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Pesquisa : E01.370.225.500.373 [Categoria DeCS]
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  1 / 1602 MEDLINE  
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[PMID]:29382818
[Au] Autor:Sánchez-Iranzo H; Galardi-Castilla M; Minguillón C; Sanz-Morejón A; González-Rosa JM; Felker A; Ernst A; Guzmán-Martínez G; Mosimann C; Mercader N
[Ad] Endereço:Development of the Epicardium and Its Role during Regeneration Group, Centro Nacional de Investigaciones Cardiovasculares (CNIC-ISCIII), Melchor Fernández Almagro 3, 28029, Madrid, Spain.
[Ti] Título:Tbx5a lineage tracing shows cardiomyocyte plasticity during zebrafish heart regeneration.
[So] Source:Nat Commun;9(1):428, 2018 01 30.
[Is] ISSN:2041-1723
[Cp] País de publicação:England
[La] Idioma:eng
[Ab] Resumo:During development, mesodermal progenitors from the first heart field (FHF) form a primitive cardiac tube, to which progenitors from the second heart field (SHF) are added. The contribution of FHF and SHF progenitors to the adult zebrafish heart has not been studied to date. Here we find, using genetic tbx5a lineage tracing tools, that the ventricular myocardium in the adult zebrafish is mainly derived from tbx5a cells, with a small contribution from tbx5a SHF progenitors. Notably, ablation of ventricular tbx5a -derived cardiomyocytes in the embryo is compensated by expansion of SHF-derived cells. In the adult, tbx5a expression is restricted to the trabeculae and excluded from the outer cortical layer. tbx5a-lineage tracing revealed that trabecular cardiomyocytes can switch their fate and differentiate into cortical myocardium during adult heart regeneration. We conclude that a high degree of cardiomyocyte cell fate plasticity contributes to efficient regeneration.
[Mh] Termos MeSH primário: Ventrículos do Coração/citologia
Miocárdio/citologia
Miócitos Cardíacos/citologia
Regeneração/genética
Proteínas com Domínio T-Box/genética
Peixe-Zebra/genética
[Mh] Termos MeSH secundário: Animais
Animais Geneticamente Modificados
Diferenciação Celular
Linhagem da Célula/genética
Rastreamento de Células
Embrião não Mamífero
Regulação da Expressão Gênica no Desenvolvimento
Genes Reporter
Proteínas de Fluorescência Verde/genética
Proteínas de Fluorescência Verde/metabolismo
Ventrículos do Coração/crescimento & desenvolvimento
Ventrículos do Coração/metabolismo
Proteínas Luminescentes/genética
Proteínas Luminescentes/metabolismo
Miocárdio/metabolismo
Miócitos Cardíacos/metabolismo
Cadeias Leves de Miosina/genética
Cadeias Leves de Miosina/metabolismo
Organogênese/genética
Células-Tronco/citologia
Células-Tronco/metabolismo
Proteínas com Domínio T-Box/deficiência
Peixe-Zebra/crescimento & desenvolvimento
Peixe-Zebra/metabolismo
[Pt] Tipo de publicação:JOURNAL ARTICLE; RESEARCH SUPPORT, NON-U.S. GOV'T
[Nm] Nome de substância:
0 (Luminescent Proteins); 0 (Myosin Light Chains); 0 (T-Box Domain Proteins); 0 (T-box transcription factor 5); 0 (red fluorescent protein); 147336-22-9 (Green Fluorescent Proteins)
[Em] Mês de entrada:1803
[Cu] Atualização por classe:180307
[Lr] Data última revisão:
180307
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:180201
[St] Status:MEDLINE
[do] DOI:10.1038/s41467-017-02650-6


  2 / 1602 MEDLINE  
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[PMID]:29335514
[Au] Autor:Kaiser M; Jug F; Julou T; Deshpande S; Pfohl T; Silander OK; Myers G; van Nimwegen E
[Ad] Endereço:Biozentrum, University of Basel and Swiss Institute of Bioinformatics, Klingelbergstrasse 50/70, 4056, Basel, Switzerland.
