RÉSUMÉ
BACKGROUND: Macrophages are the major resident immune cells in human airways coordinating responses to infection and injury. In cystic fibrosis (CF), neutrophils are recruited to the airways shortly after birth, and actively exocytose damaging enzymes prior to chronic infection, suggesting a potential defect in macrophage immunomodulatory function. Signaling through the exhaustion marker programmed death protein 1 (PD-1) controls macrophage function in cancer, sepsis, and airway infection. Therefore, we sought to identify potential associations between macrophage PD-1 and markers of airway disease in children with CF. METHODS: Blood and bronchoalveolar lavage fluid (BALF) were collected from 45 children with CF aged 3 to 62 months and structural lung damage was quantified by computed tomography. The phenotype of airway leukocytes was assessed by flow cytometry, while the release of enzymes and immunomodulatory mediators by molecular assays. RESULTS: Airway macrophage PD-1 expression correlated positively with structural lung damage, neutrophilic inflammation, and infection. Interestingly, even in the absence of detectable infection, macrophage PD-1 expression was elevated and correlated with neutrophilic inflammation. In an in vitro model mimicking leukocyte recruitment into CF airways, soluble mediators derived from recruited neutrophils directly induced PD-1 expression on recruited monocytes/macrophages, suggesting a causal link between neutrophilic inflammation and macrophage PD-1 expression in CF. Finally, blockade of PD-1 in a short-term culture of CF BALF leukocytes resulted in improved pathogen clearance. CONCLUSION: Taken together, these findings suggest that in early CF lung disease, PD-1 upregulation associates with airway macrophage exhaustion, neutrophil takeover, infection, and structural damage.
Sujet(s)
Mucoviscidose , Enfant , Humains , Récepteur-1 de mort cellulaire programmée , Poumon , Inflammation , Bactéries/métabolisme , Marqueurs biologiques/métabolisme , MacrophagesRÉSUMÉ
BACKGROUND: In this pilot study, we investigated whether induced sputum (IS) could serve as a viable alternative to bronchoalveolar lavage (BAL) and yield robust inflammatory biomarkers in toddlers with cystic fibrosis (CF) featuring minimal structural lung disease. METHODS: We collected IS, BAL (right middle lobe and lingula), and blood, and performed chest computed tomography (CT) scans from 2-year-olds with CF (N = 11), all within a single visit. Inflammatory biomarkers included 20 soluble immune mediators and neutrophil elastase (NE), as well as frequency and phenotype of T cells, monocytes/macrophages, and neutrophils. RESULTS: At the molecular level, nine mediators showed similar levels in IS and BAL (CXCL1, CXCL8, IL-1α, IL-1RA, IL-6, CCL2, CXCL10, M-CSF, VEGF-A), four were higher in IS than in BAL (CXCL5, IL-1ß, CXCL11, TNFSF10), and two were present in IS, but undetectable in BAL (IL-10, IFN-γ). Meanwhile, soluble NE had lower activity in IS than in BAL. At the cellular level, T-cell frequency was lower in IS than in BAL. Monocytes/macrophages were dominant in IS and BAL with similar frequencies, but differing expression of CD16 (lower in IS), CD115, and surface-associated NE (higher in IS). Neutrophil frequency and phenotype did not differ between IS and BAL. Finally, neutrophil frequency in IS correlated positively with air trapping. CONCLUSIONS: IS collected from 2-year-olds with CF yields biomarkers of early airway inflammation with good agreement with BAL, notably with regard to molecular and cellular outcomes related to neutrophils and monocytes/macrophages.
Sujet(s)
Mucoviscidose , Expectoration , Marqueurs biologiques , Lavage bronchoalvéolaire , Liquide de lavage bronchoalvéolaire , Humains , Granulocytes neutrophiles , Projets pilotesRÉSUMÉ
Cystic fibrosis (CF) airways feature high extracellular levels of the IL-1 family of proinflammatory mediators. These mediators are cleavage products of caspase-1, the final protease in the inflammasome cascade. Due to the proven chronic presence of reprogrammed neutrophils in the CF airway lumen, understanding inflammasome signaling in these cells is of great importance to understand how disease is perpetuated in this milieu. Here, we hypothesized that CF airway neutrophils contribute to chronic inflammation, in part, via the packaging of inflammasome-inducing signals in extracellular vesicles (EVs). We confirmed that CF airway fluid is enriched in IL-1α, IL-1ß, and IL-18, and that CF airway neutrophils up-regulate the activating receptor IL-1R1. Meanwhile, down-modulatory signals such as IL-1R2 and IL-1RA are unchanged. Active caspase-1 itself is present in CF airway fluid EVs, with neutrophil-derived EVs being most enriched. Using a transmigration model of CF airway inflammation, we show that CF airway fluid EVs are necessary and sufficient to induce primary granule exocytosis by naïve neutrophils (hallmark of reprogramming) and concomitantly activate caspase-1 and IL-1ß production by these cells and that the addition of triple-combination highly effective CFTR modulator therapy does not abrogate these effects. Finally, EVs from activated neutrophils can deliver active caspase-1 to primary tracheal epithelial cells and induce their release of IL-1α. These findings support the existence of a feed-forward inflammatory process by which reprogrammed CF airway neutrophils bypass 2-step control of inflammasome activation in neighboring cells (naïve neutrophils and epithelial cells) via the transfer of bioactive EVs.
