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1.
Nat Commun ; 11(1): 6287, 2020 12 08.
Artigo em Inglês | MEDLINE | ID: mdl-33293533

RESUMO

Mammalian cells exhibit remarkable diversity in cell size, but the factors that regulate establishment and maintenance of these sizes remain poorly understood. This is especially true for skeletal muscle, comprised of syncytial myofibers that each accrue hundreds of nuclei during development. Here, we directly explore the assumed causal relationship between multinucleation and establishment of normal size through titration of myonuclear numbers during mouse neonatal development. Three independent mouse models, where myonuclear numbers were reduced by 75, 55, or 25%, led to the discovery that myonuclei possess a reserve capacity to support larger functional cytoplasmic volumes in developing myofibers. Surprisingly, the results revealed an inverse relationship between nuclei numbers and reserve capacity. We propose that as myonuclear numbers increase, the range of transcriptional return on a per nuclear basis in myofibers diminishes, which accounts for both the absolute reliance developing myofibers have on nuclear accrual to establish size, and the limits of adaptability in adult skeletal muscle.


Assuntos
Núcleo Celular , Tamanho Celular , Músculo Esquelético/crescimento & desenvolvimento , Células Satélites de Músculo Esquelético/citologia , Animais , Feminino , Masculino , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Camundongos , Camundongos Transgênicos , Modelos Animais , Proteínas Musculares/genética , Proteínas Musculares/metabolismo , Músculo Esquelético/citologia , Células Satélites de Músculo Esquelético/metabolismo
2.
Nat Commun ; 11(1): 6288, 2020 12 08.
Artigo em Inglês | MEDLINE | ID: mdl-33293572

RESUMO

Muscle fibers are the largest cells in the body, and one of its few syncytia. Individual cell sizes are variable and adaptable, but what governs cell size has been unclear. We find that muscle fibers are DNA scarce compared to other cells, and that the nuclear number (N) adheres to the relationship N = aVb where V is the cytoplasmic volume. N invariably scales sublinearly to V (b < 1), making larger cells even more DNA scarce. N scales linearly to cell surface in adult humans, in adult and developing mice, and in mice with genetically reduced N, but in the latter the relationship eventually fails when they reach adulthood with extremely large myonuclear domains. Another exception is denervation-atrophy where nuclei are not eliminated. In conclusion, scaling exponents are remarkably similar across species, developmental stages and experimental conditions, suggesting an underlying scaling law where DNA-content functions as a limiter of muscle cell size.


Assuntos
Núcleo Celular/química , Tamanho Celular , DNA/análise , Fibras Musculares Esqueléticas/citologia , Músculo Esquelético/crescimento & desenvolvimento , Adulto , Animais , Biópsia , Citoplasma , Feminino , Voluntários Saudáveis , Humanos , Microscopia Intravital , Masculino , Camundongos , Microscopia Confocal , Fibras Musculares Esqueléticas/química , Músculo Esquelético/citologia , Músculo Esquelético/patologia , Análise de Célula Única , Adulto Jovem
3.
Nat Commun ; 11(1): 6148, 2020 12 01.
Artigo em Inglês | MEDLINE | ID: mdl-33262337

RESUMO

Sustained proliferation is a significant driver of cancer progression. Cell-cycle advancement is coupled with cell size, but it remains unclear how multiple cells interact to control their volume in 3D clusters. In this study, we propose a mechano-osmotic model to investigate the evolution of volume dynamics within multicellular systems. Volume control depends on an interplay between multiple cellular constituents, including gap junctions, mechanosensitive ion channels, energy-consuming ion pumps, and the actomyosin cortex, that coordinate to manipulate cellular osmolarity. In connected cells, we show that mechanical loading leads to the emergence of osmotic pressure gradients between cells with consequent increases in cellular ion concentrations driving swelling. We identify how gap junctions can amplify spatial variations in cell volume within multicellular spheroids and, further, describe how the process depends on proliferation-induced solid stress. Our model may provide new insight into the role of gap junctions in breast cancer progression.