[Ti] Título:Monitoring single-cell gene regulation under dynamically controllable conditions with integrated microfluidics and software.
[So] Source:Nat Commun;9(1):212, 2018 01 15.
[Is] ISSN:2041-1723
[Cp] País de publicação:England
[La] Idioma:eng
[Ab] Resumo:Much is still not understood about how gene regulatory interactions control cell fate decisions in single cells, in part due to the difficulty of directly observing gene regulatory processes in vivo. We introduce here a novel integrated setup consisting of a microfluidic chip and accompanying analysis software that enable long-term quantitative tracking of growth and gene expression in single cells. The dual-input Mother Machine (DIMM) chip enables controlled and continuous variation of external conditions, allowing direct observation of gene regulatory responses to changing conditions in single cells. The Mother Machine Analyzer (MoMA) software achieves unprecedented accuracy in segmenting and tracking cells, and streamlines high-throughput curation with a novel leveraged editing procedure. We demonstrate the power of the method by uncovering several novel features of an iconic gene regulatory program: the induction of Escherichia coli's lac operon in response to a switch from glucose to lactose.
[Mh] Termos MeSH primário: Regulação Bacteriana da Expressão Gênica
Técnicas Analíticas Microfluídicas/métodos
Análise de Célula Única/métodos
Software
[Mh] Termos MeSH secundário: Algoritmos
Rastreamento de Células/instrumentação
Rastreamento de Células/métodos
Escherichia coli/citologia
Escherichia coli/efeitos dos fármacos
Escherichia coli/genética
Glucose/farmacologia
Óperon Lac/genética
Lactose/farmacologia
Análise de Célula Única/instrumentação
[Pt] Tipo de publicação:JOURNAL ARTICLE
[Nm] Nome de substância:
IY9XDZ35W2 (Glucose); J2B2A4N98G (Lactose)
[Em] Mês de entrada:1803
[Cu] Atualização por classe:180305
[Lr] Data última revisão:
180305
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:180117
[St] Status:MEDLINE
[do] DOI:10.1038/s41467-017-02505-0


  3 / 1602 MEDLINE  
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[PMID]:29228311
[Au] Autor:Hinrichsen M; Lenz M; Edwards JM; Miller OK; Mochrie SGJ; Swain PS; Schwarz-Linek U; Regan L
[Ad] Endereço:Department of Molecular Biophysics and Biochemistry, Yale University, 266 Whitney Avenue, New Haven, CT06511, USA.
[Ti] Título:A new method for post-translationally labeling proteins in live cells for fluorescence imaging and tracking.
[So] Source:Protein Eng Des Sel;30(12):771-780, 2017 12 01.
[Is] ISSN:1741-0134
[Cp] País de publicação:England
[La] Idioma:eng
[Ab] Resumo:We present a novel method to fluorescently label proteins, post-translationally, within live Saccharomycescerevisiae. The premise underlying this work is that fluorescent protein (FP) tags are less disruptive to normal processing and function when they are attached post-translationally, because target proteins are allowed to fold properly and reach their final subcellular location before being labeled. We accomplish this post-translational labeling by expressing the target protein fused to a short peptide tag (SpyTag), which is then covalently labeled in situ by controlled expression of an open isopeptide domain (SpyoIPD, a more stable derivative of the SpyCatcher protein) fused to an FP. The formation of a covalent bond between SpyTag and SpyoIPD attaches the FP to the target protein. We demonstrate the general applicability of this strategy by labeling several yeast proteins. Importantly, we show that labeling the membrane protein Pma1 in this manner avoids the mislocalization and growth impairment that occur when Pma1 is genetically fused to an FP. We also demonstrate that this strategy enables a novel approach to spatiotemporal tracking in single cells and we develop a Bayesian analysis to determine the protein's turnover time from such data.