Sujet(s)
Mucoviscidose , Vésicules extracellulaires , Caspases , Protéine CFTR , Humains , Inflammasomes , Inflammation , Antagoniste du récepteur à l'interleukine-1 , Interleukine-18 , Granulocytes neutrophiles , Peptide hydrolases , Récepteur à l'interleukine-1 de type IIRÉSUMÉ
Cystic fibrosis (CF) lung disease is dominated by the recruitment of myeloid cells (neutrophils and monocytes) from the blood which fail to clear the lung of colonizing microbes. In prior in vitro studies, we showed that blood neutrophils migrated through the well-differentiated lung epithelium into the CF airway fluid supernatant (ASN) mimic the dysfunction of CF airway neutrophils in vivo, including decreased bactericidal activity despite an increased metabolism. Here, we hypothesized that, in a similar manner to neutrophils, blood monocytes undergo significant adaptations upon recruitment to CFASN. To test this hypothesis, primary human blood monocytes were transmigrated in our in vitro model into the ASN from healthy control (HC) or CF subjects to mimic in vivo recruitment to normal or CF airways, respectively. Surface phenotype, metabolic and bacterial killing activities, and transcriptomic profile by RNA sequencing were quantified post-transmigration. Unlike neutrophils, monocytes were not metabolically activated, nor did they show broad differences in activation and scavenger receptor expression upon recruitment to the CFASN compared to HCASN. However, monocytes recruited to CFASN showed decreased bactericidal activity. RNASeq analysis showed strong effects of transmigration on monocyte RNA profile, with differences between CFASN and HCASN conditions, notably in immune signaling, including lower expression in the former of the antimicrobial factor ISG15, defensin-like chemokine CXCL11, and nitric oxide-producing enzyme NOS3. While monocytes undergo qualitatively different adaptations from those seen in neutrophils upon recruitment to the CF airway microenvironment, their bactericidal activity is also dysregulated, which could explain why they also fail to protect CF airways from infection.
Sujet(s)
Adaptation physiologique/génétique , Microenvironnement cellulaire/génétique , Mucoviscidose/génétique , Poumon/anatomopathologie , Monocytes/anatomopathologie , Transcription génétique/génétique , Adulte , Cellules cultivées , Femelle , Humains , Mâle , Granulocytes neutrophiles/anatomopathologie , Transduction du signal/physiologieRÉSUMÉ
Bullous pemphigoid (BP) is an autoimmune blistering disease characterized by recruitment of leucocytes into skin and release of damaging enzymes, resulting in epidermal detachment and blister formation. To better understand the role of leukotriene B4 (LTB4) and other inflammatory factors in BP pathophysiology, we conducted microscopic and immunohistochemical analyses of preserved skin biopsy sections and conducted flow cytometry and ELISA analyses of matched blood and blister fluid from BP patients. Neutrophils predominated in BP blister fluid, which also contained monocytes/macrophages and T cells, but few to no eosinophils and B cells. In contrast, BP skin histology showed a different pattern, with abundant neutrophils but eosinophils being the predominant immune cell type. LTB4 pathway and neutrophil activation markers were prevalent in BP skin lesions and strongly associated with perivascular neutrophils. Blister fluid neutrophils, monocytes/macrophages and eosinophils all exhibited increased surface expression of leukotriene A4 hydrolase and neutrophil elastase (P = .002 for both). Blister fluid was also enriched in interleukins (IL)-1α, IL-1ß, IL-8, IL-10, IL-18, monocyte colony-stimulating factor (M-CSF) and vascular endothelial growth factor (VEGF). Our findings suggest differential leucocyte recruitment from blood into dermis and from dermis into blister, which correlates with disease activity, and presents potential new treatment opportunities for BP.
Sujet(s)
Exsudats et transsudats/cytologie , Leucotriène B4/métabolisme , Pemphigoïde bulleuse/sang , Pemphigoïde bulleuse/anatomopathologie , Peau/anatomopathologie , Sujet âgé , Sujet âgé de 80 ans ou plus , Granulocytes éosinophiles , Epoxide hydrolase/métabolisme , Exsudats et transsudats/métabolisme , Femelle , Cytométrie en flux , Humains , Interleukines/métabolisme , Leukocyte elastase/métabolisme , Facteur de stimulation des colonies de macrophages/métabolisme , Macrophages/enzymologie , Mâle , Adulte d'âge moyen , Monocytes/enzymologie , Infiltration par les neutrophiles , Granulocytes neutrophiles/enzymologie , Pemphigoïde bulleuse/immunologie , Facteurs raciaux , Facteurs sexuels , Peau/métabolisme , Facteur de croissance endothéliale vasculaire de type A/métabolismeRÉSUMÉ
BACKGROUND/AIMS: The term idiopathic short stature (ISS) describes short stature of unknown, but likely polygenic, etiology. This study aimed to identify genetic polymorphisms associated with the ISS phenotype, and with growth response to supplemental GH. METHODS: Using a case-control analysis we compared the prevalence of "tall" versus "short" alleles at 52 polymorphic loci (17 in growth-related candidate genes, 35 identified in prior genome-wide association studies of adult height) in 94 children with ISS followed in the Genetics and Neuroendocrinology of Short Stature International Study, versus 143 controls from the Fels Longitudinal Study. RESULTS: Four variants were nominally associated with ISS using a genotypic model, confirmed by a simultaneous confident inference approach: compared with controls children with ISS had lower odds of "tall" alleles (odds ratio, 95% CI) for GHR (0.52, 0.29-0.96); rs2234693/ESR1 (0.50, 0.25-0.98); rs967417/BMP2 (0.39, 0.17-0.93), and rs4743034/ZNF462 (0.40, 0.18-0.89). Children with ISS also had lower odds of the "tall" allele (A) at the IGFBP3 -202 promoter polymorphism (rs2855744; 0.40, 0.20-0.80) in the simultaneous confident inference analysis. A significant association with 1st-year height SD score increase during GH treatment was observed with rs11205277, located near 4 known genes: MTMR11, SV2A, HIST2H2AA3, and SF3B4; the latter, in which heterozygous mutations occur in Nager acrofacial dysostosis, appears the most relevant gene. CONCLUSIONS: In children with ISS we identified associations with "short" alleles at a number of height-related loci. In addition, a polymorphic variant located near SF3B4 was associated with the GH treatment response in our cohort. The findings in our small study warrant further investigation.
