RESUMEN
Neutrophils were long considered to be a short-lived homogenous cell population, limited to their role as first responders in anti-bacterial and -fungal immunity. While it is true that neutrophils are first to infiltrate the site of infection to eliminate pathogens, growing evidence suggests their functions could extend beyond those of basic innate immune cells. Along with their well-established role in pathogen elimination, utilizing effector functions such as phagocytosis, degranulation, and the deployment of neutrophil extracellular traps (NETs), neutrophils have recently been shown to possess antigen-presenting capabilities. Moreover, the identification of different subtypes of neutrophils points to a multifactorial heterogeneous cell population with great plasticity in which some subsets have enhanced pro-inflammatory characteristics, while others seem to behave as immunosuppressors. Interestingly, the aberrant presence of activated neutrophils with a pro-inflammatory profile in several systemic and organ-specific autoimmune diseases such as systemic lupus erythematosus (SLE), rheumatoid arthritis (RA), systemic sclerosis (SSc), multiple sclerosis (MS), and type 1 diabetes (T1D) could potentially be exploited in novel therapeutic strategies. The full extent of the involvement of neutrophils, and more specifically that of their various subtypes, in the pathophysiology of autoimmune diseases is yet to be elucidated.
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Enfermedades Autoinmunes , Trampas Extracelulares , Lupus Eritematoso Sistémico , Humanos , Neutrófilos , AutoinmunidadRESUMEN
BACKGROUND: In chronic obstructive pulmonary disease (COPD), exacerbations cause acute inflammatory flare-ups and increase the risk for hospitalization and mortality. Exacerbations are common in all disease stages and are often caused by bacterial infections e.g., non-typeable Heamophilus influenzae (NTHi). Accumulating evidence also associates vitamin D deficiency with the severity of COPD and exacerbation frequency. However, it is still unclear whether vitamin D deficiency when combined with cigarette smoking would worsen and prolong exacerbations caused by repeated infections with the same bacterial strain. METHODS: Vitamin D sufficient (VDS) and deficient (VDD) mice were exposed to nose-only cigarette smoke (CS) for 14 weeks and oropharyngeally instilled with NTHi at week 6, 10 and 14. Three days after the last instillation, mice were assessed for lung function, tissue remodeling, inflammation and immunity. The impact of VDD and CS on inflammatory cells and immunoglobulin (Ig) production was also assessed in non-infected animals while serum Ig production against NTHi and dsDNA was measured in COPD patients before and 1 year after supplementation with Vitamin D3. RESULTS: VDD enhanced NTHi eradication, independently of CS and complete eradication was reflected by decreased anti-NTHi Ig's within the lung. In addition, VDD led to an increase in total lung capacity (TLC), lung compliance (Cchord), MMP12/TIMP1 ratio with a rise in serum Ig titers and anti-dsDNA Ig's. Interestingly, in non-infected animals, VDD exacerbated the CS-induced anti-NTHi Ig's, anti-dsDNA Ig's and inflammatory cells within the lung. In COPD patients, serum Ig production was not affected by vitamin D status but anti-NTHi IgG increased after vitamin D3 supplementation in patients who were Vitamin D insufficient before treatment. CONCLUSION: During repeated infections, VDD facilitated NTHi eradication and resolution of local lung inflammation through production of anti-NTHi Ig, independently of CS whilst it also promoted autoantibodies. In COPD patients, vitamin D supplementation could be protective against NTHi infections in vitamin D insufficient patients. Future research is needed to decipher the determinants of dual effects of VDD on adaptive immunity. TRAIL REGISTRATION: ClinicalTrials, NCT00666367. Registered 23 April 2008, https://www.clinicaltrials.gov/ct2/show/study/NCT00666367 .
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Fumar Cigarrillos/efectos adversos , Infecciones por Haemophilus/complicaciones , Haemophilus influenzae/inmunología , Pulmón/microbiología , Neumonía/complicaciones , Deficiencia de Vitamina D/metabolismo , Animales , Modelos Animales de Enfermedad , Infecciones por Haemophilus/metabolismo , Infecciones por Haemophilus/microbiología , Pulmón/metabolismo , Pulmón/patología , Masculino , Ratones , Ratones Endogámicos C57BL , Neumonía/metabolismoRESUMEN
RXR signaling is predicted to have a major impact in macrophages, but neither the biological consequence nor the genomic basis of its ligand activation is known. Comprehensive genome-wide studies were carried out to map liganded RXR-mediated transcriptional changes, active binding sites, and cistromic interactions in the context of the macrophage genome architecture. The macrophage RXR cistrome has 5200 genomic binding sites, which are not impacted by ligand. Active enhancers are characterized by PU.1 binding, an increase of enhancer RNA, and P300 recruitment. Using these features, 387 liganded RXR-bound enhancers were linked to 226 genes, which predominantly reside in CTCF/cohesin-limited functional domains. These findings were molecularly validated using chromosome conformation capture (3C) and 3C combined with sequencing (3C-seq), and we show that selected long-range enhancers communicate with promoters via stable or RXR-induced loops and that some of the enhancers interact with each other, forming an interchromosomal network. A set of angiogenic genes, including Vegfa, has liganded RXR-controlled enhancers and provides the macrophage with a novel inducible program.