Assuntos
Neoplasias da Mama/fisiopatologia , Proliferação de Células , Junções Comunicantes/química , Esferoides Celulares/citologia , Neoplasias da Mama/química , Neoplasias da Mama/patologia , Linhagem Celular Tumoral , Tamanho Celular , Progressão da Doença , Feminino , Humanos , Pressão Osmótica , Esferoides Celulares/química
4.
Cell Mol Biol (Noisy-le-grand) ; 66(7): 202-206, 2020 Oct 31.
Artigo em Inglês | MEDLINE | ID: mdl-33287943

RESUMO

The pandemic diseases caused by SARS-CoV-2 are now threatening human health and survival. Early diagnosis and isolation of mild or asymptomatic COVID-19 patients is important to control the spread of SARS-CoV-2. In this study, we investigate the potential clinical utility of lymphocyte CPD for early diagnosis of COVID-19. To investigate the potential of lymphocyte cell population data (lymphocyte CPD) for use in early diagnosis of coronavirus disease 2019 (COVID-19). Lymphocyte CPD of healthy control (n = 51), common cold patients (n = 49) and mild COVID-19 patients (n = 126) were generated using hematology analyzer. The parameters were subjected to sensitivity and specificity analysis to determine their suitability as biomarkers for early diagnosis of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection. Normality analysis showed that lymphocyte CPD followed a normal distribution. There were no significant differences in white blood cells (WBC) and lymphocyte (LY#) counts as well as the neutrophil-to-lymphocyte ratio (NLR) among the groups (p > 0.05). Lymphocyte volume standard deviation (LV-SD), lymphocyte conductivity standard deviation (LC-SD) and lymphocyte light scatter standard deviation (LS-SD) were significantly higher in the COVID-19 group than in common cold and control groups (p < 0.05). The corresponding mean lymphocyte light scattering  (MLS) was significantly reduced in the COVID-19 group, relative to the common cold group, but was significantly increased, when compared with the control group (p < 0.05). Moreover, there was no significant difference in mean lymphocyte volume (MLV) between the COVID-19 group and the common cold or control group (p > 0.05), but it was significantly higher in the common cold group than in the control group (p < 0.05). At a cutoff value ≥ 16.38, LS-SD was more sensitive and specific than other lymphocyte CPD parameters. At a cutoff value ≥ 11.89, LC-SD achieved 84.4 % sensitivity, 87.5 % specificity, and an area under the curve (AUC) of 0.888. However, at a cutoff value ≥ 15.95, LS-SD reached 81.3 % sensitivity, 75 % specificity and an AUC of 0.876. These results suggest that lymphocyte CPD parameters have great diagnostic potential for SARS-CoV-2 infection and can be used for early diagnosis of the disease.


Assuntos
/diagnóstico , Contagem de Linfócitos , Adulto , /patologia , Tamanho Celular , Diagnóstico Precoce , Feminino , Humanos , Linfócitos/citologia , Linfócitos/patologia , Masculino , Pessoa de Meia-Idade , Sensibilidade e Especificidade
5.
Nat Commun ; 11(1): 5808, 2020 11 16.
Artigo em Inglês | MEDLINE | ID: mdl-33199701

RESUMO

Skeletal muscle promotes metabolic balance by regulating glucose uptake and the stimulation of multiple interorgan crosstalk. We show here that the catalytic activity of Vav2, a Rho GTPase activator, modulates the signaling output of the IGF1- and insulin-stimulated phosphatidylinositol 3-kinase pathway in that tissue. Consistent with this, mice bearing a Vav2 protein with decreased catalytic activity exhibit reduced muscle mass, lack of proper insulin responsiveness and, at much later times, a metabolic syndrome-like condition. Conversely, mice expressing a catalytically hyperactive Vav2 develop muscle hypertrophy and increased insulin responsiveness. Of note, while hypoactive Vav2 predisposes to, hyperactive Vav2 protects against high fat diet-induced metabolic imbalance. These data unveil a regulatory layer affecting the signaling output of insulin family factors in muscle.


