ADHESIVE INTERACTIONS BETWEEN FIBROBLASTS AND POLYMORPHONUCLEAR NEUTROPHILS INVITRO

Although the in vivo interaction between polymorphonuclear neutrophils (PMN) and fibroblasts may be important, these pathways have not been well studied. We have investigated the adherence of PMN to monolayers of human fetal lung fibroblasts, using a microtiter plate assay based upon the uptake by cells of the vital stain Rose Bengal. Stimulation with phorbol myristate acetate (PMA) caused a significant increase of adherence over basal levels which was rapid in onset and plateaued at 5 min. Adhesion was dependent on the leucocyte integrin family of glycoproteins, notably on Mac-1, since monoclonal antibodies toward the beta-chain (CD18) and alpha-chain (CD11b) of Mac-1 almost completely suppressed PMA-induced PMN adhesion (88% and 77% inhibition, respectively). Adhesion was also inhibited by the peptides RGDS and GRGDS (24.2% and 26.6%, respectively using 1 mM peptide). Prestimulation of fibroblasts for longer time periods (5 and 24 h) with interleukin 1-alpha and tumor necrosis factor-alpha, but not transforming growth factor-beta, also resulted in a significant increase in adhesion of unstimulated PMN (after 24 h preincubation, 10 U/ml IL1-alpha stimulated adhesion by 179% of control, 500 U/ml TNF-alpha by 157%). This indicated that there are both PMN- and fibroblast-dependent pathways for PMN adhesion. Components of the extracellular matrix of fibroblasts do not appear to play important roles in the adhesion process since addition of fibronectin and type IV collagen, or of purified antibodies to fibronectin and types I and IV collagen, did not affect PMA-induced PMN adhesion. Adhesion of PMN to fibroblasts was influenced by charge since neuraminidase treatment (which removes negatively charged groups on the cell surface) stimulated adherence in the absence of stimulus, and polyanion ligands were capable of modulating PMA-induced adherence. Given this evidence that PMN possess mechanism by which they adhere to fibroblasts, it is now relevant to establish the importance of such interactions in modifying fibroblast activities in inflammatory disorders.