[Mh] Termos MeSH primário: Rastreamento de Células/métodos
Corantes Fluorescentes/metabolismo
Engenharia de Proteínas/métodos
Processamento de Proteína Pós-Traducional
Análise de Célula Única/métodos
Espectrometria de Fluorescência/métodos
[Mh] Termos MeSH secundário: Corantes Fluorescentes/análise
Corantes Fluorescentes/química
Saccharomyces cerevisiae/genética
Saccharomyces cerevisiae/metabolismo
[Pt] Tipo de publicação:JOURNAL ARTICLE; RESEARCH SUPPORT, N.I.H., EXTRAMURAL; RESEARCH SUPPORT, NON-U.S. GOV'T
[Nm] Nome de substância:
0 (Fluorescent Dyes)
[Em] Mês de entrada:1802
[Cu] Atualização por classe:180223
[Lr] Data última revisão:
180223
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:171212
[St] Status:MEDLINE
[do] DOI:10.1093/protein/gzx059


  4 / 1602 MEDLINE  
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[PMID]:29175323
[Au] Autor:Yamauchi K; Li J; Morikawa K; Liu L; Shirayoshi Y; Nakatsuji N; Elliott DA; Hisatome I; Suemori H
[Ad] Endereço:Laboratory of Embryonic Stem Cell Research, Department of Regeneration Science and Engineering, Institute for Frontier Life and Medical Sciences, Kyoto University, 53 Kawahara-cho, Shogoin, Sakyo-ku, Kyoto 606-8507, Japan.
[Ti] Título:Isolation and characterization of ventricular-like cells derived from NKX2-5 and MLC2v double knock-in human pluripotent stem cells.
[So] Source:Biochem Biophys Res Commun;495(1):1278-1284, 2018 01 01.
[Is] ISSN:1090-2104
[Cp] País de publicação:United States
[La] Idioma:eng
[Ab] Resumo:Human pluripotent stem cell (hPSC)-derived cardiomyocytes (CMs) are a promising source for cell transplantation into the damaged heart, which has limited regenerative ability. Many methods have been developed to obtain large amounts of functional CMs from hPSCs for therapeutic applications. However, during the differentiation process, a mixed population of various cardiac cells, including ventricular, atrial, and pacemaker cells, is generated, which hampers the proper functional analysis and evaluation of cell properties. Here, we established NKX2-5 and MLC2v hPSC double knock-ins that allow for labeling, tracing, purification, and analysis of the development of ventricular cells from early to late stages. As with the endogenous transcriptional activities of these genes, MLC2v-mCherry expression following NKX2-5-eGFP expression was observed under previously established culture conditions, which mimic the in vivo cardiac developmental process. Patch-clamp and microelectrode array electrophysiological analyses showed that the NKX2-5 and MLC2v double-positive cells possess ventricular-like properties. The results demonstrate that the NKX2-5 and MLC2v hPSCs provide a powerful model system to capture region-specific cardiac differentiation from early to late stages. Our study would facilitate subtype-specific cardiac development and functional analysis using the hPSC-derived sources.
