ABSTRACT
Mucus hypersecretion contributes to the morbidity and mortality in chronic obstructive pulmonary disease (COPD), cystic fibrosis, acute asthma, and bronchiectasis. At present, no effective therapy exists for hypersecretion in any disease. Because the physiology of secretion is presented elsewhere in this meeting, this work will focus on the first therapy of hypersecretion, which involves the epidermal growth factor receptor (EGFR) cascade. The methods used in the studies discussed here are described in detail in the original publications (see references). RESULTS
EGFR is a 170 kDa membrane glycoprotein that is expressed in fetal airways and has been shown to be important in branching morphogenesis in airways (2). 315
The study of EGFR has been explored extensively in malignant tumors, where it has been implicated in cell multiplication (and thus tumor growth). Constitutive expression of EGFR is low in the airways of healthy adults (3) and in pathogen-free animals (1). Perhaps this absence of constitutive expression of EGFR made it more difficult to discover the role of the EGFR cascade in mucus cell production in the airways. There are limited studies of EGFR expression in inflammatory diseases. For example, Puddicombe et al. (4) have reported increased EGFR im-munostaining in biopsies of asthmatics, and the authors implicated this pathway in bronchial epithelial repair and “ remodeling.” Mucus cells were not implicated, and the mechanism of EGFR activation was not identified.We hypothesized that a growth factor could be involved in airway secretory cell production. We found that EGFR expression is low in airways of pathogen-free rats (1), but we discovered that stimulation of the airways with tumor necrosis factor alpha (TNFa) induces EGFR in airway epithelial cells, but mucin production was not increased (Fig. 1). However, when we stimulated EGFR with its ligands (e.g., EGF, TGFa), mucin production occurred in the following sequence: in controls, the tracheal epithelium contained few goblet or pregoblet cells. The majority of the cells were ciliated, basal, and so-called nongranulated secretory cells. When TNFa was instilled in the airways, EGFR expression occurred, but the distribution of cell types was essentially unaltered. However, activation of EGFR by EGF or TGFa produced mucins and thereby profoundly changed the distribution of epithelial cells. There was an increase in pregoblet and goblet cells and a marked decrease in the number of nongranulated secretory cells, resulting in no change in the total number of epithelial cells (Table 1). We conclude that (1) mucin-containing goblet cells are derived from nongranulated secretory cells, and (2) goblet cells are formed by differentiation of precursor cells, rather than by the growth of new epithelial cells (multiplication of cells).Most importantly, mucin production by TNFa followed by the EGFR ligand TGFa was inhibited dose-dependently by pretreatment with a selective inhibitor of EGFR tyrosine kinase (Fig. 2A). From these studies we conclude that the activation of EGFR causes mucin gene and protein expression. The findings were the first to provide a mechanism and a strategy for therapy in hypersecretory diseases. Role of EGFR Cascade in Asthmatic Hypersecretion
Mucus hypersecretion contributes to asthma mortality (7-9). When we discovered that EGFR activation induces mucin production in airways, we hypothesized that this pathway could be involved in hypersecretion in asthma, so we investigated its role in experimental asthma induced in pathogen-free rats. Intraperito-neal injection of ovalbumin (OVA) alone did not increase the number of goblet cells, but when this was followed by airway instillation of OVA (“ active sensitization” ), the number of goblet cells was increased markedly (Fig. IB); the num-
ber of ciliated and basal cells was unchanged (Table 1). Immunohistochemical studies with an antibody to EGFR showed no staining in control tracheas, but actively sensitized rats showed EGFR staining in the cells that stained positively with Alcian blue/PAS (a stain for mucus glyconconjugates). Because OVA induced EGFR expression, we studied the effect of pretreatment with a selective EGFR inhibitor, BIBX1522. The inhibitor completely prevented OVA-induced goblet cell growth (1). From these results we conclude that inhibition of EGFR prevents goblet cell production due to allergen in vivo.
