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1.
Elife ; 112022 11 08.
Artigo em Inglês | MEDLINE | ID: mdl-36346018

RESUMO

While dysregulation of adipocyte endocrine function plays a central role in obesity and its complications, the vast majority of adipokines remain uncharacterized. We employed bio-orthogonal non-canonical amino acid tagging (BONCAT) and mass spectrometry to comprehensively characterize the secretome of murine visceral and subcutaneous white and interscapular brown adip ocytes. Over 600 proteins were identified, the majority of which showed cell type-specific enrichment. We here describe a metabolic role for leucine-rich α-2 glycoprotein 1 (LRG1) as an obesity-regulated adipokine secreted by mature adipocytes. LRG1 overexpression significantly improved glucose homeostasis in diet-induced and genetically obese mice. This was associated with markedly reduced white adipose tissue macrophage accumulation and systemic inflammation. Mechanistically, we found LRG1 binds cytochrome c in circulation to dampen its pro-inflammatory effect. These data support a new role for LRG1 as an insulin sensitizer with therapeutic potential given its immunomodulatory function at the nexus of obesity, inflammation, and associated pathology.


Assuntos
Adipocinas , Resistência à Insulina , Animais , Camundongos , Inflamação , Insulina , Obesidade , Camundongos Obesos , Glicoproteínas/genética
2.
Cell Rep Med ; 3(9): 100747, 2022 09 20.
Artigo em Inglês | MEDLINE | ID: mdl-36130482

RESUMO

In a randomized crossover study in humans, high fructose feeding reduced glucose uptake in brown fat without affecting the tissue's oxidative capacity. These effects were independent of alterations in the gut microbiome.


Assuntos
Frutose , Microbioma Gastrointestinal , Tecido Adiposo Marrom , Estudos Cross-Over , Glucose , Humanos
3.
Int Heart J ; 62(2): 381-389, 2021 Mar 30.
Artigo em Inglês | MEDLINE | ID: mdl-33731514

RESUMO

Extracellular vesicles (EV) that are derived from endothelial progenitor cells (EPC) have been determined to be a novel therapy for acute myocardial infarction, with a promise for immediate "off-the-shelf" delivery. Early experience suggests delivery of EVs from allogeneic sources is safe. Yet, clinical translation of this therapy requires assurances of both EV stability following cryopreservation and absence of an adverse immunologic response to EVs from allogeneic donors. Thus, more bioactivity studies on allogeneic EVs after cold storage are necessary to establish quality standards for its widespread clinical use. Thus, in this study, we aimed to demonstrate the safety and efficacy in delivering cryopreserved EVs in allogeneic recipients as a therapy for acute myocardial infarction.In this present study, we have analyzed the cardioprotective effects of allogeneic EPC-derived EVs after storage at -80°C for 2 months, using a shear-thinning gel (STG) as an in vivo delivery vehicle. EV size, proteome, and nucleic acid cargo were observed to remain steady through extended cryopreservation via nanoparticle tracking analysis, mass spectrometry, and nanodrop analysis, respectively. Fresh and previously frozen EVs in STG were delivered intramyocardially in a rat model of myocardial infarction (MI), with both showing improvements in contractility, angiogenesis, and scar thickness in comparison to phosphate-buffered saline (PBS) and STG controls at 4 weeks post-MI. Pathologic analyses and flow cytometry revealed minimal inflammatory and immune upregulation upon exposure of tissue to EVs pooled from allogeneic donor cells.Allogeneic EPC-EVs have been known to elicit minimal immune activity and retain therapeutic efficacy after at least 2 months of cryopreservation in a post-MI model.


