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Abstract Background The loss of alveolar walls is a hallmark of emphysema. As fibroblasts play an important role in the maintenance of alveolar structure, a change in fibroblast phenotype could be involved in the pathogenesis of this disease. In a previous study we found a reduced in vitro proliferation rate and number of population doublings of parenchymal lung fibroblasts from patients with emphysema and we hypothesized that these findings could be related to a premature cellular aging of these cells. In this study, we therefore compared cellular senescence markers and expression of respective genes between lung fibroblasts from patients with emphysema and control patients without COPD. Methods Primary lung fibroblasts were obtained from 13 patients with moderate to severe lung emphysema (E) and 15 controls (C) undergoing surgery for lung tumor resection or volume reduction (n = 2). Fibroblasts (8E/9C) were stained for senescence-associated β-galactosidase (SA-β-Gal). In independent cultures, DNA from lung fibroblasts (7E/8C) was assessed for mean telomere length. Two exploratory 12 k cDNA microarrays were used to assess gene expression in pooled fibroblasts (3E/3C). Subsequently, expression of selected genes was evaluated by quantitative PCR (qPCR) in fibroblasts of individual patients (10E/9C) and protein concentration was analyzed in the cell culture supernatant. Results The median (quartiles) percentage of fibroblasts positive for SA-β-Gal was 4.4 (3.2;4.7) % in controls and 16.0 (10.0;24.8) % in emphysema (p = 0.001), while telomere length was not different. Among the candidates for differentially expressed genes in the array (factor ≥ 3), 15 were upregulated and 121 downregulated in emphysema. qPCR confirmed the upregulation of insulin-like growth factor-binding protein (IGFBP)-3 and IGFBP-rP1 (p = 0.029, p = 0.0002), while expression of IGFBP-5, -rP2 (CTGF), -rP4 (Cyr61), FOSL1, LOXL2, OAZ1 and CDK4 was not different between groups. In line with the gene expression we found increased cell culture supernatant concentrations of IGFBP-3 (p = 0.006) in emphysema. Conclusion These data support the hypothesis that premature aging of lung fibroblasts occurs in emphysema, via a telomere-independent mechanism. The upregulation of the senescence-associated IGFBP-3 and -rP1 in emphysema suggests that inhibition of the action of insulin and insulin-like growth factors could be involved in the reduced in vitro-proliferation rate.

Background: The abundance of online content contributed by patients is a rich source of insight about the lived experience of disease. Patients share disease experiences with other members of the patient and caregiver community and do so using their own lexicon of words and phrases. This lexicon and the topics that are communicated using words and phrases belonging to the lexicon help us better understand disease burden. Insights from social media may ultimately guide clinical development in ways that ensure that future treatments are fit for purpose from the patient's perspective., Objective: We sought insights into the patient experience of chronic obstructive pulmonary disease (COPD) by analyzing a substantial corpus of social media content. The corpus was sufficiently large to make manual review and manual coding all but impossible to perform in a consistent and systematic fashion. Advanced analytics were applied to the corpus content in the search for associations between symptoms and impacts across the entire text corpus., Methods: We conducted a retrospective, cross-sectional study of 5663 posts sourced from open blogs and online forum posts published by COPD patients between February 2016 and August 2019. We applied a novel neural network approach to identify a lexicon of community words and phrases used by patients to describe their symptoms. We used this lexicon to explore the relationship between COPD symptoms and disease-related impacts., Results: We identified a diverse lexicon of community words and phrases for COPD symptoms, including gasping, wheezy, mucus-y, and muck. These symptoms were mentioned in association with specific words and phrases for disease impact such as frightening, breathing discomfort, and difficulty exercising. Furthermore, we found an association between mucus hypersecretion and moderate disease severity, which distinguished mucus from the other main COPD symptoms, namely breathlessness and cough., Conclusions: We demonstrated the potential of neural networks and advanced analytics to gain patient-focused insights about how each distinct COPD symptom contributes to the burden of chronic and acute respiratory illness. Using a neural network approach, we identified words and phrases for COPD symptoms that were specific to the patient community. Identifying patterns in the association between symptoms and impacts deepened our understanding of the patient experience of COPD. This approach can be readily applied to other disease areas., (©Tobe Che Benjamin Freeman, Raul Rodriguez-Esteban, Juergen Gottowik, Xing Yang, Veit Johannes Erpenbeck, Mathias Leddin. Originally published in JMIR Medical Informatics (https://medinform.jmir.org), 11.11.2021.)

Background: Prostaglandin D 2 (PGD 2 ) signaling via prostaglandin D 2 receptor 2 (DP 2 ) contributes to atopic and non-atopic asthma. Inhibiting DP 2 has shown therapeutic benefit in certain subsets of asthma patients, improving eosinophilic airway inflammation. PGD 2 metabolites prolong the inflammatory response in asthmatic patients via DP 2 signaling. The role of PGD 2 metabolites on eosinophil and ILC2 activity is not fully understood., Methods: Eosinophils and ILC2s were isolated from peripheral blood of atopic asthmatic patients. Eosinophil shape change, ILC2 migration and IL-5/IL-13 cytokine secretion were measured after stimulation with seven PGD 2 metabolites in presence or absence of the selective DP 2 antagonist fevipiprant., Results: Selected metabolites induced eosinophil shape change with similar nanomolar potencies except for 9α,11β-PGF 2 . Maximal values in forward scatter of eosinophils were comparable between metabolites. ILC2s migrated dose-dependently in the presence of selected metabolites except for 9α,11β-PGF 2 with EC 50 values ranging from 17.4 to 91.7 nM. Compared to PGD 2 , the absolute cell migration was enhanced in the presence of Δ 12 -PGD 2 , 15-deoxy-Δ 12,14 -PGD 2 , PGJ 2 , Δ 12 -PGJ 2 and 15-deoxy-Δ 12,14 -PGJ 2 . ILC2 cytokine production was dose dependent as well but with an average sixfold reduced potency compared to cell migration (IL-5 range 108.1 to 526.9 nM, IL-13 range: 125.2 to 788.3 nM). Compared to PGD 2 , the absolute cytokine secretion was reduced in the presence of most metabolites. Fevipiprant dose-dependently inhibited eosinophil shape change, ILC2 migration and ILC2 cytokine secretion with (sub)-nanomolar potencies., Conclusion: Prostaglandin D 2 metabolites initiate ILC2 migration and IL-5 and IL-13 cytokine secretion in a DP 2 dependent manner. Our data indicate that metabolites may be important for in vivo eosinophil activation and ILC2 migration and to a lesser extent for ILC2 cytokine secretion., (© 2021. The Author(s).)

