Activation of the complement system in the adult respiratory distress syndrome.

The adult respiratory distress syndrome (ARDS) is an acute pulmonary disorder characterized by the accumulation of neutrophils within the lower respiratory tract. Because activation of the complement system can generate C5a, a potent neutrophil chemoattractant, complement activation was assessed in both serum and bronchoalveolar lavage fluid obtained from 10 patients with ARDS and compared with that from normal control subjects. Crossed immunoelectrophoresis was used to determine activation of the complement components C3 and properdin factor B (PFB), and radioimmunoassay was used to determine the presence of C5a. Complement activation was not detected either in the plasma or in the lung epithelial lining fluid of the control subjects. In contrast, evidence of C3 activation was found in the plasma of 50% of the patients with ARDS when initially studied; likewise, C3 activation, PFB activation, and C5a could all be detected in the epithelial lining fluid of all patients with ARDS with a single exception. Follow-up bronchoalveolar lavages revealed decreased amounts of C3 activation and C5a 2 to 7 days after the onset of ARDS, and the complement activation had resolved when the patients with ARDS had completely recovered. To determine if the C5a in bronchoalveolar lavage fluid could be responsible for the influx of neutrophils observed in ARDS, epithelial lining fluids obtained from both normal control subjects and from patients with ARDS were fractionated by molecular sieve chromatography. Two distinct fractions of chemotactic activity were found in the ARDS bronchoalveolar lavage fluid.(ABSTRACT TRUNCATED AT 250 WORDS)

Complement component C5 is required for release of alveolar macrophage-derived neutrophil chemotactic activity.

The influx of neutrophils into the alveolar structures can be induced by stimulation of the resident lung phagocyte, the alveolar macrophage, to release a potent neutrophil chemoattractant(s). We hypothesized that the fifth component of complement (C5) on the cell surface may be required for activation of the alveolar macrophage to release neutrophil chemotactic activity. C5 was identified on guinea pig alveolar macrophages by epifluorescent microscopy, flow cytometry, and enzyme-linked immunoabsorbent assay of eluted macrophages. When cultured for 4 h with stimuli that induce the release of chemotactic activity or for 24 h without added stimuli, purified Fab' fragments of a goat anti-C5 antibody significantly inhibited the ability of macrophages to release chemotactic activity as determined by a blindwell chamber method (p less than 0.001, all comparisons). This inhibition of chemotactic activity was not detected when anti-C5 antibody was added after the culture period. In contrast, anti-C3 antibody had no inhibitory effect at 4 h or at 24 h (p greater than 0.2, all comparisons). Partial characterization of released chemotactic activity revealed it was of low molecular weight, partially lipid soluble, and not inhibited by C5a chemotactic factor inactivator. These studies suggest that C5 may have a regulatory role in the release of chemotactic activity by alveolar macrophages.

Role of chemotactic factor inactivator in modulating alveolar macrophage-derived neutrophil chemotactic activity.

The stimulated alveolar macrophage is a potent source of neutrophil chemotactic activity. The release of this chemotactic activity can be inhibited by pretreating alveolar macrophages with anti-C5 antibody. We hypothesized that C5a, a fragment cleaved from C5 when C5 is activated, might activate the alveolar macrophage to release neutrophil chemotactic activity and that chemotactic factor inactivator, a serum inhibitor of C5a, could decrease this release. Activated complement components including C5a were found to stimulate guinea pig macrophages to release chemotactic activity into their culture supernatants at levels that were significantly higher than the chemotactic activity of C5a alone (P less than 0.001). Chemotactic factor inactivator was found to cause a marked reduction in the chemotactic activity released by macrophages stimulated with phagocytic and nonphagocytic stimuli (P less than 0.001, all comparisons). These data indicate that C5a can stimulate alveolar macrophages to release chemotactic activity in vitro, and that chemotactic factor inactivator may play a role in modulating this process.

Complement activation in cystic fibrosis respiratory fluids: in vivo and in vitro generation of C5a and chemotactic activity.

Experiments performed in vitro have demonstrated that leukocyte neutral proteases produce an important mediator of inflammation, C5a, by proteolysis of the C5 component of the complement system. Cystic fibrosis (CF) lung fluids were characterized by high levels of neutrophils (39% of total cells versus 2% in normals) and contained significantly elevated amounts of elastolytic activity (mean 17.7 ng/micrograms total protein) compared to the lung fluids obtained from normal volunteers (0.2 ng elastolytic activity/micrograms protein, p = 0.001). The objective of these studies was to determine if complement activation and complement-derived chemotactic activity are present in CF lung fluids. C3c peptide representing activation of C3 could not be identified in the bronchial-alveolar lung lavage fluids of normal subjects but was readily identified by means of crossed immunoelectrophoresis in CF lung fluids (n = 9, mean 49% of C3); the mean level of C3 was decreased in CF lung specimens. Chemotactic activity was significantly elevated in lung fluids of the CF patients when compared to normal lung fluids. Using gel-filtration chromatography and a sensitive radioimmunoassay the chemotaxin present in CF specimens was identified as the anaphylatoxin C5a. C5a levels in the bronchial-alveolar lavage fluids of CF patients was inversely related to volume in liters expired in 1 s of a forced expiratory maneuver expressed as a percent of vital capacity determined from a forced expiratory maneuver (r = -0.72). Because there was a direct relationship between the total elastolytic activity present in CF airways and the concentration of C5a (r = 0.97, p = 0.03), it was postulated that airway proteases with elastolytic activity also cleave C5, nonimmunologically producing C5a. Detailed inhibition assays revealed that much of the total elastolytic activity had the inhibition profile of a serine proteinase. The levels of the serine proteinases were closely correlated with the numbers of neutrophilic leukocytes present per ml of lavage fluid (r = 0.7, p = 0.05). However, inhibitors of leukocyte serine proteases did not prevent the generation of additional chemotactic activity and the proteolysis of radiolabeled C5 substrate was not prevented by inhibitors of neutrophil elastase. Although the purified metalloelastase of Pseudomonas aeruginosa was active on cell-bound and free C5 yielding C5a, inhibition of this bacterial protease in CF lung fluids only partially blocked cleavage of the alpha- and beta-chains of C5.(ABSTRACT TRUNCATED AT 400 WORDS)

Interleukin-2-induced chemotaxis of human T-lymphocytes.

Interleukin-2 (IL-2), a growth factor for T-lymphocytes, has been postulated to cause the accumulation of T-lymphocytes at sites of inflammation by inducing proliferation of these cells. We hypothesized that IL-2 might also serve to attract T-lymphocytes to inflammatory sites. To test this hypothesis, human T-lymphocytes were purified from the peripheral blood of normal volunteers by rosetting with neuraminidase-treated sheep red blood cells and tested for chemotactic activity by using a blind-well chamber technique. Purified IL-2 caused a greater than 20-fold attraction of T-lymphocytes compared with medium alone (P less than 0.001). This attraction was shown to be chemotactic rather than chemokinetic by checkerboard analysis. The T-lymphocyte chemotaxis could be completely inhibited by adsorption of the IL-2 with an IL-2-dependent cell line, and could be neutralized by monoclonal anti-IL-2 antibody. Further specificity of IL-2-directed chemotaxis was demonstrated by using species-specific IL-2. Mouse IL-2 was ineffective at promoting human T-lymphocyte chemotaxis. These data suggest that IL-2 may be responsible for the localized accumulation of T-lymphocytes both by causing attraction of these cells and by modulating their proliferation.