Dominique Bullens

IL-17 mRNA in sputum of asthmatic patients: linking T cell driven inflammation and granulocytic influx?

BackgroundThe role of Th2 cells (producing interleukin (IL-)4, IL-5 and IL-13) in allergic asthma is well-defined. A distinct proinflammatory T cell lineage has recently been identified, called Th17 cells, producing IL-17A, a cytokine that induces CXCL8 (IL-8) and recruits neutrophils. Neutrophilic infiltration in the airways is prominent in severe asthma exacerbations and may contribute to airway gland hypersecretion, bronchial hyper-reactivity and airway wall remodelling in asthma.Aimto study the production of IL-17 in asthmatic airways at the mRNA level, and to correlate this with IL-8 mRNA, neutrophilic inflammation and asthma severity.MethodsWe obtained airway cells by sputum induction from healthy individuals (n = 15) and from asthmatic patients (n = 39). Neutrophils were counted on cytospins and IL-17A and IL-8 mRNA expression was quantified by real-time RT-PCR (n = 11 controls and 33 asthmatics).ResultsSputum IL-17A and IL-8 mRNA levels are significantly elevated in asthma patients compared to healthy controls. IL-17 mRNA levels are significantly correlated with CD3γ mRNA levels in asthmatic patients and mRNA levels of IL-17A and IL-8 correlated with each other and with sputum neutrophil counts. High sputum IL-8 and IL-17A mRNA levels were also found in moderate-to-severe (persistent) asthmatics on inhaled steroid treatment.ConclusionThe data suggest that Th17 cell infiltration in asthmatic airways links T cell activity with neutrophilic inflammation in asthma.

The role of interleukin-17 during acute rejection after lung transplantation

Acute rejection (AR) is an important complication that can occur after lung transplantation and constitutes a risk factor for bronchiolitis obliterans syndrome, which is characterised by a neutrophilic airway inflammation. The specific aim of this study was to investigate the role of interleukin (IL)-17, which promotes chemotaxis of neutrophils by inducing IL-8 production, in AR. Cell differentials, mRNA and protein levels were quantified in bronchoalveolar lavages (BALs) taken from patients at 28 and 90 days after lung transplantation. The patients rejection status was assessed by transbronchial biopsy. An AR was found in nine out of the 26 patients examined, 28 days after transplantation. The number of BAL neutrophils and lymphocytes were increased in these patients. IL-17 mRNA and protein levels in the BAL were increased in patients with AR. Analysis of BAL obtained at day 90 after transplantation, demonstrated that the increase in IL-17 had disappeared, whereas the increase in neutrophils and lymphocytes persisted. These data showed that interleukin-17 is temporarily upregulated in bronchoalveolar lavage during acute rejection. The number of lymphocytes and neutrophils are increased in bronchoalveolar lavage during acute rejection and may persist up to 2 months after acute rejection. These findings suggest that interleukin-17 is important in the pathophysiology of acute lung rejection.

Defective CD4+CD25+ regulatory T cell functioning in collagen-induced arthritis: an important factor in pathogenesis, counter-regulated by endogenous IFN-γ

Mice with a deficiency in IFN-γ or IFN-γ receptor (IFN-γR) are more susceptible to collagen-induced arthritis (CIA), an experimental autoimmune disease that relies on the use of complete Freunds adjuvant (CFA). Here we report that the heightened susceptibility of IFN-γR knock-out (KO) mice is associated with a functional impairment of CD4+CD25+ Treg cells. Treatment of wild-type mice with depleting anti-CD25 antibody after CFA-assisted immunisation with collagen type II (CII) significantly accelerated the onset of arthritis and increased the severity of CIA. This is an indication of a role of Treg cells in the effector phase of CIA. IFN-γR deficiency did not affect the number of CD4+CD25+ T cells in the central and peripheral lymphoid tissues. In addition, CD4+CD25+ T cells isolated from naive IFN-γR KO mice had a normal potential to suppress T cell proliferation in vitro. However, after immunisation with CII in CFA, the suppressive activity of CD4+CD25+ T cells became significantly more impaired in IFN-γR-deficient mice. Moreover, expression of the mRNA for Foxp3, a highly specific marker for Treg cells, was lower. We further demonstrated that the effect of endogenous IFN-γ, which accounts for more suppressive activity in wild-type mice, concerns both Treg cells and accessory cells. Our results demonstrate that the decrease in Treg cell activity in CIA is counter-regulated by endogenous IFN-γ.