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Elementos de Facilitación Genéticos , Macrófagos/metabolismo , Neovascularización Fisiológica/fisiología , Receptores X Retinoide/metabolismo , Animales , Células Cultivadas , Regulación del Desarrollo de la Expresión Génica/efectos de los fármacos , Histonas/metabolismo , Ligandos , Macrófagos/citología , Macrófagos/efectos de los fármacos , Ratones , Compuestos Orgánicos/química , Compuestos Orgánicos/metabolismo , Compuestos Orgánicos/farmacología , ARN/metabolismo , Transcripción Genética/efectos de los fármacosRESUMEN
AIMS/HYPOTHESIS: MicroRNAs (miRNAs) are a novel class of potential biomarkers emerging in many diseases, including type 1 diabetes. Here, we aim to analyse a panel of circulating miRNAs in non-obese diabetic (NOD) mice and individuals with type 1 diabetes. METHODS: We adopted standardised methodologies for extracting miRNAs from small sample volumes to evaluate a profiling panel of mature miRNAs in paired plasma and laser-captured microdissected immune-infiltrated islets of recently diabetic and normoglycaemic NOD mice. Moreover, we validated the findings during disease progression and remission after anti-CD3 therapy in NOD mice, as well as in individuals with type 1 diabetes. RESULTS: Plasma levels of five miRNAs were downregulated in diabetic vs normoglycaemic mice. Of those, miR-409-3p was also downregulated in situ in the immune islet infiltrates of diabetic mice, suggesting an association with disease pathogenesis. Target-prediction tools linked miR-409-3p to immune- and metabolism-related signalling molecules. In situ miR-409-3p expression correlated with insulitis severity, and CD8+ central memory T cells were found to be enriched in miR-409-3p. Plasma miR-409-3p levels gradually decreased during diabetes development and improved with disease remission after anti-CD3 antibody therapy. Finally, plasma miR-409-3p levels were lower in people recently diagnosed with type 1 diabetes compared with a non-diabetic control group, and levels were inversely correlated with HbA1c levels. CONCLUSIONS/INTERPRETATION: We propose that miR-409-3p may represent a new circulating biomarker of islet inflammation and type 1 diabetes severity.
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Diabetes Mellitus Tipo 1/genética , Ratones Endogámicos NOD/genética , MicroARNs/genética , Animales , Biomarcadores/metabolismo , Femenino , Citometría de Flujo , Perfilación de la Expresión Génica , Humanos , Ratones , Reacción en Cadena en Tiempo Real de la PolimerasaRESUMEN
OBJECTIVE: Tracking the autoreactive T-cell migration in the pancreatic region after labeling with fluorinated nanoparticles (1,2-dioleoyl-sn-glycero-3-phosphoethanolamine-N-[3-(2-pyridyldithio)propionate]-perfluoro-15-crown-5-ether nanoparticles, PDP-PFCE NPs) in a diabetic murine model using 19F MRI. MATERIALS AND METHODS: Synthesis of novel PDP-PFCE fluorine tracer was performed for in vitro labeling of T cells. Labeling conditions were optimized using different PDP-PFCE NPs concentrations. For in vivo 19F MRI, mice were longitudinally followed after adoptive transfer of activated, autoreactive, labeled T cells in NOD.SCID mice. RESULTS: Established MR protocols were used for challenging T cell labeling to track inflammation in a model of diabetes after successful labeling of CD4+ and CD8+ T cells with PDP-PFCE NPs. However, T cells were difficult to be detected in vivo after their engraftment in animals. DISCUSSION: We showed successful in vitro labeling of T cells using novel fluorinated liposomal nanoparticles. However, insufficient and slow accumulation of labeled T cells and subsequent T cell proliferation in the pancreatic region remains as limitations of in vivo cell imaging by 19F MRI.
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Traslado Adoptivo , Diabetes Mellitus Experimental/diagnóstico por imagen , Diabetes Mellitus Experimental/inmunología , Diabetes Mellitus Tipo 1/diagnóstico por imagen , Diabetes Mellitus Tipo 1/inmunología , Linfocitos T/citología , Animales , Linfocitos T CD4-Positivos/citología , Linfocitos T CD8-positivos/citología , Proliferación Celular , Modelos Animales de Enfermedad , Flúor/química , Inflamación , Isótopos/química , Liposomas/química , Imagen por Resonancia Magnética , Ratones , Ratones Endogámicos NOD , Ratones SCID , Nanopartículas/química , Bazo/metabolismo , TransgenesRESUMEN
AIMS/HYPOTHESIS: The initial avascular period following islet transplantation seriously compromises graft function and survival. Enhancing graft revascularisation to improve engraftment has been attempted through virus-based delivery of angiogenic triggers, but risks associated with viral vectors have hampered clinical translation. In vitro transcribed mRNA transfection circumvents these risks and may be used for improving islet engraftment. METHODS: Mouse and human pancreatic islet cells were transfected with mRNA encoding the angiogenic growth factor vascular endothelial growth factor A (VEGF-A) before transplantation under the kidney capsule in mice. RESULTS: At day 7 post transplantation, revascularisation of grafts transfected with Vegf-A (also known as Vegfa) mRNA was significantly higher compared with non-transfected or Gfp mRNA-transfected controls in mouse islet grafts (2.11- and 1.87-fold, respectively) (vessel area/graft area, mean ± SEM: 0.118 ± 0.01 [n = 3] in Vegf-A mRNA transfected group (VEGF) vs 0.056 ± 0.01 [n = 3] in no RNA [p < 0.05] vs 0.063 ± 0.02 [n = 4] in Gfp mRNA transfected group (GFP) [p < 0.05]); EndoC-bH3 grafts (2.85- and 2.48-fold. respectively) (0.085 ± 0.02 [n = 4] in VEGF vs 0.030 ± 0.004 [n = 4] in no RNA [p < 0.05] vs 0.034 ± 0.01 [n = 5] in GFP [p < 0.05]); and human islet grafts (3.17- and 3.80-fold, respectively) (0.048 ± 0.013 [n = 3] in VEGF vs 0.015 ± 0.0051 [n = 4] in no RNA [p < 0.01] vs 0.013 ± 0.0046 [n = 4] in GFP [p < 0.01]). At day 30 post transplantation, human islet grafts maintained a vascularisation benefit (1.70- and 1.82-fold, respectively) (0.049 ± 0.0042 [n = 8] in VEGF vs 0.029 ± 0.0052 [n = 5] in no RNA [p < 0.05] vs 0.027 ± 0.0056 [n = 4] in GFP [p < 0.05]) and a higher beta cell volume (1.64- and 2.26-fold, respectively) (0.0292 ± 0.0032 µl [n = 7] in VEGF vs 0.0178 ± 0.0021 µl [n = 5] in no RNA [p < 0.01] vs 0.0129 ± 0.0012 µl [n = 4] in GFP [p < 0.001]). CONCLUSIONS/INTERPRETATION: Vegf-A mRNA transfection before transplantation provides a promising and safe strategy to improve engraftment of islets and other cell-based implants.