Assuntos
Células Progenitoras Endoteliais/citologia , Vesículas Extracelulares/patologia , Transplante de Células-Tronco Hematopoéticas/métodos , Infarto do Miocárdio/terapia , Miócitos Cardíacos/patologia , Animais , Células Cultivadas , Criopreservação , Modelos Animais de Doenças , Humanos , Infarto do Miocárdio/patologia , Ratos
4.
J Thorac Cardiovasc Surg ; 159(5): 1825-1835.e2, 2020 05.
Artigo em Inglês | MEDLINE | ID: mdl-31353103

RESUMO

BACKGROUND: Extracellular vesicles (EVs) are promising therapeutics for cardiovascular disease, but poorly-timed delivery might hinder efficacy. We characterized the time-dependent response to endothelial progenitor cell (EPC)-EVs within an injectable shear-thinning hydrogel (STG+EV) post-myocardial infarction (MI) to identify when an optimal response is achieved. METHODS: The angiogenic effects of prolonged hypoxia on cell response to EPC-EV therapy and EV uptake affinity were tested in vitro. A rat model of acute MI via left anterior descending artery ligation was created and STG+EV was delivered via intramyocardial injections into the infarct border zone at time points corresponding to phases of post-MI inflammation: 0 hours (immediate), 3 hours (acute inflammation), 4 days (proliferative), and 2 weeks (fibrosis). Hemodynamics 4 weeks post-treatment were compared across treatment and control groups (phosphate buffered saline [PBS], shear-thinning gel). Scar thickness and ventricular diameter were assessed histologically. The primary hemodynamic end point was end systolic elastance. The secondary end point was scar thickness. RESULTS: EPC-EVs incubated with chronically versus acutely hypoxic human umbilical vein endothelial cells resulted in a 2.56 ± 0.53 versus 1.65 ± 0.15-fold increase (P = .05) in a number of vascular meshes and higher uptake of EVs over 14 hours. End systolic elastance improved with STG+EV therapy at 4 days (0.54 ± 0.08) versus PBS or shear-thinning gel (0.26 ± 0.03 [P = .02]; 0.23 ± 0.02 [P = .01]). Preservation of ventricular diameter (6.20 ± 0.73 mm vs 8.58 ± 0.38 mm [P = .04]; 9.13 ± 0.25 mm [P = .01]) and scar thickness (0.89 ± 0.05 mm vs 0.62 ± 0.03 mm [P < .0001] and 0.58 ± 0.05 mm [P < .0001]) was significantly greater at 4 days, compared wit PBS and shear-thinning gel controls. CONCLUSIONS: Delivery of STG+EV 4 days post-MI improved left ventricular contractility and preserved global ventricular geometry, compared with controls and immediate therapy post-MI. These findings suggest other cell-derived therapies can be optimized by strategic timing of therapeutic intervention.


Assuntos
Células Progenitoras Endoteliais/transplante , Vesículas Extracelulares/transplante , Hemodinâmica , Infarto do Miocárdio/cirurgia , Miocárdio/patologia , Neovascularização Fisiológica , Tempo para o Tratamento , Adamantano/química , Animais , Hipóxia Celular , Proliferação de Células , Células Cultivadas , Modelos Animais de Doenças , Células Progenitoras Endoteliais/metabolismo , Vesículas Extracelulares/metabolismo , Fibrose , Géis , Células Endoteliais da Veia Umbilical Humana/metabolismo , Células Endoteliais da Veia Umbilical Humana/patologia , Humanos , Ácido Hialurônico/química , Mediadores da Inflamação/metabolismo , Infarto do Miocárdio/metabolismo , Infarto do Miocárdio/patologia , Infarto do Miocárdio/fisiopatologia , Miocárdio/metabolismo , Ratos Wistar , Fatores de Tempo , beta-Ciclodextrinas/química
6.
Macromol Biosci ; 19(1): e1800248, 2019 01.
Artigo em Inglês | MEDLINE | ID: mdl-30259658

RESUMO

Shear-thinning hydrogels are useful for biomedical applications, from 3D bioprinting to injectable biomaterials. Although they have the appropriate properties for injection, it may be advantageous to decouple injectability from the controlled release of encapsulated therapeutics. Toward this, composites of hydrogels and encapsulated microgels are introduced with microgels that are fabricated via microfluidics. The microgel cross-linker controls degradation and entrapped molecule release, and the concentration of microgels alters composite hydrogel rheological properties. For the treatment of myocardial infarction (MI), interleukin-10 (IL-10) is encapsulated in microgels and released from composites. In a rat model of MI, composites with IL-10 reduce macrophage density after 1 week and improve scar thickness, ejection fraction, cardiac output, and the size of vascular structures after 4 weeks when compared to saline injection. Improvements are also observed with the composite without IL-10 over saline, emphasizing the role of injectable hydrogels alone on tissue repair.