Human type 2 cytotoxic T (Tc2) cells are enriched in severe eosinophilic asthma and can contribute to airway eosinophilia. PGD 2 and its receptor PGD 2 receptor 2 (DP2) play important roles in Tc2 cell activation, including migration, cytokine production, and survival. In this study, we revealed novel, to our knowledge, functions of the PGD 2 /DP2 axis in Tc2 cells to induce tissue-remodeling effects and IgE-independent PGD 2 autocrine production. PGD 2 upregulated the expression of tissue-remodeling genes in Tc2 cells that enhanced the fibroblast proliferation and protein production required for tissue repair and myofibroblast differentiation. PGD 2 stimulated Tc2 cells to produce PGD 2 using the routine PGD 2 synthesis pathway, which also contributed to TCR-dependent PGD 2 production in Tc2 cells. Using fevipiprant, a specific DP2 antagonist, we demonstrated that competitive inhibition of DP2 not only completely blocked the cell migration, adhesion, proinflammatory cytokine production, and survival of Tc2 cells triggered by PGD 2 but also attenuated the tissue-remodeling effects and autocrine/paracrine PGD 2 production in Tc2 induced by PGD 2 and other stimulators. These findings further confirmed the anti-inflammatory effect of fevipiprant and provided a better understanding of the role of Tc2 cells in the pathogenesis of asthma., (Copyright © 2021 The Authors.)

This drug-drug interaction study determined the effect of cyclosporine, an inhibitor of organic anion transporting polypeptide (OATP) 1B3 and P-gp, on the pharmacokinetics (PK) of fevipiprant, an oral, highly selective, competitive antagonist of the prostaglandin D 2 receptor 2 and a substrate of the two transporters. The concomitant administration of an intravenous microdose of stable isotope-labeled fevipiprant provided the absolute bioavailability of fevipiprant as well as mechanistic insights into its PK and sensitivity to drug interactions. Liquid chromatography-mass spectrometry/mass spectrometry was used to measure plasma and urine concentrations. Geometric mean ratios [90% confidence interval (CI)] for oral fevipiprant with or without cyclosporine were 3.02 (2.38, 3.82) for C max , 2.50 (2.17, 2.88) for AUC last , and 2.35 (1.99, 2.77) for AUC inf The geometric mean ratios (90% CI) for fevipiprant intravenous microdose with or without cyclosporine were 1.04 (0.86, 1.25) for C max , 2.04 (1.83, 2.28) for AUC last , and 1.95 (1.76, 2.16) for AUC inf The absolute bioavailability for fevipiprant was approximately 0.3 to 0.4 in the absence and 0.5 in the presence of cyclosporine. The intravenous microdose allowed differentiation between systemic and presystemic effects of cyclosporine on fevipiprant, demonstrating a small (approximately 1.2-fold) presystemic effect of cyclosporine and a larger (approximately twofold) effect on systemic elimination of fevipiprant. Uptake by OATP1B3 appears to be the rate-limiting step in the hepatic elimination of fevipiprant, whereas P-gp does not have a relevant effect on oral absorption. SIGNIFICANCE STATEMENT: The drug interaction investigated here with cyclosporine, an inhibitor of several drug transporters, provides a refined quantitative understanding of the role of active transport processes in liver and intestine for the absorption and elimination of fevipiprant as well as the basis to assess the need for dose adjustment in the presence of transporter inhibitors. The applied intravenous microdose approach presents a strategy to maximize learnings from a trial, limit the number and duration of clinical trials, and enhance mechanistic drug-drug interaction understanding., (Copyright © 2020 by The American Society for Pharmacology and Experimental Therapeutics.)

Rhinovirus (RV) infections are a major cause of exacerbations in patients with asthma. Experimental RV challenges can provide insight into the pathophysiology of viral exacerbations. Previous reports, investigating mild or moderate asthma patients, have shown an upregulation in type 2 inflammation post RV infection, however, studies specifically involving asthma patients taking inhaled corticosteroids have concentrated on symptoms and lung function, rather than the inflammatory response. Eleven moderate asthma patients were inoculated with RV. Cold symptoms and asthma control were assessed at baseline and post infection. Nasal epithelial lining fluid and bronchial alveolar lavage (BAL) fluid were collected at baseline and 4 days post infection for assessment of inflammatory proteins. Patients suffered increased cold symptoms and decreased asthma control within 7 days of infection. Antiviral mechanisms were induced following inoculation, with increases in interferon -α, β, γ and λ, as well as CXCL10 and CXCL11. Type 2 inflammatory cytokines were also significantly elevated post RV infection in both nasal and bronchial samples. In BAL, epithelial derived IL-25 and IL-33 levels strongly correlated with Th2 cytokines, IL-4, IL-5 and IL-13. We show how experimental rhinovirus challenge regulates lung and nasal biomarkers in asthma patients taking inhaled corticosteroids. These biomarkers could be used to evaluate the effects of novel drugs for asthma., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

Background: Omalizumab, a humanized recombinant monoclonal anti-IgE antibody, proved to be effective in patients with chronic spontaneous urticaria (CSU), including severe and treatment-refractory CSU. Here, we report omalizumab's effect on gene expression in skin biopsies from CSU patients enrolled in a double-blind, placebo-controlled study., Methods: Chronic spontaneous urticaria patients (18-75 years) were randomized to either 300 mg omalizumab (n = 20) or placebo (n = 10) administered s.c. every 4 weeks for 12 weeks (NCT01599637). Lesional and nonlesional skin biopsies were collected from the same area of consenting patients and assessed at baseline and on Day 85 compared with skin biopsies from the same area of 10 untreated healthy volunteers (HVs). Gene expression data were generated using Affymetrix HG-U133Plus2.0 microarrays. Statistical analyses were performed using R packages., Results: At baseline, 63 transcripts were differentially expressed between lesional and nonlesional skin. Two-thirds of these lesional signatures were also differentially expressed between lesional and HV skin. Upon treatment with omalizumab, >75% of lesional signatures changed to reflect nonlesional skin expression levels (different vs placebo, P < 0.01). Transcripts upregulated in lesional skin (vs nonlesional and/or HV skin) suggested increased mast cell/leukocyte infiltration (FCER1G, C3AR1, CD93, S100A8, and S100A9), increased oxidative stress, vascularization (CYR61), and skin repair events (KRT6A, KRT16). Lesional signatures were not modulated by treatment in nonresponders (defined based on UAS7 longitudinal changes ≥16)., Conclusion: Omalizumab, in treatment responders, reverted transcriptional signatures associated with CSU lesion phenotype to reflect nonlesional/HV expression levels; this is consistent with observed omalizumab-mediated clinical improvement observed in patients with CSU., (© 2018 EAACI and John Wiley and Sons A/S. Published by John Wiley and Sons Ltd.)

