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3.
Nat Genet ; 49(8): 1192-1201, 2017 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-28628108

RESUMO

Few monogenic causes for severe manifestations of common allergic diseases have been identified. Through next-generation sequencing on a cohort of patients with severe atopic dermatitis with and without comorbid infections, we found eight individuals, from four families, with novel heterozygous mutations in CARD11, which encodes a scaffolding protein involved in lymphocyte receptor signaling. Disease improved over time in most patients. Transfection of mutant CARD11 expression constructs into T cell lines demonstrated both loss-of-function and dominant-interfering activity upon antigen receptor-induced activation of nuclear factor-κB and mammalian target of rapamycin complex 1 (mTORC1). Patient T cells had similar defects, as well as low production of the cytokine interferon-γ (IFN-γ). The mTORC1 and IFN-γ production defects were partially rescued by supplementation with glutamine, which requires CARD11 for import into T cells. Our findings indicate that a single hypomorphic mutation in CARD11 can cause potentially correctable cellular defects that lead to atopic dermatitis.


Assuntos
Proteínas Adaptadoras de Sinalização CARD/genética , Dermatite Atópica/genética , Mutação em Linhagem Germinativa , Guanilato Ciclase/genética , Sistema ASC de Transporte de Aminoácidos/metabolismo , Estudos de Coortes , Análise Mutacional de DNA , Dermatite Atópica/imunologia , Feminino , Genes Dominantes , Glutamina/metabolismo , Humanos , Células Jurkat , Ativação Linfocitária , Masculino , Alvo Mecanístico do Complexo 1 de Rapamicina , Antígenos de Histocompatibilidade Menor/metabolismo , Complexos Multiproteicos/metabolismo , NF-kappa B/metabolismo , Linhagem , Linfócitos T/imunologia , Linfócitos T/metabolismo , Serina-Treonina Quinases TOR/metabolismo
5.
J Allergy Clin Immunol ; 125(5): 995-1000, 2010 May.
Artigo em Inglês | MEDLINE | ID: mdl-20381849

RESUMO

BACKGROUND: There is little knowledge about clinical variables associated with vitamin D (VitD) insufficiency in asthmatic children. OBJECTIVE: We sought to investigate disease variables associated with VitD insufficiency in patients with childhood asthma and interaction of VitD with corticosteroid-mediated anti-inflammatory responses. METHODS: We analyzed 25-hydroxyvitamin D serum levels in 100 asthmatic children to investigate relationships between 25-hydroxyvitamin D levels and patients' characteristics. We determined VitD's effects on dexamethasone (DEX) induction of mitogen-activated protein kinase phosphatase 1 and IL-10 in PBMCs. RESULTS: The median 25-hydroxyvitamin D serum level was 31 ng/mL. Forty-seven percent of subjects had VitD levels in the insufficient range (<30 ng/mL), whereas 17% were VitD deficient (<20 ng/mL). Log(10) IgE (P = .01, rho = -0.25) and the number of positive aeroallergen skin prick test responses (P = .02, rho = -0.23) showed a significant inverse correlation with VitD levels, whereas FEV(1) percent predicted (P = .004, rho = 0.34) and FEV(1)/forced vital capacity ratio (P = .01, rho = 0.30) showed a significant positive correlation with VitD levels. The use of inhaled steroids (P = .0475), use of oral steroids (P = .02), and total steroid dose (P = .001) all showed significant inverse correlations with VitD levels. The amount of mitogen-activated protein kinase phosphatase 1 and IL10 mRNA induced by VitD plus DEX was significantly greater than that induced by DEX alone (P < .01). In an experimental model of steroid resistance in which DEX alone did not inhibit T-cell proliferation, addition of VitD to DEX resulted in significant dose-dependent suppression of cell proliferation. CONCLUSIONS: Corticosteroid use and worsening airflow limitation are associated with lower VitD serum levels in asthmatic patients. VitD enhances glucocorticoid action in PBMCs from asthmatic patients and enhances the immunosuppressive function of DEX in vitro.