Sujet(s)
Locus génétiques , Troubles de la croissance , Hormone de croissance humaine/administration et posologie , Polymorphisme génétique , Adolescent , Enfant , Femelle , Études de suivi , Étude d'association pangénomique , Troubles de la croissance/traitement médicamenteux , Troubles de la croissance/génétique , Troubles de la croissance/physiopathologie , Humains , Études longitudinales , MâleRÉSUMÉ
Neutrophils are recruited to the airways of patients with acute respiratory distress syndrome (ARDS) where they acquire an activated pro-survival phenotype with an enhanced respiratory burst thought to contribute to ARDS pathophysiology. Our in vitro model enables blood neutrophil transepithelial migration into cell-free tracheal aspirate fluid from patients to recapitulate the primary airway neutrophil phenotype observed in vivo. Neutrophils transmigrated through our model toward airway fluid from children with lower respiratory viral infections coinfected with bacteria had elevated levels of neutrophil activation markers but paradoxically exhibited an inability to kill bacteria and a defective respiratory burst compared with children without bacterial coinfection. The airway fluid from children with bacterial coinfections had higher levels of neutrophil elastase activity, as well as myeloperoxidase levels compared to children without bacterial coinfection. Neutrophils transmigrated into the aspirate fluid from children with bacterial coinfection showed decreased respiratory burst and killing activity against H. influenzae and S. aureus compared to those transmigrated into the aspirate fluid from children without bacterial coinfection. Use of a novel transmigration model recapitulates this pathological phenotype in vitro that would otherwise be impossible in a patient, opening avenues for future mechanistic and therapeutic research.
Sujet(s)
Infections bactériennes , Co-infection , Granulocytes neutrophiles , Stimulation du métabolisme oxydatif/immunologie , , Maladies virales , Adolescent , Infections bactériennes/immunologie , Infections bactériennes/microbiologie , Infections bactériennes/anatomopathologie , Infections bactériennes/virologie , Enfant , Enfant d'âge préscolaire , Co-infection/immunologie , Co-infection/microbiologie , Co-infection/anatomopathologie , Co-infection/virologie , Femelle , Haemophilus influenzae/immunologie , Humains , Nourrisson , Nouveau-né , Mâle , Granulocytes neutrophiles/immunologie , Granulocytes neutrophiles/anatomopathologie , /immunologie , /microbiologie , /anatomopathologie , /virologie , Infections de l'appareil respiratoire/immunologie , Infections de l'appareil respiratoire/microbiologie , Infections de l'appareil respiratoire/anatomopathologie , Infections de l'appareil respiratoire/virologie , Staphylococcus aureus/immunologie , Maladies virales/immunologie , Maladies virales/microbiologie , Maladies virales/anatomopathologie , Maladies virales/virologieRÉSUMÉ
BACKGROUND: Resistin is an immunometabolic mediator that is elevated in several inflammatory disorders. A ligand for Toll-like receptor 4, resistin modulates the recruitment and activation of myeloid cells, notably neutrophils. Neutrophils are major drivers of cystic fibrosis (CF) lung disease, in part due to the release of human neutrophil elastase- and myeloperoxidase-rich primary granules, leading to tissue damage. Here we assessed the relationship of resistin to CF lung disease. METHODS: Resistin levels were measured in plasma and sputum from three retrospective CF cohorts spanning a wide range of disease. We also assessed the ability of neutrophils to secrete resistin upon activation in vitro. Finally, we constructed a multivariate model assessing the relationship between resistin levels and lung function. RESULTS: Plasma resistin levels were only marginally higher in CF than in healthy control subjects. By contrast, sputum resistin levels were very high in CF, reaching 50-100 fold higher levels than in plasma. Among CF patients, higher plasma resistin levels were associated with allergic bronchopulmonary aspergillosis, and higher sputum resistin levels were associated with CF-related diabetes. Mechanistically, in vitro release of neutrophil primary granules was concomitant with resistin secretion. Overall, sputum resistin levels were negatively correlated with CF lung function, independently of other variables (age, sex, and genotype). CONCLUSIONS: Our data establish relationships between resistin levels in the plasma and sputum of CF patients that correlate with disease status, and identify resistin as a novel mechanistic link between neutrophilic inflammation and lung disease in CF.
