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Background: Subsets of patients with severe asthma remain symptomatic despite prolonged, high-dose glucocorticoid therapy. We hypothesized that the clinical glucocorticoid sensitivity of these asthmatics is reflected in differences in peripheral blood dendritic cell subsets., Objective: To compare peripheral blood leucocyte populations using flow cytometry at baseline and after 2 weeks of systemic glucocorticoid (steroid) treatment to identify immunological differences between steroid-sensitive (SS) and steroid-resistant (SR) asthmatics., Methods: Adult severe asthmatics (SS n = 12; SR n = 23) were assessed for their response to 2 weeks of therapy with oral prednisolone. Peripheral blood was obtained before and after therapy and stained for lymphocyte (CD3, CD19, CD4, CD8 and Foxp3) and dendritic cell markers (Lineage negative [CD3, CD14, CD16, CD19, CD20, CD56], HLA-DR+, CD304, CD11c, ILT3 and CD86)., Results: A higher median frequency of myeloid DCs (mDCs) but not plasmacytoid DCs (pDCs) was observed in the blood of SR as compared to SS asthmatics (P = .03). Glucocorticoid therapy significantly increased median B cell, but not T cell numbers in both cohorts, with a trend for increased numbers of Foxp3+ Tregs in SS (P = .07), but not SR subjects. Oral prednisolone therapy significantly reduced the median numbers and frequencies of total DCs and pDCs in both SS and SR asthmatics. Interestingly, the expression of HLA-DR and ILT3 was also reduced on pDCs in all patients. In contrast, therapy increased the median frequency of mDCs in SS, but reduced it in SR asthmatics., Conclusions: Myeloid DC frequency is elevated in SR compared with SS asthmatics, and mDC shows a differential response to oral prednisolone therapy., (© 2017 The Authors Clinical & Experimental Allergy Published by John Wiley & Sons Ltd.)

Background: Mild asthmatics who smoke cigarettes may develop unstable disease and neutrophilic infiltration of the airways, features more usually associated with severe asthmatic disease. The mechanisms giving rise to this response remain unclear., Objective: To address the hypothesis that smoking increases bronchial mucosal production of IL-17A which acts on bronchial epithelial cells directly and in concert with other environmental stimuli to induce the production of IL-6 and neutrophil chemotaxins., Methods: IL-17A, IL-8, IL-6, neutrophils and eosinophils were detected and quantified by immunohistochemistry in endobronchial biopsy sections from smoking and non-smoking asthmatics. Human tracheal epithelial cells (HTEpC) were cultured with IL-17A in the presence/absence of cigarette smoke extract (CSE) and aeroallergens lacking intrinsic protease activity, and IL-6 and IL-8 production measured in vitro., Results: Expression of IL-17A, IL-6 and IL-8 and neutrophil numbers was significantly elevated in the bronchial mucosa of the asthmatic smokers compared to the non-smokers. Expression of IL-17A correlated with that of IL-8 and neutrophil numbers. In the smoking asthmatics, eosinophil numbers also correlated with expression of IL-8 and IL-17A. Exposure of HTEpC cells to both CSE and IL-17A increased expression of IL-6 and IL-8 in a concentration-dependent and synergistic manner. Co-stimulation with CSE, IL-17A and aeroallergens further increased IL-6 and IL-8 production synergistically., Conclusions: The data support the hypothesis that asthmatic smokers develop neutrophilic inflammation of the airways propagated at least partly by smoke-induced production of IL-17A which together with smoke and other environmental stimuli acts on airways epithelial cells to induce neutrophil chemotaxins., (© 2017 John Wiley & Sons Ltd.)

