Mucous cell hyperplasia is common in patients with inflammatory airway diseases including asthma, COPD, bronchiectasis, and cystic fibrosis. Despite the contribution of increased mucus to airway obstruction and bacterial infection, mechanisms regulating mucous cell hyperplasia have not been elucidated. Herein, we propose that interleukin 13 (IL-13) is a key regulatory molecule controlling development of mucous cell hyperplasia. IL-13 is released by inflammatory cells at sites of injury or inflammation. It binds surface receptors of airway epithelial cells, and rapidly induces differentiation to a mucous phenotype, followed by proliferation of the mucous-producing cells. This proliferation is controlled by an IL-13-initiated intracellular signaling cascade in which phosphatidylinositol 3' kinase (Pl 3' kinase) plays a central role. The specific hypothesis to be addressed is that IL-13 elicits mucous cell proliferation via two pathways that converge to activate a single enzyme, phosphatidylinositol 3' kinase (Pl 3' kinase). These pathways include the autocrine/paracrine interaction of transforming growth factor alpha (TGF-a) with the epidermal growth factor receptor (EGF-R), and the IL-13 receptor-mediated activation of the insulin receptor substrate 2 (IRS-2). Enhanced Fl 3' kinase enzymatic activity stimulates proliferation via activation of downstream regulatory cell-cycle enzymes (e.g. p70-S6k, cdk2), resulting in an increase of mucus-producing cells. Normal human bronchial epithelial (NHBE) cells will be used as an in vitro model to address the above hypothesis. Specific culture conditions will maintain these cells so as to recreate injured epithelium in early stages of regeneration, or to model cells in the differentiated state utilizing air/liquid interface cultures. The experiments will address the hypothesis by determining: whether IL- 13 induces differentiation to a mucous phenotype [AIM 1] as well as proliferation of epithelial cells [AIM 2]; the proliferative requirement for both TGF-a acting on the EGF-R and activation of IRS-2 [AIM 3]; the central role for P1 3' kinase in this proliferation [AIM 4]; and, preliminarily, the intracellular mechanism(s) governing P1 3' kinase induced proliferation: phosphorylation, translocation, interaction with scaffolding proteins, and activation of cell cycle-regulating enzymes [AIM 5]. These studies will demonstrate novel intracellular mechanisms governing development of mucous cell hyperplasia in human airway epithelial cells.