Assuntos
Biocatálise , Homeostase , Metabolismo , Músculo Esquelético/crescimento & desenvolvimento , Músculo Esquelético/metabolismo , Proteínas Proto-Oncogênicas c-vav/metabolismo , Transdução de Sinais , Adipócitos Brancos/efeitos dos fármacos , Adipócitos Brancos/metabolismo , Tecido Adiposo Marrom/metabolismo , Animais , Biocatálise/efeitos dos fármacos , Composição Corporal/efeitos dos fármacos , Peso Corporal/efeitos dos fármacos , Linhagem Celular , Tamanho Celular/efeitos dos fármacos , Genótipo , Glucose/farmacologia , Homeostase/efeitos dos fármacos , Insulina/metabolismo , Fator de Crescimento Insulin-Like I/metabolismo , Camundongos Endogâmicos C57BL , Camundongos Knockout , Células Musculares/citologia , Células Musculares/efeitos dos fármacos , Células Musculares/metabolismo , Músculo Esquelético/efeitos dos fármacos , Tamanho do Órgão/efeitos dos fármacos , Fosfatidilinositol 3-Quinases/metabolismo , Fosforilação/efeitos dos fármacos , Proteínas Proto-Oncogênicas c-akt/metabolismo , Transdução de Sinais/efeitos dos fármacos , Regulação para Cima/efeitos dos fármacos , Proteínas rac1 de Ligação ao GTP/metabolismo
6.
Science ; 370(6514)2020 10 16.
Artigo em Inglês | MEDLINE | ID: mdl-33060329

RESUMO

Biological systems tailor their properties and behavior to their size throughout development and in numerous aspects of physiology. However, such size scaling remains poorly understood as it applies to cell mechanics and mechanosensing. By examining how the Drosophila pupal dorsal thorax epithelium responds to morphogenetic forces, we found that the number of apical stress fibers (aSFs) anchored to adherens junctions scales with cell apical area to limit larger cell elongation under mechanical stress. aSFs cluster Hippo pathway components, thereby scaling Hippo signaling and proliferation with area. This scaling is promoted by tricellular junctions mediating an increase in aSF nucleation rate and lifetime in larger cells. Development, homeostasis, and repair entail epithelial cell size changes driven by mechanical forces; our work highlights how, in turn, mechanosensitivity scales with cell size.


Assuntos
Epitélio/fisiologia , Mecanotransdução Celular , Fibras de Estresse/fisiologia , Estresse Mecânico , Animais , Caderinas/metabolismo , Tamanho Celular , Proteínas de Drosophila/metabolismo , Drosophila melanogaster , Células Epiteliais/fisiologia , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Miosina Tipo II/metabolismo , Proteínas Nucleares/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , Transativadores/metabolismo
7.
Nat Commun ; 11(1): 5319, 2020 10 21.
Artigo em Inglês | MEDLINE | ID: mdl-33087700

RESUMO

Arterial networks enlarge in response to increase in tissue metabolism to facilitate flow and nutrient delivery. Typically, the transition of a growing artery with a small diameter into a large caliber artery with a sizeable diameter occurs upon the blood flow driven change in number and shape of endothelial cells lining the arterial lumen. Here, using zebrafish embryos and endothelial cell models, we describe an alternative, flow independent model, involving enlargement of arterial endothelial cells, which results in the formation of large diameter arteries. Endothelial enlargement requires the GEF1 domain of the guanine nucleotide exchange factor Trio and activation of Rho-GTPases Rac1 and RhoG in the cell periphery, inducing F-actin cytoskeleton remodeling, myosin based tension at junction regions and focal adhesions. Activation of Trio in developing arteries in vivo involves precise titration of the Vegf signaling strength in the arterial wall, which is controlled by the soluble Vegf receptor Flt1.


Assuntos
Células Endoteliais/citologia , Células Endoteliais/fisiologia , Fatores de Troca do Nucleotídeo Guanina/fisiologia , Fator A de Crescimento do Endotélio Vascular/fisiologia , Remodelação Vascular/fisiologia , Animais , Animais Geneticamente Modificados , Tamanho Celular , Células Cultivadas , Fatores de Troca do Nucleotídeo Guanina/genética , Células Endoteliais da Veia Umbilical Humana , Humanos , Modelos Cardiovasculares , Fator de Crescimento Placentário/genética , Fator de Crescimento Placentário/fisiologia , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/fisiologia , Transdução de Sinais , Fator A de Crescimento do Endotélio Vascular/genética , Receptor 1 de Fatores de Crescimento do Endotélio Vascular/genética , Receptor 1 de Fatores de Crescimento do Endotélio Vascular/fisiologia , Remodelação Vascular/genética , Peixe-Zebra/embriologia , Peixe-Zebra/genética , Peixe-Zebra/fisiologia , Proteínas de Peixe-Zebra/genética , Proteínas de Peixe-Zebra/fisiologia , Proteínas rac1 de Ligação ao GTP/genética , Proteínas rac1 de Ligação ao GTP/fisiologia
9.
Sci Rep ; 10(1): 15989, 2020 09 29.
Artigo em Inglês | MEDLINE | ID: mdl-32994529