Background: RELM-β has been implicated in airways inflammation and remodelling in murine models. Its possible functions in human airways are largely unknown. The aim was to address the hypothesis that RELM-β plays a role in extracellular matrix deposition in asthmatic airways., Methods: The effects of RELM-β gene deficiency were studied in a model of allergen exposure in mice sensitised and challenged with Aspergillus fumigatus (Af). RELM-β expression was investigated in bronchial biopsies from asthmatic patients. Direct regulatory effects of RELM-β on human lung fibroblasts were examined using primary cultures and the MRC5 cell line in vitro., Results: Sensitisation and challenge of wild-type mice with Af-induced release of RELM-β with a time course coincident with that of procollagen in the airways. Af-induced expression of mRNA encoding some, but not all ECM in the lung parenchyma was attenuated in RELM-β-/- mice. RELM-β expression was significantly increased in the bronchial submucosa of human asthmatics compared with controls, and its expression correlated positively with that of fibronectin and α-smooth muscle actin. In addition to epithelial cells, macrophages, fibroblasts and vascular endothelial cells formed the majority of cells expressing RELM-β in the submucosa. Exposure to RELM-β increased TGF-β1, TGF-β2, collagen I, fibronectin, smooth muscle α-actin, laminin α1, and hyaluronan and proteoglycan link protein 1 (Hapl1) production as well as proliferation by human lung fibroblasts in vitro. These changes were associated with activation of ERK1/2 in MRC5 cells., Conclusion: The data are consistent with the hypothesis that elevated RELM-β expression in asthmatic airways contributes to airways remodelling at least partly by increasing fibroblast proliferation and differentiation with resulting deposition of extracellular matrix proteins., (© 2014 John Wiley & Sons Ltd.)

Background: Interleukin-25 has been implicated in the pathogenesis of asthma from studies on human asthmatics and in murine asthma models., Objectives: In this study, we hypothesized that chronic exposure of the airways to IL-25 alone is able to induce pathogenic changes observed in animal models of asthma., Methods: We performed a detailed comparison of the dynamics of development of cellular infiltration, cytokine expression and airways remodelling and hyperresponsiveness in mice sensitized and challenged with OVA, a classical model of allergic asthma and those exposed to IL-25 alone., Results: Intranasal challenge of BALB/c mice with IL-25 alone induced a delayed (compared with OVA-challenge), predominantly eosinophilic and lymphocytic infiltration into the airways lumen, along with increased production of Th2-type cytokines, chemokines and collagen, secretion of epithelial mucus, goblet cell hyperplasia, deposition of subepithelial collagen, airways smooth muscle cell hyperplasia and angiogenesis. Correspondingly, IL-25 as well as OVA challenge both induced airways hyperresponsiveness and increased lung tissue damping. In contrast, IL-25 exposure did not increase IgE or IgG1 production., Conclusions and Clinical Relevance: The data suggest that chronic airways exposure to IL-25 alone is sufficient to induce allergen- and IgE-independent, asthma-like airways inflammation, remodelling and hyperresponsiveness in mice. Thus, IL-25 is a key molecular target in asthma, irrespective of the coexistence of IgE-dependent mechanisms., (© 2014 John Wiley & Sons Ltd.)

Several epidemiological studies have evaluated potential associations between allergy and risk of malignancy. It remains clear that the relationship between allergy and cancer is complex. Three hypotheses have been proposed to account for observed relationships: these are chronic inflammation, immunosurveillance, prophylaxis, and we propose adding a fourth: inappropriate T-helper 2 (Th2) immune skewing. Each of these attempts to explain either the increased or decreased risk of different cancer types in 'allergic' patients reported in the literature. All four hypotheses are based on known mechanisms of allergic inflammation and/or IgE antibody functions, and uphold the view of an immunological basis for the relationship between allergy and malignancies. This review summarizes and draws conclusions from the epidemiological literature examining the relationships between specific types of cancer and allergic diseases. Particular emphasis is placed on the most recent contributions to the field, and on consideration of the allergic immune mechanisms that may influence positive or negative associations., (© 2013 John Wiley & Sons Ltd.)