Assuntos
Asma/tratamento farmacológico , Dexametasona/uso terapêutico , Glucocorticoides/uso terapêutico , Deficiência de Vitamina D/etiologia , Vitamina D/análogos & derivados , Criança , Pré-Escolar , Dexametasona/farmacologia , Fosfatase 1 de Especificidade Dupla/metabolismo , Feminino , Glucocorticoides/farmacologia , Humanos , Interleucina-10/metabolismo , Ativação Linfocitária , Masculino , Reação em Cadeia da Polimerase , Linfócitos T/imunologia , Linfócitos T/metabolismo , Vitamina D/sangue
6.
J Allergy Clin Immunol ; 122(3): 550-9.e3, 2008 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-18774390

RESUMO

BACKGROUND: The cause of corticosteroid-resistant (CR) asthma is unknown. OBJECTIVE: We sought to perform gene microarray analyses by using bronchoalveolar lavage (BAL) cells from well-characterized subjects with CR asthma and subject with corticosteroid-sensitive (CS) asthma to elucidate the differential expression of genes that contribute to the development of corticosteroid resistance. METHODS: The patients were characterized as having CR or CS asthma based on FEV(1) percent predicted improvement after a 1-week course of oral prednisone. Expression of selected gene targets was verified by means of real-time PCR and ELISA. RESULTS: Microarray analyses demonstrated significantly higher levels (>3-fold increase, P < .05) of transcripts for TNF-alpha, IL-1 alpha, IL-1 beta, IL-6, CXCL1, CXCL2, CXCL3, CXCL8 (IL-8), CCL3, CCL4, and CCL20 in BAL cells of subjects with CR asthma. These findings, confirmed by means of RT-PCR in additional BAL samples, were consistent with classical macrophage activation by bacterial products. In contrast, markers of alternatively activated macrophages, arginase I and CCL24, were decreased. Genes associated with activation of the LPS signaling pathway (early growth response 1, dual-specificity phosphatase 2, molecule possessing ankyrin repeats induced by LPS, and TNF-alpha-induced protein 3) were significantly increased in BAL samples from subjects with CR asthma (P < .05). These patients had significantly higher amounts (1444.0 +/- 457.3 pg/mg total protein) of LPS in BAL fluid than seen in subjects with CS asthma (270.5 +/- 216.0 pg, P < .05), as detected by using the LAL assay and confirmed by means of gas chromatographic/mass spectrometric analysis. Prolonged exposure to LPS induced functional steroid resistance to dexamethasone in normal human monocytes, as demonstrated by persistently increased IL-6 levels in the presence of dexamethasone. CONCLUSIONS: Classical macrophage activation and induction of LPS signaling pathways along with high endotoxin levels detected in BAL fluid from subjects with CR asthma suggest that LPS exposure might contribute to CR asthma.


Assuntos
Asma/imunologia , Citocinas/metabolismo , Glucocorticoides/uso terapêutico , Lipopolissacarídeos/imunologia , Ativação de Macrófagos , Prednisona/uso terapêutico , Asma/tratamento farmacológico , Asma/genética , Líquido da Lavagem Broncoalveolar/citologia , Líquido da Lavagem Broncoalveolar/imunologia , Células Cultivadas , Citocinas/genética , Citocinas/imunologia , Dexametasona/farmacologia , Resistência a Medicamentos/genética , Perfilação da Expressão Gênica , Humanos , Interleucina-6/biossíntese , Lipopolissacarídeos/análise , Macrófagos/imunologia , Macrófagos/metabolismo , Monócitos/efeitos dos fármacos , Monócitos/imunologia , Monócitos/metabolismo , Análise de Sequência com Séries de Oligonucleotídeos
7.
Pediatr Pulmonol ; 43(9): 916-23, 2008 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-18668688

RESUMO

Asthma is a leading chronic childhood illness in the US. To gain further insight into the pathophysiology of childhood asthma, we studied markers of airway inflammation and possible triggers such as bacterial lipopolysaccharide (LPS) in 18 children with chronic asthma and persistent wheezing who underwent clinically indicated bronchoscopy and bronchoalveolar lavage (BAL). We predominantly found neutrophilic airway inflammation associated with increased levels of IL-8, metalloproteinase (MMP)-9, tissue inhibitor of metalloproteinase (TIMP-1) and MMP-9/TIMP-1 ratio. A significant correlation was found between levels of LPS in BAL and airway neutrophils in BAL from a subgroup of children who had a tendency of increased levels of MMP-9 and TIMP-1, suggesting that increased LPS levels in BAL may contribute to chronic airway inflammation and early remodeling. Our data highlight the importance of defining chronic triggers of early airway inflammation in children and characterizing their inflammation, considering the use of bronchoscopy and BAL. Increased knowledge of airway inflammation in children may help prevent a more severe asthma phenotype and lead to environmental control measures and new treatment strategies to intervene against the establishment of irreversible inflammation.