RESUMO

In many species, excitable cells preserve their physiological properties despite significant variation in physical size across time and in a population. For example, neurons in crustacean central pattern generators generate similar firing patterns despite several-fold increases in size between juveniles and adults. This presents a biophysical problem because the electrical properties of cells are highly sensitive to membrane area and channel density. It is not known whether specific mechanisms exist to sense membrane area and adjust channel expression to keep a consistent channel density, or whether regulation mechanisms that sense activity alone are capable of compensating cell size. We show that destabilising effects of growth can be specifically compensated by feedback mechanism that senses average calcium influx and jointly regulate multiple conductances. However, we further show that this class of growth-compensating regulation schemes is necessarily sensitive to perturbations that alter the expression of subsets of ion channel types. Targeted perturbations of specific ion channels can trigger a pathological response of the regulation mechanism and a failure of homeostasis. Our findings suggest that physiological regulation mechanisms that confer robustness to growth may be specifically vulnerable to deletions or mutations that affect subsets of ion channels.


Assuntos
Crustáceos/genética , Canais Iônicos/genética , Neurônios/fisiologia , Animais , Proteínas de Artrópodes/genética , Proteínas de Artrópodes/metabolismo , Tamanho Celular , Crustáceos/metabolismo , Retroalimentação Fisiológica , Deleção de Genes , Homeostase , Canais Iônicos/metabolismo , Modelos Teóricos
11.
PLoS One ; 15(9): e0234835, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32881864

RESUMO

In this study, the physiological values of volumes of plasma, cells, total blood and the F blood factors were identified in 24 adult tree shrews (Tupaia belangeri; 12 male and 12 female; average BW of 123.9±19.19 g). The two-compartment model method of Evans Blue dye was used to obtain the plasma volume and the venous hematocrit was measured by microhematocrit method. To establish the relationship between body weight (BW) and blood volume of tree shrews, We performed linear fitting for these two datasets. Results were analyzed according to gender and weight (<120g vs.>120g). Statistical significance was assessed using the unpaired student t test and one-way ANOVA. The average volumes per 100g body weight of plasma, red blood cell (RBC) and total blood were 5.42±0.543, 3.24±0.445, and 8.66±0.680ml respectively. The mean body hematocrit, cardiac hematocrit, jugular vein hematocrit, femoral vein hematocrit, and tail vein hematocrit was 37.43±4.096, 39.72±3.219, 43.04±4.717, 40.84±3.041, and 38.71±3.442% respectively. The F cardiac was 0.94±0.072, F jugular vein 0.88±0.118, F femoral vein 0.92±0.111, and the F tail vein 0.97±0.117. Blood volume (ml) was 85.89103×BW (kg). This is the first study to provide the parameters of plasma volume, cell volume, total blood volume and F factor and a baseline for future research on blood physiology of tree shrews.


Assuntos
Tupaiidae/sangue , Animais , Volume Sanguíneo , Peso Corporal , Tamanho Celular , Feminino , Hematócrito , Masculino , Volume Plasmático , Tupaiidae/fisiologia
12.
Mol Cell ; 79(6): 876-877, 2020 09 17.
Artigo em Inglês | MEDLINE | ID: mdl-32946761

RESUMO

In this issue of Molecular Cell, Jalihal et al. (2020) show that cell volume changes upon osmotic stress result in rapid and reversible condensation of numerous multivalent proteins.


Assuntos
Proteínas , Tamanho Celular , Pressão Osmótica
13.
Toxicology ; 442: 152545, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32755642

