In addition to a direct proinflammatory role, IL-13 has been demonstrated to induce a goblet cell metaplastic phenotype in the airway epithelium in vivo. We have studied the direct effects of IL-13 (and IL-4) on well-differentiated, air-liquid interface cultures of human bronchial epithelial cells (HBEs) and provide a quantitative assessment of the development of a mucus hypersecretory phenotype induced by these cytokines. Using Alcian blue staining of goblet cells and immunohistochemical detection of MUC5AC, we found that IL-13 (and IL-4) induced increases in the goblet cell density (GCD) of the HBE cultures. The effects of these cytokines were critically dependent on concentration: 1 ng/ml routinely induced a 5- to 10-fold increase in GCD that was associated with a hypersecretory ion transport phenotype. Paradoxically, 10 ng/ml of either cytokine induced a profound reduction in GCD. Removal of EGF from the culture media or treatment of the cells with AG-1478 [a potent inhibitor of EGF receptor tyrosine kinase (EGFR-TK)] demonstrated that the EGFR-TK pathway was key to the regulation of the basal GCD but that it was not involved in the IL-13-driven increase. The IL-13-driven increase in GCD was, however, sensitive to inhibition of MEK (PD-98059, U-0126), p38 MAPK (SB-202190), and phosphatidylinositol (PtdIns) 3-kinase (LY-294002). These data support the concept that IL-13 is in part able to induce a mucus hypersecretory phenotype through a direct interaction with the airway epithelium and that MAP kinase and PtdIns 3-kinase signaling pathways are involved.

We have investigated the effects of IL-13 on the ion transport characteristics of human bronchial epithelial cells cultured at an apical-air interface. Ussing chamber studies indicated that 48 h pretreatment with IL-13 or IL-4 significantly reduced the basal short-circuit current (Isc) and inhibited the amiloride-sensitive current by Ͼ98%. Furthermore, the Isc responses were increased by more than six-and twofold over control values when stimulated with UTP or forskolin, respectively, after cytokine treatment. The IL-13-enhanced response to UTP/ionomycin was sensitive to bumetanide and DIDS and was reduced in a low-chloride, bicarbonate-free solution. Membrane permeablization studies indicated that IL-13 induced the functional expression of an apical Ca 2ϩ -activated anion conductance and that changes in apical or basolateral K ϩ conductances could not account for the increased Isc responses to UTP or ionomycin. The results indicate that IL-13 converts the human bronchial epithelium from an absorptive to a secretory phenotype that is the result of loss of amiloride-sensitive current and an increase in a DIDS-sensitive apical anion conductance.calcium-activated chloride channel; hypersecretion; asthma; interleukin-4 THE AIRWAY EPITHELIUM acts as a barrier protecting the lung from inhaled substances and has developed specifically for this purpose. It serves to regulate airway surface liquid volume and composition, mucus secretion, and cilia beat to maintain a sterile lung through effective mucociliary clearance. The airway epithelium is also in the ideal location to interact with the immune system when it becomes exposed to potentially harmful substances (17,26). The bronchial epithelium is a tissue comprising a heterogeneous cell population, including ciliated columnar cells, goblet cells, submucosal glands, serous cells, and basal cells. There are only a few reports of the effects of inflammatory stimuli on the functioning of the intact epithelium (1,7,13,25). With the exception of submucosal glands, the bronchial epithelium can be modeled in vitro to display a differentiated mucociliary phenotype with the ion transport characteristics of the native tissue. To date, there is only one report of the effects of inflammatory stimuli on the ion transport function of the human airway epithelium (13). Galietta and colleagues (13) described the effects of the T-helper (Th) 1 cytokines interferon-␥ (IFN-␥) and tumor necrosis factor-␣ (TNF-␣) on the ion transport characteristics of human bronchial epithelial cells (HBECs) and demonstrated that TNF-␣ was without effect, although the basal amiloride-sensitive short-circuit current (I sc ) was reduced by IFN-␥ and agonist-stimulated anion-secretion was enhanced.Currently, there are no published reports of the effects of Th2 cytokines on the ion transport characteristics of the human airway epithelium. In this study, we report the effects of the Th2 cytokine interleukin (IL)-13 on the ion transport phenotype of the human bronchial epithelium. Increased IL-13 production is rec...

. Characterization of cigarette smoke-induced inflammatory and mucus hypersecretory changes in rat lung and the role of CXCR2 ligands in mediating this effect.

Measurement of the magnitude and kinetics of exocytosis from intact epithelia has historically been difficult. Using well-differentiated cultures of human bronchial epithelial cells, we describe the use of transepithelial impedance analysis to enable the real-time quantification of mucin secretagogue-induced changes in membrane capacitance (surface area) and conductance. ATPgammaS, UTP, ionomycin, and PMA induced robust increases in total cellular capacitance that were demonstrated to be dominated by a specific increase in apical membrane surface area. The UTP-induced increase in capacitance occurred in parallel with goblet cell emptying and the secretion of mucin and was associated with decreases in apical and basolateral membrane resistances. The magnitude and kinetics of the capacitance increases were dependent on the agonist and the sidedness of the stimulation. The peak increase in capacitance induced by UTP was approximately 30 mucin granule fusions per goblet cell. Secretagogue-induced decreases in apical membrane resistance were independent of exocytosis, although each of the secretagogues induced profound reductions in basolateral membrane resistance. Transepithelial impedance analysis offers the potential to study morphological and conductance changes in cultured human bronchial epithelial cells.