Asthma is characterised by chronic airway inflammation, airway obstruction and hyper-responsiveness. The inflammatory cascade in asthma comprises a complex interplay of genetic factors, the airway epithelium, and dysregulation of the immune response.Prostaglandin D 2 (PGD 2 ) is a lipid mediator, predominantly released from mast cells, but also by other immune cells such as T H 2 cells and dendritic cells, which plays a significant role in the pathophysiology of asthma. PGD 2 mainly exerts its biological functions via two G-protein-coupled receptors, the PGD 2 receptor 1 (DP 1 ) and 2 (DP 2 ). The DP 2 receptor is mainly expressed by the key cells involved in type 2 immune responses, including T H 2 cells, type 2 innate lymphoid cells and eosinophils. The DP 2 receptor pathway is a novel and important therapeutic target for asthma, because increased PGD 2 production induces significant inflammatory cell chemotaxis and degranulation via its interaction with the DP 2 receptor. This interaction has serious consequences in the pulmonary milieu, including the release of pro-inflammatory cytokines and harmful cationic proteases, leading to tissue remodelling, mucus production, structural damage, and compromised lung function. This review will discuss the importance of the DP 2 receptor pathway and the current understanding of its role in asthma.

Unbiased Biomarkers for the Prediction of Respiratory Disease Outcomes (U-BIOPRED) was initiated in the first year of the Innovative Medicines Initiative (IMI). It was an ambitious plan to tackle the understanding of asthma through an integration of clinical and multi-'omics approaches that necessitated the bringing together of industry, academic, and patient representatives because it was too large to be managed by any one of the partners in isolation. It was a novel experience for all concerned. In this review, we describe the main features of the U-BIOPRED experience from the industry perspective. We list some of the key advantages and learnings from the perspective of the authors, and also improvements that we feel could be made in future projects., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

Fevipiprant is a novel oral prostaglandin D 2 receptor 2 (DP 2 ; also known as CRTh2) antagonist, which is currently in development for the treatment of severe asthma and atopic dermatitis. We investigated the absorption, distribution, metabolism, and excretion properties of fevipiprant in healthy subjects after a single 200-mg oral dose of [ 14 C]-radiolabeled fevipiprant. Fevipiprant and metabolites were analyzed by liquid chromatography coupled to tandem mass spectrometry and radioactivity measurements, and mechanistic in vitro studies were performed to investigate clearance pathways and covalent plasma protein binding. Biotransformation of fevipiprant involved predominantly an inactive acyl glucuronide (AG) metabolite, which was detected in plasma and excreta, representing 28% of excreted drug-related material. The AG metabolite was found to covalently bind to human plasma proteins, likely albumin; however, in vitro covalent binding to liver protein was negligible. Excretion was predominantly as unchanged fevipiprant in urine and feces, indicating clearance by renal and possibly biliary excretion. Fevipiprant was found to be a substrate of transporters organic anion transporter 3 (OAT3; renal uptake), multidrug resistance gene 1 (MDR1; possible biliary excretion), and organic anion-transporting polypeptide 1B3 (OATP1B3; hepatic uptake). Elimination of fevipiprant occurs via glucuronidation by several uridine 5'-diphospho glucuronosyltransferase (UGT) enzymes as well as direct excretion. These parallel elimination pathways result in a low risk of major drug-drug interactions or pharmacogenetic/ethnic variability for this compound., (Copyright © 2017 by The American Society for Pharmacology and Experimental Therapeutics.)

This article contains data on clinical endpoints (Peak Flow Expiratory Rate, fractional exhaled nitric oxide and total IgE serum levels) and plasma pharmacokinetic parameters concerning the use of the oral CRTh2 antagonist QAW039 (fevipiprant) in mild to moderate asthma patients. Information on experimental design and methods on how this data was obtained is also described. Further interpretation and discussion of this data can be found in the article "The oral CRTh2 antagonist QAW039 (fevipiprant): a phase II study in uncontrolled allergic asthma" (Erpenbeck et al., in press) [1].

Background: There is an unmet medical need for allergic asthma patients who are uncontrolled on conventional therapies. The aim of this study was to collect efficacy and safety data for QAW039, an oral chemoattractant receptor-homologous molecule expressed on Th2 cells (CRTh2) receptor antagonist, for the treatment of asthma., Methods: This was an exploratory phase II, double-blind, randomized, placebo-controlled multi-center study. Patients with mild-to-moderate uncontrolled allergic asthma (N = 170) were either without or weaned off inhaled corticosteroids (ICS) and long-acting β-agonists (LABA) and randomized (1:1) to QAW039 (500 mg once daily) or to placebo for 28 days., Results: Overall, 157 patients completed the study. There were no significant differences between QAW039 and placebo for trough forced expiratory volume in 1 s (FEV1) or Asthma control questionnaire (ACQ) in the total population. Subgroup analyses demonstrated that patients with a FEV1 <70% of predicted at baseline treated with QAW039 had significant improvement compared with placebo in trough FEV1 (QAW039- Placebo [Δ] = 207 mL; 90% confidence interval [CI]: 96, 319; P = 0.002) and ACQ7 (Δ = -0.41; 90%CI: -0.69, -0.13; P = 0.009). QAW039 reached a mean maximum concentration (Cmax) of 3440 ng/mL on day 28 at a median Tmax of 1 h (range 0.5-4 h). Most adverse events (AEs) were mild/moderate and balanced between both groups, with no serious AEs., Conclusions: In the general study population, no improvement in lung function was observed with QAW039. However, a subgroup analysis revealed that patients with greater severity of airflow limitation (FEV1 < 70%) had improved lung function and asthma control when treated with QAW039. QAW039 also demonstrated a favorable safety profile., Trials Registration: ClinicalTrials.govNCT01253603., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

We evaluated the pharmacokinetics (PK), safety, and tolerability of a novel oral CRTh2 antagonist, fevipiprant (QAW039), in healthy subjects. Peak concentrations of fevipiprant in plasma were observed 1-3 hours postdosing. Concentrations declined in a multiexponential manner, followed by an apparent terminal phase (t1/2 , ∼20 hours). Steady state was achieved in 4 days with <2-fold accumulation. Elimination was partly by renal excretion (≤30% of the dose) and glucuronidation. Food had minimal impact on the PK of fevipiprant, and it was well tolerated at single and multiple oral doses up to 500 mg/day. No dose-dependent adverse events were observed, and all the events were mild or moderate in severity. Systemic concentrations were sufficiently high to achieve relevant target occupancy, considering in vitro pharmacology data. In summary, the data support further development as a once-daily oral therapy for allergic diseases., (© 2016, The Authors. Clinical Pharmacology in Drug Development Published by Wiley Periodicals, Inc. on behalf of The American College of Clinical Pharmacology.)