RESUMO

The Organisation for Economic Co-operation and Development has listed thirteen engineered nanomaterials (ENM) in order to investigate their toxicity on human health. Silicon dioxide (SiO2) and titanium dioxide (TiO2) are included on that list and we added indium tin oxide (ITO) nanoparticles (NPs) to our study, which is not listed on OECD suggested ENM to be investigated, however ITO NPs has a high potential of industrial production. We evaluate the physicochemical properties of SiO2 NPs (10-20 nm), TiO2 nanofibers (NFs; 3 µm length) and ITO NPs (<50 nm) and the impact of protein-corona formation on cell internalization. Then, we evaluated the toxicity of uncoated ENM on human lung epithelial cells exposed to 10 and 50 µg/cm2 for 24 h. TiO2 NFs showed the highest capability to adsorb proteins onto the particle surface followed by SiO2 NPs and ITO NPs after acellular incubation with fetal bovine serum. The protein adsorption had no impact on Alizarin Red S conjugation, intrinsic properties for reactive oxygen (ROS) formation or cell uptake for all types of ENM. Moreover, TiO2 NFs induced highest cell alterations in human lung epithelial cells exposed to 10 and 50 µg/cm2 while ITO NPs induced moderated cytotoxicity and SiO2 NPs caused even lower cytotoxicity under the same conditions. DNA, proteins and lipids were mainly affected by TiO2 NFs followed by SiO2 NPs with toxic effects in protein and lipids while limited variations were detected after exposure to ITO NPs on spectra analyzed by Fourier Transform Infrared Spectroscopy.


Assuntos
Nanoestruturas/química , Nanoestruturas/toxicidade , Coroa de Proteína/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Células A549 , Tamanho Celular , Grânulos Citoplasmáticos/metabolismo , Grânulos Citoplasmáticos/ultraestrutura , Células Epiteliais/metabolismo , Humanos , Metabolismo dos Lipídeos/efeitos dos fármacos , Mucosa Respiratória/citologia , Mucosa Respiratória/metabolismo , Dióxido de Silício/química , Dióxido de Silício/metabolismo , Dióxido de Silício/toxicidade , Propriedades de Superfície , Titânio/química , Titânio/metabolismo , Titânio/toxicidade , Cicatrização/efeitos dos fármacos
14.
Dev Cell ; 54(3): 297-298, 2020 08 10.
Artigo em Inglês | MEDLINE | ID: mdl-32781022

RESUMO

In this issue of Developmental Cell, Mukherjee et al. (2020) investigate control of nuclear growth by live imaging of early embryogenesis, perturbations of blastomere dimensions, and reconstitution in vitro. The authors uncover new mechanisms of nuclear size scaling by the amount of inherited perinuclear ER and duration of interphase.


Assuntos
Blastômeros , Núcleo Celular , Tamanho Celular , Retículo Endoplasmático , Interfase
15.
Sci Rep ; 10(1): 12810, 2020 07 30.
Artigo em Inglês | MEDLINE | ID: mdl-32732908

RESUMO

Current research on surface modifications has yielded advanced implant biomaterials. Various implant surface modifications have been shown to be promising in improving bone target cell response, but more comprehensive studies whether certain implant surface modifications can directly target cell behavioural features such as morphogenesis and proliferation are needed. Here, we studied the response of primary alveolar bone cells on various implant surface modifications in terms of osteoblast morphology and proliferation in vitro. Analyses of surface modifications led to surface-related test parameters including the topographical parameters micro-roughness, texture aspect and surface enlargement as well as the physicochemical parameter surface wettability. We compared osteoblast morphology and proliferation towards the above-mentioned parameters and found that texture aspect and surface enlargement but not surface roughness or wettability exhibited significant impact on osteoblast morphology and proliferation. Detailed analysis revealed osteoblast proliferation as a function of cell morphology, substantiated by an osteoblast size- and morphology-dependent increase in mitotic activity. These findings show that implant surface topography controls cell behavioural morphology and subsequently cell proliferation, thereby opening the road for cell instructive biomaterials.


Assuntos
Materiais Biocompatíveis , Proliferação de Células , Osteoblastos/citologia , Osteoblastos/fisiologia , Próteses e Implantes , Processo Alveolar/citologia , Tamanho Celular , Células Cultivadas , Humanos , Mitose/fisiologia , Propriedades de Superfície , Molhabilidade
16.
Mol Cell ; 79(6): 978-990.e5, 2020 09 17.
Artigo em Inglês | MEDLINE | ID: mdl-32857953

RESUMO

Processing bodies (PBs) and stress granules (SGs) are prominent examples of subcellular, membraneless compartments that are observed under physiological and stress conditions, respectively. We observe that the trimeric PB protein DCP1A rapidly (within ∼10 s) phase-separates in mammalian cells during hyperosmotic stress and dissolves upon isosmotic rescue (over ∼100 s) with minimal effect on cell viability even after multiple cycles of osmotic perturbation. Strikingly, this rapid intracellular hyperosmotic phase separation (HOPS) correlates with the degree of cell volume compression, distinct from SG assembly, and is exhibited broadly by homo-multimeric (valency ≥ 2) proteins across several cell types. Notably, HOPS sequesters pre-mRNA cleavage factor components from actively transcribing genomic loci, providing a mechanism for hyperosmolarity-induced global impairment of transcription termination. Our data suggest that the multimeric proteome rapidly responds to changes in hydration and molecular crowding, revealing an unexpected mode of globally programmed phase separation and sequestration.


