INTERLEUKIN-8 IS A MAJOR COMPONENT OF PLEURAL LIQUID CHEMOTACTIC ACTIVITY IN A RABBIT MODEL OF ENDOTOXIN PLEURISY

Gram-negative endotoxin induces production of the potent chemotactic f actor interleukin-8 (IL-8) in vitro; however, the importance of IL-8 i n endotoxin-induced inflammation in vivo is unknown. We asked whether IL-8 is an important contributor to chemotactic activity in acute infl ammatory liquids formed in response to endotoxin, and, if present, wha t concentrations of IL-8 antigen are generated. For these studies, we cloned and expressed rabbit recombinant IL-8 (rrIL-8), developed speci fic anti-rabbit IL-8 monoclonal antibodies (mAb), and then used these reagents to develop assays to detect rabbit IL-8 bioactivity and measu re rabbit IL-8 antigen. Escherichia coli endotoxin (20 ng/ml, n = 4, o r 2,000 ng/ml, n = 4) was instilled into the pleural space of eight ra bbits for 6 h. Rabbit IL-8 bioactivity in the endotoxin pleurisy sampl es was assayed by measuring the migration of rabbit neutrophils toward the pleural liquid under two different conditions: 1) after addition of an anti-IL-8 neutralizing mAb and 2) after desensitization of the n eutrophils to rrIL-8. Addition of the anti-IL-8 mAb decreased neutroph il migration toward the pleural liquid by 65 +/- 13 and 75 +/- 22% (me an +/- SE, after 20 and 2,000 ng/ml endotoxin, respectively; P < 0.01 compared with a control mAb). Desensitization of neutrophils to rrIL-8 decreased their migration toward the pleural liquid by 72 +/- 5% (P = 0.03, compared with exposure of neutrophils to buffer alone). Rabbit IL-8 antigen was measured by enzyme-linked immunosorbent assay in infl ammatory pleural liquids at physiologically active concentrations (2.3 +/- 0.5 and 6.9 +/- 7.4 ng/ml, means +/- SE, 6 h after 20 and 2,000 n g/ml endotoxin, respectively). We conclude that IL-8 is an important c hemotaxin generated in vivo during endotoxin-induced inflammation.