal signaling pathway regulates the loss of mesendodermal competence in the prospective ectoderm.
Assuntos
Desenvolvimento Embrionário/genética , Endoderma/embriologia , Proteínas de Homeodomínio/genética , Mesoderma/embriologia , Fatores de Transcrição/genética , Proteínas de Peixe-Zebra/genética , Peixe-Zebra , Animais , Animais Geneticamente Modificados , Ectoderma/embriologia , Ectoderma/metabolismo , Embrião não Mamífero , Endoderma/metabolismo , Regulação da Expressão Gênica no Desenvolvimento , Proteínas de Homeodomínio/metabolismo , Mesoderma/metabolismo , Proteína Nodal/metabolismo , Transdução de Sinais/genética , Fatores de Transcrição/metabolismo , Peixe-Zebra/embriologia , Peixe-Zebra/genética , Proteínas de Peixe-Zebra/metabolismoRESUMO
Cripto-1 is a protein expressed during embryonal development and has been linked to several malignant processes in cancer. Since the discovery of cripto-1 in the late 1980s, it has become a subject of biomarker investigation in several types of cancer which in many cases relies on immunolocalization of cripto-1 using antibodies. Investigating cripto-1 expression and localization in primary glioblastoma cells, we discovered nonspecific binding of cripto-1 antibody to the extracellular matrix Geltrex. A panel of four cripto-1 antibodies was investigated with respect to their binding to the Geltrex matrix and to the cripto-1 positive control cells NTERA2. The cripto-1 expression was varied for the different antibodies with respect to cellular localization and fixation methods. To further elaborate on these findings, we present a systematic review of cripto-1 antibodies found in the literature and highlight some possible cross reactants with data on sequence alignments and structural comparison of EGF domains.
Assuntos
Anticorpos Monoclonais/imunologia , Anticorpos Monoclonais/metabolismo , Matriz Extracelular/metabolismo , Proteínas Ligadas por GPI/imunologia , Glioblastoma/patologia , Peptídeos e Proteínas de Sinalização Intercelular/imunologia , Proteínas de Neoplasias/imunologia , Sequência de Aminoácidos , Anticorpos Monoclonais/classificação , Movimento Celular , Glioblastoma/imunologia , Glioblastoma/metabolismo , Humanos , Células Tumorais CultivadasRESUMO
Congenital heart defects are the most common birth defects in humans, and those that affect the proper alignment of the outflow tracts and septation of the ventricles are a highly significant cause of morbidity and mortality in infants. A late differentiating population of cardiac progenitors, referred to as the anterior second heart field (AHF), gives rise to the outflow tract and the majority of the right ventricle and provides an embryological context for understanding cardiac outflow tract alignment and membranous ventricular septal defects. However, the transcriptional pathways controlling AHF development and their roles in congenital heart defects remain incompletely elucidated. Here, we inactivated the gene encoding the transcription factor MEF2C in the AHF in mice. Loss of Mef2c function in the AHF results in a spectrum of outflow tract alignment defects ranging from overriding aorta to double-outlet right ventricle and dextro-transposition of the great arteries. We identify Tdgf1, which encodes a Nodal co-receptor (also known as Cripto), as a direct transcriptional target of MEF2C in the outflow tract via an AHF-restricted Tdgf1 enhancer. Importantly, both the MEF2C and TDGF1 genes are associated with congenital heart defects in humans. Thus, these studies establish a direct transcriptional pathway between the core cardiac transcription factor MEF2C and the human congenital heart disease gene TDGF1. Moreover, we found a range of outflow tract alignment defects resulting from a single genetic lesion, supporting the idea that AHF-derived outflow tract alignment defects may constitute an embryological spectrum rather than distinct anomalies.
Assuntos
Fator de Crescimento Epidérmico/fisiologia , Regulação da Expressão Gênica no Desenvolvimento , Glicoproteínas de Membrana/fisiologia , Proteínas de Neoplasias/fisiologia , Animais , Animais Recém-Nascidos , Modelos Animais de Doenças , Fator de Crescimento Epidérmico/genética , Feminino , Deleção de Genes , Coração/embriologia , Cardiopatias Congênitas/genética , Comunicação Interventricular/genética , Ventrículos do Coração , Humanos , Hibridização In Situ , Fatores de Transcrição MEF2/genética , Fatores de Transcrição MEF2/fisiologia , Masculino , Glicoproteínas de Membrana/genética , Camundongos , Morfogênese/genética , Proteínas de Neoplasias/genética , Organogênese , Análise de Sequência de RNA , Distribuição Tecidual , Transcrição Gênica , Transposição dos Grandes Vasos/genéticaRESUMO
Induced pluripotent stem cells (iPS cells) are a prospective resource for regenerative biomedicine. iPS cells can differentiate into any type of stem, progenitor and somatic cells to help replace structures within damaged organs or tissues. However, iPS cells themselves, can produce malignant tumors if they are injected into the body of an immunocompatible or immunodeficient recipient. Thus, it is necessary to detect any residual iPS cells content in biomedical cell products obtained from iPS cells and destined for transplantation. In this article we describe searches for iPS cells in heterogeneous cell mixtures, using two different methods-quantitative RT-PCR and droplet digital PCR (ddPCR). In experiments with various heterogeneous mixtures, including mixtures with neural stem cells, we found that the OCT4, TDGF1 and LIN28 genes are the best markers for such a search, and droplet digital PCR provides the greatest measurement accuracy, which is 0.002%. Thus, we have confirmed the advantage of using droplet digital PCR in the search for pluripotent stem cells in heterogeneous cell mixtures. We hope that this data can be useful for biosafety control in regenerative biomedicine.