Assuntos
Endorribonucleases/genética , Precursores de RNA/genética , Estresse Fisiológico/genética , Transativadores/genética , Terminação da Transcrição Genética , Animais , Tamanho Celular , Sobrevivência Celular/genética , Humanos , Pressão Osmótica/fisiologia , Proteoma/genética
17.
Nat Commun ; 11(1): 4038, 2020 08 12.
Artigo em Inglês | MEDLINE | ID: mdl-32788587

RESUMO

Asparaginyl-tRNA synthetase1 (NARS1) is a member of the ubiquitously expressed cytoplasmic Class IIa family of tRNA synthetases required for protein translation. Here, we identify biallelic missense and frameshift mutations in NARS1 in seven patients from three unrelated families with microcephaly and neurodevelopmental delay. Patient cells show reduced NARS1 protein, impaired NARS1 activity and impaired global protein synthesis. Cortical brain organoid modeling shows reduced proliferation of radial glial cells (RGCs), leading to smaller organoids characteristic of microcephaly. Single-cell analysis reveals altered constituents of both astrocytic and RGC lineages, suggesting a requirement for NARS1 in RGC proliferation. Our findings demonstrate that NARS1 is required to meet protein synthetic needs and to support RGC proliferation in human brain development.


Assuntos
Aspartato-tRNA Ligase/deficiência , Aspartato-tRNA Ligase/genética , Córtex Cerebral/patologia , Microcefalia/genética , Células-Tronco Neurais/patologia , Organoides/patologia , Aminoacil-RNA de Transferência/genética , Adolescente , Adulto , Sequência de Bases , Diferenciação Celular , Proliferação de Células , Tamanho Celular , Sobrevivência Celular , Criança , Família , Feminino , Fibroblastos/metabolismo , Fibroblastos/patologia , Células HEK293 , Humanos , Células-Tronco Pluripotentes Induzidas/metabolismo , Antígeno Ki-67/metabolismo , Masculino , Mutação/genética , Células-Tronco Neurais/metabolismo , Neuroglia/metabolismo , Linhagem , Adulto Jovem
18.
Nat Commun ; 11(1): 3955, 2020 08 07.
Artigo em Inglês | MEDLINE | ID: mdl-32769998

RESUMO

Cellular therapy to treat heart failure is an ongoing focus of intense research, but progress toward structural and functional recovery remains modest. Engineered augmentation of established cellular effectors overcomes impediments to enhance reparative activity. Such 'next generation' implementation includes delivery of combinatorial cell populations exerting synergistic effects. Concurrent isolation and expansion of three distinct cardiac-derived interstitial cell types from human heart tissue, previously reported by our group, prompted design of a 3D structure that maximizes cellular interaction, allows for defined cell ratios, controls size, enables injectability, and minimizes cell loss. Herein, mesenchymal stem cells (MSCs), endothelial progenitor cells (EPCs) and c-Kit+ cardiac interstitial cells (cCICs) when cultured together spontaneously form scaffold-free 3D microenvironments termed CardioClusters. scRNA-Seq profiling reveals CardioCluster expression of stem cell-relevant factors, adhesion/extracellular-matrix molecules, and cytokines, while maintaining a more native transcriptome similar to endogenous cardiac cells. CardioCluster intramyocardial delivery improves cell retention and capillary density with preservation of cardiomyocyte size and long-term cardiac function in a murine infarction model followed 20 weeks. CardioCluster utilization in this preclinical setting establish fundamental insights, laying the framework for optimization in cell-based therapeutics intended to mitigate cardiomyopathic damage.