Sujet(s)
Mucoviscidose/métabolisme , Débits expiratoires forcés/physiologie , Résistine/métabolisme , Expectoration/métabolisme , Adolescent , Adulte , Marqueurs biologiques/métabolisme , Mucoviscidose/diagnostic , Mucoviscidose/physiopathologie , Évolution de la maladie , Femelle , Études de suivi , Humains , Poumon/physiopathologie , Mâle , Adulte d'âge moyen , Pronostic , Tests de la fonction respiratoire , Études rétrospectives , Jeune adulteRÉSUMÉ
RATIONALE: Neutrophils are recruited to the airways of individuals with cystic fibrosis (CF). In adolescents and adults with CF, airway neutrophils actively exocytose the primary granule protease elastase (NE), whose extracellular activity correlates with lung damage. During childhood, free extracellular NE activity is measurable only in a subset of patients, and the exocytic function of airway neutrophils is unknown. OBJECTIVES: To measure NE exocytosis by airway neutrophils in relation to free extracellular NE activity and lung damage in children with CF. METHODS: We measured lung damage using chest computed tomography coupled with the Perth-Rotterdam Annotated Grid Morphometric Analysis for Cystic Fibrosis scoring system. Concomitantly, we phenotyped blood and BAL fluid leukocytes by flow and image cytometry, and measured free extracellular NE activity using spectrophotometric and Förster resonance energy transfer assays. Children with airway inflammation linked to aerodigestive disorder were enrolled as control subjects. MEASUREMENTS AND MAIN RESULTS: Children with CF but not disease control children harbored BAL fluid neutrophils with high exocytosis of primary granules, before the detection of bronchiectasis. This measure of NE exocytosis correlated with lung damage (R = 0.55; P = 0.0008), whereas the molecular measure of free extracellular NE activity did not. This discrepancy may be caused by the inhibition of extracellular NE by BAL fluid antiproteases and its binding to leukocytes. CONCLUSIONS: NE exocytosis by airway neutrophils occurs in all children with CF, and its cellular measure correlates with early lung damage. These findings implicate live airway neutrophils in early CF pathogenesis, which should instruct biomarker development and antiinflammatory therapy in children with CF.
Sujet(s)
Mucoviscidose/physiopathologie , Exocytose/physiologie , Lésion pulmonaire/physiopathologie , Granulocytes neutrophiles/métabolisme , Pancreatic elastase/métabolisme , Enfant d'âge préscolaire , Femelle , Humains , Nourrisson , MâleRÉSUMÉ
BACKGROUND: Airway neutrophils are abundant in some children with severe asthma, but their functions are poorly understood. OBJECTIVE: To characterize that the inflammatory airway environment of children with neutrophil-predominant severe asthma promotes neutrophil survival and disrupts neutrophil-associated innate immune defenses. METHODS: Sixty-seven children with severe asthma refractory to high-dose inhaled corticosteroid treatment undergoing bronchoscopy with bronchoalveolar lavage (BAL) for clinical indications were stratified into neutrophil "high" versus "low" groups on the basis of BAL differential counts. Neutrophil activation markers, functional assays, and phenotyping studies were performed, as well as airway macrophage functional assays. Results were compared with those from children with moderate asthma treated with inhaled corticosteroids. RESULTS: Children with neutrophil-predominant severe asthma had increased markers of neutrophil activation/degranulation and a greater magnitude of airway proinflammatory cytokine and chemokine release. Primary neutrophils exposed to BAL of these children exhibited greater phagocytic capability and greater neutrophil extracellular trap formation, but a more impaired respiratory burst. Despite greater abundance of airway TGF-ß1, the neutrophils were not more apoptotic. Instead, neutrophils had a highly proinflammatory phenotype associated with a number of surface markers that regulate neutrophil activation, recruitment/migration, and granule release. Airway macrophages from children with neutrophil-predominant severe asthma were also more proinflammatory with impaired phagocytosis and increased apoptosis. CONCLUSIONS: Children with neutrophil-predominant severe asthma have proinflammatory neutrophils with enhanced survival. Airway macrophages are also proinflammatory and dysfunctional and may contribute to global innate immune impairment. Therapies that target neutrophils and related inflammation may be warranted in this subset of children.
Sujet(s)
Asthme/immunologie , Granulocytes neutrophiles/immunologie , Adolescent , Hormones corticosurrénaliennes/usage thérapeutique , Adulte , Asthme/traitement médicamenteux , Liquide de lavage bronchoalvéolaire/cytologie , Liquide de lavage bronchoalvéolaire/immunologie , Enfant , Cytokines/immunologie , Résistance aux substances , Femelle , Cellules HL-60 , Humains , Macrophages/immunologie , Mâle , Phagocytose , Phénotype , Jeune adulteRÉSUMÉ
Recruitment of neutrophils to the airways, and their pathological conditioning therein, drive tissue damage and coincide with the loss of lung function in patients with cystic fibrosis (CF). So far, these key processes have not been adequately recapitulated in models, hampering drug development. Here, we hypothesized that the migration of naïve blood neutrophils into CF airway fluid in vitro would induce similar functional adaptation to that observed in vivo, and provide a model to identify new therapies. We used multiple platforms (flow cytometry, bacteria-killing, and metabolic assays) to characterize functional properties of blood neutrophils recruited in a transepithelial migration model using airway milieu from CF subjects as an apical chemoattractant. Similarly to neutrophils recruited to CF airways in vivo, neutrophils migrated into CF airway milieu in vitro display depressed phagocytic receptor expression and bacterial killing, but enhanced granule release, immunoregulatory function (arginase-1 activation), and metabolic activities, including high Glut1 expression, glycolysis, and oxidant production. We also identify enhanced pinocytic activity as a novel feature of these cells. In vitro treatment with the leukotriene pathway inhibitor acebilustat reduces the number of transmigrating neutrophils, while the metabolic modulator metformin decreases metabolism and oxidant production, but fails to restore bacterial killing. Interestingly, we describe similar pathological conditioning of neutrophils in other inflammatory airway diseases. We successfully tested the hypothesis that recruitment of neutrophils into airway milieu from patients with CF in vitro induces similar pathological conditioning to that observed in vivo, opening new avenues for targeted therapeutic intervention.
