Characterization of COPD based on the peripheral neutrophil proteome

COPD (chronic obstructive pulmonary disease) is a degenerative chronic inflammation in the lungs with tissue damage as consequence. Nearly all COPD cases in the Western world are caused by cigarette smoking, however, approximately 15% of all smokers develop COPD and point out that the development of COPD has an environmental and genetic background. In bronchial alveolar lavage (BAL) of COPD patients a high percentage of neutrophilic granulocytes are present which probably play a major role in the tissue damage in the lungs. When this inflammation is present no drugs are successful in damping this process.

COPD is a heterogeneous disease with many phenotypes. However, the characterization is solely based on spirometry data (table 1). The GOLD (Global Initiative for Chronic Obstructive Lung Disease) classification is based on the FEV1 (Forced Expired Volume in 1 second) and FVC (Forced Vital Capacity). The decrease of FEV1 and FVC are a consequence of the inflammation present in the lungs and are not a good marker for the inflammation itself. The inflammation in the lungs has also systemic effects. For instance, peripheral neutrophils can be primed (pre-activated) phenotype or higher levels of several cytokines like TNF and GM-CSF are present. The presence of these cytokines might be different between the several phenotypes of COPD, however, the short half-life of these cytokines makes it very difficult to measure. These cytokines have a systemic effect on for instance the neutrophils and based on this feature we are interested whether we can characterize COPD based on the systemic effects in COPD patients.

Table 1: GOLD classification COPD (Pauwels et. al, Am. J. Respir. Crit. Care. Med., 2001 and www.goldcopd.org)

We hypothesize that the systemic inflammation has an effect on peripheral neutrophil protein expression and that this can be used to characterize COPD.

The general protein expression of neutrophils from COPD patients we investigate with 2 dimensional (2D-) gel electrophoresis (figure 1). With this technique we are able to separate several hundreds of proteins (a proteome) from neutrophils isolated from COPD patients and compare them with neutrophils from other COPD patients, neutrophils from healthy controls and neutrophils from in-vitro stimulated neutrophils. The latter we do to compare a proteome from COPD patients with neutrophils stimulated in-vitro with for instance TNFalpha.

Figure 1: Example of a 2D-gel of neutrophils

To find differences in membrane protein expression of neutrophils from COPD patients we will measure approximately 20 proteins by FACS analysis. This is a easy and rapid method to measure membrane protein expression of known protein. Beside this characterization of COPD we are interested in the specific functions of the neutrophil in-vitro and in-vivo.

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Jeroen Langereis, PhD-student