Julia Barkans

Chronic hyperplastic sinusitis: association of tissue eosinophilia with mRNA expression of granulocyte-macrophage colony-stimulating factor and interleukin-3.

BACKGROUND We investigated the association among tissue eosinophilia, cellular infiltration, and cytokine mRNA expression in chronic hyperplastic sinusitis (CHS). METHODS Percutaneous biopsies of the maxillary sinuses and nasal polyps were performed in 12 adult patients (six men and six women) of whom seven were nonallergic and 11 were asthmatic. Tissues were compared with biopsy specimens from the inferior and middle turbinates of normal control subjects. RESULTS Histologically, an eosinophil-predominant inflammatory infiltrate was seen in 10 of 12 patients, whereas a mild to moderate neutrophilic infiltrate was seen in 4 of 12 patients. As determined by immunocytochemistry, diseased tissues and normal control tissues differed significantly in terms of the number of activated (EG2+) eosinophils (p = 0.005) but not in terms of CD3+ or CD4+ T lymphocytes, elastase-positive neutrophils or CD68+ macrophages. The number of eosinophils did not correlate with that of any other cell type. By in situ hybridization, CHS tissues showed significantly higher numbers of granulocyte-macrophage colony-stimulating factor (GM-CSF) and interleukin (IL)-3 mRNA-positive cells than normal control tissues (p = 0.002 and 0.0005, respectively) per high-powered field. There was a significant correlation between the number of infiltrating EG2+ eosinophils and cells that expressed mRNA for GM-CSF (r = 0.60, p = 0.041) or IL-3 (r = 0.69, p = 0.013). Furthermore, epithelial cells did not show detectable mRNA expression for GM-CSF or IL-3. No significant correlation was found between IL-5 mRNA expression and infiltrating EG2+ eosinophils in diseased tissues. However, the IL-5 density was significantly higher in the five patients with CHS who had positive allergy skin test results than in the seven patients with negative skin test results (p = 0.017) or in normal control subjects. CONCLUSIONS Our data support a role for GM-CSF and IL-3 in the eosinophilia characteristic of CHS and show that IL-5 mRNA expression is not a prominent feature of nonallergic inflammation. The cellular sources of GM-CSF and IL-3 in CHS remain to be definitely determined.

Kinetics of cell infiltration and cytokine messenger RNA expression after intradermal challenge with allergen and tuberculin in the same atopic individuals

BACKGROUND Previous studies, in which one time point was used, have shown that cells infiltrating skin biopsy specimens taken during allergen-induced late-phase responses (LPR) had a TH2-like (interleukin-4 [IL]-4 and IL-5 mRNA+) cytokine profile, whereas in delayed-type hypersensitivity (DTH) there was a predominant TH1-type pattern. OBJECTIVE The study was designed to examine the kinetics of accumulation of inflammatory cells and cells expressing mRNA for TH2- or TH1-type cytokines in LPR and DTH elicited simultaneously in the same subjects. METHODS Immunocytochemistry (alkaline phosphatase anti-alkaline phosphatase technique) and in situ hybridization were used to analyze skin biopsy specimens taken during allergen-induced LPR. RESULTS In LPR elevated numbers of CD3+ and CD4+ cells, eosinophils, neutrophils, and IL-4 and IL-5 mRNA+ cells were detected as early as 1 hour after allergen challenge, with a peak at 6 hours, which was maintained for up to 96 hours. A small but significant delayed increase in macrophages, CD8+ and CD25+ cells, and IL-2 and interferon-gamma mRNA+ cells was also observed, but only at the 48-hour and 96-hour time points. In contrast, in DTH the numbers of CD3+, CD4+, and mRNA+ cells for IL-2 and interferon-gamma were not elevated until 24 hours after challenge and peaked at 48 hours after injection. At 48 hours there was an additional small but significant increase in IL-4 and IL-5 mRNA+ cells. For both LPR and DTH the kinetics of the increases in inflammatory cells and cytokine mRNA-expressing cells paralleled the clinical response. CONCLUSIONS In LPR accumulation of T cells and granulocytes, together with cells expressing mRNA encoding for TH2-type cytokines, is relatively rapid (i.e., within 1 to 6 hours), whereas in DTH the T cell/macrophage infiltration and appearance of cells expressing TH1-type cytokines are not apparent until 24 to 48 hours. In LPR there is a TH1-type (or possibly TH0) component at 48 to 96 hours, and in DTH there is an additional TH2/TH0 response.

