Benoit Wallaert

Increased secretion of tumor necrosis factor α and interleukin-6 by alveolar macrophages consecutive to the development of the late asthmatic reaction

The late asthmatic reaction (LAR), consecutive to bronchial allergen challenge, is characterized both by the influx of various cells in proximal and distal airways and by the enhancement of bronchial hyperresponsiveness. However, the exact conditions for the development of the inflammatory reaction during the LAR remain to be specified. Since monokines play a key role in inflammatory processes, particularly in the lung, the production of tumor necrosis factor-alpha (TNF-alpha), interleukin; 1-beta (IL-1-beta) and interleukin-6 (IL-6) by alveolar macrophages (AM), collected 18 to 20 hours after exposure to allergen, was evaluated in 15 allergic subjects with asthma submitted to a challenge test with Dermatophagoides pteronyssinus (N = 6) or with wheat flour (N = 9) and in three healthy subjects. After bronchial provocation test, four patients presented no bronchial response (group 1), and six patients, a single early reaction (group 2). In contrast, five patients developed successively an immediate plus a late response (group 3). The monokine production was compared to that from nine allergic subjects with asthma studied at baseline (group 0) and from 11 unchallenged healthy subjects (control subjects). Measurements of cytokines were evaluated for TNF-alpha and IL-1-beta by a specific immunoradiometric assay, whereas IL-6 levels were appreciated by the proliferation of 7TD1 cells. No detectable amounts of TNF-alpha, IL-1-beta, and IL-6 were in bronchial alveolar lavage fluid, even after a tenfold concentration. In contrast, a significant increase of TNF-alpha (10,642 +/- 3127 U/ml) and IL-6 (1250 +/- 427 U/ml) concentrations was noted in AM supernatants from patients exhibiting an LAR (group 3) compared to cells recovered from groups 2, 1, and 0 and to challenged or unchallenged control subjects (805 +/- 244, 995 +/- 521, 1269 +/- 524, 688 +/- 85, and 445 +/- 74 pg of TNF-alpha per milliliter, respectively; 190 +/- 64, 114 +/- 91, 242 +/- 95, 80 +/- 9, and 54 +/- 19 U/ml of IL-6 per milliliter, respectively). No modification of IL-1-beta contents could be detected between the different groups. A significant correlation was detected between concentrations of TNF and IL-6 (r = 0.92; p less than 0.001). These results demonstrate TNF-alpha and IL-6 secretion by AM consecutively to the development of LAR in allergic subjects with asthma, confirming that AMs are activated after allergen challenge.(ABSTRACT TRUNCATED AT 400 WORDS)

IL-9 and its receptor in allergic and nonallergic lung disease: Increased expression in asthma

BACKGROUND Bronchial asthma is a chronic inflammatory disease associated with genetic components. Recently IL-9 has been reported as a candidate gene for asthma and to be associated with bronchial hyperresponsiveness and elevated levels of total serum IgE. OBJECTIVE To investigate the contribution of IL-9 to the pathogenesis of asthma, we examined the expression of IL-9 and its receptor (IL-9R) in bronchial tissue from subjects with atopic asthma (n = 10), chronic bronchitis (n = 11), and sarcoidosis (n = 9) and from atopic (n = 7) and nonatopic (n = 10) healthy control subjects. METHODS Bronchial biopsy specimens were examined for the presence of IL-9 and IL-9R protein and messenger RNA (mRNA) by immunocytochemistry and in situ hybridization, respectively. To phenotype the cells expressing IL-9 in asthmatic tissue, combined in situ hybridization and immunocytochemistry was also performed. RESULTS There was a highly significant difference (P <.001) in the expression of IL-9 mRNA in asthmatic airways (20.6 +/- 4.0 cells/mm of basement membrane) compared with chronic bronchitis (5.6 +/- 4.4), sarcoidosis (2.5 +/- 1.8), atopic control subjects (7.7 +/- 2.2), and healthy control subjects (2.7 +/- 2.3). The number of IL-9 immunoreactive cells was also greater in asthmatic patients compared with the other groups (P <.05). Although the level of IL-9R mRNA expression did not differ in any of the groups (P >.05), IL-9R immunoreactivity was significantly higher in asthmatic compared with control subjects. Furthermore, IL-9 mRNA expression levels were also significantly correlated with FEV(1) (P <.05) and the airway responsiveness to methacholine producing a 20% fall in FEV(1) (P <. 01). The cells expressing IL-9 mRNA in asthmatic tissue were CD3(+) lymphocytes (68%), major basic protein(+) eosinophils (16%), and elastase(+) neutrophils (8%). CONCLUSION The results of this study demonstrate the potential of IL-9 to be a marker for atopic asthma and furthermore suggest an important role for this cytokine in the pathophysiologic mechanisms of this disease.

miR-199a-5p Is upregulated during fibrogenic response to tissue injury and mediates TGFbeta-induced lung fibroblast activation by targeting caveolin-1.

