Isolation and partial characterization of a human alveolar macrophage-derived neutrophil-activating factor.

Abstract

Human alveolar macrophages (AM) were obtained from eight normal volunteers using fiberoptic bronchoscopic lavage to explore potential interrelationships among leukocytes in pulmonary defense against infection. AM placed in monolayer tissue cultures released material into culture supernatants with the capacity to enhance the bactericidal capacity of human neutrophils. Neutrophils preexposed to supernatants killed Pseudomonas aeruginosa from 70 to 90% more efficiently than control cells (P < 0.02). AM culture supernatants contained this material by 4 h of incubation, and in vitro stimulation of AM cultures with heat-killed P. aeruginosa further increased its production.Gel filtration of AM culture supernatants with a G-50 Sephadex column allowed isolation of a 6,000-D neutrophilactivating factor (NAF) that was resistant to heat (56°C, 30 min). The isoelectric point of NAF, as determined by chromatofocusing, was -7.6. Enzyme digestion of NAF specimens, prepared sequentially by gel filtration and chromatofocusing, demonstrated 50-70% loss of activity after incubations with trypsin, chymotrypsin, and neuraminidase. NAF was only minimally chemotactic and eluted from Sephadex G-50 with particles of a different molecular size than those of AMderived chemotactic factors (i.e., -10,000 D and <500 D). Preincubation of neutrophils with NAF resulted in greater release of superoxide anion upon their subsequent stimulation by either bacterial phagocytosis or by phorbol myristate acetate, as compared with control neutrophils stimulated in a like manner. These studies indicate that human AM secrete a heatstable, low molecular weight basic protein, with the capacity to enhance oxidative microbicidal activity of neutrophils.

A novel neutrophil-activating factor produced by human mononuclear phagocytes.

“…Some of the above reports as well as our present data clearly indicate that the neutrophil-activating peptides released by mononuclear phagocytes are distinct from the two well-known monokines, IL-1 and tumor necrosis factor (TNF) . Several of the NAF-like molecules were shown to be considerably smaller than either monokine (26,28,30,31), and to exert effects that could not be obtained with recombinant and highly purified monokines (31) . In addition, amino acid sequencing of purified NAF (8) showed no significant structural homology with IL-la and ß and TNF-a.…”

Cellular events in alveolitis and the evolution of pulmonary fibrosis

“…Prolonged cigarette smoking, in addition to other, well known risks, is often associated with the accumulation of pulmonary alveolar macrophages that are larger than in nonsmokers (Miiller and Hirschberg 1984), have an enhanced cytotoxic potential for normal lung parenchymal cells in vitro ) and, mainly with the help of complement component C5 (Robbins et al 1987), can attract (Hunninghake etal. 1978;Hunninghake and Crystal 1983) and activate (Pennington et al 1985) neutrophils. It seems safe, therefore, to accept that cigarette smoking may damage the alveolar epithelium (Jones et al 1980), at least in part, by the cytotoxic effects of phagocytes.…”

Macrophage‐granulocyte interactions in allergic diseases

“…We have previously demonstrated that alveolar macrophages (AM) placed in monolayer tissue cultures release a substance which enhances the bactericidal capacity of human neutrophils [32]. Neutrophils pre-exposed to supernatants killed Pseudomonas aeruginosa 70-90% more efficiently than control cells.…”