Background: Unlike other IL-17 family members, the Th2-derived cytokine IL-25 (IL-17E) induces (promotes) Th2 responses. One or both of the two receptors for IL-25 (IL-17RA, IL-17RB) is expressed on inflammatory cells and tissue structural cells, suggesting that in addition to promoting Th2-type inflammation IL-25 may also act on structural cells at sites of Th2-type inflammation such as in the asthmatic bronchial mucosa to promote remodelling changes., Objective: Our previous studies showed elevated expression of IL-25 and IL-17RB immunoreactivity in asthmatic airways with co-localization of the latter to endothelial cells. We therefore hypothesized that IL-25 acts on endothelial cells through this receptor to induce production of the key angiogenic and remodelling cytokine basic fibroblast growth factor (bFGF)., Methods: Polymerase chain reaction (PCR) immunocytochemistry/immunohistochemistry and ELISA were employed to detect expression of IL-17RB, IL-17RA and bFGF by human vascular endothelial cells (HUVEC) and immunoreactivity for IL-25 and bFGF in asthmatic bronchial biopsies. Receptor-blocking antibodies, PCR and an in vitro angiogenesis assay were used to investigate whether IL-25 acts on IL-17RB or IL-17RA to induce bFGF expression and angiogenesis. PCR was also employed to investigate the signalling pathways involved in IL-25-mediated bFGF expression., Results: HUVEC constitutively expressed IL-17RB, IL-17RA and bFGF. Production of the latter was further increased by IL-25, but attenuated after blockade of the IL-17RB, but not the IL-17RA receptor. Neutralization of endogenous VEGF and bFGF completely abrogated IL-25-induced angiogenesis which was also inhibited by blocking IL-17RB, but not IL-17RA. The PI3K-specific inhibitor LY294002 also completely attenuated IL-25-induced bFGF expression. Immunoreactivity for IL-25 and bFGF was elevated in the asthmatic bronchial mucosa and the expression of each correlated with the other., Conclusions and Clinical Relevance: Our data support the hypothesis that IL-25 contributes to elevated bFGF in asthmatic airways by acting on the endothelial cell IL-17RB receptor through PI3K-signalling pathways. Targeting the pathways might benefit therapy of airways remodelling., (© 2012 Blackwell Publishing Ltd.)

Background: IgE antibodies, sequestered into tissues and retained locally by the high-affinity IgE receptor, FcɛRI, on powerful effector cells such as mast cells, macrophages and eosinophils, may offer improvements in the therapy of solid tumours. The chimeric antibody, MOv18 IgE, against the human ovarian carcinoma antigen, folate receptor α (FRα), is more effective than its IgG1 counterpart in xenograft models of ovarian cancer. Although MOv18 IgE binds to a single epitope on FRα and cannot cross-link IgE receptors on basophils, there remains a risk that components in the circulation of ovarian cancer patients might cross-link FRα-MOv18-IgE-receptor-FcɛRI complexes on basophils to cause type I hypersensitivity., Objective: To assess the propensity for MOv18 used in a therapeutic setting to cause FcɛRI-mediated type I hypersensitivity., Methods: As validated readouts of the potential for MOv18 to cause FcɛRI-mediated type I hypersensitivity we measured release of a granule-stored mediator from a rat basophilic leukaemia cell line RBL SX-38 stably transfected with human tetrameric (αβγ2) FcɛRI, and induction of CD63 on blood basophils from patients with ovarian carcinoma and healthy controls ex vivo., Results: Serum FRα levels were increased in ovarian cancer patients compared with healthy controls. MOv18 IgE alone, or in the presence of its antigen recombinant human FRα, or of healthy volunteer (n=14) or ovarian carcinoma patient (n=32) sera, did not induce RBL SX-38 cell degranulation. Exposure to FRα-expressing ovarian tumour cells at target-to-effector ratios expected within tumours induced degranulation. MOv18 IgE did not induce expression of CD63 in blood basophils from either healthy volunteers (n=6), or cancer patients, despite detectable levels of circulating FRα (n=5)., Conclusion and Clinical Relevance: These encouraging data are compatible with the hypothesis that, when ovarian carcinoma patients are treated with MOv18, FcɛRI-mediated activation of effector cells occurs within the tumour mass but not in the circulation mandating, with due caution, further pre-clinical studies., (© 2011 Blackwell Publishing Ltd.)