As miRNAs are associated with normal cellular processes, deregulation of miRNAs is thought to play a causative role in many complex diseases. Nevertheless, the precise contribution of miRNAs in fibrotic lung diseases, especially the idiopathic form (IPF), remains poorly understood. Given the poor response rate of IPF patients to current therapy, new insights into the pathogenic mechanisms controlling lung fibroblasts activation, the key cell type driving the fibrogenic process, are essential to develop new therapeutic strategies for this devastating disease. To identify miRNAs with potential roles in lung fibrogenesis, we performed a genome-wide assessment of miRNA expression in lungs from two different mouse strains known for their distinct susceptibility to develop lung fibrosis after bleomycin exposure. This led to the identification of miR-199a-5p as the best miRNA candidate associated with bleomycin response. Importantly, miR-199a-5p pulmonary expression was also significantly increased in IPF patients (94 IPF versus 83 controls). In particular, levels of miR-199a-5p were selectively increased in myofibroblasts from injured mouse lungs and fibroblastic foci, a histologic feature associated with IPF. Therefore, miR-199a-5p profibrotic effects were further investigated in cultured lung fibroblasts: miR-199a-5p expression was induced upon TGFβ exposure, and ectopic expression of miR-199a-5p was sufficient to promote the pathogenic activation of pulmonary fibroblasts including proliferation, migration, invasion, and differentiation into myofibroblasts. In addition, we demonstrated that miR-199a-5p is a key effector of TGFβ signaling in lung fibroblasts by regulating CAV1, a critical mediator of pulmonary fibrosis. Remarkably, aberrant expression of miR-199a-5p was also found in unilateral ureteral obstruction mouse model of kidney fibrosis, as well as in both bile duct ligation and CCl4-induced mouse models of liver fibrosis, suggesting that dysregulation of miR-199a-5p represents a general mechanism contributing to the fibrotic process. MiR-199a-5p thus behaves as a major regulator of tissue fibrosis with therapeutic potency to treat fibroproliferative diseases.

Quality of life in nasal polyposis

BACKGROUND Nasal polyposis (NP) is a frequent inflammatory chronic disease of the upper respiratory tract, which may impair quality of life (QOL). The NP impact, which is frequently associated with lower respiratory disorders, has never before been studied. OBJECTIVE We initiated this prospective study to establish internal validity and reliability of the generic SF-36 questionnaire in NP and to determine to what level daily functioning becomes impaired as a result of NP. METHODS Forty-nine consecutive patients with NP were included. They were assessed for the severity of nasal symptoms and underwent pulmonary function tests. The QOL profiles in patients with NP were compared with those of patients with perennial rhinitis (n = 111) and healthy subjects (n = 116). RESULTS Cronbachs coefficient alpha demonstrated the high reliability and validity of the SF-36 questionnaire for patients with NP (alpha =.89). NP impaired QOL more than perennial allergic rhinitis (P <.05). The impairment of QOL was greater when NP was associated with asthma (P <.05). SF-36 scores appeared highly correlated to pulmonary function (FEV1, maximal midexpiratory flow, forced vital capacity), suggesting relationships between QOL in NP and associated bronchial obstruction. Severity of nasal symptoms were not related to QOL scales. In addition, sequential evaluations of QOL, nasal symptoms, and pulmonary function were performed 10 months after the first evaluation in 28 patients with NP. These evaluations demonstrated that NP treatment either with nasal steroids or endonasal ethmoidectomy significantly improved both nasal symptoms and QOL without significant change of pulmonary function. CONCLUSION Our study clearly demonstrated that the SF-36 questionnaire presented a high internal validity and reliability in patients with NP. NP impaired QOL to a greater degree than perennial allergic rhinitis. QOL improvement after NP treatment is related to nasal symptoms improvement.

Hierarchical cluster and survival analyses of antisynthetase syndrome: phenotype and outcome are correlated with anti-tRNA synthetase antibody specificity.

