BackgroundAsthma is characterized by variable airflow obstruction, airway inflammation, airway hyper-responsiveness and airway remodelling. Airway smooth muscle (ASM) hyperplasia is a feature of airway remodelling and contributes to bronchial wall thickening. We sought to investigate the expression levels of chemokines in primary cultures of ASM cells from asthmatics vs healthy controls and to assess whether differentially expressed chemokines (i) promote fibrocyte (FC) migration towards ASM and (ii) are increased in blood from subjects with asthma and in sputum samples from those asthmatics with bronchial wall thickening.MethodsChemokine concentrations released by primary ASM were measured by MesoScale Discovery platform. The chemokine most highly expressed by ASM from asthmatics compared with healthy controls was confirmed by ELISA, and expression of its cognate chemokine receptor by FCs was examined by immunofluorescence and flow cytometry. The role of this chemokine in FC migration towards ASM was investigated by chemotaxis assays.ResultsChemokine (C-C motif) ligand 2 (CCL2) levels were increased in primary ASM supernatants from asthmatics compared with healthy controls. CCR2 was expressed on FCs. Fibrocytes migrated towards recombinant CCL2 and ASM supernatants. These effects were inhibited by CCL2 neutralization. CCL2 levels were increased in blood from asthmatics compared with healthy controls, and sputum CCL2 was increased in asthmatics with bronchial wall thickening.ConclusionsAirway smooth muscle-derived CCL2 mediates FC migration and potentially contributes to the development of ASM hyperplasia in asthma.

BackgroundFibrocytes are bone marrow–derived CD34+ collagen I–positive cells present in peripheral blood that develop α-smooth muscle actin expression and contractile activity in tissue culture. They are implicated in the pathogenesis of tissue remodeling and fibrosis in both patients with asthma and those with idiopathic pulmonary fibrosis. Targeting fibrocyte migration might therefore offer a new approach for the treatment of these diseases. Ion channels play key roles in cell function, but the ion-channel repertoire of human fibrocytes is unknown.ObjectiveWe sought to examine whether human fibrocytes express the KCa3.1 K+ channel and to determine its role in cell differentiation, survival, and migration.MethodsFibrocytes were cultured from the peripheral blood of healthy subjects and patients with asthma. Whole-cell patch-clamp electrophysiology was used for the measurement of ion currents, whereas mRNA and protein were examined to confirm channel expression. Fibrocyte migration and proliferation assays were performed in the presence of KCa3.1 ion-channel blockers.ResultsHuman fibrocytes cultured from the peripheral blood of both healthy control subjects and asthmatic patients expressed robust KCa3.1 ion currents together with KCa3.1 mRNA and protein. Two specific and distinct KCa3.1 blockers (TRAM-34 and ICA-17043) markedly inhibited fibrocyte migration in transwell migration assays. Channel blockers had no effect on fibrocyte growth, apoptosis, or differentiation in cell culture.ConclusionsThe K+ channel KCa3.1 plays a key role in human fibrocyte migration. Currently available KCa3.1-channel blockers might therefore attenuate tissue fibrosis and remodeling in patients with diseases such as idiopathic pulmonary fibrosis and asthma through the inhibition of fibrocyte recruitment.

Myofibroblasts are mesenchyme-derived cells responsible for tissue repair after injury. Resident populations of myofibroblasts are present throughout the lung. In addition, it is likely that myofibroblast progenitors (fibrocytes) can migrate to the lung from the circulation during injury. The relationship and interdependencies among myofibroblasts, fibroblasts, and myocytes within the airway wall remain poorly understood. Myofibroblasts are likely to be present in primary culture systems derived from airway wall tissue. The phenotyping of cells in such cultures is complicated by the lack of specific markers for these cell types. Important responses including migration, synthetic function, and the regulation of matrix, in the normal and asthmatic airway previously considered to be driven by airway myocytes may in fact at least in part be due to responses of myofibroblast populations.

There is evidence supporting a role for the nociceptin/orphanin FQ (N/OFQ; NOP) receptor and its endogenous ligand N/OFQ in the modulation of neurogenic inflammation, airway tone and calibre. We hypothesized that NOP receptor activation has beneficial effects upon asthma immunopathology and airway hyperresponsiveness. Therefore, the expression and function of N/OFQ and the NOP receptor were examined in healthy and asthmatic human airway tissues. The concept was further addressed in an animal model of allergic asthma. EXPERIMENTAL APPROACHNOP receptor expression was investigated by quantitative real-time PCR. Sputum N/OFQ was determined by RIA. N/OFQ function was tested using several assays including proliferation, migration, collagen gel contraction and wound healing. The effects of N/OFQ administration in vivo were studied in ovalbumin (OVA)-sensitized and challenged mice. KEY RESULTSNOP receptors were expressed on a wide range of human and mouse immune and airway cells. Eosinophils expressed N/OFQprecursor mRNA and their number correlated with N/OFQ concentration. N/OFQ was found in human sputum and increased in asthma. Additionally, in asthmatic human lungs N/OFQ immunoreactivity was elevated. NOP receptor activation inhibited migration of immunocytes and increased wound healing in airway structural cells. Furthermore, N/OFQ relaxed spasmogenstimulated gel contraction. Remarkably, these findings were mirrored in OVA-mice where N/OFQ treatment before or during sensitization substantially reduced airway constriction and immunocyte trafficking to the lung, in particular eosinophils. N/OFQ also reduced inflammatory mediators and mucin production. CONCLUSIONS AND IMPLICATIONSWe demonstrated a novel dual airway immunomodulator/bronchodilator role for N/OFQ and suggest targeting this system as an innovative treatment for asthma.