Human alveolar macrophages (AMs) were obtained by bronchoscopy from 11 healthy adult subjects and placed into tissue culture for 24 hr. Brief preexposure (15 min) of human neutrophils to AM culture supernatants led to a greater than twofold increase in neutrophil killing of a serum-resistant strain of Pseudomonas aeruginosa (P less than 0.02). No increase in phagocytosis of 35S-labeled Pseudomonas could be detected for neutrophils preexposed to AM supernatants. However, upon exposure to bacteria, neutrophils preincubated with AM supernatants generated significantly more (P less than 0.05) superoxide anion than controls. This suggested that AM supernatants enhanced neutrophil oxidative bactericidal capacity. The material in AM supernatants which enhanced neutrophilic killing of Pseudomonas was less than 10,000 daltons in mass and heat-stable (56 C for 30 min). Release of this material was partially inhibitable by exposure of AMs to cycloheximide in tissue cultures. These data suggest that an AM-mediated amplification of neutrophil bactericidal capacity might be important in the defense of the human lung.

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.

Conflicting reports exist regarding the activity of natural killer (NK) cells in resting lung. We speculated that use of cells from differing lung compartments in past studies might have contributed to these discrepancies. To address this issue, a compartmental analysis of relative NK activity of airspace (bronchoalveolar lavage) cells versus interstitial lung cells was conducted in guinea pigs. Non-pulmonary (spleen) cells were also assayed for NK activity. For assays, cells were obtained from each compartment in the same host, then assayed concomitantly using a chromium release assay with the tumor target cell line, K562. Virtually identical NK cell activity was demonstrated by cells derived from airspaces and lung interstitium. This activity was significantly less (p less than 0.05) than that of spleen cells, and its expression required periods of effector: target cell contact which were considerably longer (16 hr) than those needed for spleen cells (4 hr). Further experiments documented the capacity for augmentation of airspace NK cell activity by intratracheal instillation of interferon inducers poly I:C and carboxymethyl acridanone. We conclude that resting guinea pig pulmonary NK cells have low activity relative to systemic cells, that no significant difference in activity exists among NK cells in airspace and interstitial compartments, and that local activation of airspace NK cells is possible using intratracheal interferon inducers.