U-BIOPRED is a European Union consortium of 20 academic institutions, 11 pharmaceutical companies and six patient organisations with the objective of improving the understanding of asthma disease mechanisms using a systems biology approach.This cross-sectional assessment of adults with severe asthma, mild/moderate asthma and healthy controls from 11 European countries consisted of analyses of patient-reported outcomes, lung function, blood and airway inflammatory measurements.Patients with severe asthma (nonsmokers, n=311; smokers/ex-smokers, n=110) had more symptoms and exacerbations compared to patients with mild/moderate disease (n=88) (2.5 exacerbations versus 0.4 in the preceding 12 months; p<0.001), with worse quality of life, and higher levels of anxiety and depression. They also had a higher incidence of nasal polyps and gastro-oesophageal reflux with lower lung function. Sputum eosinophil count was higher in severe asthma compared to mild/moderate asthma (median count 2.99% versus 1.05%; p=0.004) despite treatment with higher doses of inhaled and/or oral corticosteroids.Consistent with other severe asthma cohorts, U-BIOPRED is characterised by poor symptom control, increased comorbidity and airway inflammation, despite high levels of treatment. It is well suited to identify asthma phenotypes using the array of "omic" datasets that are at the core of this systems medicine approach., (Copyright ©ERS 2015.)

Background: It has frequently been speculated that pruritus and skin lesions develop after topical exposure to aeroallergens in sensitized patients with atopic dermatitis (AD)., Objective: We sought to study cutaneous reactions to grass pollen in adult patients with AD with accompanying clear IgE sensitization to grass allergen in an environmental challenge chamber using a monocenter, double-blind, placebo-controlled study design., Methods: Subjects were challenged on 2 consecutive days with either 4000 pollen grains/m(3) of Dactylis glomerata pollen or clean air. The severity of AD was assessed at each study visit up to 5 days after challenge by (objective) scoring of AD (SCORAD). Additionally, air-exposed and non-air-exposed skin areas were each scored using local SCORAD scoring and investigator global assessments. Levels of a series of serum cytokines and chemokines were determined by using a Luminex-based immunoassay. The primary end point of the study was the change in objective SCORAD scores between prechallenge and postchallenge values., Results: Exposure to grass pollen induced a significant worsening of AD. A pronounced eczema flare-up of air-exposed rather than covered skin areas occurred. In grass pollen-exposed subjects a significantly higher increase in CCL17, CCL22, and IL-4 serum levels was observed., Conclusions: This study demonstrates that controlled exposure to airborne allergens of patients with a so-called extrinsic IgE-mediated form of AD induced a worsening of cutaneous symptoms., (Copyright © 2015 American Academy of Allergy, Asthma & Immunology. Published by Elsevier Inc. All rights reserved.)

IgE is central to the pathophysiology of allergic asthma. Omalizumab, a humanized anti-IgE mAb, specifically binds free IgE and interrupts the allergic cascade by preventing binding of IgE with its high-affinity FcεRI receptors on mast cells, antigen-presenting cells, and other inflammatory cells. The clinical efficacy of omalizumab has been well documented in a number of clinical trials that involve adults, adolescents, and children with moderate-to-severe and severe allergic asthma. In these studies, omalizumab reduced exacerbations, asthma symptoms, inhaled corticosteroid and rescue medication use, and improved quality of life relative to placebo or best standard of care. Similar benefits have been reported in observational studies in "real-world" populations of patients. Results from recent pooled data from randomized clinical trials and from a large prospective cohort study provide reassurance about the long-term safety of omalizumab. Omalizumab dosing is individualized according to body weight and serum-IgE level, and recent adjustments to the dosing algorithm in Europe have enabled more patients to be eligible for treatment. Ongoing and future research is investigating the optimal duration of therapy, accurate predictors of response to treatment, and efficacy in nonatopic asthma as well as other IgE-mediated conditions., (Copyright © 2014 American Academy of Allergy, Asthma & Immunology. Published by Elsevier Inc. All rights reserved.)

Background: When first approved in the European Union (EU), the omalizumab dosing table had upper bodyweight and IgE limits of 150 kg and 700 IU/mL, respectively. In this study, we assessed the safety, pharmacokinetics (PK) and pharmacodynamics (PD) of omalizumab in patients with IgE/bodyweight combinations above those in the original dosing table., Methods: A multicentre, open-label, parallel-group study assessed the safety, PK and PD of omalizumab in 32 patients with mild-to-moderate allergic (IgE-mediated) asthma. Patients received two subcutaneous injections of omalizumab at one of three dosage levels (450, 525, or 600 mg), chosen according to baseline IgE (300-2000 IU/mL) and bodyweight (40-150 kg), with a 14-day interval between injections., Results: Overall, 69 adverse events (AEs), none of them serious, were reported by 26 (81.3%) patients. Analysis of laboratory measurements, vital signs and ECG data revealed no adverse findings of clinical relevance. The PK profile was consistent with previous data for lower doses. Mean maximum decrease of free IgE from screening was ≥99% for all three doses, and mean free IgE concentrations remained <25 ng/mL for at least 2 weeks after the second dose. The reductions in free IgE were consistent with levels previously associated with clinical improvements., Conclusions: The safety and PK/PD findings from this study are consistent with previous data, and supported the extension of the omalizumab dosing table to include those patients with higher IgE/bodyweight combinations. Clinical trial registry and registration number: clinicaltrials.gov (NCT00546143)., (Copyright © 2014 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Question: Inflammatory cell numbers are important endpoints in clinical studies relying on endobronchial biopsies. Assumption-based bidimensional (2D) counting methods are widely used, although theoretically design-based stereologic three-dimensional (3D) methods alone offer an unbiased quantitative tool. We assessed the method agreement between 2D and 3D counting designs in practice when applied to identical samples in parallel., Materials and Methods: Biopsies from segmental bronchi were collected from healthy non-smokers (n = 7) and smokers (n = 7), embedded and sectioned exhaustively. Systematic uniform random samples were immunohistochemically stained for macrophages (CD68) and T-lymphocytes (CD3), respectively. In identical fields of view, cell numbers per volume unit (NV) were assessed using the physical disector (3D), and profiles per area unit (NA) were counted (2D). For CD68+ cells, profiles with and without nucleus were separately recorded. In order to enable a direct comparison of the two methods, the zero-dimensional CD68+/CD3+-ratio was calculated for each approach. Method agreement was tested by Bland-Altmann analysis., Results: In both groups, mean CD68+/CD3+ ratios for NV and NA were significantly different (non-smokers: 0.39 and 0.68, p<0.05; smokers: 0.49 and 1.68, p<0.05). When counting only nucleated CD68+ profiles, mean ratios obtained by 2D and 3D counting were similar, but the regression-based Bland-Altmann analysis indicated a bias of the 2D ratios proportional to their magnitude. This magnitude dependent deviation differed between the two groups., Conclusions: 2D counts of cell and nuclear profiles introduce a variable size-dependent bias throughout the measurement range. Because the deviation between the 3D and 2D data was different in the two groups, it precludes establishing a 'universal conversion formula'.