Assuntos
Marcadores Genéticos , Células-Tronco Embrionárias Humanas/citologia , Células-Tronco Pluripotentes Induzidas/citologia , Linhagem Celular , Técnicas de Cocultura , Contenção de Riscos Biológicos , Proteínas Ligadas por GPI/genética , Células-Tronco Embrionárias Humanas/química , Humanos , Células-Tronco Pluripotentes Induzidas/química , Peptídeos e Proteínas de Sinalização Intercelular/genética , Proteínas de Neoplasias/genética , Fator 3 de Transcrição de Octâmero/genética , Proteínas de Ligação a RNA/genética , Reação em Cadeia da Polimerase em Tempo Real , Medicina RegenerativaRESUMO
BACKGROUND: Accumulating evidence has highlighted the potential role of long non-coding RNAs (lncRNAs) in the biological behaviors of glioblastoma stem cells (GSCs). Here, we elucidated the function and possible molecular mechanisms of the effect of lncRNA-SOX2OT on the biological behaviors of GSCs. RESULTS: Real-time PCR demonstrated that SOX2OT expression was up-regulated in glioma tissues and GSCs. Knockdown of SOX2OT inhibited the proliferation, migration and invasion of GSCs, and promoted GSCs apoptosis. MiR-194-5p and miR-122 were down-regulated in human glioma tissues and GSCs, and miR-194-5p and miR-122 respectively exerted tumor-suppressive functions by inhibiting the proliferation, migration and invasion of GSCs, while promoting GSCs apoptosis. Knockdown of SOX2OT significantly increased the expression of miR-194-5p and miR-122 in GSCs. Dual-luciferase reporter assay revealed that SOX2OT bound to both miR-194-5p and miR-122. SOX3 and TDGF-1 were up-regulated in human glioma tissues and GSCs. Knockdown of SOX3 inhibited the proliferation, migration and invasion of GSCs, promoted GSCs apoptosis, and decreased TDGF-1 mRNA and protein expression through direct binding to the TDGF-1 promoter. Over-expression of miR-194-5p and miR-122 decreased the mRNA and protein expression of SOX3 by targeting its 3'UTR. Knockdown of TDGF-1 inhibited the proliferation, migration and invasion of GSCs, promoted GSCs apoptosis, and inhibited the JAK/STAT signaling pathway. Furthermore, SOX3 knockdown also inhibited the SOX2OT expression through direct binding to the SOX2OT promoter and formed a positive feedback loop. CONCLUSION: This study is the first to demonstrate that the SOX2OT-miR-194-5p/miR-122-SOX3-TDGF-1 pathway forms a positive feedback loop and regulates the biological behaviors of GSCs, and these findings might provide a novel strategy for glioma treatment.
Assuntos
Neoplasias Encefálicas/patologia , Proteínas Ligadas por GPI/genética , Glioblastoma/patologia , Peptídeos e Proteínas de Sinalização Intercelular/genética , MicroRNAs/genética , Proteínas de Neoplasias/genética , RNA Longo não Codificante/genética , Fatores de Transcrição SOXB1/genética , Regulação para Cima , Animais , Neoplasias Encefálicas/genética , Neoplasias Encefálicas/metabolismo , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células , Retroalimentação Fisiológica , Proteínas Ligadas por GPI/metabolismo , Regulação Neoplásica da Expressão Gênica , Técnicas de Silenciamento de Genes , Glioblastoma/genética , Glioblastoma/metabolismo , Humanos , Peptídeos e Proteínas de Sinalização Intercelular/metabolismo , Camundongos , Proteínas de Neoplasias/metabolismo , Transplante de Neoplasias , Células-Tronco Neoplásicas/metabolismo , Células-Tronco Neoplásicas/patologia , Fatores de Transcrição SOXB1/metabolismoRESUMO
Cripto is a small glycosylphosphatidylinisitol (GPI)-anchored and secreted oncofetal protein that plays important roles in regulating normal physiological processes, including stem cell differentiation, embryonal development, and tissue growth and remodeling, as well as pathological processes such as tumor initiation and progression. Cripto functions as a co-receptor for TGF-ß ligands such as Nodal, GDF1, and GDF3. Soluble and secreted forms of Cripto also exhibit growth factor-like activity and activate SRC/MAPK/PI3K/AKT pathways. Glucose-Regulated Protein 78 kDa (GRP78) binds Cripto at the cell surface and has been shown to be required for Cripto signaling via both TGF-ß and SRC/MAPK/PI3K/AKT pathways. To provide a comprehensive overview of the scientific literature related to Cripto, we performed, for the first time, a bibliometric analysis of the biological roles of Cripto as reported in the scientific literature covering the last 10 years. We present different fields of knowledge in comprehensive areas of research on Cripto, ranging from basic to translational research, using a keyword-driven approach. Our ultimate aim is to aid the scientific community in conducting targeted research by identifying areas where research has been conducted so far and, perhaps more importantly, where critical knowledge is still missing.