The clinical phenotype and evolution of antisynthetase syndrome (ASS) are heterogeneous. This study was therefore undertaken to identify subgroups of ASS patients with similar clinico-biological features and outcomes. This retrospective multicentric study included 233 consecutive patients with three different anti-aminoacyl-tRNA-synthetase antibodies (anti-ARS): anti-Jo1 (n=160), anti-PL7 (n=25) and anti-PL12 (n=48). To characterise ASS patients, bivariate, multiple correspondence (MCA), cluster and survival analyses were performed. Interstitial lung disease (ILD) and myositis were the most common ASS manifestations. However, their respective frequencies were correlated to anti-ARS specificity: ILD was more frequent (80% and 88% vs 67%, p=0.014) whereas myositis was less common (44% and 47% vs 74%, p<0.001) in patients with anti-PL7 and anti-PL12 compared to anti-Jo1. The MCA suggested that anti-PL7 and anti-PL12 phenotypes were close to one another and distinct from anti-Jo1. The clustering analysis confirmed these data, identifying subgroups strongly defined by the anti-ARS specificity and other clinical features. Cluster 1 (n=175, 86% of anti-Jo1) defined patients with the most diffuse phenotype, whereas patients from cluster 2 (n=48, 96% of anti-PL12 and anti-PL7) exhibited a disease more restricted to the lung. Patient survival was also conditioned by the anti-ARS specificity, and was significantly lower in patients with anti-PL7/12 rather than anti-Jo1 (p=0.012). Other factors associated with poor survival were mostly related to pulmonary involvement, including severe dyspnea (p=0.003) and isolated ILD (p=0.009) at diagnosis. In patients with ASS, the phenotype and the survival were correlated with the anti-ARS specificity.

Expression of E-Selectin, ICAM-1 and VCAM-1 on Bronchial Biopsies from Allergic and Non-Allergic Asthmatic Patients

The bronchial mucosa of asthmatic patients is characterized by a large influx of eosinophils, monocytes and lymphocytes. Leucocyte migration and accumulation is thought to involve adhesion molecules, as shown in an animal of allergic asthma and in humans with other allergic diseases. The aim of this study was to evaluate the expression of E-selectin, intercellular adhesion molecule-1 (ICAM-1) and vascular cellular adhesion molecule-1 (VCAM-1) on endothelium and epithelium in bronchial biopsies obtained from patients with allergic (n = 17) and non-allergic asthma (n = 18). Bronchial biopsies were taken in asthmatic patients and control subjects (n = 10) by fiberoptic bronchoscopy and embedded in paraffin. The cellular infiltrate was evaluated by May-Grünwald-Giemsa staining. Adhesion molecule expression was analyzed by immunohistochemistry using mouse monoclonal antibodies; the results were expressed as the percentage of positive cells. For each patient, we evaluated the severity of asthma as defined by the AAS score and the treatment. In controls, low expression of ICAM-1 was observed on the epithelium and endothelium (9.6 +/- 2.7 and 11.2 +/- 4.1%, respectively), while E-selectin and VCAM-1 were not expressed. In patients with allergic asthma, a significant increase of ICAM-1 expression was observed on epithelium and endothelium (28 +/- 5.3 and 35.6 +/- 5%, respectively), whereas E-selectin (17.4 +/- 4.8%) and VCAM-1 (12.8 +/- 3.6%) were overexpressed only on endothelium. In allergic asthmatic patients, adhesion molecule expression on endothelium was correlated with eosinophil and total leucocyte infiltrate (p < 0.05). In contrast, adhesion molecule expression in biopsies from patients with non-allergic asthma (14.1 +/- 5.2 and 15.3 +/- 3.6% for ICAM-1 expression on epithelium and endothelium, respectively) was not significantly different from the control subjects.(ABSTRACT TRUNCATED AT 250 WORDS)

Interstitial lung disease and anti-Jo-1 antibodies: difference between acute and gradual onset

Aim: A multicentre retrospective study was undertaken to examine patients with interstitial lung disease (ILD) with the initial clinical manifestation of an anti-synthetase syndrome (anti-Jo-1 antibodies), and to analyse the characteristics and long-term outcome of these patients according to their clinical presentation (acute or gradual onset), treatment and adverse events related to treatment. Methods: 32 patients, 15 (47%) presenting with acute onset and associated respiratory insufficiency (group A) and 17 (53%) with gradual onset (group G) were examined. Myositis was diagnosed at admission in only 31% of cases and was observed during follow-up in 56% of cases, but the prevalence did not differ between the two groups. Results: Fever and radiological patterns including diffuse patchy ground-glass opacities, basal irregular lines and consolidation on high-resolution CT scan were more frequent in group A than in group G. More patients in group G had neutrophils in the bronchoalveolar lavage fluid and autoantibodies other than anti-Jo-1 (rheumatoid factor, anti SSa/SSb) than in group A. The percentage of patients in whom the ILD improved at 3 months was significantly higher in group A than in group G (13/15 vs 9/17; p = 0.006). In contrast, after 12 months, most patients with ILD progression were in group A and were treated with corticosteroids alone. A combination of corticosteroids and an immunosuppressive drug was required in most cases (84%) at the end of the follow-up period. Severe adverse effects of treatment were observed and varicella zoster virus infection was frequent. Conclusions: Early testing for anti-synthetase antibodies, particularly anti-Jo-1, and creatine kinase determination are useful procedures in patients presenting with ILD. Treatment with corticosteroids and immunosuppressive drugs is required in most patients. At the end of the study, around two-thirds of patients had stable ILD while the other third had disease progression with respiratory insufficiency.