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Células Secretoras de Insulina/citología , Islotes Pancreáticos/citología , Neovascularización Fisiológica , ARN Mensajero/genética , Transfección , Factor A de Crecimiento Endotelial Vascular/genética , Animales , Supervivencia Celular , Humanos , Insulina/metabolismo , Células Secretoras de Insulina/trasplante , Trasplante de Islotes Pancreáticos , RatonesRESUMEN
As cancer is becoming more and more a chronic disease, a large proportion of patients is confronted with devastating side effects of certain anti-cancer drugs. The most common neurological complications are painful peripheral neuropathies. Chemotherapeutics that interfere with microtubules, including plant-derived vinca-alkaloids such as vincristine, can cause these chemotherapy-induced peripheral neuropathies (CIPN). Available treatments focus on symptom alleviation and pain reduction rather than prevention of the neuropathy. The aim of this study was to investigate the potential of specific histone deacetylase 6 (HDAC6) inhibitors as a preventive therapy for CIPN using multiple rodent models for vincristine-induced peripheral neuropathies (VIPN). HDAC6 inhibition increased the levels of acetylated α-tubulin in tissues of rodents undergoing vincristine-based chemotherapy, which correlates to a reduced severity of the neurological symptoms, both at the electrophysiological and the behavioral level. Mechanistically, disturbances in axonal transport of mitochondria is considered as an important contributing factor in the pathophysiology of VIPN. As vincristine interferes with the polymerization of microtubules, we investigated whether disturbances in axonal transport could contribute to VIPN. We observed that increasing α-tubulin acetylation through HDAC6 inhibition restores vincristine-induced defects of axonal transport in cultured dorsal root ganglion neurons. Finally, we assured that HDAC6-inhibition offers neuroprotection without interfering with the anti-cancer efficacy of vincristine using a mouse model for acute lymphoblastic leukemia. Taken together, our results emphasize the therapeutic potential of HDAC6 inhibitors with beneficial effects both on vincristine-induced neurotoxicity, as well as on tumor proliferation.
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Antineoplásicos/efectos adversos , Histona Desacetilasa 6/antagonistas & inhibidores , Neoplasias/tratamiento farmacológico , Fármacos Neuroprotectores/farmacología , Enfermedades del Sistema Nervioso Periférico/tratamiento farmacológico , Vincristina/efectos adversos , Animales , Antineoplásicos/farmacología , Transporte Axonal/efectos de los fármacos , Células Cultivadas , Modelos Animales de Enfermedad , Ganglios Espinales/efectos de los fármacos , Ganglios Espinales/metabolismo , Histona Desacetilasa 6/metabolismo , Inhibidores de Histona Desacetilasas/farmacología , Masculino , Ratones Endogámicos NOD , Ratones SCID , Microtúbulos/efectos de los fármacos , Microtúbulos/metabolismo , Mitocondrias/efectos de los fármacos , Mitocondrias/metabolismo , Neoplasias/enzimología , Enfermedades del Sistema Nervioso Periférico/inducido químicamente , Enfermedades del Sistema Nervioso Periférico/enzimología , Tubulina (Proteína)/metabolismoRESUMEN
AIMS/HYPOTHESIS: Hypoxia in the initial days after islet transplantation leads to considerable loss of islet mass and contributes to disappointing outcomes in the clinical setting. The aim of the present study was to investigate whether co-transplantation of human non-endothelial bone marrow-derived multipotent adult progenitor cells (MAPCs), which are non-immunogenic and can secrete angiogenic growth factors during the initial days after implantation, could improve islet engraftment and survival. METHODS: Islets (150) were co-transplanted, with or without human MAPCs (2.5 × 105) as separate or composite pellets, under the kidney capsule of syngeneic alloxan-induced diabetic C57BL/6 mice. Blood glucose levels were frequently monitored and IPGTTs were carried out. Grafts and serum were harvested at 2 and 5 weeks after transplantation to assess outcome. RESULTS: Human MAPCs produced high amounts of angiogenic growth factors, including vascular endothelial growth factor, in vitro and in vivo, as demonstrated by the induction of neo-angiogenesis in the chorioallantoic membrane assay. Islet-human MAPC co-transplantation as a composite pellet significantly improved the outcome of islet transplantation as measured by the initial glycaemic control, diabetes reversal rate, glucose tolerance and serum C-peptide concentration compared with the outcome following transplantation of islets alone. Histologically, a higher blood vessel area and density in addition to a higher vessel/islet ratio were detected in recipients of islet-human MAPC composites. CONCLUSIONS/INTERPRETATION: The present data suggest that co-transplantation of mouse pancreatic islets with human MAPCs, which secrete high amounts of angiogenic growth factors, enhance islet graft revascularisation and subsequently improve islet graft function.
