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Protein tyrosine phosphatase 1B (PTP1B) is a non-receptor tyrosine phosphatase best known for its role in regulating insulin and leptin signalling. Recently, knowledge on the role of PTP1B as a major regulator of multiple signalling pathways involved in cell growth, proliferation, viability and metabolism has expanded, and PTP1B is recognised as a therapeutic target in several human disorders, including diabetes, obesity, cardiovascular diseases and hematopoietic malignancies. The function of PTP1B in the immune system was largely overlooked until it was discovered that PTP1B negatively regulates the Janus kinase-a signal transducer and activator of the transcription (JAK/STAT) signalling pathway, which plays a significant role in modulating immune responses. PTP1B is now known to determine the magnitude of many signalling pathways that drive immune cell activation and function. As such, PTP1B inhibitors are being developed and tested in the context of inflammation and autoimmune diseases. Here, we provide an up-to-date summary of the molecular role of PTP1B in regulating immune cell function and how targeting its expression and/or activity has the potential to change the outcomes of immune-mediated and inflammatory disorders.
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Proteína Tirosina Fosfatase não Receptora Tipo 1 , Transdução de Sinais , Humanos , Proteína Tirosina Fosfatase não Receptora Tipo 1/metabolismo , Proteína Tirosina Fosfatase não Receptora Tipo 1/genética , Animais , Inflamação/metabolismo , Inflamação/imunologia , Diabetes Mellitus/imunologia , Diabetes Mellitus/metabolismoRESUMO
OBJECTIVE: Atherosclerosis is a chronic inflammatory process induced by the influx and entrapment of excess lipoproteins into the intima media of arteries. Previously, our lab demonstrated that systemic PTP1B inhibition protects against atherosclerosis in preclinical LDLR-/- models. Similarly, it was shown that myeloid-specific PTP1B ablation decreases plaque formation and ameliorates dyslipidaemia in the ApoE-/- model of atherosclerosis. We hypothesized that the relevant improvements in dyslipidaemia following modification of PTP1B activation may either result from changes in hepatic cholesterol biosynthesis and/or increased uptake and degradation by liver-resident macrophages. We examined this in animal models and patients with coronary artery disease. METHODS: In this study, we determined the cholesterol-lowering effect of myeloid-PTP1B deletion in mice fed a high-fat high-cholesterol diet and examined effects on total cholesterol levels and lipoprotein profiles. We also determined the effects of PTP1B inhibition to oxLDL-C challenge on foam cell formation and cholesterol efflux in human monocytes/macrophages. RESULTS: We present evidence that myeloid-PTP1B deficiency significantly increases the affinity of Kupffer cells for ApoB containing lipoproteins, in an IL10-dependent manner. We also demonstrate that PTP1B inhibitor, MSI-1436, treatment decreased foam cell formation in Thp1-derived macrophages and increased macrophage cholesterol efflux to HDL in an AMPK-dependent manner. We present evidence of three novel and distinct mechanisms regulated by PTP1B: an increase in cholesterol efflux from foam cells, decreased uptake of lipoproteins into intra-lesion macrophages in vitro and a decrease of circulating LDL-C and VLDL-C in vivo. CONCLUSIONS: Overall, these results suggest that myeloid-PTP1B inhibition has atheroprotective effects through improved cholesterol handling in atherosclerotic lesions, as well as increased reverse cholesterol transport. Trial registration Research registry, researchregistry 3235. Registered 07 November 2017, https://www.researchregistry.com/browse-the-registry#home/registrationdetails/5a01d0fce7e1904e93e0aac5/ .