Background: Omalizumab has been shown to suppress responses to inhaled allergens in allergic asthma patients with pretreatment immunoglobulin E (IgE) ≤700 IU/ml. To extend current dosing tables, we evaluated the potential of high omalizumab doses to block allergen-induced bronchoconstriction in patients with higher IgE levels., Methods: Asthmatic adults (18-65 years; body weight 40-150 kg) were divided into groups according to screening IgE (group 1: 30-300 IU/ml; group 2: 700-2,000 IU/ml) and randomized 2:1 to omalizumab/placebo every 2 or 4 weeks for 12-14 weeks. Allergen bronchoprovocation (ABP) testing was performed before treatment and at weeks 8 and 16. The primary efficacy endpoint, the early-phase allergic response (EAR), was defined as the maximum percentage drop in forced expiratory volume in 1 s during the first 30 min after ABP. Serum free IgE was determined as a pharmacodynamic endpoint, and the exhaled fractional concentration of nitric oxide (FE(NO)) was an exploratory endpoint., Results: Fifty patients were included in the study. Omalizumab improved EAR; at week 8, EAR was 23.1% for placebo, 9.3% in group 1 (p = 0.018 versus placebo) and 5.6% in group 2 (p < 0.001). At week 16, EAR was 20%, 11.8% (p = 0.087) and 5.1% (p < 0.001), respectively. Free IgE decreased in groups 1 and 2 and remained <50 ng/ml in all patients during weeks 6-16. Omalizumab completely suppressed FE(NO) increases after ABP in both groups., Conclusions: Omalizumab blocked early asthmatic responses over a broad range of IgE/body weight combinations. Extending the dosing tables enables omalizumab to benefit a wider range of patients., (Copyright © 2012 S. Karger AG, Basel.)

Background: Allergen-containing subpollen particles (SPP) are released from whole plant pollen upon contact with water or even high humidity. Because of their size SPP can preferentially reach the lower airways where they come into contact with surfactant protein (SP)-D. The aim of the present study was to investigate the influence of SP-D in a complex three-dimensional human epithelial airway model, which simulates the most important barrier functions of the epithelial airway. The uptake of SPP as well as the secretion of pro-inflammatory cytokines was investigated., Methods: SPP were isolated from timothy grass and subsequently fluorescently labeled. A human epithelial airway model was built by using human Type II-pneumocyte like cells (A549 cells), human monocyte derived macrophages as well as human monocyte derived dendritic cells. The epithelial cell model was incubated with SPP in the presence and absence of surfactant protein D. Particle uptake was evaluated by confocal microscopy and advanced computer-controlled analysis. Finally, human primary CD4+ T-Cells were added to the epithelial airway model and soluble mediators were measured by enzyme linked immunosorbent assay or bead array., Results: SPP were taken up by epithelial cells, macrophages, and dendritic cells. This uptake coincided with secretion of pro-inflammatory cytokines and chemokines. SP-D modulated the uptake of SPP in a cell type specific way (e.g. increased number of macrophages and epithelial cells, which participated in allergen particle uptake) and led to a decreased secretion of pro-inflammatory cytokines., Conclusion: These results display a possible mechanism of how SP-D can modulate the inflammatory response to inhaled allergen.

Background: Pulmonary surfactant protein D (SP-D) is considered as a candidate biomarker for the functional integrity of the lung and for disease progression, which can be detected in serum. The origin of SP-D in serum and how serum concentrations are related to pulmonary concentrations under inflammatory conditions is still unclear., Methods: In a cross-sectional study comprising non-smokers (n=10), young--(n=10), elderly smokers (n=20), and smokers with COPD (n=20) we simultaneously analysed pulmonary and serum SP-D levels with regard to pulmonary function, exercise, repeatability and its quaternary structure by native gel electrophoresis. Statistical comparisons were conducted by ANOVA and post-hoc testing for multiple comparisons; repeatability was assessed by Bland-Altman analysis., Results: In COPD, median (IQR) pulmonary SP-D levels were lower (129(68) ng/ml) compared to smokers (young: 299(190), elderly: 296(158) ng/ml; p<0.01) and non-smokers (967(708) ng/ml; p<0.001). The opposite was observed in serum, with higher concentrations in COPD (140(89) ng/ml) as compared to non-smokers (76(47) ng/ml; p<0.01). SP-D levels were reproducible and correlated with the degree of airway obstruction in all smokers. In addition, smoking lead to disruption of the quaternary structure., Conclusions: Pulmonary and serum SP-D levels are stable markers influenced by smoking and related to airflow obstruction and disease state. Smaller subunits of pulmonary SP-D and the rapid increase of serum SP-D levels in COPD due to exercise support the translocation hypothesis and its use as a COPD biomarker., Trial Registration: no interventional trial.

Pollen starch granules (PSGs) are allergen particles that get into contact with pulmonary surfactant and phagocytes in the terminal airways. In this study, the effects of surfactant protein D (SP-D) on the interaction of PSGs with phagocytes and on the pulmonary clearance of PSGs were determined. Fluorescently labeled PSGs were incubated in vitro with murine lung macrophages or dendritic cells (DCs) ± recombinant rat SP-D (rrSP-D). In addition, the effect of SP-D on uptake of PSGs by lung macrophages and DCs was studied in vivo. Furthermore, PSGs were instilled in Balb/c mice and the effects of SP-D on total lung clearance were assessed by optical imaging. SP-D treatment increased the number of PSG-positive macrophages and DCs in vitro. Furthermore, SP-D accelerated uptake/binding by alveolar macrophages and reduced the number of PSG-positive tissue macrophages and DCs at 24 hours. However, SP-D did not affect total lung clearance of PSGs and it did not enhance the T-cell proliferation induced by PSG-positive DCs. In conclusion, SP-D increased PSG-positive cells in vitro and accelerated PSG binding/uptake in vivo. The observed effects were limited to cellular clearance mechanisms and did not affect the total clearance of PSGs from the lung.

Background: Allergen-containing subpollen particles (SPP) are released from whole plant pollen upon contact with water or even high humidity. Because of their size SPP can preferentially reach the lower airways where they come into contact with surfactant protein (SP)-D. Our previous work demonstrated that SP-D increases the uptake of SPP by alveolar macrophages. In the present study, we investigated the uptake of SPP in human primary epithelial cells and the potential modulation by SP-D. The patho-physiological consequence was evaluated by measurement of pro-inflammatory mediators., Methods: SPP were isolated from timothy grass and subsequently fluorescently labelled. Human primary bronchial epithelial cells were incubated with SPP or polystyrene particles (PP) in the presence and absence of surfactant protein D. In addition, different sizes and surface charges of the PP were studied. Particle uptake was evaluated by flow cytometry and confocal microscopy. Soluble mediators were measured by enzyme linked immunosorbent assay or bead array., Results: SPP were taken up by primary epithelial cells in a dose dependent manner. This uptake was coincided with secretion of Interleukin (IL)-8. SP-D increased the fraction of bronchial epithelial cells that bound SPP but not the fraction of cells that internalized SPP. SPP-induced secretion of IL-8 was further increased by SP-D. PP were bound and internalized by epithelial cells but this was not modulated by SP-D., Conclusions: Epithelial cells bind and internalize SPP and PP which leads to increased IL-8 secretion. SP-D promotes attachment of SPP to epithelial cells and may thus be involved in the inflammatory response to inhaled allergen.