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Diabetes Mellitus Tipo 1/terapia , Trasplante de Células Madre/métodos , Células Madre/citología , Adulto , Animales , Glucemia/fisiología , Células Cultivadas , Diabetes Mellitus Experimental/fisiopatología , Diabetes Mellitus Experimental/terapia , Diabetes Mellitus Tipo 1/fisiopatología , Modelos Animales de Enfermedad , Femenino , Humanos , Islotes Pancreáticos/metabolismo , Masculino , Ratones Endogámicos C57BL , Persona de Mediana Edad , Neovascularización Fisiológica/fisiología , Células Madre/fisiologíaRESUMEN
AIMS/HYPOTHESIS: Type 1 diabetes is an endocrine disease where a long preclinical phase, characterised by immune cell infiltration in the islets of Langerhans, precedes elevated blood glucose levels and disease onset. Although several studies have investigated the role of the immune system in this process of insulitis, the importance of the beta cells themselves in the initiation of type 1 diabetes is less well understood. The aim of this study was to investigate intrinsic differences present in the islets from diabetes-prone NOD mice before the onset of insulitis. METHODS: The islet transcriptome and proteome of 2-3-week-old mice was investigated by microarray and 2-dimensional difference gel electrophoresis (2D-DIGE), respectively. Subsequent analyses using sophisticated pathway analysis and ranking of differentially expressed genes and proteins based on their relevance in type 1 diabetes were performed. RESULTS: In the preinsulitic period, alterations in general pathways related to metabolism and cell communication were already present. Additionally, our analyses pointed to an important role for post-translational modifications (PTMs), especially citrullination by PAD2 and protein misfolding due to low expression levels of protein disulphide isomerases (PDIA3, 4 and 6), as causative mechanisms that induce beta cell stress and potential auto-antigen generation. CONCLUSIONS/INTERPRETATION: We conclude that the pancreatic islets, irrespective of immune differences, may contribute to the initiation of the autoimmune process. DATA AVAILABILITY: All microarray data are available in the ArrayExpress database ( www.ebi.ac.uk/arrayexpress ) under accession number E-MTAB-5264.
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Islotes Pancreáticos/metabolismo , Análisis de Secuencia por Matrices de Oligonucleótidos/métodos , Estado Prediabético/metabolismo , Proteómica/métodos , Animales , Hidrolasas/genética , Hidrolasas/metabolismo , Ratones , Ratones Endogámicos C57BL , Ratones Endogámicos NOD , Modelos Teóricos , Estado Prediabético/patología , Desiminasas de la Arginina Proteica , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Electroforesis Bidimensional Diferencial en GelRESUMEN
Pro-inflammatory cytokines contribute to pancreatic beta cell apoptosis in type 1 diabetes at least in part by inducing endoplasmic reticulum (ER) stress and the consequent unfolded protein response (UPR). It remains to be determined what causes the transition from "physiological" to "apoptotic" UPR, but accumulating evidence indicates that signaling by the ER transmembrane protein IRE1α is critical for this transition. IRE1α activation is regulated by both intra-ER and cytosolic cues. We evaluated the role for the presently discovered cytokine-induced and IRE1α-interacting protein ubiquitin D (UBD) on the regulation of IRE1α and its downstream targets. UBD was identified by use of a MAPPIT (mammalian protein-protein interaction trap)-based IRE1α interactome screen followed by comparison against functional genomic analysis of human and rodent beta cells exposed to pro-inflammatory cytokines. Knockdown of UBD in human and rodent beta cells and detailed signal transduction studies indicated that UBD modulates cytokine-induced UPR/IRE1α activation and apoptosis. UBD expression is induced by the pro-inflammatory cytokines interleukin (IL)-1ß and interferon (IFN)-γ in rat and human pancreatic beta cells, and it is also up-regulated in beta cells of inflamed islets from non-obese diabetic mice. UBD interacts with IRE1α in human and rodent beta cells, modulating IRE1α-dependent activation of JNK and cytokine-induced apoptosis. Our data suggest that UBD provides a negative feedback on cytokine-induced activation of the IRE1α/JNK pro-apoptotic pathway in cytokine-exposed beta cells.
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Apoptosis , Endorribonucleasas/metabolismo , Células Secretoras de Insulina/metabolismo , Proteínas Quinasas JNK Activadas por Mitógenos/metabolismo , Proteínas Serina-Treonina Quinasas/metabolismo , Ubiquitinas/metabolismo , Anciano , Anciano de 80 o más Años , Animales , Western Blotting , Línea Celular , Línea Celular Tumoral , Células Cultivadas , Citocinas/farmacología , Endorribonucleasas/genética , Femenino , Expresión Génica/efectos de los fármacos , Células HEK293 , Humanos , Células Secretoras de Insulina/efectos de los fármacos , Masculino , Persona de Mediana Edad , Unión Proteica , Proteínas Serina-Treonina Quinasas/genética , Interferencia de ARN , Ratas , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Ubiquitinas/genética , Adulto JovenRESUMEN
OBJECTIVE: To evaluate if the up-regulation of vascular endothelial growth factor strengthens the protective effect of amniotic fluid stem cells in a renal ischemia-reperfusion injury model. DESIGN: Randomized animal study. SETTINGS: University research laboratory. SUBJECTS: A total of 40 males 12-week-old Wistar rats were subjected to ischemia-reperfusion and assigned to four groups: amniotic fluid stem cells, vascular endothelial growth factor-amniotic fluid stem cells in two different doses, and vehicle. Ten animals were used as sham-controls. INTERVENTION: Six hours after induction of renal ischemia-reperfusion injury, amniotic fluid stem cells, vascular endothelial growth factor-amniotic fluid stem cells in two different doses, or vehicle were injected intraarterially. MEASUREMENTS AND MAIN RESULTS: Analyses were performed at 24 hours, 48 hours, and 2 months after treatment. Outcome measures included serum creatinine, urine microprotenuira, and immunohistomorphometric analyses. Vascular endothelial growth factor-amniotic fluid stem cells induced a significantly higher nephroprotection than amniotic fluid stem cells. This effect was mediated mainly by immunomodulation, which led to lower macrophage infiltration and higher presence of regulatory T cell after ischemia-reperfusion injury. At medium term, it inhibited the progression toward chronic kidney disease. Vascular endothelial growth factor-amniotic fluid stem cells can worsen the ischemia-reperfusion injury when delivered in a high dose. CONCLUSIONS: Up-regulation of vascular endothelial growth factor enhances the therapeutic effect of human amniotic fluid stem cells in rats with renal ischemia-reperfusion injury, mainly by mitogenic, angiogenic, and anti-inflammatory mechanisms.