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Aterosclerose , Dislipidemias , Humanos , Camundongos , Animais , Proteínas Quinases Ativadas por AMP , Aterosclerose/patologia , Colesterol/metabolismo , Homeostase , Camundongos KnockoutRESUMO
Streptozotocin (STZ) is widely used to induce experimental diabetes in murine models. However, the ability to induce diabetic nephropathy (DN) is more challenging. It has been recommended to inject STZ at multiple low doses within 15 min after dissolution due to its alleged instability. However, some studies suggest that STZ is stable for days due to equilibration of its two anomers (α and ß), 90 min after dissolution, and that this anomer-equilibrated STZ leads to higher survival rates and persistent hyperglycaemia with minimal weight loss. The aim of this study was to determine an optimal dose of anomer-equilibrated STZ to induce kidney tubular damage and compare it with the more commonly used freshly prepared STZ. We hypothesised that anomer-equilibrated STZ provides a better, reproducible experimental model of diabetes-induced kidney damage with improved animal welfare. Body measurements, fasting glycaemia, insulinemia and renal histology were assessed in male C57Bl/6J at two and six months of age treated with fresh (50 mg/kg) or anomer-equilibrated (dose ranging 35-50 mg/kg) STZ or vehicle control. We demonstrated a dose-dependent effect of anomer-equilibrated STZ on the induction of hypo-insulinaemia and hyperglycaemia, as well as body weight in two-month-old mice. Interestingly, in six-month-old mice STZ leads to body weight loss, independently of STZ preparation mode. Anomer-equilibrated STZ provoked moderate to severe kidney tubule structural damage, resulting in significant kidney hypertrophy, whereas freshly prepared STZ only caused mild alterations. In conclusion, our study proposes that anomer-equilibrated STZ provides a robust murine model of diabetes and early-stage diabetic nephropathy, which can be used to test therapeutic approaches to treat and/or prevent renal damage.
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Diabetes Mellitus Experimental , Nefropatias Diabéticas , Hiperglicemia , Camundongos , Masculino , Animais , Nefropatias Diabéticas/patologia , Estreptozocina , Camundongos Endogâmicos C57BL , Diabetes Mellitus Experimental/patologia , Rim/patologia , Hiperglicemia/patologiaRESUMO
Aortic valve stenosis is the most common valve disease in the western world. Central to the pathogenesis of this disease is the growth of new blood vessels (angiogenesis) within the aortic valve allowing infiltration of immune cells and development of intra-valve inflammation. Identifying the cellular mediators involved in this angiogenesis is important as this may reveal new therapeutic targets which could ultimately prevent the progression of aortic valve stenosis. Aortic valves from patients undergoing surgery for aortic valve replacement or dilation of the aortic arch were examined both ex vivo and in vitro. We now demonstrate that the anti-angiogenic protein, soluble fms-like tyrosine kinase 1 (sFlt1), a non-signalling soluble receptor for vascular endothelial growth factor, is constitutively expressed in non-diseased valves. sFlt-1 expression was, however, significantly reduced in aortic valve tissue from patients with aortic valve stenosis while protein markers of hypoxia were simultaneously increased. Exposure of primary-cultured valve interstitial cells to hypoxia resulted in a decrease in the expression of sFlt-1. We further reveal using a bioassay that siRNA knock-down of sFlt1 in valve interstitial cells directly results in a pro-angiogenic environment. Finally, incubation of aortic valves with sphingosine 1-phosphate, a bioactive lipid-mediator, increased sFlt-1 expression and inhibited angiogenesis within valve tissue. In conclusion, this study demonstrates that sFlt1 expression is directly correlated with angiogenesis in aortic valves and the observed decrease in sFlt-1 expression in aortic valve stenosis could increase valve inflammation, promoting disease progression. This could be a viable therapeutic target in treating this disease.
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Estenose da Valva Aórtica , Receptor 1 de Fatores de Crescimento do Endotélio Vascular , Humanos , Receptor 1 de Fatores de Crescimento do Endotélio Vascular/genética , Receptor 1 de Fatores de Crescimento do Endotélio Vascular/metabolismo , Fator A de Crescimento do Endotélio Vascular/metabolismo , Estenose da Valva Aórtica/metabolismo , Valva Aórtica/patologia , Inflamação/patologia , Hipóxia/metabolismoRESUMO
Macrophages play a key role in tissue development and homeostasis, innate immune defence against microbes or tumours, and restoring homeostasis through tissue regeneration following infection or injury. The ability to adopt such diverse functions is due to their heterogeneous nature, which is driven largely by their developmental origin and their response to signals they encounter from the microenvironment. The most well-characterized signals driving macrophage phenotype and function are biochemical and metabolic. However, the way macrophages sense and respond to their extracellular biophysical environment is becoming increasingly recognized in the field of mechano-immunology. These biophysical cues can be signals from tissue components, such as the composition and charge of extracellular matrix or topography, elasticity, and stiffness of the tissue surrounding cells; and mechanical forces such as shear stress or stretch. Macrophages are important in determining whether a disease resolves or becomes chronic. Ageing and diseases such as cancer or fibrotic disorders are associated with significant changes in the tissue biophysical environment, and this provides signals that integrate with those from biochemical and metabolic stimuli to ultimately dictate the overall function of macrophages. This review provides a brief overview of macrophage polarization, followed by a selection of commonly recognized physiological and applied biophysical stimuli impacting macrophage activity, and the potential signalling mechanisms driving downstream responses. The effects of biophysical cues on macrophages' function in homeostasis and disease and the associated clinical implications are also highlighted.