Assuntos
Asma/imunologia , Neutrófilos/fisiologia , Polissacarídeos Bacterianos/imunologia , Sons Respiratórios/imunologia , Adolescente , Líquido da Lavagem Broncoalveolar/química , Criança , Pré-Escolar , Feminino , Humanos , Lactente , Masculino , Polissacarídeos Bacterianos/análise
8.
Curr Allergy Asthma Rep ; 8(3): 188-94, 2008 May.
Artigo em Inglês | MEDLINE | ID: mdl-18589836

RESUMO

Atopic dermatitis (AD) is a chronic, pruritic, inflammatory skin disease affecting more than 10% of all children. Sensitization to foods triggers isolated skin symptoms in about 30% of children. These symptoms include immediate reactions within minutes after ingesting food without exacerbation of AD and early and late exacerbations of AD. It is important to identify clinically relevant sensitizations to foods using skin prick tests, a specific IgE blood test (ImmunoCAP; Phadia, Portage, MI, USA), and double-blind, placebo-controlled food challenges to initiate appropriate dietary interventions and avoid unnecessary dietary restrictions. Children with AD triggered by food allergens demonstrate a distinct immune response upon stimulation of their peripheral blood mononuclear cells with food allergen. A defective skin barrier and increased intestinal permeability appear to facilitate allergen sensitization. Appropriate skin care to maintain skin barrier function and dietary avoidance of highly allergenic foods during infancy may help to prevent allergen sensitization, thereby reducing the severity of AD and food allergies.


Assuntos
Dermatite Atópica/etiologia , Hipersensibilidade Alimentar/complicações , Dermatite Atópica/diagnóstico , Dermatite Atópica/terapia , Hipersensibilidade Alimentar/epidemiologia , Humanos , Interleucina-10/genética , Polimorfismo de Nucleotídeo Único , Prevalência , Fatores de Risco
9.
J Allergy Clin Immunol ; 120(5): 1065-72, 2007 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-17900681

RESUMO

BACKGROUND: Steroid-resistant (SR) asthma is characterized by airway inflammation that fails to resolve despite treatment with corticosteroids, raising concerns that resistance to steroid therapy in asthma could lead to airway remodeling. OBJECTIVE: We sought to determine whether SR asthma is accompanied by decreased airflow reversibility and could lead to airway remodeling. METHODS: Spirometric results were evaluated for 40 asthmatic patients defined as having SR or steroid-sensitive (SS) asthma on the basis of a 1-week course of oral prednisone. Twenty-three asthmatic patients underwent bronchoscopy with collection of bronchoalveolar lavage (BAL) fluid to analyze markers of airway remodeling in BAL fluid and cells. RESULTS: Prednisone significantly improved FEV(1) percent predicted in SS asthma (62.0% +/- 10.9% [mean +/- SD] to 79.4% +/- 11.3%, P < .001) but not in SR asthma (66.9% +/- 10.0% to 65.9% +/- 12.1%). The bronchodilator response was significantly greater in the SS than in the SR group (Delta FEV(1) percent predicted, 33.5% +/- 22.5% vs 15.2% +/- 7.9%; P = .001), regardless of inhaled corticosteroid use. No difference in amounts of matrix metalloproteinase (MMP) 9, PMN elastase, or vascular endothelial growth factor was found in BAL fluid from both groups. Tissue inhibitor of metalloproteinases (TIMP) 1 levels were, however, significantly less in BAL fluid of patients with SR asthma compared with those in patients with SS asthma (921.9 +/- 313.4 vs 2267.0 +/- 456.8 pg/mL, P < .05), resulting in significantly higher MMP-9/TIMP-1 ratios in the BAL fluid of patients with SR asthma (0.24 +/- 0.04 vs 0.11 +/- 0.03, P < .01). Finally, dexamethasone treatment induced TIMP-1 mRNA in BAL fluid cells from patients with SS asthma (P < .01) but not in cells from patients with SR asthma. CONCLUSION: Bronchodilator reversibility is impaired in SR asthma and is associated with a shift in MMP-9/TIMP-1 ratio caused by inability of steroids to enhance TIMP-1 production, potentially promoting proteolytic activity in airways of patients with SR asthma and contributing to chronic airway remodeling. CLINICAL IMPLICATIONS: SR asthma might lead to irreversible airways disease.