Assuntos
Microambiente Celular , Miocárdio/patologia , Cicatrização , Animais , Animais Recém-Nascidos , Capilares/patologia , Agregação Celular , Morte Celular , Linhagem da Célula , Tamanho Celular , Citoproteção , Células Progenitoras Endoteliais/citologia , Feminino , Ventrículos do Coração/diagnóstico por imagem , Ventrículos do Coração/patologia , Ventrículos do Coração/fisiopatologia , Humanos , Processamento de Imagem Assistida por Computador , Recém-Nascido , Células-Tronco Mesenquimais/citologia , Camundongos Endogâmicos NOD , Infarto do Miocárdio/patologia , Infarto do Miocárdio/fisiopatologia , Miócitos Cardíacos/citologia , Estresse Oxidativo , Comunicação Parácrina , Ratos Sprague-Dawley , Transcrição Genética
19.
PLoS Comput Biol ; 16(8): e1008044, 2020 08.
Artigo em Inglês | MEDLINE | ID: mdl-32797044

RESUMO

Genetic studies have recently highlighted the importance of fat distribution, as well as overall adiposity, in the pathogenesis of obesity-associated diseases. Using a large study (n = 1,288) from 4 independent cohorts, we aimed to investigate the relationship between mean adipocyte area and obesity-related traits, and identify genetic factors associated with adipocyte cell size. To perform the first large-scale study of automatic adipocyte phenotyping using both histological and genetic data, we developed a deep learning-based method, the Adipocyte U-Net, to rapidly derive mean adipocyte area estimates from histology images. We validate our method using three state-of-the-art approaches; CellProfiler, Adiposoft and floating adipocytes fractions, all run blindly on two external cohorts. We observe high concordance between our method and the state-of-the-art approaches (Adipocyte U-net vs. CellProfiler: R2visceral = 0.94, P < 2.2 × 10-16, R2subcutaneous = 0.91, P < 2.2 × 10-16), and faster run times (10,000 images: 6mins vs 3.5hrs). We applied the Adipocyte U-Net to 4 cohorts with histology, genetic, and phenotypic data (total N = 820). After meta-analysis, we found that mean adipocyte area positively correlated with body mass index (BMI) (Psubq = 8.13 × 10-69, ßsubq = 0.45; Pvisc = 2.5 × 10-55, ßvisc = 0.49; average R2 across cohorts = 0.49) and that adipocytes in subcutaneous depots are larger than their visceral counterparts (Pmeta = 9.8 × 10-7). Lastly, we performed the largest GWAS and subsequent meta-analysis of mean adipocyte area and intra-individual adipocyte variation (N = 820). Despite having twice the number of samples than any similar study, we found no genome-wide significant associations, suggesting that larger sample sizes and a homogenous collection of adipose tissue are likely needed to identify robust genetic associations.


Assuntos
Adipócitos , Aprendizado de Máquina , Obesidade , Adipócitos/classificação , Adipócitos/citologia , Tecido Adiposo/fisiologia , Adulto , Índice de Massa Corporal , Tamanho Celular , Biologia Computacional/métodos , Feminino , Estudo de Associação Genômica Ampla , Humanos , Masculino , Pessoa de Meia-Idade , Redes Neurais de Computação , Obesidade/epidemiologia , Obesidade/genética , Fenótipo , Polimorfismo de Nucleotídeo Único/genética
20.
Sci Rep ; 10(1): 11991, 2020 07 20.
Artigo em Inglês | MEDLINE | ID: mdl-32686718

RESUMO

Traumatic brain injury (TBI) results in a cascade of cellular responses, which produce neuroinflammation, partly due to microglial activation. Transforming from surveying to primed phenotypes, microglia undergo considerable molecular changes. However, specific microglial profiles in rat remain elusive due to tedious methodology and limited availability of reagents. Here, we present a flow cytometry-based analysis of rat microglia 24 h after TBI using the controlled cortical impact model, validated with a bioinformatics approach. Isolated microglia are analyzed for morphological changes and their expression of activation markers using flow cytometry, traditional gating-based analysis methods and support the data by employing bioinformatics statistical tools. We use CD45, CD11b/c, and p2y12 receptor to identify microglia and evaluate their activation state using CD32, CD86, RT1B, CD200R, and CD163. The results from logic-gated flow cytometry analysis was validated with bioinformatics-based analysis and machine learning algorithms to detect quantitative changes in morphology and marker expression in microglia due to activation following TBI.


Assuntos
Biomarcadores/metabolismo , Lesões Encefálicas Traumáticas/metabolismo , Biologia Computacional , Citometria de Fluxo , Microglia/metabolismo , Animais , Lesões Encefálicas Traumáticas/patologia , Polaridade Celular , Tamanho Celular , Microglia/patologia , Ratos Sprague-Dawley
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