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Lesión Renal Aguda/terapia , Líquido Amniótico/citología , Daño por Reperfusión/terapia , Trasplante de Células Madre , Regulación hacia Arriba , Factor A de Crecimiento Endotelial Vascular/metabolismo , Animales , Progresión de la Enfermedad , Inyecciones Intraarteriales , Macrófagos/metabolismo , Ratas Wistar , Insuficiencia Renal Crónica/prevención & control , Células Madre/metabolismo , Linfocitos T Reguladores/metabolismoRESUMEN
Type 1 diabetes is due to destruction of pancreatic ß-cells. Lysine deacetylase inhibitors (KDACi) protect ß-cells from inflammatory destruction in vitro and are promising immunomodulators. Here we demonstrate that the clinically well-tolerated KDACi vorinostat and givinostat revert diabetes in the nonobese diabetic (NOD) mouse model of type 1 diabetes and counteract inflammatory target cell damage by a mechanism of action consistent with transcription factor--rather than global chromatin--hyperacetylation. Weaning NOD mice received low doses of vorinostat and givinostat in their drinking water until 100-120 d of age. Diabetes incidence was reduced by 38% and 45%, respectively, there was a 15% increase in the percentage of islets without infiltration, and pancreatic insulin content increased by 200%. Vorinostat treatment increased the frequency of functional regulatory T-cell subsets and their transcription factors Gata3 and FoxP3 in parallel to a decrease in inflammatory dendritic cell subsets and their cytokines IL-6, IL-12, and TNF-α. KDACi also inhibited LPS-induced Cox-2 expression in peritoneal macrophages from C57BL/6 and NOD mice. In insulin-producing ß-cells, givinostat did not upregulate expression of the anti-inflammatory genes Socs1-3 or sirtuin-1 but reduced levels of IL-1ß + IFN-γ-induced proinflammatory Il1a, Il1b, Tnfα, Fas, Cxcl2, and reduced cytokine-induced ERK phosphorylation. Further, NF-κB genomic iNos promoter binding was reduced by 50%, and NF-κB-dependent mRNA expression was blocked. These effects were associated with NF-κB subunit p65 hyperacetylation. Taken together, these data provide a rationale for clinical trials of safety and efficacy of KDACi in patients with autoimmune disease such as type 1 diabetes.
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Cromatina/metabolismo , Diabetes Mellitus Tipo 1/metabolismo , Inhibidores de Histona Desacetilasas/farmacología , Células Secretoras de Insulina/citología , Animales , Línea Celular , Citocinas/metabolismo , Modelos Animales de Enfermedad , Epigénesis Genética , Femenino , Factor de Transcripción GATA3/metabolismo , Histona Desacetilasas/metabolismo , Humanos , Ácidos Hidroxámicos/farmacología , Inflamación , Ratones , Ratones Endogámicos C57BL , Ratones Endogámicos NOD , Fosforilación , Regiones Promotoras Genéticas , Unión Proteica , Procesamiento Proteico-Postraduccional , Ratas , Factores de Tiempo , VorinostatRESUMEN
Pancreas injury by partial duct ligation (PDL) activates a healing response, encompassing ß-cell neogenesis and proliferation. Macrophages (MΦs) were recently shown to promote ß-cell proliferation after PDL, but they remain poorly characterized. We assessed myeloid cell diversity and the factors driving myeloid cell dynamics following acute pancreas injury by PDL. In naive and sham-operated pancreas, the myeloid cell compartment consisted mainly of two distinct tissue-resident MΦ types, designated MHC-II(lo) and MHC-II(hi) MΦs, the latter being predominant. MHC-II(lo) and MHC-II(hi) pancreas MΦs differed at the molecular level, with MHC-II(lo) MΦs being more M2-activated. After PDL, there was an early surge of Ly6C(hi) monocyte infiltration in the pancreas, followed by a transient MHC-II(lo) MΦ peak and ultimately a restoration of the MHC-II(hi) MΦ-dominated steady-state equilibrium. These intricate MΦ dynamics in PDL pancreas depended on monocyte recruitment by C-C chemokine receptor 2 and macrophage-colony stimulating factor receptor as well as on macrophage-colony stimulating factor receptor-dependent local MΦ proliferation. Functionally, MHC-II(lo) MΦs were more angiogenic. We further demonstrated that, at least in C-C chemokine receptor 2-KO mice, tissue MΦs, rather than Ly6C(hi) monocyte-derived MΦs, contributed to ß-cell proliferation. Together, our study fully characterizes the MΦ subsets in the pancreas and clarifies the complex dynamics of MΦs after PDL injury.