[Mh] Termos MeSH primário: Técnicas de Cultura Celular por Lotes/métodos
Miosinas Cardíacas/metabolismo
Rastreamento de Células/métodos
Ventrículos do Coração/citologia
Proteína Homeobox Nkx-2.5/metabolismo
Miócitos Cardíacos/citologia
Cadeias Leves de Miosina/metabolismo
Células-Tronco Pluripotentes/citologia
[Mh] Termos MeSH secundário: Miosinas Cardíacas/genética
Diferenciação Celular/fisiologia
Separação Celular/métodos
Células Cultivadas
Técnicas de Introdução de Genes
Genes Reporter/genética
Ventrículos do Coração/metabolismo
Proteína Homeobox Nkx-2.5/genética
Seres Humanos
Miócitos Cardíacos/metabolismo
Cadeias Leves de Miosina/genética
Células-Tronco Pluripotentes/metabolismo
Engenharia Tecidual/métodos
[Pt] Tipo de publicação:JOURNAL ARTICLE; RESEARCH SUPPORT, NON-U.S. GOV'T
[Nm] Nome de substância:
0 (Homeobox Protein Nkx-2.5); 0 (Myosin Light Chains); 0 (NKX2-5 protein, human); 0 (myosin light chain 2); EC 3.6.1.- (Cardiac Myosins)
[Em] Mês de entrada:1712
[Cu] Atualização por classe:180105
[Lr] Data última revisão:
180105
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:171128
[St] Status:MEDLINE


  5 / 1602 MEDLINE  
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[PMID]:28464210
[Au] Autor:Odeleye AOO; Castillo-Avila S; Boon M; Martin H; Coopman K
[Ad] Endereço:Centre for Biological Engineering, Loughborough University, Loughborough LE11 3TU, United Kingdom.
[Ti] Título:Development of an optical system for the non-invasive tracking of stem cell growth on microcarriers.
[So] Source:Biotechnol Bioeng;114(9):2032-2042, 2017 09.
[Is] ISSN:1097-0290
[Cp] País de publicação:United States
[La] Idioma:eng
[Ab] Resumo:The emergence of medicinal indications for stem cell therapies has seen a need to develop the manufacturing capacity for adherent cells such as mesenchymal stem cells (MSCs). One such development is in the use of microcarriers, which facilitate enhanced cell densities for adherent stem cell cultures when compared with 2D culture platforms. Given the variety of stem cell expansion systems commercially available, novel methods of non-invasive and automated monitoring of cell number, confluence, and aggregation, within disparate environments, will become imperative to process control, ensuring reliable and consistent performance. The in situ epi-illumination of mouse embryonic fibroblasts and human mesenchymal stem cells attached to Cytodex 1 and 3 microcarriers was achieved using a bespoke microscope. Robust image processing techniques were developed to provide quantitative measurements of confluence, aggregate recognition, and cell number, without the need for fluorescent labeling or cell detachment. Large datasets of cells counted on individual microcarriers were statistically analyzed and compared with NucleoCounter measurements, with an average difference of less than 7% observed from days 0 to 6 of a 12-day culture noted, prior to the onset of aggregation. The developed image acquisition system and post-processing methodologies were successfully applied to dynamically moving colonized microcarriers. The proposed system offers a novel method of cell identification at the individual level, to consistently and accurately assess viable cell number, confluence, and cell distribution, while also minimizing the variability inherent in the current invasive means by which cells adhered to microcarriers are analyzed. Biotechnol. Bioeng. 2017;114: 2032-2042. © 2017 The Authors. Biotechnology and Bioengineering Published by Wiley Periodicals, Inc.
[Mh] Termos MeSH primário: Rastreamento de Células/instrumentação
Aumento da Imagem/instrumentação
Transplante de Células-Tronco Mesenquimais/instrumentação
Células Mesenquimais Estromais/citologia
Células Mesenquimais Estromais/fisiologia
Microscopia/instrumentação
Dispositivos Ópticos
[Mh] Termos MeSH secundário: Crescimento Celular
Células Cultivadas
Seres Humanos
Aumento da Imagem/métodos
Miniaturização
Reconhecimento Automatizado de Padrão/métodos
Reprodutibilidade dos Testes
Sensibilidade e Especificidade
[Pt] Tipo de publicação:EVALUATION STUDIES; JOURNAL ARTICLE; RESEARCH SUPPORT, NON-U.S. GOV'T
[Em] Mês de entrada:1711
[Cu] Atualização por classe:171226
[Lr] Data última revisão:
171226
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:170503
[St] Status:MEDLINE
[do] DOI:10.1002/bit.26328


  6 / 1602 MEDLINE  
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[PMID]:29215830
[Au] Autor:Poveshchenko AF; Solovieva AO; Zubareva KE; Strunkin DN; Gricyk OB; Poveshchenko OV; Shurlygina AV; Konenkov VI
[Ti] Título:[Study of migration and distribution of bone marrow cells transplanted animals with B16 melanoma ].