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Atherosclerosis is a chronic inflammatory disease characterized by lipid accumulation and plaque formation in arterial vessel walls. Atherosclerotic plaques narrow the arterial lumen to increase the risk of heart attacks, ischemic stroke and peripheral vascular disease, which are major and worldwide health and economic burdens. Macrophage accumulation within plaques is characteristic of all stages of atherosclerosis and their presence is a potential marker of disease activity and plaque stability. Macrophages engulf lipids and modified lipoproteins to form foam cells that express pro-inflammatory and chemotactic effector molecules, stress inducing factors and reactive oxygen species. They control plaque stability and rupture through secretion of metalloproteinases and extracellular matrix degradation. Although macrophages can worsen disease by propagating inflammation, they can stabilize atherosclerotic plaques through tissue remodeling, promoting the formation of a fibrous cap, clearing apoptotic cells to prevent necrotic core formation and through vascular repair. In atherosclerosis, macrophages respond to dyslipidaemia, cytokines, dying cells, metabolic factors, lipids, physical stimuli and epigenetic factors and exhibit heterogeneity in their activation depending on the stimuli they receive. Understanding these signals and the pathways driving macrophage function within developing and established plaques and how they can be pharmacologically modulated, represents a strategy for the prevention and treatment of atherosclerosis. This review focusses on the current understanding of factors controlling macrophage heterogeneity and function in atherosclerosis. Particular attention is given to the macrophage intracellular signaling pathways and transcription factors activated by biochemical and biophysical stimuli within plaques, and how they are integrated to regulate plaque formation and stability.
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Aterosclerose , Placa Aterosclerótica , Humanos , Ativação de Macrófagos , Placa Aterosclerótica/metabolismo , Aterosclerose/metabolismo , Transdução de Sinais , LipoproteínasRESUMO
A treatment with direct healing effects on the gastrointestinal epithelial barrier is desirable for inflammatory bowel disease (IBD). Interleukin-27 (IL-27) is an immunoregulatory cytokine, and oral delivery is an effective treatment in murine models of IBD. We aimed to define IL-27 effects on the human gastrointestinal epithelial barrier. We characterised gene and protein expression of permeability mediators in a human colon-derived organoid model. Functional permeability was determined in an organoid-derived 2D monolayer by transepithelial electrical resistance. IL-27 effects on epithelial innate immune responses were assessed through expression of cytokines, anti-microbial peptides and MUC genes. IL-27 effects on wound healing and proliferation were determined in human colon epithelial cell lines. IL-27 led to restoration of permeability regulation following inflammatory cytokine insult (p = 0.001), associated with differential expression of tight junction mediators with decrease in claudin 2 (p = 0.024) and increase in claudin 4 (p < 0.001), E-cadherin (p < 0.001) and zona occludens (p = 0.0014). IL-27 evoked differential gene expression of epithelial-derived innate immune responses (reduced IL1B and IL18, and increased IL33, HBD1, MUC1 and MUC2; p < 0.012). IL-27 induced epithelial barrier wound healing through restitution (p < 0.001), and increased proliferation (p < 0.001) following injury. Overall, IL-27 provokes mucosal healing of the human gastrointestinal epithelial barrier.
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Extracorporeal Shock Wave Therapy (ESWT) is used clinically in various disorders including chronic wounds for its pro-angiogenic, proliferative, and anti-inflammatory effects. However, the underlying cellular and molecular mechanisms driving therapeutic effects are not well characterized. Macrophages play a key role in all aspects of healing and their dysfunction results in failure to resolve chronic wounds. We investigated the role of ESWT on macrophage activity in chronic wound punch biopsies from patients with non-healing venous ulcers prior to, and two weeks post-ESWT, and in macrophage cultures treated with clinical shockwave intensities (150-500 impulses, 5 Hz, 0.1 mJ/mm2). Using wound area measurements and histological/immunohistochemical analysis of wound biopsies, we show ESWT enhanced healing of chronic ulcers associated with improved wound angiogenesis (CD31 staining), significantly decreased CD68-positive macrophages per biopsy area and generally increased macrophage activation. Shockwave treatment of macrophages in culture significantly boosted uptake of apoptotic cells, healing-associated cytokine and growth factor gene expressions and modulated macrophage morphology suggestive of macrophage activation, all of which contribute to wound resolution. Macrophage ERK activity was enhanced, suggesting one mechanotransduction pathway driving events. Collectively, these in vitro and in vivo findings reveal shockwaves as important regulators of macrophage functions linked with wound healing. This immunomodulation represents an underappreciated role of clinically applied shockwaves, which could be exploited for other macrophage-mediated disorders.
