NEUTRALIZATION OF IL-8 INHIBITS NEUTROPHIL INFLUX IN A RABBIT MODEL OF ENDOTOXIN-INDUCED PLEURISY

Although the potent neutrophil chemotaxin, IL-8, is a known product of endotoxin-stimulated cells in vitro, the contribution of IL-8 to neut rophil recruitment in Gram-negative endotoxin inflammation in vivo is unknown. To determine whether neutralization of IL-8 would decrease en dotoxin-induced neutrophil influx, we generated neutralizing mAbs to r abbit rIL-8 for use in our rabbit model of endotoxin-induced pleurisy. One mAb, ARIL8.2, specifically inhibited both rabbit rIL-8-induced ch emotactic activity and activation of the rabbit IL-8 receptor transfec ted in 293 cells. Anesthetized rabbits with in-dwelling pleural cathet ers received either neutralizing mAb (ARIL8.2; 1 mg/kg) or irrelevant isotype-matched mAb (anti-HIV gp120) i.v. 1 h before as well as intrap leurally (20 mu g/ml) at the time of intrapleural instillation of Esch erichia coil endotoxin (200 ng bilaterally). ARIL8.2 blocked 77% of en dotoxin-induced neutrophil influx (21 +/- 2 (SE) x 10(6) (ARIL8.2) vs 91 +/- 15 X 10(6) (anti-gp120) (p < 0.0001)). By Western analysis, a b and corresponding to rabbit IL-8 was detected in the pleural liquid of rabbits in both groups. By ELISA, however, the concentration of free, unbound IL-8 in the pleural liquid was significantly less in the ARIL 8.2 group than in the anti-gp120 group for at least 4 h, confirming th at ARIL8.2 bound the IL-8 generated in vivo during that time. We concl ude that neutralization of IL-8 profoundly inhibits neutrophil recruit ment in endotoxin-induced pleurisy indicating that IL-8 is a major che motactic factor in this model of acute inflammation.