Pulmonary infection and a robust inflammatory response are dominant clinical features of cystic fibrosis (CF), and comprise the major cause of morbidity and mortality in CF patients. It has been suggested that abnormal airway epithelial cells and abnormal immune responses collaborate and result in severe chronic lung disease. The contribution of airway epithelia to the inflammatory response in CF is being extensively studied, however much less is known about the role of primary immune cells in mediating the hyper- responsiveness observed in CF lung disease. Neither is it known if immune cells have a direct contribution to the lung phenotype or if the exaggerated inflammatory response is merely secondary to the primary epithelial defect. Recent reports suggest that CFTR may have an important role in the normal function of both macrophages and neutrophils. Additionally, our preliminary data suggest that the macrophage may play an important role in the hyper-responsiveness of the CF airway. The mechanism(s) underlying the hyper-responsiveness of CFTR-/- macrophages is not known. It is known that signal transduction in response to LPS is mediated by Toll-like receptor 4 (TLR4), which binds to LPS specifically. We have found that upon LPS stimulation, CF macrophages and express higher amounts of TLR4 on their membrane when compared to WT macrophages. In concert with accessory LPS-binding proteins including MD-2 and CD14, and TLR4 signaling plays a major role in the activation of the innate immune response to PA. These findings suggest that immune cells directly contribute to the exaggerated immune response in CF. In order to investigate this hypothesis we propose: i.) to investigate if hematopoietic engraftment of CFTR+ cells will ameliorate the hyper-inflammatory immune response in CFTR-/- mice. We will use both in vitro assays, as well as, in vivo studies to determine if CFTR null immune cells play a primary role in the abnormal immune response in CFTR-/- mice and whether WT immune cells can rectify this response;ii.) we will dissect the roles of CFTR+ epithelial cells and CFTR+ macrophages in the in vivo response to LPS in the chronically inflamed lung using a gene complementation strategy. We will determine if CFTR-/- mice with macrophage specific CFTR expression versus epithelial-cell specific CFTR expression has an inflammatory response that is similar to CFTR-/-, CFTR-/+ or WT mice and iii.) lastly, we will examine if the lack of CFTR is directly responsible for the exaggerated immune response to LPS in macrophages by blocking CFTR in WT cells and by enhancing expression of CFTR in CFTR-/- cells
The ultimate goal of this project is to apply insights gained from these studies to improve the clinical management of people with cystic fibrosis and develop new therapies to treat this disease.
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