Evaluation of airway inflammation by quantitative Th1/Th2 cytokine mRNA measurement in sputum of asthma patients

Background: Asthma is a chronic inflammatory disorder of the airways driven by T cell activation. Th2 cells and their cytokines are thought to play a role in the pathophysiology of allergic as well as non-allergic asthma. Methods: Airway cells were obtained by sputum induction from 15 healthy and 39 asthmatic individuals and the airway T cell cytokine profiles (interleukin (IL)-4, IL-5, IL-13, IL-10 and interferon (IFN)-γ) at the mRNA level were studied by real time RT-PCR. Results: Asthma patients had increased expression of IL-5 (p = 0.001) and IL-13 (p = 0.03) mRNA in sputum compared with non-asthmatic controls. IL-4 mRNA and IFN-γ mRNA were detectable in the sputum of 44% and 21% of patients, respectively, but not in controls. Sputum IL-10 mRNA levels did not differ significantly between patients and controls. Sputum mRNA expression levels of IL-4, IL-5, and IL-13 were significantly correlated with the percentage of eosinophils and were higher in subjects with allergic asthma than in those with non-allergic asthma (p = 0.03, p = 0.02 and p = 0.0002, respectively); they did not differ between mild asthmatic subjects and those with moderate to severe asthma. In contrast, IFN-γ mRNA expression was higher in non-allergic than in allergic patients (p = 0.04) and higher in patients with moderate to severe asthma than in those with mild asthma (p<0.01). Sputum IL-5 mRNA levels (but not the other cytokine mRNA levels) were also correlated with exhaled nitric oxide (eNO) and with bronchial hyperreactivity expressed as the histamine concentration resulting in a 20% decrease in forced expiratory volume in 1 second. Conclusion: Real time RT-PCR analysis of mRNA in induced sputum confirms a predominance of Th2 cytokines in both allergic and non-allergic asthma. IL-5 levels reflect eosinophil infiltration as well as eNO levels and hyperreactivity, and levels of the Th1 cytokine IFN-γ indicate asthma severity. The technique is a promising tool for use in further studies of asthma severity and disease activity.

The Role of the IL23/IL17 Axis in Bronchiolitis Obliterans Syndrome After Lung Transplantation

Bronchiolitis obliterans syndrome (BOS) is the leading cause of death after lung transplantation. Treatment is challenging, as the precise pathophysiology remains unclear.

Sinonasal pathology in nonallergic asthma and COPD: 'united airway disease' beyond the scope of allergy.

Background:  In contrast to the epidemiological and clinical association between allergic rhinitis and asthma, upper airway inflammation is less characterized in patients with nonatopic asthma and virtually unexplored in chronic obstructive pulmonary disease (COPD). Here, sinonasal pathology is studied in patients with allergic asthma, nonallergic asthma and COPD.

Reciprocal changes of Foxp3 expression in blood and intestinal mucosa in IBD patients responding to infliximab.

Background: Immune therapies may act in inflammatory bowel diseases (IBD) by modulating regulatory T cells (Tregs). Therefore, we investigated the effect of infliximab (IFX) therapy on Forkhead box protein3 (Foxp3) T cells in blood and intestinal mucosa from Crohns disease (CD) and ulcerative colitis (UC). Methods: Forty patients with active IBD (23 CD / 17 UC) were treated with IFX 5 mg/kg intravenously at weeks 0, 2, 6, and each 8 weeks thereafter. Blood samples were obtained before every infusion and T‐lymphocyte subsets were characterized by flow cytometry. Foxp3 expression in intestinal biopsies from 43 patients with active IBD (19 CD / 24 UC) before and after IFX infusion and from 6 controls were assessed by quantitative reverse‐transcription polymerase chain reaction and immunohistochemistry. Plasma C‐reactive protein (CRP), clinical response, and endoscopic healing data were collected in parallel. Results: IFX therapy resulted in a significant and sustained relative increase of CD4+CD25+Foxp3+ Treg and of CD4+CD25−Foxp3+ Treg cells in peripheral blood (both P < 0.0001 compared to baseline), particularly in responders (both P < 0.05 compared to nonresponders). The change in CRP over time inversely correlated with the increase of CD25+Foxp3+ cells (P < 0.001, r = −0.39) and durable clinical response was associated with a sustained increase of circulating Foxp3+ cells. Surprisingly, IFX therapy downregulated mucosal mRNA and protein expression of Foxp3 in UC and CD responders (both P < 0.001) but not in nonresponders. Conclusions: IFX therapy has opposite effects in Foxp3+ Treg cells in blood and gut mucosa, which suggests a redistribution of this important T‐cell subset. Inflamm Bowel Dis 2010