Application of monoclonal antibodies against major basic protein (BMK-13) and eosinophil cationic protein (EG1 and EG2) for quantifying eosinophils in bronchial biopsies from atopic asthma.

A monoclonal antibody prepared against the eosinophil major basic protein (MBP) was compared with the anti‐eosinophil cationic protein (ECP) antibodies (Eg1 and EG2) in immunostaining of bronchial biopsies from atopic asthma and controls. Anti‐MBP (designated BMK‐13) did not cross‐react with other eosinophil basic proteins (i.e. ECP, eosinophil peroxidase [EPO] or eosinophil‐derived neurotoxin [EDN]) and stained more than 98% of peripheral blood eosinophils irrespective of their degree of activation. EG2 stained 15% of resting and 75% of activated eosinophils; EG1 recognized 74% and 78% of resting and activated cells, respectively.

Activin and transforming growth factor-β signaling pathways are activated after allergen challenge in mild asthma

BACKGROUND Both transforming growth factor (TGF)-beta(1) and activin-A have been implicated in airway remodeling in asthma, but the modulation of their specific signaling pathways after disease activation remains undefined. OBJECTIVE To define the expression kinetics of TGF-beta(1), activin-A ligands, and follistatin (a natural activin inhibitor), their type I and type II receptors (activin-like kinase[ALK]-1, ALK-5, ALK-4, TbetaRII, and ActRIIA/RIIB) and activation of signaling (via phosphorylated (p) Smad2), in the asthmatic airway after allergen challenge. METHODS Immunohistochemistry was performed on bronchial biopsies from 15 mild atopic patients with asthma (median age, 25 years; median FEV(1)% predicted, 97%) at baseline and 24 hours after allergen inhalation. Functional effects of activin-A were evaluated by using cultured normal human bronchial epithelial (NHBE) cells. RESULTS pSmad2(+) epithelial cells increased at 24 hours (P = .03), and pSmad2 was detected in submucosal cells. No modulation of activin-A, follistatin, or TGF-beta(1) expression was demonstrated. Activin receptor(+) cells increased after allergen challenge: ALK-4 in epithelium (P = .04) and submucosa (P = .04), and ActRIIA in epithelium (P = .01). The TGF-beta receptor ALK-5 expression was minimal in the submucosa at baseline and after challenge and was downregulated in the epithelium after challenge (P = .02), whereas ALK-1 and TbetaRII expression in the submucosa increased after allergen challenge (P = .03 and P = .004, respectively). ALK-1 and ALK-4 expression by T cells was increased after allergen challenge. Activin-A induced NHBE cell proliferation, was produced by NHBE cells in response to TNF-alpha, and downregulated TNF-alpha and IL-13-induced chemokine production by NHBE cells. CONCLUSION Both TGF-beta and activin signaling pathways are activated on allergen provocation in asthma. Activin-A may contribute to resolution of inflammation.

Natural killer T cells in bronchial biopsies from human allergen challenge model of allergic asthma

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Modulation of human eosinophil chemotaxis and adhesion by anti-allergic drugs in vitro