Sujet(s)
Mucoviscidose/immunologie , Granulocytes neutrophiles/immunologie , Animaux , Composés azabicycliques/pharmacologie , Benzoates/pharmacologie , Cellules sanguines , Cellules de la moelle osseuse , Cellules cultivées , Chimiotaxie des leucocytes , Milieux de culture conditionnés/pharmacologie , Mucoviscidose/anatomopathologie , Exocytose/effets des médicaments et des substances chimiques , Cytométrie en flux , Glycolyse , Humains , Leukocyte elastase/métabolisme , Leucotriène B4/pharmacologie , Lipopolysaccharides/pharmacologie , Metformine/pharmacologie , Souris , Activation des neutrophiles , Granulocytes neutrophiles/effets des médicaments et des substances chimiques , Granulocytes neutrophiles/métabolisme , Granulocytes neutrophiles/anatomopathologie , Consommation d'oxygène , Pinocytose , Pseudomonas aeruginosa , Appareil respiratoire/immunologie , Appareil respiratoire/anatomopathologie , Expectoration/immunologie , Migration transendothéliale et transépithéliale/effets des médicaments et des substances chimiquesRÉSUMÉ
PURPOSE: Variants in the gene encoding Programmed Cell Death-1 (PDCD1) have been associated with susceptibility to Systemic Lupus Erythematosus and other autoimmune diseases. Given that clinically distinct autoimmune phenotypes share common genetic susceptibility factors, variants in PDCD-1 were tested for a possible association with Juvenile Idiopathic Arthritis (JIA). METHODS: Four Single Nucleotide Polymorphisms (SNPS) in the PDCD1 gene were genotyped and analyzed: rs7421861, rs11568821, rs10204525, and rs7568402 in 834 cases and 855 controls of Northern European ancestry. Each variant was examined for possible associations with JIA and then analyzed for association with JIA categories. RESULTS: PDCD1 variants showed no association with JIA in the cohort overall (rs7421861 p=0.63, rs11568821 p=0.13, rs10204525 p=0.31, and rs7568402 p=0.45). Stratification by JIA categories indicated a significant association between systemic JIA and PDCD1 rs7568402 (OR=0.53, p=0.0027), which remained significant after 10,000 permutations, but was not replicated in an independent multi-ethnic systemic JIA cohort. A nominal association between enthesitis-related arthritis and rs115668821 was also observed (OR=0.22, p=0.012). CONCLUSION: Unlike other multiple autoimmune disease associated genetic variants, there was no association between PDCD1 variants and JIA or JIA categories.
Sujet(s)
Arthrite juvénile/génétique , Prédisposition génétique à une maladie/génétique , Récepteur-1 de mort cellulaire programmée/génétique , Maladies auto-immunes/génétique , Auto-immunité/génétique , Études cas-témoins , Enfant , Femelle , Génotype , Humains , Mâle , Polymorphisme de nucléotide simpleRÉSUMÉ
BACKGROUND: Severe asthma in children is a heterogeneous disorder associated with variable responses to corticosteroid treatment. Criterion standards for corticosteroid responsiveness assessment in children are lacking. OBJECTIVE: This study sought to characterize systemic corticosteroid responses in children with severe asthma after treatment with intramuscular triamcinolone and to identify phenotypic and molecular predictors of an intramuscular triamcinolone response. METHODS: Asthma-related quality of life, exhaled nitric oxide, blood eosinophils, lung function, and inflammatory cytokine and chemokine mRNA gene expression in peripheral blood mononuclear cells were assessed in 56 children with severe asthma at baseline and 14 days after intramuscular triamcinolone injection. The Asthma Control Questionnaire was used to classify children with severe asthma into corticosteroid response groups. RESULTS: Three groups of children with severe asthma were identified: controlled severe asthma, children who achieved control after triamcinolone, and children who did not achieve control. At baseline, these groups were phenotypically similar. After triamcinolone, discordance between symptoms, lung function, exhaled nitric oxide, and blood eosinophils was noted. Clinical phenotypic predictors were of limited utility in predicting the triamcinolone response, whereas systemic mRNA expression of inflammatory cytokines and chemokines related to IL-2, IL-10, and TNF signaling pathways, namely, AIMP1, CCR2, IL10RB, and IL5, strongly differentiated children who failed to achieve control with triamcinolone administration. CONCLUSIONS: Systemic corticosteroid responsiveness in children with severe asthma is heterogeneous. Alternative prediction models that include molecular endotypic as well as clinical phenotypic features are needed to identify which children derive the most clinical benefit from systemic corticosteroid step-up therapy given the potential side effects.