[So] Source:Patol Fiziol Eksp Ter;61(2):10-21, 2017 Apr-Jun.
[Is] ISSN:0031-2991
[Cp] País de publicação:Russia (Federation)
[La] Idioma:rus
[Ab] Resumo:Purpose. Reveal features migration and distribution of syngeneic bone marrow cells (BMC) and subpopulations (MSC) after transplantation into the recipient carrier B16 melanoma bodies. Methods. We used mouse male and female C57BL/6 mice. Induction of Tumor Growth: B16 melanoma cells implanted subcutaneously into right hind paw of female C57BL/6 mice at a dose of 2.5 x 105 cells / mouse. migration study in vivo distribution and BMC and MSC was performed using genetic markers - Y-chromosome specific sequence line male C57Bl/6 syngeneic intravenous transplantation in females using the polymerase chain reaction (PCR) in real time on Authorized Termal Cycler - Light Cycler 480 II / 96 (Roche). Introduction suspension of unseparated bone marrow cells, mesenchymal stem cells from donor to recipient male mice (syngeneic recipient female C57BL/6), followed by isolation of recipients of organs was performed at regular intervals, then of organ recipients isolated DNA. Results. It was shown that bone marrow cells positive for Y-chromosome in migrate lymphoid (lymph nodes, spleen, bone marrow) or in non-lymphoid organs (liver, heart, brain, skin) syngeneic recipients. In addition to the migration of cells from the bone marrow to other organs, there is a way back migration of cells from the circulation to the bone marrow. B16 melanoma stimulates the migration of transplanted MSCs and BMC in bone marrow. It is found that tumor growth enhanced migration of transplanted bone marrow cells, including populations of MSCs in the bone marrow. In the early stages of tumor formation MSC migration activity higher than the BMC. In the later stages of tumor formation undivided population of bone marrow cells migrate to the intense swelling compared with a population of MSCs. Conclusion. The possibility of using bone marrow MSCs for targeted therapy of tumor diseases, because migration of MSCs in tumor tissue can be used to effectively deliver anticancer drugs.
[Mh] Termos MeSH primário: Transplante de Medula Óssea
Movimento Celular
Rastreamento de Células/métodos
Melanoma/metabolismo
Transplante de Células-Tronco Mesenquimais
Neoplasias Experimentais/metabolismo
[Mh] Termos MeSH secundário: Aloenxertos
Animais
Melanoma/patologia
Camundongos
Neoplasias Experimentais/patologia
[Pt] Tipo de publicação:JOURNAL ARTICLE
[Em] Mês de entrada:1712
[Cu] Atualização por classe:171221
[Lr] Data última revisão:
171221
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:171208
[St] Status:MEDLINE


  7 / 1602 MEDLINE  
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[PMID]:28743800
[Au] Autor:Ghosh A; Syed SM; Tanwar PS
[Ad] Endereço:Gynaecology Oncology Group, School of Biomedical Sciences and Pharmacy, University of Newcastle, Callaghan, New South Wales, 2308, Australia.
[Ti] Título: genetic cell lineage tracing reveals that oviductal secretory cells self-renew and give rise to ciliated cells.
[So] Source:Development;144(17):3031-3041, 2017 09 01.