Allergic rhinitis (AR) affects more than 20% of the population in the United Kingdom and western Europe and represents a major cause of morbidity that includes interference with usual daily activities and impairment of sleep quality. This guidance prepared by the Standards of Care Committee (SOCC) of the British Society for Allergy and Clinical Immunology (BSACI) is for the management of AR in patients that have failed to achieve adequate relief of symptoms despite treatment with intranasal corticosteroids and/or antihistamines. The guideline is based on evidence and is for use by both adult physicians and paediatricians practising allergy. During the development of these guidelines, all BSACI members were included in the consultation process using a web-based system. Their comments and suggestions were carefully considered by the SOCC. Where evidence was lacking, consensus was reached by the experts on the committee. Included in this guideline are indications and contraindications for immunotherapy, criteria for patient selection, the evidence for short- and long-term efficacy of subcutaneous and sublingual immunotherapy, and discussion on safety and the different modes of immunotherapy including, pre-seasonal and co-seasonal treatments. There are sections on children, allergen standardization, vaccines used in the United Kingdom, oral allergy syndrome, cost effectiveness of immunotherapy and practical considerations of undertaking immunotherapy including recommendations on who should undertake immunotherapy and dosing schedules. Finally, there is discussion on potential biomarkers of response to immunotherapy, the use of component-resolved diagnostics, novel approaches, alternative routes and potential areas for future research., (© 2011 Blackwell Publishing Ltd.)

Background: Concomitant methotrexate (MTX) therapy of oral corticosteroid (CS)-dependent asthmatics has been shown to spare CS therapy, but the mechanism is unknown. In a previous report, we showed that MTX increases T cell inhibition by CS. In this report we focus on effects of MTX on immunoglobulin concentrations and their possible clinical relevance., Objective: To monitor changes in circulating leucocytes and Ig in a group of these patients during MTX therapy, and to relate these changes to clinical 'response' as defined by oral CS reduction., Methods: Sixteen severe asthmatics dependent on oral prednisolone 15 (7.5-25) mg/day in addition to high dose inhaled CS were treated with MTX 15 mg intramuscularly, weekly for 28 weeks. Prednisolone dosages were maintained constant for 12 weeks then reduced systematically over the next 16 weeks provided that asthma control did not deteriorate. Patients were classified a priori as 'responders' or 'non-responders' to MTX (reduction of initial oral prednisolone requirement by >or=50% or <50%, respectively). Patients were followed-up for a further 12 weeks after MTX withdrawal. Serum Ig and differential blood leucocyte counts were measured at baseline, 12, 28 and 40 weeks., Results: MTX therapy allowed significant, but individually variable, reductions in oral prednisolone dosages (P<0.00001) without alteration of lung function or symptoms. This was associated with significant reductions in mean serum concentrations of Ig of all classes, which reversed following MTX withdrawal. Reductions in IgE and IgG were significantly greater in the MTX 'responders' as compared with 'non-responders', and changes in IgE, IgG and IgM correlated with changes in prednisolone requirements. Differential blood leucocyte counts showed no significant variation., Conclusion: MTX therapy reduced oral CS requirements in these severe asthmatics to a degree which correlated with reduced circulating Ig but not lymphopaenia, suggesting a possible cause and effect relationship. These reductions might also contribute to the documented incidence of opportunistic infection in these circumstances.

