AGONIST-INDUCED LIPOXIN A(4) GENERATION - DETECTION BY A NOVEL LIPOXIN A(4)-ELISA

Lipoxin A(4) (LXA(4)) possesses potent bioactions. To facilitate its detection, an enzyme linked immunosorbent assay (ELISA) was developed that proved sensitive and selective. Quantitation by ELISA of LXA, generated from cellular sources strongly correlated (r = 0.99) with values obtained by high-pressure liquid chromatography (HPLC). We used this LXA(4)-ELISA to examine parameters influencing LXA(4) generation from endogenous substrates during human platelet neutrophil (PLT-PMN) interactions in vitro. Agonist induced LXA(4) production was clearly evident at a PLT-PMN ratio of 10:1, and recombinant human granulocyte/monocyte colony stimulating factor-priming of PMN augmented LXA(4) generation 5-6 fold. The chemotactic peptide formylmethionyl-leucyl-phenylalanine, platelet-derived growth factor and arachidonic acid (20:4n-6) each stimulated formation of immunoreactive LXA(4) (iLXA(4)) in these co-incubations. The presence of iLXA(4) was also evaluated in vivo in aspirin-sensitive asthmatic patients who, in a randomized, double-blind crossover design, underwent nasal lavage after they each ingested a predetermined threshold dose of aspirin or placebo. Aspirin challenge provoked statistically significant increases in iLXA(4) in each patient (P < 0.005). These results validate the use of a solid-phase ELISA for detection of LXA(4). Furthermore, the use of this ELISA has allowed the first documentation of iLXA(4) formation in human subjects with aspirin-sensitive asthma following specific antigenic challenge.