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Macrófagos/inmunología , Macrófagos/patología , Monocitos/inmunología , Monocitos/patología , Páncreas/inmunología , Páncreas/lesiones , Animales , Antígenos Ly/metabolismo , Movimiento Celular/inmunología , Proliferación Celular , Microambiente Celular/inmunología , Antígenos de Histocompatibilidad Clase II/metabolismo , Ligadura , Activación de Macrófagos , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Ratones Transgénicos , Células Mieloides/clasificación , Células Mieloides/inmunología , Células Mieloides/patología , Páncreas/patología , Conductos Pancreáticos/lesiones , Receptor de Factor Estimulante de Colonias de Macrófagos/metabolismo , Regeneración/inmunologíaRESUMEN
The biologically active form of vitamin D, 1,25-dihydroxyvitamin D3 [1,25(OH)2D3], is able to promote the generation of tolerogenic mature dendritic cells (mDCs) with an impaired ability to activate autoreactive T cells. These cells could represent a reliable tool for the promotion or restoration of Ag-specific tolerance through vaccination strategies, for example in type 1 diabetes patients. However, successful transfer of 1,25(OH)2D3-treated mDCs (1,25D3-mDCs) depends on the capacity of 1,25(OH)2D3 to imprint a similar tolerogenic profile in cells derived from diabetes-prone donors as from diabetes-resistant donors. In this study, we examined the impact of 1,25(OH)2D3 on the function and phenotype of mDCs originating from healthy (C57BL/6) and diabetes-prone (NOD) mice. We show that 1,25(OH)2D3 is able to imprint a phenotypic tolerogenic profile on DCs derived from both mouse strains. Both NOD- and C57BL/6-derived 1,25D3-mDCs decreased the proliferation and activation of autoreactive T cells in vitro, despite strain differences in the regulation of cytokine/chemokine expression. In addition, 1,25D3-mDCs from diabetes-prone mice expanded CD25(+)Foxp3(+) regulatory T cells and induced intracellular IL-10 production by T cells in vitro. Furthermore, 1,25D3-mDCs exhibited an intact functional migratory capacity in vivo that favors homing to the liver and pancreas of adult NOD mice. More importantly, when cotransferred with activated CD4(+) T cells into NOD.SCID recipients, 1,25D3-mDCs potently dampened the proliferation of autoreactive donor T cells in the pancreatic draining lymph nodes. Altogether, these results argue for the potential of 1,25D3-mDCs to restore Ag-specific immune tolerance and arrest autoimmune disease progression in vivo.
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Células Dendríticas/inmunología , Tolerancia Inmunológica/efectos de los fármacos , Vitamina D/análogos & derivados , Animales , Quimiotaxis de Leucocito/efectos de los fármacos , Quimiotaxis de Leucocito/inmunología , Células Dendríticas/citología , Células Dendríticas/efectos de los fármacos , Citometría de Flujo , Técnica del Anticuerpo Fluorescente , Tolerancia Inmunológica/inmunología , Activación de Linfocitos/efectos de los fármacos , Activación de Linfocitos/inmunología , Ratones , Ratones Endogámicos C57BL , Ratones Endogámicos NOD , Ratones Transgénicos , Fenotipo , Reacción en Cadena en Tiempo Real de la Polimerasa , Vitamina D/farmacologíaRESUMEN
Type 1 diabetes is an autoimmune disease with a strong inflammatory component. The cytokines interleukin-1ß and interferon-γ contribute to beta cell apoptosis in type 1 diabetes. These cytokines induce endoplasmic reticulum stress and the unfolded protein response (UPR), contributing to the loss of beta cells. IRE1α, one of the UPR mediators, triggers insulin degradation and inflammation in beta cells and is critical for the transition from "physiological" to "pathological" UPR. The mechanisms regulating inositol-requiring protein 1α (IRE1α) activation and its signaling for beta cell "adaptation," "stress response," or "apoptosis" remain to be clarified. To address these questions, we combined mammalian protein-protein interaction trap-based IRE1α interactome and functional genomic analysis of human and rodent beta cells exposed to pro-inflammatory cytokines to identify novel cytokine-induced regulators of IRE1α. Based on this approach, we identified N-Myc interactor (NMI) as an IRE1α-interacting/modulator protein in rodent and human pancreatic beta cells. An increased expression of NMI was detected in islets from nonobese diabetic mice with insulitis and in rodent or human beta cells exposed in vitro to the pro-inflammatory cytokines interleukin-1ß and interferon-γ. Detailed mechanistic studies demonstrated that NMI negatively modulates IRE1α-dependent activation of JNK and apoptosis in rodent and human pancreatic beta cells. In conclusion, by using a combined omics approach, we identified NMI induction as a novel negative feedback mechanism that decreases IRE1α-dependent activation of JNK and apoptosis in cytokine-exposed beta cells
Asunto(s)
Endorribonucleasas/metabolismo , Células Secretoras de Insulina/metabolismo , Interferón gamma/metabolismo , Interleucina-1beta/metabolismo , Péptidos y Proteínas de Señalización Intracelular/metabolismo , Proteínas Quinasas JNK Activadas por Mitógenos/metabolismo , Complejos Multienzimáticos/metabolismo , Proteínas Serina-Treonina Quinasas/metabolismo , Respuesta de Proteína Desplegada/fisiología , Anciano , Animales , Apoptosis/fisiología , Endorribonucleasas/genética , Células HEK293 , Humanos , Células Secretoras de Insulina/citología , Interferón gamma/genética , Interleucina-1beta/genética , Péptidos y Proteínas de Señalización Intracelular/genética , Proteínas Quinasas JNK Activadas por Mitógenos/genética , Ratones , Persona de Mediana Edad , Complejos Multienzimáticos/genética , Proteínas Serina-Treonina Quinasas/genética , Ratas , Ratas WistarRESUMEN
