Sputum stimulation of human neutrophil degradation of lung proteoglycans

Abstract

Purpose: Neutrophil-mediated matrix degradation was proposed to be a pathogenetic factor in bronchiectasis. In this study we test for the degradation of lung proteoglycans in a model matrix when neutrophils are activated by pro-inflammatory mediators in bronchial secretions. Methods: Proteoglycans isolated from rat lungs were entrapped in 5 % polyacrylamide gel beads as a model matrix for incubation with either normal human neutrophils and/or sputum sol from patients with bronchiectasis. When proteoglycans were degraded, fragments containing glycosaminoglycans released into the incubation medium and were assayed for hexuronate content. Proteoglycan degradation is indicated when there was an increase in hexuronate content when compared with control incubations. The data represent mean ± SD of five independent experiments performed in duplicate. Results: Blank incubations of neutrophils and sputum sol gave hexuronate concentration of (6.52±1.02) μg/ml and (8.72±1.25) μg/ml respectively. Single incubations with gel beads showed increase hexuronate contents of (8.47±1.18) μg/ml and (10.04±1.94) μg/ml respectively. This indicated that both sputum sol from patients with bronchiectasis and neutrophils from normal volunteers exhibited basal proteoglycan-degrading activity. Combined incubation of sputum sol and neutrophils with gel beads showed further increase in hexuronate concentration to (27.95±1.70) μg/ml. The stimulation was neutralized to (16.42±0.32) μg/ml by antibody against TNFα, a pro-inflammatory mediator found in sputum sol of patients with bronchoectasis. Eglin C, a serine protease inhibitors of neutrophil elastase and cathepsin G, was found to lower neutrophil-mediated proteoglycan degradation to hexuronate concentration of (15.07±0.66) μg/ml in the medium. The results suggest that serine protease secreted by neutrophils may degrade proetoglycans in a model matrix and this secretion is stimulated by TNFα in bronchial secretions of patients with bronchiectasis. Conclusions: Our preliminary experiment suggest that sputum sol are able to stimulate neutrophil-mediated proteoglycan degradation. Besides, TNFα in sputum sol are able to enhance the effect of neutrophil-mediated proteoglycan degradation. We postulate that cytokine may play a role in this process. Clinical Implications: Understanding of mechanisms of tissue degradation can contribute to therapeutic strategy that prevents long term lung damage leading to respiratory failure.