Elevated expression of both mRNA and protein levels of IL-17A in sputum of stable Cystic Fibrosis patients

BackgroundT helper 17 (Th17) cells can recruit neutrophils to inflammatory sites through production of IL-17, which induces chemokine release. IL-23 is an important inducer of IL-17 and IL-22 production. Our aim was to study the role of Th17 cells in cystic fibrosis (CF) lung disease by measuring IL-17 protein and mRNA levels and IL-22 and IL-23 mRNA in sputum of clinically stable CF patients and by comparing these levels with healthy controls.MethodsSputum induction was performed in adult CF patients outside of an exacerbation and healthy control subjects. IL-17A protein levels were measured in supernatants with cytometric bead array (CBA) and RNA was isolated and quantitative RT-PCR was performed for IL-17A, IL-22 and IL-23.ResultsWe found significantly higher levels of IL-17A protein and mRNA levels (both: p < 0.0001) and IL-23 mRNA levels (p < 0.0001) in the sputum of CF group as compared to controls. We found very low levels of IL-22 mRNA in the CF group. The levels of IL-17 and IL-23 mRNA were higher in patients chronically infected with Pseudomonas aeruginosa (P. aeruginosa) as compared to those who were not chronically infected with P. aeruginosa. The presence of Staphylococcus aureus (S. aureus) on sputum did not affect the IL-17 or IL-23 levels. There was no correlation between IL-17 or IL-23 levels and FEV1 nor sputum neutrophilia.ConclusionThe elevated levels of IL-17 and IL-23 might indicate that Th17 cells are implicated in the persistent neutrophil infiltration in CF lung disease and chronic infection with P. aeruginosa.

Impact of lipoteichoic acid modification on the performance of the probiotic Lactobacillus rhamnosus GG in experimental colitis.

While some probiotic strains might have adjuvant effects in the therapy for inflammatory bowel diseases (IBD), these effects remain controversial and cannot be generalized. In this study, a dltD mutant of the model probiotic Lactobacillus rhamnosus GG (LGG), having a drastic modification in its lipoteichoic acid (LTA) molecules, was analysed for its effects in an experimental colitis model. Dextran sulphate sodium (DSS) was used to induce either moderate to severe or mild chronic colitis in mice. Mice received either phosphate‐buffered saline (PBS), LGG wild‐type or the dltD mutant via the drinking water. Macroscopic parameters, histological abnormalities, cytokine and Toll‐like receptor (TLR) expression were analysed to assess disease activity. LGG wild‐type did not show efficacy in the different experimental colitis set‐ups. This wild‐type strain even seemed to exacerbate the severity of colitic parameters in the moderate to severe colitis model compared to untreated mice. In contrast, mice treated with the dltD mutant showed an improvement of some colitic parameters compared to LGG wild‐type‐treated mice in both experimental models. In addition, treatment with the dltD mutant correlated with a significant down‐regulation of Toll‐like receptor‐2 expression and of downstream proinflammatory cytokine expression in the colitic mice. These results show that molecular cell surface characteristics of probiotics are crucial when probiotics are considered for use as supporting therapy in IBD.

Original article: Sinonasal pathology in nonallergic asthma and COPD: ‘united airway disease’ beyond the scope of allergy

Background:  In contrast to the epidemiological and clinical association between allergic rhinitis and asthma, upper airway inflammation is less characterized in patients with nonatopic asthma and virtually unexplored in chronic obstructive pulmonary disease (COPD). Here, sinonasal pathology is studied in patients with allergic asthma, nonallergic asthma and COPD.