Background: Concomitant methotrexate (MTX) therapy of severe, oral corticosteroid-dependent asthmatics has been shown to be corticosteroid sparing, but the mechanism is unknown. We hypothesized that MTX therapy of these patients increases the susceptibility of their T cells to corticosteroid inhibition., Objective: To measure prednisolone inhibition of lectin-induced proliferation of peripheral blood T cells from a group of these patients before, during and following MTX therapy., Methods: Eighteen severe asthmatics (median (range) age 56 (33-68) years, FEV1 61 (38-69)% predicted, dependent on oral prednisolone 15 (7.5-25) mg/day in addition to high-dose, inhaled corticosteroids) were treated with MTX 15 mg intramuscularly, weekly for 28 weeks. After 12 weeks of MTX, oral prednisolone dosages were reduced systematically over 16 weeks, provided that asthma control did not deteriorate. Patients were followed for a further 12 weeks after MTX withdrawal. Concentration-dependent, prednisolone inhibition of lectin-induced proliferation of peripheral blood T cells was measured just prior to MTX therapy (week 1) and at weeks 12, 28 and 40, and IC50 concentrations were interpolated., Results: By week 28 of MTX therapy, patients were able to reduce oral prednisolone dosages from (median, SIQR) 15 (10-20.5) to 5.9 (1.4-9.4) mg/day (P<0.01) without alteration of lung function and symptoms, while median IC50 values for prednisolone inhibition of peripheral blood T cell proliferation were reduced from 49 (21-144) to 4 (1-9) nm (P<0.0001). These increased again to 15 (9.4-25.7) mg/day and 36 (18-67) nm, respectively, following MTX withdrawal. A correlation (P<0.01) was observed between percentage reductions in prednisolone dosages in vivo and fold changes in prednisolone IC50in vitro between weeks 12 and 28., Conclusion: This effect of MTX may at least partly account for its oral corticosteroid-sparing effect in severe asthma.

Background: Annexin-1 (ANXA1, lipocortin 1) is a pleiotrophic protein produced by many cell types including peripheral blood leucocytes. Although it has been shown to inhibit "macroscopic" inflammatory processes in animal models, its direct effects on antigen-activated human T cells have not been studied., Objective: To test the hypothesis that ANXA1-derived peptides inhibit antigen-driven prototype Th1 and Th2-type human T cell responses of clinical relevance and lectin-driven responses in vitro., Methods: Peripheral blood mononuclear cells (PBMC) were isolated from 14 atopic subjects sensitized to house dust mite allergen (Dermatophagoides pteronyssinus, Der p) and purified protein derivative (PPD) of Mycobacterium tuberculosis. PBMC (1 x 106/mL) were cultured with phytohaemagglutinin (PHA; 5 microg/mL; 4 days), Der p (25 microg/mL; 6 days), PPD (10 microg/mL, 6 days) or medium control. Two ANXA1-derived peptides, Ac2-26 and AF-2 (5-500 microM), were assessed for possible inhibition of PHA-and antigen-induced T cell proliferation (measured by 3H-thymidine uptake), while Ac2-26 was assessed for inhibition of Der p-induced interleukin (IL)-5 release and PPD-induced interferon-gamma (IFN-gamma) release (measured by ELISA). Comparison was made with dexamethasone as an established inhibitory control. Endogenous production by PBMC of cell surface-associated and intracellular ANXA1 in response to PHA, Der p and PPD in the presence and absence of dexamethasone was measured by specific ELISA., Results: Both PHA- and antigen-induced T cellular proliferation were inhibited by dexamethasone. Although neither ANXA1-derived peptide significantly altered PHA-induced proliferation, both effected concentration-dependent reductions in antigen-induced proliferation, Ac2-26 being the more potent. Peptides of identical amino acid composition to Ac2-26 and AF-2, but of random sequence, were ineffective at equivalent concentrations. In addition, Ac2-26 and dexamethasone inhibited Der p-induced IL-5 release and PPD-induced IFN-gamma release in a concentration-dependent fashion. Endogenous ANXA1 was detectable in PBMC, but at concentrations approximately 104-fold lower, in molar terms, than the effective concentrations of the exogenously added, ANXA1-derived inhibitory peptides. Endogenous production was not significantly altered by any of the T cell stimuli employed in this study, in the presence or absence of dexamethasone., Conclusion: In prototype Th1 and Th2-type human T cell responses, ANXA1-derived peptides can inhibit antigen-driven cellular proliferation and cytokine production.