Assuntos
Materiais Biocompatíveis , Hidrogéis , Interleucina-10 , Animais , Materiais Biocompatíveis/química , Materiais Biocompatíveis/farmacocinética , Materiais Biocompatíveis/farmacologia , Preparações de Ação Retardada/química , Preparações de Ação Retardada/farmacocinética , Preparações de Ação Retardada/farmacologia , Modelos Animais de Doenças , Hidrogéis/química , Hidrogéis/farmacocinética , Hidrogéis/farmacologia , Interleucina-10/química , Interleucina-10/farmacocinética , Interleucina-10/farmacologia , Infarto do Miocárdio/metabolismo , Ratos
7.
Cardiovasc Res ; 114(7): 1029-1040, 2018 06 01.
Artigo em Inglês | MEDLINE | ID: mdl-29566124

RESUMO

Aims: Previous studies have demonstrated improved cardiac function following myocardial infarction (MI) after administration of endothelial progenitor cells (EPCs) into ischaemic myocardium. A growing body of literature supports paracrine effectors, including extracellular vesicles (EVs), as the main mediators of the therapeutic benefits of EPCs. The direct use of paracrine factors is an attractive strategy that harnesses the effects of cell therapy without concerns of cell engraftment or viability. We aim to reproduce the beneficial effects of EPC treatment through delivery of EPC-derived EVs within a shear-thinning gel (STG) for precise localization and sustained delivery. Methods and results: EVs were harvested from EPCs isolated from adult male Rattus norvegicus (Wistar) rats and characterized by electron microscopy, nanoparticle tracking analysis (NTA), and mass spectrometry. EVs were incorporated into the STG and injected at the border zone in rat models of MI. Haemodynamic function, angiogenesis, and myocardial remodelling were analyzed in five groups: phosphate buffered saline (PBS) control, STG control, EVs in PBS, EVs in STG, and EPCs in STG. Electron microscopy and NTA of EVs showed uniform particles of 50-200 nm. EV content analysis revealed several key angiogenic mediators. EV uptake by endothelial cells was confirmed and followed by robust therapeutic angiogenesis. In vivo animal experiments demonstrated that delivery of EVs within the STG resulted in increased peri-infarct vascular proliferation, preservation of ventricular geometry, and improved haemodynamic function post-MI. Conclusions: EPC-derived EVs delivered into ischaemic myocardium via an injectable hydrogel enhanced peri-infarct angiogenesis and myocardial haemodynamics in a rat model of MI. The STG greatly increased therapeutic efficiency and efficacy of EV-mediated myocardial preservation.


Assuntos
Proteínas Angiogênicas/metabolismo , Micropartículas Derivadas de Células/transplante , Células Progenitoras Endoteliais/transplante , Ácido Hialurônico/química , Infarto do Miocárdio/cirurgia , Neovascularização Fisiológica , Transplante de Células-Tronco/métodos , Função Ventricular Esquerda , Animais , Micropartículas Derivadas de Células/metabolismo , Micropartículas Derivadas de Células/ultraestrutura , Células Cultivadas , Modelos Animais de Doenças , Células Progenitoras Endoteliais/metabolismo , Células Progenitoras Endoteliais/ultraestrutura , Células Endoteliais da Veia Umbilical Humana/metabolismo , Humanos , Hidrogéis , Masculino , Infarto do Miocárdio/metabolismo , Infarto do Miocárdio/patologia , Infarto do Miocárdio/fisiopatologia , Miocárdio/metabolismo , Miocárdio/patologia , Comunicação Parácrina , Ratos Wistar , Recuperação de Função Fisiológica , Transdução de Sinais , Fatores de Tempo , Pressão Ventricular
8.
J Invest Dermatol ; 138(1): 141-149, 2018 01.
Artigo em Inglês | MEDLINE | ID: mdl-28927893

RESUMO

Melanoma cells share many biological properties with neural crest stem cells. Here we show that the homeodomain transcription factor MSX1, which is significantly correlated with melanoma disease progression, reprograms melanocytes and melanoma cells toward a neural crest precursor-like state. MSX1-reprogrammed normal human melanocytes express the neural crest marker p75 and become multipotent. MSX1 induces a phenotypic switch in melanoma, which is characterized by an oncogenic transition from an E-cadherin-high nonmigratory state toward a ZEB1-high invasive state. ZEB1 up-regulation is responsible for the MSX1-induced migratory phenotype in melanoma cells. Depletion of MSX1 significantly inhibits melanoma metastasis in vivo. These results show that neural crest-like reprogramming achieved by a single factor is a critical process for melanoma progression.


