Agonist-induced lipoxin A4 generation: detection by a novel lipoxin A4-ELISA.
Lipoxin A4 (LXA4) possesses potent bioactions. To facilitate its detection, an enzyme-linked immunosorbent assay (ELISA) was developed that proved sensitive and selective. Quantitation by ELISA of LXA4 generated from cellular sources strongly correlated (r = 0.99) with values obtained by high-pressure liquid chromatography (HPLC). We used this LXA4-ELISA to examine parameters influencing LXA4 generation from endogenous substrates during human platelet-neutrophil (PLT-PMN) interactions in vitro. Agonist-induced LXA4 production was clearly evident at a PLT-PMN ratio of 10:1, and recombinant human granulocyte/monocyte colony stimulating factor-priming of PMN augmented LXA4 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 LXA4 (iLXA4) in these co-incubations. The presence of iLXA4 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 iLXA4 in each patient (P < 0.005). These results validate the use of a solid-phase ELISA for detection of LXA4. Furthermore, the use of this ELISA has allowed the first documentation of iLXA4 formation in human subjects with aspirin-sensitive asthma following specific antigenic challenge.
Levy
B D
BD
Department of Medicine, Brigham and Women's Hospital, Boston, Massachusetts.
Bertram
S
S
Tai
H H
HH
Israel
E
E
Fischer
A
A
Drazen
J M
JM
Serhan
C N
CN
D000328:Adult; D001241:Aspirin; D001249:Asthma / Q000139:chemically induced / Q000378:metabolism; D001826:Body Fluids / Q000737:chemistry; D004311:Double-Blind Method; D004797:Enzyme-Linked Immunosorbent Assay; D006801:Humans; D006893:Hydroxyeicosatetraenoic Acids / Q000032:analysis* / Q000096:biosynthesis; D044045:Lipoxins; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity
D006893:Hydroxyeicosatetraenoic Acids; D044045:Lipoxins; C040527:lipoxin A4; D001241:Aspirin