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Tratamento por Ondas de Choque Extracorpóreas/métodos , Macrófagos/fisiologia , Mecanotransdução Celular , Úlcera Varicosa/radioterapia , Cicatrização/efeitos da radiação , Adulto , Idoso , Idoso de 80 Anos ou mais , Animais , Doença Crônica , Feminino , Humanos , Macrófagos/efeitos da radiação , Masculino , Pessoa de Meia-Idade , Úlcera Varicosa/metabolismo , Úlcera Varicosa/patologiaRESUMO
Factor XIII (FXIII) is a transglutaminase that promotes thrombus stability by cross-linking fibrin. The cellular form, a homodimer of the A subunits, denoted FXIII-A, lacks a classical signal peptide for its release; however, we have shown that it is exposed on activated platelets. Here we addressed whether monocytes expose intracellular FXIII-A in response to stimuli. Using flow cytometry, we demonstrate that FXIII-A antigen and activity are up-regulated on human monocytes in response to stimulation by IL-4 and IL-10. Higher basal levels of the FXIII-A antigen were noted on the membrane of the monocytic cell line THP-1, but activity was significantly enhanced following stimulation with IL-4 and IL-10. In contrast, treatment with lipopolysaccharide did not upregulate exposure of FXIII-A in THP-1 cells. Quantification of the FXIII-A activity revealed a significant increase in THP-1 cells in total cell lysates following stimulation with IL-4 and IL-10. Following fractionation, the largest pool of FXIII-A was membrane associated. Monocytes were actively incorporated into the fibrin mesh of model thrombi. We found that stimulation of monocytes and THP-1 cells with IL-4 and IL-10 stabilized FXIII-depleted thrombi against fibrinolytic degradation, via a transglutaminase-dependent mechanism. Our data suggest that monocyte-derived FXIII-A externalized in response to stimuli participates in thrombus stabilization.
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Fator XIIIa/metabolismo , Monócitos/metabolismo , Trombose/metabolismo , Voluntários Saudáveis , Humanos , Células THP-1/metabolismoRESUMO
Thyroid diseases, associated with either increased or decreased concentrations of circulating thyroid hormones, are prevalent in both human and veterinary populations. Hypothyroidism is a differential diagnosis for many medical problems as the disease presents with nonspecific clinical signs that can include lethargy, weight gain, cold intolerance, and dermatologic manifestations such as alopecia. Alopecia is a frequently reported problem in captive nonhuman primates (NHP), and hypothyroidism is considered to be a differential diagnosis. However, thyroid function test results in NHP using total T4 (TT4) and free T4 (FT4) assays are difficult to interpret without accurate reference intervals (RI) for comparison. As a consequence, hypothyroidism may be underdiagnosed in these species. The objective of this study was to establish RI for TT4 and FT4 in healthy populations of cynomolgus macaques ( n = 133; age range 2.6 to 24.7 y) and rhesus macaques ( n = 172; age range 0.8 to 31.0 y). Serum samples were collected across a 14-y period during routine anesthetic events in clinically healthy animals, and TT4 and FT4 concentrations were measured using commercially available immunoassays. The RI established for TT4 and FT4 were 5.1 to 14.9 ug/dL and 0.48 to 1.17 ng/dL for cynomolgus macaques, and 3.9 to 14.7 ug/dL and 0.36 to 1.12 ng/dL for rhesus macaques. Significant differences in thyroid hormone concentrations were found between Indian and Chinese origin rhesus, and between Mauritian and other origin cynomolgus. In addition, juvenile and subadult rhesus exhibited significantly higher FT4 and TT4 concentrations than did older animals. Individual RI were established for subgroups with adequately different thyroid hormone concentrations. These results will allow a more thorough diagnostic evaluation of cynomolgus and rhesus macaques with clinical signs consistent with thyroid disease and will ultimately be a refinement in NHP medicine.