[Is] ISSN:1477-9129
[Cp] País de publicação:England
[La] Idioma:eng
[Ab] Resumo:The epithelial lining of the fallopian tube is vital for fertility, providing nutrition to gametes and facilitating their transport. It is composed of two major cell types: secretory cells and ciliated cells. Interestingly, human ovarian cancer precursor lesions primarily consist of secretory cells. It is unclear why secretory cells are the dominant cell type in these lesions. Additionally, the underlying mechanisms governing fallopian tube epithelial homoeostasis are unknown. In the present study, we showed that across the different developmental stages of mouse oviduct, secretory cells are the most frequently dividing cells of the oviductal epithelium. genetic cell lineage tracing showed that secretory cells not only self-renew, but also give rise to ciliated cells. Analysis of a Wnt reporter mouse model and various Wnt target genes showed that the Wnt signaling pathway is involved in oviductal epithelial homoeostasis. By developing two triple-transgenic mouse models, we showed that Wnt/ß-catenin signaling is essential for self-renewal as well as the differentiation of secretory cells. In summary, our results provide mechanistic insight into oviductal epithelial homoeostasis.
[Mh] Termos MeSH primário: Linhagem da Célula
Autorrenovação Celular
Rastreamento de Células
Cílios/metabolismo
Oviductos/citologia
Oviductos/secreção
[Mh] Termos MeSH secundário: Animais
Diferenciação Celular
Proliferação Celular
Epitélio/patologia
Tubas Uterinas/patologia
Feminino
Predisposição Genética para Doença
Seres Humanos
Camundongos Endogâmicos C57BL
Neoplasias Ovarianas/patologia
Fator de Transcrição PAX8/metabolismo
Fatores de Risco
Via de Sinalização Wnt
beta Catenina/metabolismo
[Pt] Tipo de publicação:JOURNAL ARTICLE; RESEARCH SUPPORT, NON-U.S. GOV'T
[Nm] Nome de substância:
0 (PAX8 Transcription Factor); 0 (Pax8 protein, mouse); 0 (beta Catenin)
[Em] Mês de entrada:1710
[Cu] Atualização por classe:171126
[Lr] Data última revisão:
171126
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:170727
[St] Status:MEDLINE
[do] DOI:10.1242/dev.149989


  8 / 1602 MEDLINE  
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[PMID]:28465336
[Au] Autor:Farrell DL; Weitz O; Magnasco MO; Zallen JA
[Ad] Endereço:Howard Hughes Medical Institute and Developmental Biology Program, Sloan Kettering Institute, New York, NY 10065, USA.
[Ti] Título:SEGGA: a toolset for rapid automated analysis of epithelial cell polarity and dynamics.
[So] Source:Development;144(9):1725-1734, 2017 05 01.
[Is] ISSN:1477-9129
[Cp] País de publicação:England
[La] Idioma:eng
[Ab] Resumo:Epithelial remodeling determines the structure of many organs in the body through changes in cell shape, polarity and behavior and is a major area of study in developmental biology. Accurate and high-throughput methods are necessary to systematically analyze epithelial organization and dynamics at single-cell resolution. We developed SEGGA, an easy-to-use software for automated image segmentation, cell tracking and quantitative analysis of cell shape, polarity and behavior in epithelial tissues. SEGGA is free, open source, and provides a full suite of tools that allow users with no prior computational expertise to independently perform all steps of automated image segmentation, semi-automated user-guided error correction, and data analysis. Here we use SEGGA to analyze changes in cell shape, cell interactions and planar polarity during convergent extension in the embryo. These studies demonstrate that planar polarity is rapidly established in a spatiotemporally regulated pattern that is dynamically remodeled in response to changes in cell orientation. These findings reveal an unexpected plasticity that maintains coordinated planar polarity in actively moving populations through the continual realignment of cell polarity with the tissue axes.