Assuntos
Transformação Celular Neoplásica/patologia , Reprogramação Celular/fisiologia , Fator de Transcrição MSX1/fisiologia , Melanócitos/patologia , Melanoma/patologia , Neoplasias Cutâneas/patologia , Animais , Antígenos CD/metabolismo , Caderinas/metabolismo , Diferenciação Celular/fisiologia , Linhagem Celular Tumoral , Movimento Celular , Derme/citologia , Derme/patologia , Progressão da Doença , Células HEK293 , Células-Tronco Embrionárias Humanas , Humanos , Estimativa de Kaplan-Meier , Neoplasias Hepáticas/patologia , Neoplasias Hepáticas/secundário , Fator de Transcrição MSX1/genética , Melanoma/mortalidade , Melanoma/secundário , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCID , Proteínas do Tecido Nervoso/metabolismo , Crista Neural/fisiologia , Interferência de RNA , RNA Interferente Pequeno/metabolismo , Receptores de Fator de Crescimento Neural/metabolismo , Neoplasias Cutâneas/mortalidade , Ensaios Antitumorais Modelo de Xenoenxerto , Homeobox 1 de Ligação a E-box em Dedo de Zinco/metabolismo
10.
J Neurosci ; 33(47): 18381-95, 2013 Nov 20.
Artigo em Inglês | MEDLINE | ID: mdl-24259563

RESUMO

The transcription factor, ΔFosB, is robustly and persistently induced in striatum by several chronic stimuli, such as drugs of abuse, antipsychotic drugs, natural rewards, and stress. However, very few studies have examined the degree of ΔFosB induction in the two striatal medium spiny neuron (MSN) subtypes. We make use of fluorescent reporter BAC transgenic mice to evaluate induction of ΔFosB in dopamine receptor 1 (D1) enriched and dopamine receptor 2 (D2) enriched MSNs in ventral striatum, nucleus accumbens (NAc) shell and core, and in dorsal striatum (dStr) after chronic exposure to several drugs of abuse including cocaine, ethanol, Δ(9)-tetrahydrocannabinol, and opiates; the antipsychotic drug, haloperidol; juvenile enrichment; sucrose drinking; calorie restriction; the serotonin selective reuptake inhibitor antidepressant, fluoxetine; and social defeat stress. Our findings demonstrate that chronic exposure to many stimuli induces ΔFosB in an MSN-subtype selective pattern across all three striatal regions. To explore the circuit-mediated induction of ΔFosB in striatum, we use optogenetics to enhance activity in limbic brain regions that send synaptic inputs to NAc; these regions include the ventral tegmental area and several glutamatergic afferent regions: medial prefrontal cortex, amygdala, and ventral hippocampus. These optogenetic conditions lead to highly distinct patterns of ΔFosB induction in MSN subtypes in NAc core and shell. Together, these findings establish selective patterns of ΔFosB induction in striatal MSN subtypes in response to chronic stimuli and provide novel insight into the circuit-level mechanisms of ΔFosB induction in striatum.


