CD14 expression and soluble CD14 after segmental allergen provocation in atopic asthma

Abstract

Allergic asthma is associated with the recruitment of activated inflammatory cells after allergen challenge. Surface expression of CD14 has been proposed as a marker of cell activation and differentiation. We therefore measured CD14 expression on activated macrophages and granulocytes as well as soluble CD14 (sCD14) concentrations in peripheral blood and bronchoalveolar lavage (BAL) following segmental allergen provocation (SAP) with individually standardized doses of allergen in eight patients with allergic asthma. Two segments of the right lung were challenged with allergen. Two segments of the left lung, into which saline was instilled, served as controls. CD14 expression on macrophages and granulocytes was determined by flow-cytometry and concentrations of interleukins and sCD14 were analysed by enzyme-linked immunosorbent assay (ELISA) 10 min and 18 h after challenge. Soluble CD14 concentrations remained unchanged in BAL fluid after saline challenge and 10 min after SAP, but increased significantly 18 h after SAP. Although macrophage numbers increased 18h after SAP, CD14 expression on these cells did not change. Unlike macrophages, granulocyte numbers correlated with sCD14 levels 18 h after SAP while their CD14 expression decreased significantly. Furthermore, sCD14 correlated with interleukin (IL)-13 concentrations 18 h after SAP. An increase in soluble CD14 can be observed 18 h but not 10 min after segmental allergen provocation suggesting local release of this surface antigen. Our findings imply that CD14-mediated cell activation following segmental allergen provocation could play a role in asthmatic inflammation.

IL-9 and IL-13 Production by Activated Mast Cells Is Strongly Enhanced in the Presence of Lipopolysaccharide: NF-κB Is Decisively Involved in the Expression of IL-9

“…Interestingly, inflammatory and bronchial obstructive responses to inhalation of endotoxin in asthmatic patients have been described, and it has been supposed that bacterial Ags can potentiate the action of inhalant Ags (60 -62). In asthmatics, extravasation of LPS binding protein and soluble CD14 into the bronchoalveolar compartment after allergen inhalation has also been reported, implying a role for LPS in amplification of the inflammatory response (63)(64)(65). During late asthmatic reactions, an accumulation of mast cells occurs in lung in parallel with an increased IgE level in serum that promotes IgEmediated activation of mast cells.…”

Mucosal inflammation in severe glucocorticoid-dependent asthma

“…Prior to the segmental allergen challenge, eight patients underwent an inhaled allergen challenge as previously described [17] to establish dual bronchial reactions to the inhaled allergen and to determine the individual provocative dose causing a 20% fall in FEV1 (PD20) for the respective allergen (two patients were challenged with a dose of 50 protein nitrogen unit (PNU)).…”

“…Bronchoscopy was performed as previously described [17] using an Olympus BF 1T30 (Hamburg, Germany) via the oral or nasal route following local anaesthesia with Novesine1 (Wander, Bern, Switzerland).…”

“…Determination of IL-4, IL-5, and GM-CSF IL-4 and IL-5 were assessed in BAL fluid as previously described [17]. GM-CSF in BAL fluid was assessed using monoclonal anti-human cytokine antibodies (Clone D2-23B6; PharMingen, San Diego, CA, USA) as the capture antibody and biotin-conjugated monoclonal anti-human cytokine antibodies (Clone BVD2-21C11; PharMingen) in a concentration of 2 mg .…”

“…Peripheral blood and BAL samples were processed and cells were counted by flow cytometry as described in detail previously [17]. After lysis of erythrocytes, 20 mL of either whole blood or cells from BAL were incubated in the presence of saturating concentrations of phycoerythrin (PE)-conjugated CD69 (Becton Dickinson, San Jose, CA, USA) and fluorescein isothiocyanate (FITC)-conjugated CD16 (Immunotech, Marseille, France) or PEconjugated anti-IgG (DAKO, Hamburg, Germany) and FITC-conjugated CD16, respectively, in the dark on ice for 30 min.…”