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Testes Hematológicos , Testes de Função Tireóidea , Animais , Macaca fascicularis , Macaca mulatta , Valores de ReferênciaRESUMO
The microbiome exerts considerable control over immune homeostasis and influences susceptibility to autoimmune and autoinflammatory disease (AD/AID) such as inflammatory bowel disease (IBD), multiple sclerosis (MS), type 1 diabetes (T1D), psoriasis, and uveitis. In part, this is due to direct effects of the microbiome on gastrointestinal (GI) physiology and nutrient transport, but also to indirect effects on immunoregulatory controls, including induction and stabilization of T regulatory cells (T reg). Secreted bacterial metabolites such as short-chain fatty acids (SCFA) are under intense investigation as mediators of these effects. In contrast, folate (vitamin B9), an essential micronutrient, has attracted less attention, possibly because it exerts global physiological effects which are difficult to differentiate from specific effects on the immune system. Here, we review the role of folate in AD/AID with some emphasis on sight-threatening autoimmune uveitis. Since folate is required for the generation and maintenance of T reg , we propose that one mechanism for microbiome-based control of AD/AID is via folate-dependent induction of GI tract T reg , particularly colonic T reg, via anergic T cells (T an). Hence, folate supplementation has potential prophylactic and/or therapeutic benefit in AID/AD.
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Doenças Autoimunes/imunologia , Autoimunidade , Ácido Fólico/metabolismo , Microbioma Gastrointestinal/imunologia , Inflamação/imunologia , Animais , Doenças Autoimunes/dietoterapia , Doenças Autoimunes/metabolismo , Doenças Autoimunes/microbiologia , Modelos Animais de Doenças , Ácido Fólico/administração & dosagem , Humanos , Inflamação/dietoterapia , Inflamação/metabolismo , Inflamação/microbiologia , Mucosa Intestinal/imunologia , Mucosa Intestinal/metabolismo , Mucosa Intestinal/microbiologia , Linfócitos T Reguladores/imunologia , Linfócitos T Reguladores/metabolismoRESUMO
In both sickle cell disease and malaria, red blood cells (RBCs) are phagocytosed in the spleen, but receptor-ligand pairs mediating uptake have not been identified. Here, we report that patches of high mannose N-glycans (Man5-9GlcNAc2), expressed on diseased or oxidized RBC surfaces, bind the mannose receptor (CD206) on phagocytes to mediate clearance. We find that extravascular hemolysis in sickle cell disease correlates with high mannose glycan levels on RBCs. Furthermore, Plasmodium falciparum-infected RBCs expose surface mannose N-glycans, which occur at significantly higher levels on infected RBCs from sickle cell trait subjects compared to those lacking hemoglobin S. The glycans are associated with high molecular weight complexes and protease-resistant, lower molecular weight fragments containing spectrin. Recognition of surface N-linked high mannose glycans as a response to cellular stress is a molecular mechanism common to both the pathogenesis of sickle cell disease and resistance to severe malaria in sickle cell trait.
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Anemia Falciforme/metabolismo , Eritrócitos/metabolismo , Manose/metabolismo , Fagócitos/metabolismo , Polissacarídeos/metabolismo , Membrana Eritrocítica/metabolismo , Membrana Eritrocítica/parasitologia , Eritrócitos/parasitologia , Citometria de Fluxo/métodos , Hemólise , Humanos , Ligantes , Malária Falciparum/metabolismo , Malária Falciparum/parasitologia , Glicoproteínas de Membrana/metabolismo , Fagocitose , Plasmodium falciparum/fisiologia , Ligação Proteica , Receptores Imunológicos/metabolismoRESUMO
Macrophages provide a first line of defense against microorganisms, and while some mechanisms to kill pathogens such as the oxidative burst are well described, others are still undefined or unknown. Here, we report that the Rab32 guanosine triphosphatase and its guanine nucleotide exchange factor BLOC-3 (biogenesis of lysosome-related organelles complex-3) are central components of a trafficking pathway that controls both bacterial and fungal intracellular pathogens. This host-defense mechanism is active in both human and murine macrophages and is independent of well-known antimicrobial mechanisms such as the NADPH (reduced form of nicotinamide adenine dinucleotide phosphate)-dependent oxidative burst, production of nitric oxide, and antimicrobial peptides. To survive in human macrophages, Salmonella Typhi actively counteracts the Rab32/BLOC-3 pathway through its Salmonella pathogenicity island-1-encoded type III secretion system. These findings demonstrate that the Rab32/BLOC-3 pathway is a novel and universal host-defense pathway and protects mammalian species from various pathogens.
