Regulation of the Release of Inflammatory Mediators From Lung Macrophages.

Chronic obstructive pulmonary disease (COPD) is the fifth leading cause of death in the UK and is the only common cause of death that is increasing. COPD is currently 6th in the global impact of diseases and is predicted to by the 3rd leading cause of death by 2020. In the UK for 2000-2001 the estimated cost of COPD related illness is estimated at ~£980 million of which pharmacotherapy accounts for 48% of expenditure. At present pharmacotherapy for COPD is purely symptomatic with no drugs currently available that can halt the relentless progression of this disease. Therefore, there is a need for improved therapy for the treatment of COPD. Cigarette smoking is the major risk factor in the development of COPD, yet for reasons unknown only ~15% of smokers develop this disease, suggesting there is an underlying genetic component.

COPD encompasses chronic bronchitis, small airways disease and emphysema and is associated with increased inflammatory cells in the lung including neutrophils, macrophages and CD8+ T-lymphocytes. This inflammatory infiltrate is thought to be responsible for all the pathophysiological features of COPD but the precise mechanisms underlying this inflammatory response are unknown.In particular the macrophage is thought to mediate all the pathophysiological features of this disease. To this end, macrophages will be isolated from lung parenchyma using discontinuous percoll gradients. The cells will then be cultured overnight prior to assessment of inflammatory mediator release. Cells will be stimulated by a variety of agents including but not exclusively LPS, IL-1beta or TNF-alpha. The release of inflammatory mediators into the cell culture media will be measured using ELISA techniques. Enzyme activity will also be measured in both the cells and the culture media. Cell surface expression of specific markers will be assessed using immunocytochemistry and FACS analysis. Function of macrophages will be assessed by measuring phagocytotic activity by FACS analysis and fluorimetry. Specific signal transduction pathways will be assessed by the use of specific pathway inhibitors and gene expression measured by real-time PCR. The effects of novel therapeutic agents will be tested on these outputs with the aim to identify novel therapies for COPD.