[Mh] Termos MeSH primário: Polaridade Celular
Técnicas Citológicas/métodos
Células Epiteliais/citologia
Software
[Mh] Termos MeSH secundário: Animais
Automação
Forma Celular
Rastreamento de Células
Drosophila melanogaster/citologia
Drosophila melanogaster/embriologia
Embrião não Mamífero/citologia
Células Epiteliais/metabolismo
Genótipo
Processamento de Imagem Assistida por Computador
[Pt] Tipo de publicação:JOURNAL ARTICLE; RESEARCH SUPPORT, N.I.H., EXTRAMURAL; RESEARCH SUPPORT, NON-U.S. GOV'T
[Em] Mês de entrada:1709
[Cu] Atualização por classe:171126
[Lr] Data última revisão:
171126
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:170504
[St] Status:MEDLINE
[do] DOI:10.1242/dev.146837


  9 / 1602 MEDLINE  
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[PMID]:28931067
[Au] Autor:Zarychta-Wisniewska W; Burdzinska A; Zagozdzon R; Dybowski B; Butrym M; Gajewski Z; Paczek L
[Ad] Endereço:Department of Immunology, Transplant Medicine and Internal Diseases, Transplantation Institute, Medical University of Warsaw, Warsaw, Poland.
[Ti] Título:In vivo imaging system for explants analysis-A new approach for assessment of cell transplantation effects in large animal models.
[So] Source:PLoS One;12(9):e0184588, 2017.
[Is] ISSN:1932-6203
[Cp] País de publicação:United States
[La] Idioma:eng
[Ab] Resumo:INTRODUCTION: Despite spectacular progress in cellular transplantology, there are still many concerns about the fate of transplanted cells. More preclinical studies are needed, especially on large animal models, to bridge the translational gap between basic research and the clinic. Herein, we propose a novel approach in analysis of cell transplantation effects in large animals explants using in vivo imaging system (IVIS®) or similar equipment. MATERIAL AND METHODS: In the in vitro experiment cells labeled with fluorescent membrane dyes: DID (far red) or PKH26 (orange) were visualized with IVIS®. The correlation between the fluorescence signal and cell number with or without addition of minced muscle tissue was calculated. In the ex vivo study urethras obtained from goats after intraurethral cells (n = 9) or PBS (n = 4) injections were divided into 0.5 cm cross-slices and analyzed by using IVIS®. Automatic algorithm followed or not by manual setup was used to separate specific dye signal from tissue autofluorescence. The results were verified by systematic microscopic analysis of standard 10 µm specimens prepared from slices before and after immunohistochemical staining. Comparison of obtained data was performed using diagnostic test function. RESULTS: Fluorescence signal strength in IVIS® was directly proportional to the number of cells regardless of the dye used and detectable for minimum 0.25x106 of cells. DID-derived signal was much less affected by the background signal in comparison to PKH26 in in vitro test. Using the IVIS® to scan explants in defined arrangement resulted in precise localization of DID but not PKH26 positive spots. Microscopic analysis of histological specimens confirmed the specificity (89%) and sensitivity (80%) of IVIS® assessment relative to DID dye. The procedure enabled successful immunohistochemical staining of specimens derived from analyzed slices. CONCLUSIONS: The IVIS® system under appropriate conditions of visualization and analysis can be used as a method for ex vivo evaluation of cell transplantation effects. Presented protocol allows for evaluation of cell delivery precision rate, enables semi-quantitative assessment of signal, preselects material for further analysis without interfering with the tissue properties. Far red dyes are appropriate fluorophores to cell labeling for this application.
[Mh] Termos MeSH primário: Rastreamento de Células/métodos
Corantes Fluorescentes/metabolismo
Transplante de Células-Tronco Mesenquimais
Células Mesenquimais Estromais/citologia
Imagem Molecular/métodos
Uretra/metabolismo
[Mh] Termos MeSH secundário: Animais
Proliferação Celular
Células Cultivadas
Feminino
Fluorescência
Cabras
Compostos Orgânicos/metabolismo
Uretra/citologia
[Pt] Tipo de publicação:JOURNAL ARTICLE
[Nm] Nome de substância:
0 (Fluorescent Dyes); 0 (Organic Chemicals); 0 (PKH 26)
[Em] Mês de entrada:1710
[Cu] Atualização por classe:171017
[Lr] Data última revisão:
171017
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:170921
[St] Status:MEDLINE
[do] DOI:10.1371/journal.pone.0184588


  10 / 1602 MEDLINE  
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[PMID]:28920953
[Au] Autor:Ganuza M; Hall T; Finkelstein D; Chabot A; Kang G; McKinney-Freeman S
[Ad] Endereço:Department of Hematology, St Jude Children's Research Hospital, Memphis, Tennessee 38105, USA.