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Salmonella typhi , Proteínas rab de Ligação ao GTP , Animais , Fatores de Troca do Nucleotídeo Guanina/metabolismo , Humanos , Lisossomos/metabolismo , Macrófagos/metabolismo , Mamíferos/metabolismo , Camundongos , Proteínas rab de Ligação ao GTP/metabolismoRESUMO
We specify the clinical features of a spontaneous experimental autoimmune uveitis (EAU) model, in which foreign hen-egg lysozyme (HEL) is expressed in the retina, controlled by the promoter for interphotoreceptor retinol binding protein (IRBP). We previously reported 100% P21 (post-partum day) IRBP:HEL single transgenic (sTg) mice, when crossed to transgenic T cell receptor mice (3A9) generating the double transgenic (dTg) genotype, develop EAU despite profound lymphopenia (thymic HEL-specific T cell deletion). In this work, we characterized the immune component of this model and found conventional dTg CD4+ T cells were less anergic than those from 3A9 controls. Furthermore, prior in vitro HEL-activation of 3A9 anergic T cells (Tan) rendered them uveitogenic upon adoptive transfer (Tx) to sTg mice, while antigen-experienced (AgX, dTg), but not naïve (3A9) T cells halted disease in P21 dTg mice. Flow cytometric analysis of the AgX cells elucidated the underlying pathology: FoxP3+CD25hiCD4+ T regulatory cells (Treg) comprised â¼18%, while FR4+CD73+FoxP3-CD25lo/-CD4+ Tan comprised â¼1.2% of total cells. Further Treg-enrichment (â¼80%) of the AgX population indicated FoxP3+CD25hiCD4+ Treg played a key role in EAU-suppression while FoxP3-CD25lo/-CD4+ T cells did not. Here we present the novel concept of dual immunological tolerance where spontaneous EAU is due to escape from anergy with consequent failure of Treg induction and subsequent imbalance in the [Treg:Teffector] cell ratio. The reduced numbers of Tan, normally sustaining Treg to prevent autoimmunity, are the trigger for disease, while immune homeostasis can be restored by supplementation with AgX, but not naïve, antigen-specific Treg.
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Doenças Autoimunes/imunologia , Imunoterapia Adotiva/métodos , Retina/patologia , Linfócitos T Reguladores/imunologia , Uveíte/imunologia , Animais , Células Cultivadas , Modelos Animais de Doenças , Proteínas do Olho/imunologia , Fatores de Transcrição Forkhead/metabolismo , Humanos , Tolerância Imunológica , Subunidade alfa de Receptor de Interleucina-2/metabolismo , Camundongos , Camundongos Transgênicos , Proteínas de Ligação ao Retinol/imunologia , Linfócitos T Reguladores/transplanteRESUMO
Excessive inflammation by phagocytes during Aspergillus fumigatus infection is thought to promote lung function decline in CF patients. CFTR modulators have been shown to reduce A. fumigatus colonization in vivo, however, their antifungal and anti-inflammatory mechanisms are unclear. Other treatments including azithromycin and acebilustat may dampen Aspergillus-induced inflammation due to their immunomodulatory properties. Therefore, we set out in this study to determine the effects of current CF therapies on ROS production and fungal killing, either direct or indirect by enhancing antifungal immune mechanisms in peripheral blood immune cells from CF patients upon A. fumigatus infection. Isolated peripheral blood mononuclear cells (PBMCs) and polymorphonuclear cells (PMNs) from CF patients and healthy volunteers were challenged with A. fumigatus following pre-treatment with CFTR modulators, azithromycin or acebilustat. Ivacaftor/lumacaftor treated CF and control subject PMNs resulted in a significant reduction (p < 0.05) in Aspergillus-induced ROS. For CF PBMC, Aspergillus-induced ROS was significantly reduced when pre-treated with ivacaftor alone (p < 0.01) or in combination with lumacaftor (p < 0.01), with a comparable significant reduction in control subject PBMC (p < 0.05). Azithromycin and acebilustat had no effect on ROS production by CF or control subject phagocytes. None of the treatments showed an indirect or direct antifungal activity. In summary, CFTR modulators have potential for additional immunomodulatory benefits to prevent or treat Aspergillus-induced inflammation in CF. The comparable effects of CFTR modulators observed in phagocytes from control subjects questions their exact mechanism of action.