Background: Pulmonary surfactant reduces surface tension and is present at the air-liquid interface in the alveoli where inhaled nanoparticles preferentially deposit. We investigated the effect of titanium dioxide (TiO(2)) nanosized particles (NSP) and microsized particles (MSP) on biophysical surfactant function after direct particle contact and after surface area cycling in vitro. In addition, TiO(2) effects on surfactant ultrastructure were visualized., Methods: A natural porcine surfactant preparation was incubated with increasing concentrations (50-500 microg/ml) of TiO(2) NSP or MSP, respectively. Biophysical surfactant function was measured in a pulsating bubble surfactometer before and after surface area cycling. Furthermore, surfactant ultrastructure was evaluated with a transmission electron microscope., Results: TiO(2) NSP, but not MSP, induced a surfactant dysfunction. For TiO(2) NSP, adsorption surface tension (gammaads) increased in a dose-dependent manner from 28.2 + or - 2.3 mN/m to 33.2 + or - 2.3 mN/m (p < 0.01), and surface tension at minimum bubble size (gammamin) slightly increased from 4.8 + or - 0.5 mN/m up to 8.4 + or - 1.3 mN/m (p < 0.01) at high TiO(2) NSP concentrations. Presence of NSP during surface area cycling caused large and significant increases in both gammaads (63.6 + or - 0.4 mN/m) and gammamin (21.1 + or - 0.4 mN/m). Interestingly, TiO(2) NSP induced aberrations in the surfactant ultrastructure. Lamellar body like structures were deformed and decreased in size. In addition, unilamellar vesicles were formed. Particle aggregates were found between single lamellae., Conclusion: TiO(2) nanosized particles can alter the structure and function of pulmonary surfactant. Particle size and surface area respectively play a critical role for the biophysical surfactant response in the lung.

Pulmonary surfactant is a complex mixture of lipids and proteins that reduces the surface tension at the air-liquid interface. In addition to its biophysical function, some surfactant components play an important role for the innate and adaptive immunity of the lung. A negative modulation of the surfactant function was observed in allergic asthma leading to the assumption that the therapeutic application of surfactant components might be beneficial in this disease. So far, there are a number of preclinical and already some clinical studies demonstrating various effects of different surfactant components that were administered with preventive or therapeutic aim in allergic asthma. This review summarizes the current knowledge on the possibilities to treat allergic asthma with surfactant components.

Background: There is increasing interest in testing surfactant preparations for asthma therapy. Previously, Curosurf was demonstrated to increase inflammation in allergic asthmatics. So far, little is known about the immunomodulatory effects of therapeutic surfactants, in particular concerning the interaction of surfactant components with eosinophils as key effector cells of the allergic airway inflammation. The aim of the present study was to determine the effect of different therapeutic surfactants on cellular functions of eosinophils., Methods: Eosinophils were isolated from peripheral blood of atopic volunteers and incubated with the natural animal-derived surfactants Curosurf or Alveofact or the synthetic recombinant human surfactant Venticute at different concentrations for up to 42 h., Results: Curosurf and Venticute modulated the viability of eosinophils. While incubation with Curosurf increased the number of necrotic eosinophils after 1, 20 and 42 h, Venticute increased the number of apoptotic and necrotic cells after 1 h, but there were no differences compared with control cells at later time points. All surfactant preparations increased the levels of eosinophil cationic protein after 20 h and, in addition, Curosurf enhanced eosinophil cationic protein release after 42 h. The supernatant of eosinophils induced chemotaxis against autologous eosinophils, and the presence of Curosurf, but not Alveofact or Venticute, augmented the chemotactic effect. Chemotaxis was partly blocked by inhibition of eotaxin but not by inhibition of leukotrienes or platelet-activating factor., Conclusions: Therapeutic surfactants differ in their effects on eosinophil viability and the accompanying release of inflammatory mediators and chemotactic signals. Proinflammatory effects were most pronounced for the natural surfactant Curosurf., (Copyright (C) 2009 S. Karger AG, Basel.)

Background: Cytochrome P450 monooxygenases (CYP) play an important role in the defense against inhaled toxicants, and expression of CYP enzymes may differ among various lung cells and tissue compartments., Methods: We studied the effects of tobacco smoke in volunteers and investigated gene expression of 19 CYPs and 3 flavin-containing monooxygenases, as well as isoforms of glutathione S-transferases (GST) and uridine diphosphate glucuronosyltransferases (UGT) and the microsomal epoxide hydrolase (EPHX1) in bronchoalveolar lavage cells and bronchial biopsies derived from smokers (n = 8) and nonsmokers (n = 10). We also investigated gene expression of nuclear transcription factors known to be involved in the regulation of xenobiotic metabolism enzymes., Results: Gene expression of CYP1A1, CYP1B1, CYP2S1, GSTP1, and EPHX1 was induced in bronchoalveolar lavage cells of smokers, whereas expression of CYP2B6/7, CYP3A5, and UGT2A1 was repressed. In bronchial biopsies of smokers, CYP1A1, CYP1B1, CYP2C9, GSTP1, and GSTA2 were induced, but CYP2J2 and EPHX1 were repressed. Induction of CYP1A1 and CYP1B1 transcript abundance resulted in increased activity of the coded enzyme. Finally, expression of the liver X receptor and the glucocorticoid receptor was significantly up-regulated in bronchoalveolar lavage cells of smokers., Conclusions: We found gene expression of pulmonary xenobiotic metabolizing enzymes and certain key transcription factors to be regulated in bronchoalveolar lavage cells and bronchial biopsies of smokers. The observed changes demonstrate tissue specificity in xenobiotic metabolism, with likely implications for the metabolic activation of procarcinogens to ultimate carcinogens of tobacco smoke.