Assuntos
Corticosteroides/uso terapêutico , Asma/tratamento farmacológico , Asma/patologia , Brônquios/patologia , Broncodilatadores/uso terapêutico , Resistência a Medicamentos , Adulto , Líquido da Lavagem Broncoalveolar/química , Dexametasona/uso terapêutico , Feminino , Humanos , Elastase de Leucócito/análise , Elastase de Leucócito/metabolismo , Masculino , Metaloproteinase 9 da Matriz/análise , Metaloproteinase 9 da Matriz/metabolismo , Pessoa de Meia-Idade , RNA Mensageiro/análise , RNA Mensageiro/metabolismo , Espirometria , Inibidor Tecidual de Metaloproteinase-1/análise , Inibidor Tecidual de Metaloproteinase-1/genética , Inibidor Tecidual de Metaloproteinase-1/metabolismo , Fator A de Crescimento do Endotélio Vascular/análise , Fator A de Crescimento do Endotélio Vascular/metabolismo
10.
Am J Respir Cell Mol Biol ; 27(3): 361-7, 2002 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-12204899

RESUMO

Glucocorticoid (GC) insensitivity is a challenging clinical problem associated with many chronic inflammatory disorders and life-threatening disease progression. The molecular basis of GC insensitivity, however, is unknown. Alternative splicing of the GC receptor (GCR) pre-mRNA generates a second GCR, termed GCRbeta, which does not bind GC but antagonizes the transactivating activity of the classic GCR, termed GCRalpha. GC-insensitive conditions have been associated with increased GCRbeta expression. Whether or not increased GCRbeta expression can contribute to GC insensitivity, however, remains controversial. To more precisely demonstrate the effect of GCRbeta on steroid responsiveness, we virally transduced GCRbeta cDNA into mouse DO-11.10 hybridoma cells, as mice are known to be deficient in the GCRbeta gene. We demonstrate that viral transduction of GCRbeta cDNA into mouse hybridoma cells to induce stable expression of GCRbeta results in GC insensitivity of these cells. Furthermore, in such cells GCRalpha is complexed with GCRbeta. Such heterodimer formation may account for the reduced effectiveness of GC action in cells overexpressing GCRbeta.


Assuntos
Corticosteroides/farmacologia , Hibridomas/efeitos dos fármacos , Receptores de Glucocorticoides/metabolismo , Processamento Alternativo , Animais , Western Blotting , Células Cultivadas , Dexametasona/farmacologia , Dimerização , Resistência a Medicamentos , Proteínas de Fluorescência Verde , Humanos , Hibridomas/metabolismo , Proteínas Luminescentes/genética , Proteínas Luminescentes/metabolismo , Camundongos , Receptores de Glucocorticoides/análise , Receptores de Glucocorticoides/genética , Transdução Genética
11.
Pediatr Allergy Immunol ; 13(1): 18-23, 2002 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-12000494

RESUMO

Venom immunotherapy (VIT) is able to protect insect venom-allergic patients against life-threatening sting reactions. Standardized sting challenges can be used as a diagnostic tool to check whether VIT is required. No data are available on the long-term predictive value of sting challenge tests. The purpose of this study was to investigate the long-term predictive value of sequential bee-sting challenges with respect to the ability to predict future sting reactions in bee-venom (BV) allergic children. Between 1988 and 1992, 92 BV-allergic children had been challenged with sequential bee stings at intervals of 2-6 weeks to determine the necessity of VIT. In 1996, all 92 families were followed-up using standardized telephone interviews. Until the follow-up, 61 children (66.3%) had experienced at least one natural bee sting. Based on the results of the initial challenge tests, 13 of the 61 patients had been started on VIT. Two of these 13 (15.4%) developed systemic reactions 1 year after VIT of 5 years, of which one was mild and one was severe. Among the 48 re-stung patients who were not treated with VIT, three children (6.3%) experienced mild systemic reactions, whereas 45 children reported no more than a local reaction. The long-term predictive value of sequential bee-sting challenge tests for systemic reactions in children not treated with VIT remained at a level of 93.8% (95% confidence interval: 82.8-98.7%) even over a period of more than 6 years. Based on this data, we conclude that sequential bee-sting challenges are a powerful tool to determine the necessity for VIT in BV-allergic children.


Assuntos
Venenos de Abelha/efeitos adversos , Abelhas , Adolescente , Animais , Anticorpos Anti-Idiotípicos/sangue , Anticorpos Anti-Idiotípicos/imunologia , Especificidade de Anticorpos , Venenos de Abelha/imunologia , Criança , Pré-Escolar , Dessensibilização Imunológica , Feminino , Humanos , Mordeduras e Picadas de Insetos/imunologia , Mordeduras e Picadas de Insetos/terapia , Masculino , Valor Preditivo dos Testes , Testes Cutâneos
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