Sujet(s)
Asthme/traitement médicamenteux , Cytokines/métabolisme , Granulocytes éosinophiles/anatomopathologie , Sous-unité bêta du récepteur à l'interleukine-10/métabolisme , Interleukine-5/métabolisme , Protéines tumorales/métabolisme , Protéines de liaison à l'ARN/métabolisme , Récepteurs CCR2/métabolisme , Triamcinolone/usage thérapeutique , Adolescent , Asthme/diagnostic , Biomarqueurs pharmacologiques/métabolisme , Enfant , Cytokines/génétique , Évolution de la maladie , Femelle , Humains , Sous-unité bêta du récepteur à l'interleukine-10/génétique , Interleukine-5/génétique , Mâle , Protéines tumorales/génétique , Monoxyde d'azote/métabolisme , ARN messager/analyse , Protéines de liaison à l'ARN/génétique , Récepteurs CCR2/génétique , Tests de la fonction respiratoireRÉSUMÉ
Two variants (c.[301_302delAG];[301_302delAG] and c.[150delA];[150delA]) in the PROP1 gene are the most common genetic causes of recessively inherited combined pituitary hormones deficiency (CPHD). Our objective was to analyze in detail the origin of the two most prevalent variants. In the multicentric study were included 237 patients with CPHD and their 15 relatives carrying c.[301_302delAG];[301_302delAG] or c.[150delA];[150delA] or c.[301_302delAG];[ 150delA]. They originated from 21 different countries worldwide. We genotyped 21 single-nucleotide variant markers flanking the 9.6-Mb region around the PROP1 gene that are not in mutual linkage disequilibrium in the general populations--a finding of a common haplotype would be indicative of ancestral origin of the variant. Haplotypes were reconstructed by Phase and Haploview software, and the variant age was estimated using an allelic association method. We demonstrated the ancestral origin of both variants--c.[301_302delAG] was carried on 0.2 Mb-long haplotype in a majority of European patients arising ~101 generations ago (confidence interval 90.1-116.4). Patients from the Iberian Peninsula displayed a different haplotype, which was estimated to have emerged 23.3 (20.1-29.1) generations ago. Subsequently, the data indicated that both the haplotypes were transmitted to Latin American patients ~13.8 (12.2-17.0) and 16.4 (14.4-20.1) generations ago, respectively. The c.[150delA] variant that was carried on a haplotype spanning about 0.3 Mb was estimated to appear 43.7 (38.4-52.7) generations ago. We present strong evidence that the most frequent variants in the PROP1 gene are not a consequence of variant hot spots as previously assumed, but are founder variants.
Sujet(s)
Prédisposition génétique à une maladie , Haplotypes/génétique , Protéines à homéodomaine/génétique , Hypopituitarisme/génétique , Mutation/génétique , Humains , Prévalence , LogicielRÉSUMÉ
Gonadal hypofunction is described in male and female patients with sickle cell anemia (SCA) after bone marrow transplant (BMT) and in males treated with hydroxyurea (HU). Anti-Müllerian hormone (AMH) is a serum marker of ovarian reserve. This study describes AMH and follicle-stimulating hormone (FSH) levels in female SCA subjects treated with supportive care (SCA-SC), HU (SCA-HU) and BMT (SCA-BMT). SCA (SS/Sß(0)) subjects not on HU, on HU and status-post BMT, ages 10-21 years were recruited. SCA-HU subjects were treated with HU ≥ 20 mg/kg for ≥ 12 consecutive months. SCA-BMT subjects had received busulfan and cyclophosphamide. Serum AMH and random FSH levels were obtained. Diminished ovarian reserve (DOR) was defined as AMH level <5th percentile for age-matched controls. Subjects also with FSH >40 IU/L were classified as having premature ovarian insufficiency (POI). 14 SCA-SC (14.5 ± 2.7 years), 33 SCA-HU (14.4 ± 2.4 years) and 9 SCA-BMT (14.3 ± 2.7 years) females were included. AMH was undetectable in all SCA-BMT subjects and <5th percentile in 24% of SCA-HU subjects. FSH was menopausal (>40 IU/L) in 88.9% of SCA-BMT subjects. All SCA-BMT subjects and 24% of subjects on HU had DOR; 89% of SCA-BMT subjects had POI. AMH and FSH may be useful tools in assessing ovarian reserve and function.