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Aspergilose/tratamento farmacológico , Regulador de Condutância Transmembrana em Fibrose Cística/antagonistas & inibidores , Fibrose Cística , Fagócitos , Espécies Reativas de Oxigênio , Aspergillus fumigatus , Humanos , Leucócitos MononuclearesRESUMO
BACKGROUND: Takotsubo syndrome mimics acute myocardial infarction (MI) at presentation. OBJECTIVES: To explore differences in ECG time-course that could further help distinguish the two conditions. METHODS: Serial ECG's (day 0-4) of 27 acute takotsubo and 37 MI patients, all presenting with anterior ST-elevation, were analysed for detailed morphology and timing of de/re-polarisation. All underwent cardiac magnetic resonance. RESULTS: The presenting ECG (day 0) showed significantly fewer total abnormal leads (p = .001), comparable number of ST-elevation leads but lesser total magnitude of ST-elevation (p = .003), smaller sum of positive T wave amplitude (p = .006) and lesser number of pathological Q waves (p = .005) in takotsubo vs the MI group. After day 0, takotsubo patients developed more widespread T wave inversion (p = .001, day 3) and/or deeper T waves compared to MI, (sum of the T-wave amplitude slope of change between days 0-3: -43.1 ± 9.6 vs - 16.6 ± 5.4 mm, p = .02). Although there was no difference in mean QTc between the groups on any day, between days 0-3 there was a progressive increase in QTc in takotsubo vs a decrease in MI (34.1 ± 12.2 vs -29.5 ± 9.3 ms, slope of change p < .001). There was significantly more myocardial oedema (native T1 mapping) in takotsubo vs MI (p = .02), which resulted in increased left ventricular mass index in takostubo (p = .04). CONCLUSIONS: The differences in presenting (day 0) ECG between takotsubo and MI are significant but subtle, reinforcing the importance of acute cardiac catheterisation for accurate diagnosis. During the next 3 days there is progressive increase in the depth and spread of T-waves and QTc duration in takotsubo vs MI - these may aid the diagnostic confidence in patients with bystander non-obstructive coronary disease.
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Infarto do Miocárdio , Infarto do Miocárdio com Supradesnível do Segmento ST , Cardiomiopatia de Takotsubo , Diagnóstico Diferencial , Eletrocardiografia , Humanos , Infarto do Miocárdio/diagnóstico , Infarto do Miocárdio com Supradesnível do Segmento ST/diagnóstico por imagem , Cardiomiopatia de Takotsubo/diagnóstico por imagemRESUMO
PURPOSE: Inflammation is central in disease pathophysiology and accurate methods for its detection and quantification are increasingly required to guide diagnosis and therapy. Here we explored the ability of Fast Field-Cycling Magnetic Resonance (FFC-MR) in quantifying the signal of ultra-small superparamagnetic iron oxide particles (USPIO) phagocytosed by J774 macrophage-like cells as a proof-of-principle. METHODS: Relaxation rates were measured in suspensions of J774 macrophage-like cells loaded with USPIO (0-200⯵g/ml Fe as ferumoxytol), using a 0.25â¯T FFC benchtop relaxometer and a human whole-body, in-house built 0.2â¯T FFC-MR prototype system with a custom test tube coil. Identical non-imaging, saturation recovery pulse sequence with 90° flip angle and 20 different evolution fields selected logarithmically between 80 µT and 0.2â¯T (3.4â¯kHz and 8.51â¯MHz proton Larmor frequency [PLF] respectively). Results were compared with imaging flow cytometry quantification of side scatter intensity and USPIO-occupied cell area. A reference colorimetric iron assay was used. RESULTS: The T1 dispersion curves derived from FFC-MR were excellent in detecting USPIO at all concentrations examined (0-200⯵g/ml Fe as ferumoxytol) vs. control cells, pâ¯≤â¯0.001. FFC-NMR was capable of reliably detecting cellular iron content as low as 1.12â¯ng/µg cell protein, validated using a colorimetric assay. FFC-MR was comparable to imaging flow cytometry quantification of side scatter intensity but superior to USPIO-occupied cell area, the latter being only sensitive at exposuresâ¯≥â¯10⯵g/ml USPIO. CONCLUSIONS: We demonstrated for the first time that FFC-MR is capable of quantitative assessment of intra-cellular iron which will have important implications for the use of USPIO in a variety of biological applications, including the study of inflammation.