A preponderance of eosinophils at sites of inflammation is a hallmark of helminthic infections and allergic-type diseases such as bronchial asthma (1, 2). The role of eosinophils as prime effector cells of the pathophysiology of asthma is supported by the demonstration that in vitro, activated eosinophils release granule-associated basic proteins that are cytotoxic for respiratory epithelial cells (3). In addition, eosinophils synthesize substantial amounts of lipid mediators such as leukotriene C, (LTC,) and platelet-activating factor (PAF), dti novo. These have the potential to induce the characteristic features of airway obstruction in asthma (4-6). Clinical studies in parallel have emphasized the invariable localization of activated eosinophils in bronchial biopsies from asthmatic subjects (7,8), and a direct correlation has been demonstrated between eosinophil numbers in bronchial lavage and the degree of airway hyperreactivity (9). These findings, however, are circumstantial, and further insight into the role of eosinophils in airway disease can be obtained by investigating agents that inhibit or ablate eosinophil recruitment and/or activation at sites of allergic inflammation. We have shown previously that anti-allergic agents such as sodium cromoglycate (SCG) and nedocromil sodium inhibit eosinophil activation in vitro as assessed by down-regulation of enhanced receptor expression and cytotoxicity to opsonized targets (10-12). Cetirizine, a major metabolite of hydroxyzine, is a potent and HI-receptor antagonist with a limited potential for sedation and anticholinergic side effects (13). The ability of cetirizine to attenuate eosinophil infiltration in vivo to cutaneous sites following allergen challenge ( 1416) and administration of PAF (17) suggests an anti-inflammatory mode of action that is distinct from its anti-Hi, activity. Cetirizine inhibits PAFinduced enhancement of eosinophil complement and immunogloblulin G (1gG)-dependent rossette formation and cytotoxicity to complement coated schistosomula of Schistosoma marzsoni (18). However, unlike SCG and nedocromil sodium, which have no significant effect on PAF-induced chemotaxis of eosinophils from asthmatic subjects ( 19), cetirizine inhibits eosinophilotactic responses towards PAF and formyl-methionyl leucyl phenylalanine in vitro (20,21). In addition, cetirizine selectively inhibits PAF-induced eosinophil but not neutrophil hyperadherence to plasma-coated glass (IS), suggesting a mechanism by which this drug is able to inhibit eosinophil recruitment in viivo. The adhesion of leukocytes to vascular endothelial cells represents a crucial early stage in the transvascular migration of leukocytes.

Basal Expression of Bone Morphogenetic Protein Receptor Is Reduced in Mild Asthma

RATIONALE Despite increasing recognition of bone morphogenetic protein (BMP) signaling in tissue remodeling, the expression pattern of ligands and signaling pathways remain undefined in the asthmatic airway. OBJECTIVES To determine expression of BMP ligands (BMP-2, BMP-4, and BMP-7) and type I and type II receptors (ALK-2, ALK-3, ALK-6, and BMPRII) as well as evidence for activation of BMP signaling via detection of phosphorylated Smad1/5 (pSmad1/5) expression in asthmatic airways at baseline (compared with nonasthmatic controls), and after allergen challenge. METHODS Bronchial biopsies were obtained from 6 nonasthmatic control volunteers, and 15 atopic patients with asthma (median age, 25 yr; median FEV(1)% predicted, 97%) at baseline, then at 24 hours and 7 days after allergen challenge. Expression of BMP ligands, receptors, and signaling was analyzed using immunohistochemistry. MEASUREMENTS AND MAIN RESULTS BMP ligand expression did not differ between asthmatic and control airways at baseline. Compared with the normal airway, there was significant down-regulation of ALK-2 (P = 0.001), ALK-6 (P = 0.0009), and BMPRII (P = 0.009) expression in asthma. Allergen challenge was associated with marked and sustained up-regulation of BMP-7 in airway epithelium (P = 0.017) and infiltrating inflammatory cells (P = 0.071) (predominantly in eosinophils, but also CD4(+) T cells, mast cells, and macrophages). Up-regulation of pSmad1/5 expression (P = 0.031), ALK-2 (P = 0.002), and ALK-6 (P < 0.001) was observed indicating active signaling. CONCLUSIONS BMP receptor expression is down-regulated in the asthmatic airway, which may impede repair responses. Allergen provocation increases expression of the regulatory ligand BMP-7, activates BMP signaling, and increases receptor expression, all of which may contribute to repair and control of inflammation.