Background: Neurotrophins have been implicated in the pathogenesis of asthma because of their ability to promote hyperreactivity of sensory neurons and to induce airway inflammation. Hyperreactivity of sensory nerves is one key mechanism of airway hyperreactivity that is defined as an abnormal reactivity of the airways to unspecific stimuli, such as cold air and cigarette smoke. Neurotrophins use a dual-receptor system consisting of Trk receptor tyrosine kinases and the structurally unrelated p75 neurotrophin receptor., Objective: The aim of this study was to characterize the distribution, allergen-dependent regulation, and functional relevance of the Trk receptors in allergic asthma., Methods: BALB/c mice were sensitized to ovalbumin. After provocation with ovalbumin or vehicle aerosol, respectively, Trk receptor expression was analyzed in lung tissue by means of fluorescence microscopy and quantitative RT-PCR. To assess the functional relevance of Trk receptors in asthma, we tested the effects of the intranasally administered pan-Trk receptor decoy REN1826. Allergic airway inflammation was quantified and lung function was measured by using head-out body plethysmography., Results: Trk receptors were expressed in neurons, airway smooth muscle cells, and cells of the inflammatory infiltrate surrounding the bronchi and upregulated after allergen challenge. Local application of REN1826 reduced IL-4 and IL-5 cytokine levels but had no effect on IL-13 levels or the cellular composition of bronchoalveolar lavage fluid cells. Furthermore, REN1826 decreased broncho-obstruction in response to sensory stimuli, indicating a diminished hyperreactivity of sensory nerves, but did not influence airway smooth muscle hyperreactivity in response to methacholine., Conclusion: These results emphasize the important role of Trk receptor signaling in the development of asthma., Clinical Implications: Our data indicate that blocking of Trk receptor signaling might reduce asthma symptoms.

Background: The surfactant protein SP-D has been reported to reduce bronchial hyper-responsiveness, blood eosinophilia, and T-helper type 2 cytokines in models of allergic asthma. However, little is known about the functional effect of SP-D on the early airway response upon allergen inhalation, which is an important feature of this disease., Objective: We investigated whether SP-D is able to reduce the immediate allergen-induced mediator release and the early bronchial obstruction in addition to its effects on airway inflammation and bronchial hyperresponsiveness in an Aspergillus fumigatus mouse asthma model., Methods: A. fumigatus-sensitized mice were treated with a recombinant fragment of human SP-D or placebo. Lung functions were measured in orotracheally intubated, spontaneously breathing animals using body plethysmography. In addition, passively sensitized precision-cut lung slices (PCLS) were used to determine the effect of SP-D on allergen-induced histamine release., Results: SP-D inhibited the allergen-induced early airway response and reduced airway hyperresponsiveness compared with placebo. Eosinophilia in bronchoalveolar lavage and lung tissue was reduced after SP-D treatment, possibly by reducing eotaxin levels in the lung. Furthermore, SP-D treatment reduced the allergen-induced histamine release from PCLS., Conclusion: These data suggest that SP-D not only reduces allergen-induced eosinophilic inflammation and airway hyperresponsiveness but also provides protection against early airway obstruction by inhibition of early mediator release.

Background: Allergic asthma is associated with airway inflammation and dysfunction of pulmonary surfactant. Because surfactant proteins (SP) account for immunomodulatory functions as well as biophysical functions, we hypothesized that the allergic response in asthma might be accompanied by a dysregulation of SPs., Methods: We measured levels of SP-A, SP-B, SP-C and SP-D by enzyme-linked immunosorbent assay in bronchoalveolar lavage (BAL) fluid of 23 asthma patients and 10 healthy control subjects under well-controlled conditions before and 24 h after segmental allergen provocation. These data were related to surfactant function, Th(2) cytokine levels in BAL fluid and to the degree of eosinophilic inflammation., Results: In patients with asthma, allergen challenge increased BAL levels of SP-B, SP-C and SP-D while SP-A was decreased. For SP-B and SP-D, a moderate increase was also observed after saline challenge. In contrast, no alterations were observed in healthy control subjects. Levels of SP-B and SP-C in asthmatics correlated with the ratio of small to large surfactant aggregates (SA/LA ratio) and correlated negatively with BAL surface activity. Furthermore, increased SP-C but not SP-B levels after allergen challenge correlated with eosinophil numbers, interleukin (IL)-5, and IL-13 in BAL while increased SP-D levels only correlated with eosinophil numbers., Conclusions: This study demonstrates significant alterations of all SPs in BAL fluid after allergen challenge of which SP-C was most closely related to surfactant dysfunction and the degree of the allergic inflammation.

Background: GATA-3 (GATA binding protein 3) and the proto-oncogene c-maf are Th2-regulating transcription factors that control the expression of interleukin (IL)-4 and IL-5, while T-bet (T-box expressed in T cells) is a Th1-specific transcription factor that controls the expression of interferon (IFN-gamma). Allergen provocation in asthmatics induces a Th2-dominated cytokine profile, but so far it is unknown whether the skewed cytokine expression is reflected by the expression of the respective transcriptional regulators., Objective: The aim of this study was to determine the regulation of Th1- and Th2-specific transcription factors and cytokines in 10 atopic subjects with mild asthma and 5 nonatopic healthy controls at baseline and after segmental sham and allergen challenge., Methods: The mRNA expression of GATA-3, c-maf and T-bet was determined by real-time polymerase chain reaction in bronchoalveolar lavage (BAL) cells and bronchial biopsies. The percentage of IL-4+, IL-5+ and IFN-gamma+ BAL T cells was determined by flow cytometry, and BAL levels of these cytokines were measured by ELISA., Results: In BAL cells of asthmatics, the mRNA expression of all transcription factors was increased after allergen challenge. In bronchial biopsies, the basal expression of GATA-3 was increased in asthmatics compared to healthy controls but decreased after allergen challenge. Compared to sham challenge, the percentage of IL-5+/CD4+ BAL T cells was increased after allergen challenge in asthmatics while the percentage of IFN-gamma+/CD4+ and IFN-gamma+/CD8+ T cells was decreased. Expression of c-maf mRNA in BAL cells correlated with IL-4+/CD4+ BAL cells and BAL IL-5 levels., Conclusions: Segmental allergen challenge in asthmatics leads to increased GATA-3, c-maf and T-bet expression in BAL cells but not in bronchial biopsies., (Copyright (c) 2006 S. Karger AG, Basel.)

Mast cells play a key role in allergy and asthma. They reside at the host-environment interface and are among the first cells to make contact with inhaled microorganisms and particulate antigens. Pulmonary surfactant proteins A and D (SP-A and SP-D) function in lung host defense by enhancing microbe phagocytosis and mediating other immune cell functions, but little is known about their effects on mast cells. We hypothesized that SP-A and/or SP-D modulate IgE-dependent mast cell functions. Pollen starch granules (PSG) extracted from Dactylis glomerata and coated with trinitrophenol (TNP) were used as a model of an inhaled organic particulate allergen. Our data revealed that SP-D inhibited by 50% the release of beta-hexosaminidase by peritoneal mast cells sensitized with IgE anti-TNP and stimulated with TNP-PSG. In contrast, SP-A had no effect. Furthermore, SP-D aggregated PSG in a dose-dependent manner, and this aggregation was mediated by SP-D's carbohydrate recognition domain. A single arm SP-D mutant (RrSP-Dser15,20) neither aggregated PSG nor inhibited degranulation, suggesting that multimerization of SP-D is required for maximal PSG aggregation and inhibition of PSG-induced mast cell degranulation. This study is the first to demonstrate that SP-D modulates IgE-mediated mast cell functions, which are important in asthma and allergic inflammation.