Background: T-cell production of eosinophil-active cytokines (IL-5, IL-3, GM-CSF) is thought to be fundamental to asthma pathogenesis. Inhaled aeroallergens may be one important stimulus for T-cell cytokine production in asthma., Objective: To compare the potency and efficacy of the topical anti-asthma glucocorticoids beclomethasone dipropionate (BDP) and fluticasone propionate (FP) in inhibiting allergen-driven peripheral blood T-cell proliferation and production of IL-3, IL-5 and GM-CSF mRNA and protein., Methods: Peripheral blood mononuclear cells from six atopic asthmatics sensitized to house dust mite (HDM) were cultured in the presence of HDM and serial dilutions of BDP or FP in vitro. Cellular proliferation (7 days) and culture supernatant cytokine concentrations (6 days) were measured by uptake of tritiated thymidine and ELISA, respectively. Cytokine mRNA expression (24 h) was measured in three subjects using a quantitative PCR technique., Results: Both BDP and FP inhibited allergen-induced T-cell proliferation, expression of IL-3, IL-5 and GM-CSF mRNA, and secretion of the corresponding proteins in a concentration-dependent fashion. FP was considerably more potent, but not more efficacious, in exerting these actions., Conclusions: Both BDP and FP have the potential markedly to inhibit allergen-induced T-cell production of asthma-relevant cytokines. This activity is effected at the level of T-cell proliferation and cytokine gene transcription. These properties may be key features of the anti-asthma activity of these drugs. The greater potency of FP in vitro may be responsible for its greater clinical potency.

We have measured the expression of messenger ribonucleic acid (RNA) (mRNA) encoding interleukin-5 (IL-5), IL-4, IL-2 and interferon-gamma (IFN gamma) in peripheral blood mononuclear cells (PBMC) from 10 patients with acute exacerbations of asthma and nine non-asthmatic controls. Measurements were repeated in seven of the asthmatics following 7 days of oral glucocorticoid therapy. Total RNA was extracted from the PBMC, reverse transcribed using oligo-(dT) primers and aliquots of the resulting complementary DNA (cDNA) amplified using the polymerase chain reaction (PCR) in the presence of cytokine-specific primers under non-saturating conditions. PCR products were quantified on a relative basis after Southern blotting and probing with radiolabelled internal oligonucleotide probes by computer assisted densitometry of blot autoradiographs. The relative amounts of IL-5 mRNA in PBMC from the asthmatic patients prior to glucocorticoid therapy were greater (P < 0.01) than those in PBMC from non-asthmatic controls. In contrast, there were no differences in the relative amounts of IL-4, IL-2 and IFN gamma mRNA. In the asthmatics, the relative amounts of IL-5 mRNA correlated with the peripheral blood eosinophil counts (P = 0.02). After oral glucocorticoid therapy of the asthmatics, lung function improved and the relative amounts of PBMC IL-5 mRNA were reduced (P = 0.04) and no longer differed from those in PBMC from non-asthmatic controls. Glucocorticoid therapy was not associated with significant changes in the relative amounts of PBMC IL-4, IL-2 and IFN gamma mRNA. PBMC from atopic subjects contained significantly greater quantities of IL-4 mRNA (P = 0.04) but not IL-5, IL-2 and IFN gamma mRNA compared with non-atopic subjects regardless of their asthmatic status. We conclude that PBMC of patients with acute exacerbations of asthma demonstrate elevated expression of mRNA encoding IL-5, but not IL-2, IL-4 and IFN gamma and that the clinical improvement associated with glucocorticoid therapy is associated with a reduction of IL-5 mRNA expression. We further conclude that elevated expression in PBMC of mRNA encoding IL-4 is a feature of atopy but not of asthma. These observations suggest that IL-5 synthesis by activated T-lymphocytes may be relevant to the pathogenesis of asthma, and that inhibition of this release by glucocorticoids may at least partly explain their therapeutic effect in this disease.