Sujet(s)
Drépanocytose/thérapie , Hormone antimullérienne/sang , Antidrépanocytaires/usage thérapeutique , Transplantation de moelle osseuse , Hydroxy-urée/usage thérapeutique , Insuffisance ovarienne primitive/thérapie , Adolescent , Drépanocytose/sang , Drépanocytose/complications , Drépanocytose/diagnostic , Marqueurs biologiques/sang , Busulfan/usage thérapeutique , Études cas-témoins , Enfant , Cyclophosphamide/usage thérapeutique , Femelle , Hormone folliculostimulante/sang , Hémoglobine S/métabolisme , Hétérozygote , Homozygote , Humains , Ménarche/physiologie , Agonistes myélo-ablatifs/usage thérapeutique , Réserve ovarienne/effets des médicaments et des substances chimiques , Insuffisance ovarienne primitive/sang , Insuffisance ovarienne primitive/complications , Insuffisance ovarienne primitive/diagnostic , Jeune adulteRÉSUMÉ
Bacteria colonize cystic fibrosis (CF) airways, and although T cells with appropriate Ag specificity are present in draining lymph nodes, they are conspicuously absent from the lumen. To account for this absence, we hypothesized that polymorphonuclear neutrophils (PMNs), recruited massively into the CF airway lumen and actively exocytosing primary granules, also suppress T cell function therein. Programmed death-ligand 1 (PD-L1), which exerts T cell suppression at a late step, was expressed bimodally on CF airway PMNs, delineating PD-L1(hi) and PD-L1(lo) subsets, whereas healthy control (HC) airway PMNs were uniformly PD-L1(hi). Blood PMNs incubated in CF airway fluid lost PD-L1 over time; in coculture, Ab blockade of PD-L1 failed to inhibit the suppression of T cell proliferation by CF airway PMNs. In contrast with PD-L1, arginase 1 (Arg1), which exerts T cell suppression at an early step, was uniformly high on CF and HC airway PMNs. However, arginase activity was high in CF airway fluid and minimal in HC airway fluid, consistent with the fact that Arg1 activation requires primary granule exocytosis, which occurs in CF, but not HC, airway PMNs. In addition, Arg1 expression on CF airway PMNs correlated negatively with lung function and positively with arginase activity in CF airway fluid. Finally, combined treatment with arginase inhibitor and arginine rescued the suppression of T cell proliferation by CF airway fluid. Thus, Arg1 and PD-L1 are dynamically modulated upon PMN migration into human airways, and, Arg1, but not PD-L1, contributes to early PMN-driven T cell suppression in CF, likely hampering resolution of infection and inflammation.
Sujet(s)
Arginase/immunologie , Antigène CD274/immunologie , Mucoviscidose/immunologie , Granulocytes neutrophiles/immunologie , Lymphocytes T/immunologie , Adulte , Apoptose/immunologie , Arginase/biosynthèse , Antigène CD274/antagonistes et inhibiteurs , Prolifération cellulaire , Exocytose/immunologie , Femelle , Humains , Inflammation/immunologie , Inflammation/anatomopathologie , Poumon/immunologie , Poumon/anatomopathologie , Activation des lymphocytes/immunologie , Mâle , Tests de la fonction respiratoire , Muqueuse respiratoire/cytologie , Muqueuse respiratoire/immunologie , Jeune adulteRÉSUMÉ
BACKGROUND: The mechanisms underlying glucocorticoid responsiveness are largely unknown. Although redox regulation of the glucocorticoid receptor (GR) has been reported, it has not been studied in asthmatic patients. OBJECTIVE: We characterized systemic cysteine oxidation and its association with inflammatory and clinical features in healthy children and children with difficult-to-treat asthma. We hypothesized that cysteine oxidation would be associated with increased markers of oxidative stress and inflammation, increased features of asthma severity, decreased clinically defined glucocorticoid responsiveness, and impaired GR function. METHODS: PBMCs were collected from healthy children (n = 16) and children with asthma (n = 118) aged 6 to 17 years. Children with difficult-to-treat asthma underwent glucocorticoid responsiveness testing with intramuscular triamcinolone. Cysteine, cystine, and inflammatory chemokines and reactive oxygen species generation were quantified, and expression and activity of the GR were assessed. RESULTS: Cysteine oxidation was present in children with difficult-to-treat asthma and accompanied by increased reactive oxygen species generation and increased CCL3 and CXCL1 mRNA expression. Children with the greatest extent of cysteine oxidation had more features of asthma severity, including poorer symptom control, greater medication use, and less glucocorticoid responsiveness despite inhaled glucocorticoid therapy. Cysteine oxidation also modified the GR protein by decreasing available sulfhydryl groups and decreasing nuclear GR expression and activity. CONCLUSIONS: A highly oxidized cysteine redox state promotes a posttranslational modification of the GR that might inhibit its function. Given that cysteine oxidation is prevalent in children with difficult-to-treat asthma, the cysteine redox state might represent a potential therapeutic target for restoration of glucocorticoid responsiveness in this population.
Sujet(s)
Asthme/traitement médicamenteux , Glucocorticoïdes/usage thérapeutique , Agranulocytes/immunologie , Maturation post-traductionnelle des protéines , Récepteurs aux glucocorticoïdes/immunologie , Triamcinolone/usage thérapeutique , Administration par inhalation , Adolescent , Asthme/génétique , Asthme/immunologie , Asthme/anatomopathologie , Chimiokine CCL3/génétique , Chimiokine CCL3/immunologie , Chimiokine CXCL1/génétique , Chimiokine CXCL1/immunologie , Enfant , Cystéine/composition chimique , Cystéine/immunologie , Cystine/composition chimique , Cystine/immunologie , Surveillance des médicaments , Femelle , Expression des gènes , Humains , Injections musculaires , Agranulocytes/composition chimique , Agranulocytes/effets des médicaments et des substances chimiques , Agranulocytes/anatomopathologie , Mâle , Oxydoréduction , Stress oxydatif , Culture de cellules primaires , ARN messager/génétique , ARN messager/immunologie , Récepteurs aux glucocorticoïdes/composition chimique , Récepteurs aux glucocorticoïdes/génétiqueRÉSUMÉ
BACKGROUND: Anti-Mullerian Hormone (AMH), a proposed indicator of ovarian follicle reserve in adults, has not been characterized in pediatric and adolescent females by race and/or ethnicity. OBJECTIVES: To describe AMH levels in healthy American girls and determine the influence of age and race/ethnicity on AMH. SUBJECTS: SUBJECTS aged 10-21 years were recruited from primary care settings and emergency departments. Race/ethnicity was characterized as White, Black or Hispanic. METHODS: Serum for AMH levels (ng/mL) was measured using an enzyme-linked immunosorbent assay. RESULTS: Thirty-one White, 60 Black and 24 Hispanic subjects were recruited. Mean AMH levels were 3.19 ng/mL (22.8 pmol/L) (SD 2.12) for Whites, 3.25 ng/mL (23.2 pmol/L) (SD 2.23) for Blacks and 2.97 ng/mL (21.2 pmol/L) (SD 1.75) for Hispanics. ANCOVA showed no difference in AMH levels among race/ethnicities, controlling for age (p=0.91). Age was significantly associated with AMH (p<0.001, R²=0.12). CONCLUSION: AMH levels do not vary by race/ethnicity, and AMH levels increase with age.
