A NEUTROPHIL-DERIVED PROTEOLYTIC INACTIVE ELASTASE HOMOLOG (HHBP) MEDIATES REVERSIBLE CONTRACTION OF FIBROBLASTS AND ENDOTHELIAL-CELL MONOLAYERS AND STIMULATES MONOCYTE SURVIVAL AND THROMBOSPONDIN SECRETION

Human heparin-binding protein (hHBP) is a recently discovered proteolytically inactive neutrophil elastase homologue with sequence identity to azurocidin and CAP37. The protein has antibacterial properties and chemotactic activity toward monocytes. In the present work, we show that monocytes, cultured under serum-free conditions, developed morphological changes and formed multicellular aggregates 4 h after the addition of hHBP at a concentration of 10 mug/ml. However, after prolonged incubation (11 days) with unchanged medium, the cells spread again. The hHBP-treated cells had a two- to threefold increase in survival compared to control cells, measured using trypan blue as an indicator of living cells. Differentiation of the alive cells to macrophages was detected by changes in morphology, a threefold increase in protein content, and a three- to fourfold increase in acid phosphatase activity. When monocytes in parallel experiments were labelled with [S-35]methionine de novo synthesis and secretion of thrombospondin in a dose-dependent manner was observed after 16 h, with half-maximal secretion at 2 mug hHBP/ml and a maximal 12-fold increase in secretion with respect to controls at 16 mug/ml. Supplementary labeling with [S-35]sulfate revealed that the same monocytes down-regulated the secretion of a large proteoglycan (300-400 kd), apparently also with a half-maximal decrease rate at 2 mug/ml hHBP. Exposure of confluent fibroblast and endothelial cell monolayers to hHPB (10 mug/ml) in the absence of fetal calf serum resulted in cell contraction leaving gaps between cells, the phenomenon being recognizable within 4 h after addition of hHBP. Addition of fetal calf serum to a concentration of 10% completely restored the monolayers. A unique role of hHBP in host defense involving recruitment of monocytes and a key funciton of hHBP in neutrophil extravasation in response to inflammatory chemotactic signals such as leukotriene B4, complement peptide C5a, and N-formyl-methionyl-leucyl-phenylalanine are suggested.