[Ti] Título:Lifelong haematopoiesis is established by hundreds of precursors throughout mammalian ontogeny.
[So] Source:Nat Cell Biol;19(10):1153-1163, 2017 Oct.
[Is] ISSN:1476-4679
[Cp] País de publicação:England
[La] Idioma:eng
[Ab] Resumo:Current dogma asserts that mammalian lifelong blood production is established by a small number of blood progenitors. However, this model is based on assays that require the disruption, transplantation and/or culture of embryonic tissues. Here, we used the sample-to-sample variance of a multicoloured lineage trace reporter to assess the frequency of emerging lifelong blood progenitors while avoiding the disruption, culture or transplantation of embryos. We find that approximately 719 Flk1 mesodermal precursors, 633 VE-cadherin endothelial precursors and 545 Vav1 nascent blood stem and progenitor cells emerge to establish the haematopoietic system at embryonic days (E)7-E8.5, E8.5-E11.5 and E11.5-E14.5, respectively. We also determined that the spatio-temporal recruitment of endothelial blood precursors begins at E8.5 and ends by E10.5, and that many c-Kit clusters of newly specified blood progenitors in the aorta are polyclonal in origin. Our work illuminates the dynamics of the developing mammalian blood system during homeostasis.
[Mh] Termos MeSH primário: Células Endoteliais/metabolismo
Hematopoese
Células-Tronco Hematopoéticas/metabolismo
[Mh] Termos MeSH secundário: Animais
Antígenos CD/genética
Antígenos CD/metabolismo
Caderinas/genética
Caderinas/metabolismo
Diferenciação Celular
Linhagem da Célula
Rastreamento de Células/métodos
Células Cultivadas
Técnicas de Cocultura
Células Endoteliais/transplante
Regulação da Expressão Gênica no Desenvolvimento
Genótipo
Idade Gestacional
Transplante de Células-Tronco Hematopoéticas
Integrases/genética
Integrases/metabolismo
Modelos Lineares
Proteínas Luminescentes/genética
Proteínas Luminescentes/metabolismo
Camundongos Endogâmicos C57BL
Camundongos Transgênicos
Microscopia de Fluorescência
Modelos Biológicos
Fenótipo
Proteínas Proto-Oncogênicas c-kit/genética
Proteínas Proto-Oncogênicas c-kit/metabolismo
Proteínas Proto-Oncogênicas c-vav/genética
Proteínas Proto-Oncogênicas c-vav/metabolismo
RNA não Traduzido/genética
RNA não Traduzido/metabolismo
Transdução de Sinais
Fatores de Tempo
Receptor 2 de Fatores de Crescimento do Endotélio Vascular/genética
Receptor 2 de Fatores de Crescimento do Endotélio Vascular/metabolismo
[Pt] Tipo de publicação:JOURNAL ARTICLE
[Nm] Nome de substância:
0 (Antigens, CD); 0 (Cadherins); 0 (Gt(ROSA)26Sor non-coding RNA, mouse); 0 (Luminescent Proteins); 0 (Proto-Oncogene Proteins c-vav); 0 (RNA, Untranslated); 0 (Vav1 protein, mouse); 0 (cadherin 5); EC 2.7.10.1 (Kdr protein, mouse); EC 2.7.10.1 (Proto-Oncogene Proteins c-kit); EC 2.7.10.1 (Vascular Endothelial Growth Factor Receptor-2); EC 2.7.7.- (Cre recombinase); EC 2.7.7.- (Integrases)
[Em] Mês de entrada:1710
[Cu] Atualização por classe:171114
[Lr] Data última revisão:
171114
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:170919
[St] Status:MEDLINE
[do] DOI:10.1038/ncb3607



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