RESUMO
PURPOSE: We investigated the relationships between biomarkers related to endoplasmic reticulum stress proteins (glucose-regulated protein of molecular mass 78 [GRP78] and Cripto-1 [teratocarcinoma-derived growth factor 1 protein]), pathologic response, and prognosis in locally advanced rectal cancer. MATERIALS AND METHODS: All clinical stage II and III rectal cancer patients received 50.4 Gy over 5.5 weeks, plus 5-fluorouracil (400 mg/m(2)/day) and leucovorin (20 mg/m(2)/day) bolus on days 1 to 5 and 29 to 33, and surgery was performed at 7 to 10 weeks after completion of all therapies. Expression of GRP78 and Cripto-1 proteins was determined by immunohistochemistry and was assessed in 101 patients with rectal cancer treated with neoadjuvant chemoradiotherapy (CRT). RESULTS: High expression of GRP78 and Cripto-1 proteins was observed in 86 patients (85.1%) and 49 patients (48.5%), respectively. Low expression of GRP78 protein was associated with a significantly high rate of down staging (80.0% vs. 52.3%, respectively; p=0.046) and a significantly low rate of recurrence (0% vs. 33.7%, respectively; p=0.008) compared with high expression of GRP78 protein. Mean recurrence-free survival according to GRP78 expression could not be estimated because the low expression group did not develop recurrence events but showed a significant correlation with time to recurrence, based on the log rank method (p=0.007). GRP78 also showed correlation with overall survival, based on the log rank method (p=0.045). CONCLUSION: GRP78 expression is a predictive and prognostic factor for down staging, recurrence, and survival in rectal cancer patients treated with 5-fluorouracil and leucovorin neoadjuvant CRT.
Assuntos
Quimiorradioterapia Adjuvante/métodos , Proteínas Ligadas por GPI/metabolismo , Proteínas de Choque Térmico/metabolismo , Peptídeos e Proteínas de Sinalização Intercelular/metabolismo , Terapia Neoadjuvante , Proteínas de Neoplasias/metabolismo , Neoplasias Retais/metabolismo , Neoplasias Retais/terapia , Idoso , Biomarcadores Tumorais , Chaperona BiP do Retículo Endoplasmático , Feminino , Fluoruracila/administração & dosagem , Fluoruracila/uso terapêutico , Proteínas Ligadas por GPI/genética , Proteínas de Choque Térmico/genética , Humanos , Peptídeos e Proteínas de Sinalização Intercelular/genética , Laparoscopia , Leucovorina/administração & dosagem , Leucovorina/uso terapêutico , Masculino , Pessoa de Meia-Idade , Proteínas de Neoplasias/genética , PrognósticoRESUMO
Congenital Heart Disease (CHD) is the most frequent and deadly birth defect. Patients with CHD that survive the neonatal period often progress to develop advanced heart failure requiring specialized treatment including cardiac transplantation. A full understanding of the transcriptional networks that direct cardiac progenitors during heart development will enhance our understanding of both normal cardiac function and pathological states. These findings will also have important applications for emerging therapies and the treatment of congenital heart disease. Furthermore, a number of shared transcriptional pathways or networks have been proposed to regulate the development and regeneration of tissues such as the heart. We have utilized transgenic technology to isolate and characterize cardiac progenitor cells from the developing mouse heart and have begun to define specific transcriptional networks of cardiovascular development. Initial studies identified Tdgf1 as a potential target of Nkx2-5. To mechanistically dissect the regulation of this molecular program, we utilized an array of molecular biological techniques to confirm that Nkx2-5 is an upstream regulator of the Tdgf1 gene in early cardiac development. These studies further define Nkx2-5 mediated transcriptional networks and enhance our understanding of cardiac morphogenesis.