Nitric oxide (NO) levels are increased in the exhaled air of asthmatics. As NO levels correlate with allergic airway inflammation, NO measurement has been suggested for disease monitoring. In patients with asthma, we previously demonstrated that intrabronchial treatment with a natural porcine surfactant enhanced airway inflammation after segmental allergen provocation. We studied whether local levels of NO reflect the degree of allergic airway inflammation following segmental allergen challenge with or without surfactant pretreatment. Segmental NO, as well as nitrite and nitrate in bronchoalveolar lavage (BAL) fluid, was measured before and after segmental challenge with either saline, saline plus allergen, or surfactant plus allergen in 16 patients with asthma and five healthy subjects. The data were compared with inflammatory BAL cells. Segmental NO levels were increased after instillation of saline (p < 0.05), or surfactant plus allergen in asthmatics (p < 0.05), and values were higher after surfactant plus allergen compared to saline challenge. Nitrate BAL levels were not altered after saline challenge but increased after allergen challenge (p < 0.05) and further raised by surfactant (p < 0.05), whereas nitrite levels were not altered by any treatment. Segmental NO and nitrate levels correlated with the degree of eosinophilic airway inflammation, and nitrate levels also correlated with neutrophil and lymphocyte numbers in BAL. In healthy subjects, NO, nitrite, and nitrate were unaffected. Thus, segmental NO and nitrate levels reflect the degree of allergic airway inflammation in patients with asthma. Measurement of both markers can be useful in studies using segmental allergen provocation, to assess local effects of potential immunomodulators.

Previous studies indicated potent inhibitory effects of beta2-adrenoceptor (beta2AR) activation on the immunological mediator release of mast cells (MC). Here, we studied effects of beta2AR agonists on human MC mediator release, and in particular on MC proliferation, adhesion, and migration. MC were isolated from human intestinal mucosa, purified, and cultured in the presence of stem cell factor (SCF). beta2AR activation by epinephrine, norepinephrine, and salbutamol suppressed the IgE receptor-dependent release of histamine, lipid mediators, and TNF-alpha, and inhibited SCF-dependent MC proliferation and migration. Moreover, beta2-adrenergic stimulation interfered with MC adhesion to fibronectin and human endothelial cells. Using fluorescent phallacidin, we found that beta2AR activation reduced the amount of filamentous actin (F-actin) within minutes, whereas MC stimulation by either IgE receptor cross-linking or SCF caused F-actin accumulation. Interestingly, this activation-induced F-actin increase was abolished by previous beta2-adrenergic stimulation. Finally, we demonstrated that disruption of the F-actin cytoskeleton by latrunculin B mimicked the effects of beta2AR agonists on MC adhesion and migration. Our results argue for an important role of F-actin interference in beta2AR-mediated MC inhibition. Furthermore, the data support the concept of neuroimmune interactions regulating intestinal MC distribution, density, and functionality in vivo.

Recent studies have shown that surfactant components, in particular the collectins surfactant protein (SP)-A and -D, modulate the phagocytosis of various pathogens by alveolar macrophages. This interaction might be important not only for the elimination of pathogens but also for the elimination of inhaled allergens and might explain anti-inflammatory effects of SP-A and SP-D in allergic airway inflammation. We investigated the effect of surfactant components on the phagocytosis of allergen-containing pollen starch granules (PSG) by alveolar macrophages. PSG were isolated from Dactylis glomerata or Phleum pratense, two common grass pollen allergens, and incubated with either rat or human alveolar macrophages in the presence of recombinant human SP-A, SP-A purified from patients suffering from alveolar proteinosis, a recombinant fragment of human SP-D, dodecameric recombinant rat SP-D, or the commercially available surfactant preparations Curosurf and Alveofact. Dodecameric rat recombinant SP-D enhanced binding and phagocytosis of the PSG by alveolar macrophages, whereas the recombinant fragment of human SP-D, SP-A, or the surfactant lipid preparations had no effect. In addition, recombinant rat SP-D bound to the surface of the PSG and induced aggregation. Binding, aggregation, and enhancement of phagocytosis by recombinant rat SP-D was completely blocked by EDTA and inhibited by d-maltose and to a lesser extent by d-galactose, indicating the involvement of the carbohydrate recognition domain of SP-D in these functions. The modulation of allergen phagocytosis by SP-D might play an important role in allergen clearance from the lung and thereby modulate the allergic inflammation of asthma.

The cytokine IL-6 acts via a specific receptor complex that consists of the membrane-bound IL-6 receptor (mIL-6R) or the soluble IL-6 receptor (sIL-6R) and glycoprotein 130 (gp130). In this study, we investigated the role of IL-6R components in asthma. We observed increased levels of sIL-6R in the airways of patients with allergic asthma as compared to those in controls. In addition, local blockade of the sIL-6R in a murine model of late-phase asthma after OVA sensitization by gp130-fraction constant led to suppression of Th2 cells in the lung. By contrast, blockade of mIL-6R induced local expansion of Foxp3-positive CD4+CD25+ Tregs with increased immunosuppressive capacities. CD4+CD25+ but not CD4+CD25- lung T cells selectively expressed the IL-6R alpha chain and showed IL-6-dependent STAT-3 phosphorylation. Finally, in an in vivo transfer model of asthma in immunodeficient Rag1 mice, CD4+CD25+ T cells isolated from anti-IL-6R antibody-treated mice exhibited marked immunosuppressive and antiinflammatory functions. IL-6 signaling therefore controls the balance between effector cells and Tregs in the lung by means of different receptor components. Furthermore, inhibition of IL-6 signaling emerges as a novel molecular approach for the treatment of allergic asthma.

There is increasing evidence for a role of pulmonary surfactant in asthma and allergic inflammation. In murine asthma models, recent studies have demonstrated that surfactant components downregulate the allergic inflammation. Therefore, we tested the hypothesis that in individuals with mild asthma, a natural porcine surfactant preparation (Curosurf) given before segmental allergen challenge can reduce the allergic airway inflammation. Ten patients with asthma and five healthy control subjects were treated in two segments with either Curosurf or vehicle followed by local allergen challenge. Six additional patients with asthma received Curosurf before allergen challenge in one segment as above, but the second segment was instilled with Curosurf without allergen challenge. Unexpectedly, surfactant treatment augmented the eosinophilic inflammation 24 hours after allergen challenge. A direct chemotactic effect of Curosurf was excluded. However, levels of eotaxin and interleukin-5 were increased in bronchoalveolar lavage after Curosurf treatment, whereas IFN-gamma-levels and numbers of IFN-gamma(+) T cells were decreased. Curosurf had no influence on spreading and retention of allergen determined by allergen uptake in mice. These findings demonstrate that treatment with a natural porcine surfactant results in an augmentation of the eosinophilic inflammation after allergen challenge that is more likely due to immunomodulatory effects than to biophysical properties of the surfactant.