BACKGROUND: Chronic obstructive pulmonary disease (COPD) is characterized by excessive inflammation and disturbed bacterial clearance in the airways. Although cigarette smoke (CS) exposure poses a major risk, vitamin D deficiency could potentially contribute to COPD progression. Many in vitro studies demonstrate important anti-inflammatory and antibacterial effects of vitamin D, but a direct contribution of vitamin D deficiency to COPD onset and disease progression has not been explored. METHODS: In the current study, we used a murine experimental model to investigate the combined effect of vitamin D deficiency and CS exposure on the development of COPD-like characteristics. Therefore, vitamin D deficient or control mice were exposed to CS or ambient air for a period of 6 (subacute) or 12 weeks (chronic). Besides lung function and structure measurements, we performed an in depth analysis of the size and composition of the cellular infiltrate in the airways and lung parenchyma and tested the ex vivo phagocytic and oxidative burst capacity of alveolar macrophages. RESULTS: Vitamin D deficient mice exhibited an accelerated lung function decline following CS exposure compared to control mice. Furthermore, early signs of emphysema were only observed in CS-exposed vitamin D deficient mice, which was accompanied by elevated levels of MMP-12 in the lung. Vitamin D deficient mice showed exacerbated infiltration of inflammatory cells in the airways and lung parenchyma after both subacute and chronic CS exposure compared to control mice. Furthermore, elevated levels of typical proinflammatory cytokines and chemokines could be detected in the bronchoalveolar lavage fluid (KC and TNF-α) and lung tissue (IP-10, MCP-1, IL-12) of CS-exposed vitamin D deficient mice compared to control mice. Finally, although CS greatly impaired the ex vivo phagocytic and oxidative burst function of alveolar macrophages, vitamin D deficient mice did not feature an additional defect. CONCLUSIONS: Our data demonstrate that vitamin D deficiency both accelerates and aggravates the development of characteristic disease features of COPD. As vitamin D deficiency is highly prevalent, large randomized trials exploring effects of vitamin D supplementation on lung function decline and COPD onset are needed.
Asunto(s)
Pulmón/fisiopatología , Enfermedad Pulmonar Obstructiva Crónica/etiología , Humo , Fumar/efectos adversos , Deficiencia de Vitamina D/complicaciones , Animales , Líquido del Lavado Bronquioalveolar/química , Calcio/sangre , Citocinas/metabolismo , Modelos Animales de Enfermedad , Progresión de la Enfermedad , Femenino , Mediadores de Inflamación/metabolismo , Pulmón/metabolismo , Pulmón/patología , Activación de Macrófagos , Macrófagos Alveolares , Masculino , Metaloproteinasa 12 de la Matriz/metabolismo , Ratones Endogámicos C57BL , Fagocitosis , Neumonía/etiología , Neumonía/fisiopatología , Enfermedad Pulmonar Obstructiva Crónica/diagnóstico , Enfermedad Pulmonar Obstructiva Crónica/metabolismo , Enfermedad Pulmonar Obstructiva Crónica/fisiopatología , Enfisema Pulmonar/etiología , Enfisema Pulmonar/fisiopatología , Estallido Respiratorio , Factores de Riesgo , Factores de Tiempo , Vitamina D/análogos & derivados , Vitamina D/sangre , Deficiencia de Vitamina D/diagnóstico , Deficiencia de Vitamina D/metabolismoRESUMEN
AIMS/HYPOTHESIS: Research on the pathogenesis of type 1 diabetes relies heavily on good animal models. The aim of this work was to study the translational value of animal models of type 1 diabetes to the human situation. METHODS: We compared the four major animal models of spontaneous type 1 diabetes, namely the NOD mouse, BioBreeding (BB) rat, Komeda rat and LEW.1AR1-iddm rat, by examining the immunohistochemistry and in situ RT-PCR of immune cell infiltrate and cytokine pattern in pancreatic islets, and by comparing findings with human data. RESULTS: After type 1 diabetes manifestation CD8(+) T cells, CD68(+) macrophages and CD4(+) T cells were observed as the main immune cell types with declining frequency, in infiltrated islets of all diabetic pancreases. IL-1ß and TNF-α were the main proinflammatory cytokines in the immune cell infiltrate in NOD mice, BB rats and LEW.1AR1-iddm rats, as well as in humans. The Komeda rat was the exception, with IFN-γ and TNF-α being the main cytokines. In addition, IL-17 and IL-6 and the anti-inflammatory cytokines IL-4, IL-10 and IL-13 were found in some infiltrating immune cells. Apoptotic as well as proliferating beta cells were observed in infiltrated islets. In healthy pancreases no proinflammatory cytokine expression was observed. CONCLUSIONS/INTERPRETATION: With the exception of the Komeda rat, the animal models mirror very well the situation in humans with type 1 diabetes. Thus animal models of type 1 diabetes can provide meaningful information on the disease processes in the pancreas of patients with type 1 diabetes.
