MATRIX METALLOPROTEINASE AND ELASTASE ACTIVITIES IN LPS-INDUCED ACUTELUNG INJURY IN GUINEA-PIGS

Matrix metalloproteinases (MMPs) and elastase are proteolytic enzymes specifically directed against extracellular matrix (ECM) components. T hey are secreted by inflammatory cells and may consequently contribute to the lesions of the ECM observed during acute pulmonary edema. We t herefore evaluated the MMP and elastase activities, which are secreted by cultured alveolar macrophages (AMACs) and polymorphonuclear neutro phils (PMNs) and present in the bronchoalveolar lavage (BAL) fluid in a guinea pig model of acute lung injury induced by intratracheal insti llation of lipopolysaccharide (LPS). The control group was given 0.9% NaCl. 24 h after instillation, a BAL was performed, the BAL, fluid was separated from the cells by centrifugation, and AMACs and PMNs were s eparately cultured for 24 h. In BAL fluid from LPS-treated guinea pigs , we found 1) an increase in free gelatinase activity, tested on [H-3] gelatin (0.7 +/- 0.2 mu g.200 mu l BAL fluid(-1).48 h(-1) vs. 0.2 +/- 0.1 in controls, P < 0.05), and 2) increased total gelatinase activiti es, as assessed by zymography. The molecular masses of the major gelat inase species found in BAL fluid by zymography were 92 and 68 kDa. The 92-kDa gelatinase was secreted by both AMACs and PMNs, as demonstrate d by zymography of their respective culture media. When tested on [H-3 ]elastin, the elastase activity of BAL fluid of LPS-treated animals ex hibited no increase, but when tested on a synthetic peptidic substrate [N-succinyl- (L-alanine)(3)-p-nitro anilide (SLAPN)], increased elast ase-like activity was observed (from 17 +/- 4 nmol of SLAPN.200 mu l B AL fluid(-1) 24 h(-1) in control group to 34 +/- 8 in LPS group, P < 0 .05). This increase was attributable to the activity of a metalloendop eptidase that was inhibited by the metal chelator EDTA but not by the specific tissue inhibitor of MMPs.