Chronic progressive lung infection with bacterial pathogens, particularly Pseudomonas aeruginosa, is responsible for most of the morbidity and mortality in patients with cystic fibrosis (CF). In preliminary studies, we have found that the sputum of P. aeruginosa infected patients is highly inhibitory for lymphocyte proliferative responses. The inhibitory activity present in the sputum is heat resistant. In parallel studies we have found potent, heat resistant inhibitory factor(s) for lymphocyte proliferation in P. aeruginosa culture supernatants. These inhibitory factors have been identified as phenazine pigments. The inhibitory activity of these low molecular weight pigments is not neutralized by anti-P. aeruginosa antibodies. Our first specific aim is to identify the heat resistant inhibitor(s) of lymphocyte proliferation which are present in CF sputum. To this effect, high resolution chromatography and other analytical methods will be used. The presence of inhibitors in CF sputum will be correlated with bacterial infection and functional inhibitory activity. The second specific aim is to establish the effect of P. aeruginosa phenazine pigments and inhibitors present in CF sputa on lymphocyte activation, and on phagocytic capabilities of polymorphonuclear cells (PMN's) and alveolar macrophages (AM). These methods include surface expression of activation markers on T-lymphocytes, secretion of immunoglobulins by B-lymphocytes, and phagocytosis, nitrobluetetrazolium reduction and P. aeruginosa uptake and killing by PMN's and AMs. The subsequent goal of these studies is to define alterations in cells obtained directly from CF sputum or bronchial lavage. The third specific aim is to establish whether CF lymphocytes or PMN's are more susceptible to inhibition by P. aeruginosa phenazine pigments than are normal lymphocytes or PMN's. A last specific aim is to search for possible antagonists for lymphocytes inhibitors produced by P. aeruginosa. An inhibition-neutralizing effect will be sought using antibodies to an inhibitor covalently attached to a carrier, or through competitive inhibition with similar but non-inhibitory compounds. The presence of heat resistant inhibitors in CF sputum is likely to contribute to the chronicity and progression of CF pulmonary infection. This mechanism of inhibition of pulmonary resistance needs to be understood in order to design specific therapeutic approaches to overcome it.