Assuntos
Corpo Estriado/citologia , Dopaminérgicos/farmacologia , Emoções/efeitos dos fármacos , Optogenética , Proteínas Proto-Oncogênicas c-fos/metabolismo , Animais , Antidepressivos/farmacologia , Agonistas de Receptores de Canabinoides/farmacologia , Dronabinol/farmacologia , Meio Ambiente , Regulação da Expressão Gênica/efeitos dos fármacos , Proteínas de Fluorescência Verde/genética , Técnicas In Vitro , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Neurônios/classificação , Neurônios/efeitos dos fármacos , Fosfopiruvato Hidratase/metabolismo , Receptores de Dopamina D1/genética , Receptores de Dopamina D2/genética
11.
J Neurosci ; 30(48): 16082-90, 2010 Dec 01.
Artigo em Inglês | MEDLINE | ID: mdl-21123555

RESUMO

Brain stimulation and imaging studies in humans have highlighted a key role for the prefrontal cortex in clinical depression; however, it remains unknown whether excitation or inhibition of prefrontal cortical neuronal activity is associated with antidepressant responses. Here, we examined cellular indicators of functional activity, including the immediate early genes (IEGs) zif268 (egr1), c-fos, and arc, in the prefrontal cortex of clinically depressed humans obtained postmortem. We also examined these genes in the ventral portion of the medial prefrontal cortex (mPFC) of mice after chronic social defeat stress, a mouse model of depression. In addition, we used viral vectors to overexpress channel rhodopsin 2 (a light-activated cation channel) in mouse mPFC to optogenetically drive "burst" patterns of cortical firing in vivo and examine the behavioral consequences. Prefrontal cortical tissue derived from clinically depressed humans displayed significant reductions in IEG expression, consistent with a deficit in neuronal activity within this brain region. Mice subjected to chronic social defeat stress exhibited similar reductions in levels of IEG expression in mPFC. Interestingly, some of these changes were not observed in defeated mice that escape the deleterious consequences of the stress, i.e., resilient animals. In those mice that expressed a strong depressive-like phenotype, i.e., susceptible animals, optogenetic stimulation of mPFC exerted potent antidepressant-like effects, without affecting general locomotor activity, anxiety-like behaviors, or social memory. These results indicate that the activity of the mPFC is a key determinant of depression-like behavior, as well as antidepressant responses.


Assuntos
Transtorno Depressivo/genética , Transtorno Depressivo/terapia , Genes Precoces/fisiologia , Córtex Pré-Frontal/fisiologia , Animais , Channelrhodopsins , Transtorno Depressivo/patologia , Proteína 1 de Resposta de Crescimento Precoce/biossíntese , Regulação da Expressão Gênica , Humanos , Relações Interpessoais , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Córtex Pré-Frontal/patologia
12.
Science ; 330(6002): 385-90, 2010 Oct 15.
Artigo em Inglês | MEDLINE | ID: mdl-20947769

RESUMO

The nucleus accumbens is a key mediator of cocaine reward, but the distinct roles of the two subpopulations of nucleus accumbens projection neurons, those expressing dopamine D1 versus D2 receptors, are poorly understood. We show that deletion of TrkB, the brain-derived neurotrophic factor (BDNF) receptor, selectively from D1+ or D2+ neurons oppositely affects cocaine reward. Because loss of TrkB in D2+ neurons increases their neuronal excitability, we next used optogenetic tools to control selectively the firing rate of D1+ and D2+ nucleus accumbens neurons and studied consequent effects on cocaine reward. Activation of D2+ neurons, mimicking the loss of TrkB, suppresses cocaine reward, with opposite effects induced by activation of D1+ neurons. These results provide insight into the molecular control of D1+ and D2+ neuronal activity as well as the circuit-level contribution of these cell types to cocaine reward.


Assuntos
Fator Neurotrófico Derivado do Encéfalo/metabolismo , Transtornos Relacionados ao Uso de Cocaína/metabolismo , Cocaína/farmacologia , Neurônios/metabolismo , Núcleo Accumbens/metabolismo , Receptor trkB/metabolismo , Recompensa , Transdução de Sinais , Animais , Comportamento Animal/efeitos dos fármacos , Channelrhodopsins , Condicionamento Psicológico , Luz , Camundongos , Camundongos Transgênicos , Proteína Quinase 1 Ativada por Mitógeno/metabolismo , Proteína Quinase 3 Ativada por Mitógeno/metabolismo , Modelos Biológicos , Atividade Motora/efeitos dos fármacos , Núcleo Accumbens/citologia , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Receptor trkB/genética , Receptores de Dopamina D1/metabolismo , Receptores de Dopamina D2/metabolismo
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