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Óxido Ferroso-Férrico/química , Macrófagos/metabolismo , Imageamento por Ressonância Magnética/métodos , Colorimetria , Desenho de Equipamento , Citometria de Fluxo , Humanos , Técnicas In Vitro , Inflamação/metabolismo , Imageamento por Ressonância Magnética/instrumentação , Tamanho da Partícula , Fagocitose , Estudo de Prova de Conceito , SuspensõesRESUMO
Immune privilege (IP), a term introduced to explain the unpredicted acceptance of allogeneic grafts by the eye and the brain, is considered a unique property of these tissues. However, immune responses are modified by the tissue in which they occur, most of which possess IP to some degree. The eye therefore displays a spectrum of IP because it comprises several tissues. IP as originally conceived can only apply to the retina as it contains few tissue-resident bone-marrow derived myeloid cells and is immunologically shielded by a sophisticated barrier - an inner vascular and an outer epithelial barrier at the retinal pigment epithelium. The vascular barrier comprises the vascular endothelium and the glia limitans. Immune cells do not cross the blood-retinal barrier (BRB) despite two-way transport of interstitial fluid, governed by tissue oncotic pressure. The BRB, and the blood-brain barrier (BBB) mature in the neonatal period under signals from the expanding microbiome and by 18 months are fully established. However, the adult eye is susceptible to intraocular inflammation (uveitis; frequency ~200/100,000 population). Uveitis involving the retinal parenchyma (posterior uveitis, PU) breaches IP, while IP is essentially irrelevant in inflammation involving the ocular chambers, uveal tract and ocular coats (anterior/intermediate uveitis/sclerouveitis, AU). Infections cause ~50% cases of AU and PU but infection may also underlie the pathogenesis of immune-mediated "non-infectious" uveitis. Dysbiosis accompanies the commonest form, HLA-B27-associated AU, while latent infections underlie BRB breakdown in PU. This review considers the pathogenesis of uveitis in the context of IP, infection, environment, and the microbiome.
Assuntos
Bactérias/imunologia , Microbioma Gastrointestinal , Privilégio Imunológico , Intestinos/microbiologia , Úvea/imunologia , Uveíte/imunologia , Animais , Bactérias/metabolismo , Barreira Hematoencefálica/imunologia , Barreira Hematoencefálica/metabolismo , Disbiose , Predisposição Genética para Doença , Antígeno HLA-B27/genética , Antígeno HLA-B27/imunologia , Humanos , Fatores de Risco , Linfócitos T Reguladores/imunologia , Linfócitos T Reguladores/metabolismo , Linfócitos T Reguladores/microbiologia , Úvea/metabolismo , Uveíte/genética , Uveíte/metabolismo , Uveíte/microbiologiaRESUMO
BACKGROUND: The prevalence of osteoarthritis (OA) increases with obesity, with up to two thirds of the elderly obese population affected by OA of the knee. The metabolic syndrome (MetS), frequently associated with central obesity and characterised by elevated waist circumference, raised fasting plasma glucose concentration, raised triglycerides, reduced high-density lipoproteins, and/or hypertension, is implicated in the pathogenesis of OA. This narrative review discusses the mechanisms involved in the influence of MetS on OA, with a focus on the effects on macrophages and chondrocytes. MAIN TEXT: A skewing of macrophages towards a pro-inflammatory M1 phenotype within synovial and adipose tissues is thought to play a role in OA pathogenesis. The metabolic perturbations typical of MetS are important drivers of pro-inflammatory macrophage polarisation and activity. This is mediated via alterations in the levels and activities of the cellular nutrient sensors 5' adenosine monophosphate-activated protein kinase (AMPK) and mammalian target of rapamycin complex 1 (mTORC1), intracellular accumulation of metabolic intermediates such as succinate and citrate, and increases in free fatty acids (FFAs) and hyperglycaemia-induced advanced glycation end-products (AGEs) that bind to receptors on the macrophage surface. Altered levels of adipokines, including leptin and adiponectin, further influence macrophage polarisation. The metabolic alterations in MetS also affect the cartilage through direct effects on chondrocytes by stimulating the production of pro-inflammatory and catabolic factors and possibly by suppressing autophagy and promoting cellular senescence. CONCLUSIONS: The influence of MetS on OA pathogenesis involves a wide range of metabolic alterations that directly affect macrophages and chondrocytes. The relative burden of intra-articular versus systemic adipose tissue in the MetS-associated OA remains to be clarified. Understanding how altered metabolism interacts with joints affected by OA is crucial for the development of further strategies for treating this debilitating condition, such as supplementing existing therapies with metformin and utilising ω-3 fatty acid derivatives to restore imbalances in ω-3 and ω-6 fatty acids.