Sujet(s)
Hormone antimullérienne/sang , Ethnies/statistiques et données numériques , Réserve ovarienne/physiologie , /statistiques et données numériques , Adolescent , Adulte , Facteurs âges , Enfant , Femelle , Humains , États-Unis/épidémiologie , Jeune adulteRÉSUMÉ
BACKGROUND: Although more than 100 non-HLA variants have been tested for associations with juvenile idiopathic arthritis (JIA) in candidate gene studies, only a few have been replicated. We sought to replicate reported associations of single nucleotide polymorphisms (SNPs) in the PTPN22, TNFA and MIF genes in a well-characterized cohort of children with JIA. METHODS: We genotyped and analyzed 4 SNPs in 3 genes: PTPN22 C1858T (rs2476601), TNFA G-308A, G-238A (rs1800629, rs361525) and MIF G-173C (rs755622) in 647 JIA cases and 751 healthy controls. We tested for association between each variant and JIA as well as JIA subtypes. We adjusted for multiple testing using permutation procedures. We also performed a meta-analysis that combined our results with published results from JIA association studies. RESULTS: While the PTPN22 variant showed only modest association with JIA (OR = 1.29, p = 0.0309), it demonstrated a stronger association with the RF-positive polyarticular JIA subtype (OR = 2.12, p = 0.0041). The MIF variant was not associated with the JIA as a whole or with any subtype. The TNFA-238A variant was associated with JIA as a whole (OR 0.66, p = 0.0265), and demonstrated a stronger association with oligoarticular JIA (OR 0.33, p = 0.0006) that was significant after correction for multiple testing. TNFA-308A was not associated with JIA, but was nominally associated with systemic JIA (OR = 0.33, p = 0.0089) and enthesitis-related JIA (OR = 0.40, p = 0.0144). Meta-analyses confirmed significant associations between JIA and PTPN22 (OR 1.44, p <0.0001) and TNFA-238A (OR 0.69, p < 0.0086) variants. Subtype meta-analyses of the PTPN22 variant revealed associations between RF-positive, RF-negative, and oligoarticular JIA, that remained significant after multiple hypothesis correction (p < 0.0005, p = 0.0007, and p < 0.0005, respectively). CONCLUSIONS: We have confirmed associations between JIA and PTPN22 and TNFA G-308A. By performing subtype analyses, we discovered a statistically-significant association between the TNFA-238A variant and oligoarticular JIA. Our meta-analyses confirm the associations between TNFA-238A and JIA, and show that PTPN22 C1858T is associated with JIA as well as with RF-positive, RF-negative and oligoarticular JIA.
RÉSUMÉ
OBJECTIVE: Children with childhood-onset rheumatoid arthritis (RA) include those with rheumatoid factor or anti-citrullinated protein antibody-positive juvenile idiopathic arthritis. To test the hypothesis that adult-onset RA-associated variants are also associated with childhood-onset RA, we investigated RA-associated variants at 5 loci in a cohort of patients with childhood-onset RA. We also assessed the cumulative association of these variants in susceptibility to childhood-onset RA using a weighted genetic risk score (wGRS). METHODS: A total of 155 children with childhood-onset RA and 684 healthy controls were genotyped for 5 variants in the PTPN22, TRAF1/C5, STAT4, and TNFAIP3 loci. High-resolution HLA-DRB1 genotypes were available for 149 cases and 373 controls. We tested each locus for association with childhood-onset RA via logistic regression. We also computed a wGRS for each subject, with weights based on the natural log of the published odds ratios (ORs) for the alleles investigated, and used logistic regression to test the wGRS for association with childhood-onset RA. RESULTS: Childhood-onset RA was associated with TNFAIP3 rs10499194 (OR 0.60 [95% confidence interval 0.44-0.83]), PTPN22 rs2476601 (OR 1.61 [95% confidence interval 1.11-2.31]), and STAT4 rs7574865 (OR 1.41 [95% confidence interval 1.06-1.87]) variants. The wGRS was significantly different between cases and controls (P < 2 × 10(-16) ). Individuals in the third to fifth quintiles of wGRS had a significantly increased disease risk compared to baseline (individuals in the first quintile). Higher wGRS was associated with increased risk of childhood-onset RA, especially among males. CONCLUSION: The magnitude and direction of the association between TNFAIP3, STAT4, and PTPN22 variants and childhood-onset RA are similar to those observed in RA, suggesting that adult-onset RA and childhood-onset RA share common genetic risk factors. Using a wGRS, we have demonstrated the cumulative association of RA-associated variants with susceptibility to childhood-onset RA.