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Apoptosis , Linfocitos B/patología , Citocinas/metabolismo , Diabetes Mellitus Experimental , Diabetes Mellitus Tipo 1/metabolismo , Diabetes Mellitus Tipo 1/patología , Células Secretoras de Insulina/patología , Animales , Apoptosis/inmunología , Linfocitos B/inmunología , Diabetes Mellitus Experimental/metabolismo , Diabetes Mellitus Experimental/patología , Diabetes Mellitus Tipo 1/inmunología , Regulación de la Expresión Génica , Humanos , Inmunohistoquímica , Células Secretoras de Insulina/inmunología , Células Secretoras de Insulina/metabolismo , Interferón gamma/metabolismo , Ratones , Ratones Endogámicos NOD , Ratas , Ratas Endogámicas BB , Ratas Endogámicas Lew , Reacción en Cadena en Tiempo Real de la Polimerasa , Factor de Necrosis Tumoral alfa/metabolismoRESUMEN
Tumor growth coincides with an accumulation of myeloid-derived suppressor cells (MDSCs), which exert immune suppression and which consist of two main subpopulations, known as monocytic (MO) CD11b(+) CD115(+) Ly6G(-) Ly6C(high) MDSCs and granulocytic CD11b(+) CD115(-) Ly6G(+) Ly6C(int) polymorphonuclear (PMN)-MDSCs. However, whether these distinct MDSC subsets hamper all aspects of early CD8(+) T-cell activation--including cytokine production, surface marker expression, survival, and cytotoxicity--is currently unclear. Here, employing an in vitro coculture system, we demonstrate that splenic MDSC subsets suppress antigen-driven CD8(+) T-cell proliferation, but differ in their dependency on IFN-γ, STAT-1, IRF-1, and NO to do so. Moreover, MO-MDSC and PMN-MDSCs diminish IL-2 levels, but only MO-MDSCs affect IL-2Rα (CD25) expression and STAT-5 signaling. Unexpectedly, however, both MDSC populations stimulate IFN-γ production by CD8(+) T cells on a per cell basis, illustrating that some T-cell activation characteristics are actually stimulated by MDSCs. Conversely, MO-MDSCs counteract the activation-induced change in CD44, CD62L, CD162, and granzyme B expression, while promoting CD69 and Fas upregulation. Together, these effects result in an altered CD8(+) T-cell adhesiveness to the extracellular matrix and selectins, sensitivity to FasL-mediated apoptosis, and cytotoxicity. Hence, MDSCs intricately influence different CD8(+) T-cell activation events in vitro, whereby some parameters are suppressed while others are stimulated.
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Linfocitos T CD8-positivos/inmunología , Activación de Linfocitos/inmunología , Células Mieloides/inmunología , Neoplasias/inmunología , Animales , Antígenos CD/biosíntesis , Antígenos de Diferenciación de Linfocitos T/biosíntesis , Antígenos Ly/metabolismo , Apoptosis/inmunología , Antígeno CD11b/metabolismo , Adhesión Celular/inmunología , Línea Celular , Proliferación Celular , Femenino , Granzimas/biosíntesis , Receptores de Hialuranos/biosíntesis , Factor 1 Regulador del Interferón , Interferón gamma/biosíntesis , Interleucina-2/metabolismo , Subunidad alfa del Receptor de Interleucina-2/metabolismo , Selectina L/biosíntesis , Lectinas Tipo C/biosíntesis , Glicoproteínas de Membrana/biosíntesis , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Neutrófilos/inmunología , Óxido Nítrico/metabolismo , Receptor de Factor Estimulante de Colonias de Macrófagos/metabolismo , Factor de Transcripción STAT1/metabolismo , Factor de Transcripción STAT5/metabolismo , Transducción de Señal/inmunología , Regulación hacia Arriba , Receptor fas/biosíntesisRESUMEN
An altered gut microbiota is associated with type 1 diabetes (T1D), affecting the production of short-chain fatty acids (SCFA) and glucose homeostasis. We previously demonstrated that enhancing serum acetate and butyrate using a dietary supplement (HAMSAB) improved glycemia in non-obese diabetic (NOD) mice and patients with established T1D. The effects of SCFA on immune-infiltrated islet cells remain to be clarified. Here, we performed single-cell RNA sequencing on islet cells from NOD mice fed an HAMSAB or control diet. HAMSAB induced a regulatory gene expression profile in pancreas-infiltrated immune cells. Moreover, HAMSAB maintained the expression of ß-cell functional genes and decreased cellular stress. HAMSAB-fed mice showed preserved pancreatic endocrine cell identity, evaluated by decreased numbers of poly-hormonal cells. Finally, SCFA increased insulin levels in human ß-like cells and improved transplantation outcome in NOD/SCID mice. Our findings support the use of metabolite-based diet as attractive approach to improve glucose control in T1D.
RESUMEN
Vitamin D is a fat-soluble precursor of the circulating 25-hydroxyvitamin D3 (25(OH)D3)which can be converted by the 1α-hydroxylase (1α(OH)ase) enzyme into the bioactive hormonal metabolite 1,25-dihydroxyvitamin D3 (1,25(OH)2D3), generally known to promote bone mineralization through its ability to enhance calcium absorption from the gut. Importantly, in humans, vitamin D is mainly derived from endogenous production of vitamin D3 from ultraviolet (UV) radiation exposure to the skin while a small part (<10%) is obtained via dietary intake of dairy products and fatty fish (1). Taking these factors into account, geographic distribution and seasonality, skin pigmentation, age, and lifestyle may predispose certain populations to be at a higher risk of developing vitamin D insufficiency or deficiency (2). The first valid reports correlating the importance of an adequate vitamin D status to optimal human health originate from the early part of the 20th century, when vitamin D was described to prevent and treat the bone disease rickets. Since then, the findings that vitamin D receptors (VDR) are present in many body tissues and that vitamin D metabolizing enzymes can be found in various cells outside the kidney, including the intestine, prostate, immune cells, and within the skin itself (reviewed in reference 3), have revolutionized the vitamin D business. In this review, we will mainly focus on vitamin D as a component of immune regulation and on the role of vitamin D in antigen-specific and non-specific therapies with potential relevance for type 1 diabetes (T1D).