Impact of impaired aerobic capacity on liver transplant candidates.

Background. Oxygen consumption at peak exercise (peak VO2) is the most accurate index of aerobic capacity (AC), which reflects the physical condition of an individual and is currently considered the gold standard for cardiorespiratory fitness. Evaluation of peak VO2 to identify high-risk candidates for liver transplantation (LT) may represent an interesting approach. The aims of this study were (a) to describe AC and identify factors independently associated with peak VO2; (b) to analyze the prognostic value of peak VO2 in patients referred for preliminary evaluation of LT; and (c) to provide preliminary data on the influence of peak VO2 on length of hospitalization and the need for oxygen support after LT. Results. Peak VO2 was determined in patients referred for preliminary evaluation for LT. One hundred thirty-five candidates were included. More than half had severe alterations in peak VO2. Age, gender, model-for-end-stage liver disease (MELD) score, tobacco use, and hemoglobin were independently associated with peak VO2. Candidates with severe alterations in peak VO2 had a lower 1-year survival than others. Model-for-end-stage liver disease score and peak VO2 were independently associated with survival. In patients with a MELD above 17, those with severe alterations of peak VO2 AC had lower 1-year survival than the others. Among patients who underwent LT, those with severe impairment of peak VO2 showed a trend toward a higher mean length of hospitalization after LT and had significantly longer need for oxygen support. Conclusions. Peak VO2 is severely impaired in candidates for LT and affects survival and post-LT course. Perioperative respiratory rehabilitation programs validated in lung and heart transplantation must be tested.

The MUC5B variant is associated with idiopathic pulmonary fibrosis but not with systemic sclerosis interstitial lung disease in the European Caucasian population.

A polymorphism on the MUC5B promoter (rs35705950) has been associated with idiopathic pulmonary fibrosis (IPF) but not with systemic sclerosis (SSc) with interstitial lung disease (ILD). We genotyped the MUC5B promoter in the first 142 patients of the French national prospective cohort of IPF, in 981 French patients with SSc (346 ILD), 598 Italian patients with SSc (207 ILD), 1383 French controls and 494 Italian controls. A meta-analysis was performed including all American data available. The T risk allele was present in 41.9% of the IPF patients, 10.8% of the controls (P = 2×10–44), OR 6.3 [4.6–8.7] for heterozygous patients and OR 21.7 [10.4–45.3] for homozygous patients. Prevalence of the T allele was not modified according to age, gender, smoking in IPF patients. However, none of the black patients with IPF presented the T allele. The prevalence of the T risk allele was similar between French (10%) and Italian (12%) cohorts of SSc whatever the presence of an ILD (11.1% and 13.5%, respectively). Meta-analysis confirmed the similarity between French, Italian and American cohorts of IPF or SSc-ILD. This study confirms 1) an association between the T allele risk and IPF, 2) an absence of association with SSc-ILD, suggesting different pathophysiology.

Increased intestinal permeability in bronchial asthma

T lymphocytes are a major component of bronchial inflammatory processes in asthma. Because lymphocytes have the ability to migrate from one mucosal site to another, we initiated this prospective study to demonstrate mucosal abnormalities of the digestive barrier in asthma. To establish this we studied intestinal permeability in a group of 37 patients with asthma (21 allergic and 16 nonallergic) by measuring chromium 51-labeled ethylenediaminetetraacetatic acid (CrEDTA) urinary recovery. The results were compared with those obtained in a group of 13 nonasthmatic patients with chronic obstructive pulmonary disease and 26 healthy control subjects. Urinary recovery of CrEDTA was significantly higher in patients with asthma (2.5% +/- 1.95%) than in patients with chronic obstructive pulmonary disease (1.16% +/- 0.48%) and healthy control subjects (1.36% +/- 0.14%). There was no significant difference in intestinal permeability between patients with allergic asthma (2.94% +/- 2.4%) and those with nonallergic asthma (1.92% +/- 0.9%). Intestinal permeability was not correlated with the severity of asthma as measured by FEV1. Similarly, intestinal permeability did not significantly vary according to Aas score or steroid treatment. Serum IgE values and eosinophil blood count were not correlated with intestinal permeability. Intestinal permeability was evaluated sequentially in seven patients with asthma (4 allergic and 3 nonallergic) with a mean interval of 7.6 months (range, 2 to 13 months) and did not significantly change. Our results support the hypothesis that a general defect of the whole mucosal system is present as a cause or a consequence of bronchial asthma.