Effect of pertussis toxin and B-oligomer on platelet-activating factor-induced generation of inositol phosphates in porcine alveolar macrophages.

OBJECTIVE

To test the hypothesis that platelet-activating factor (PAF) induces inositol phosphate turnover through a receptor-linked, pertussis toxin-sensitive guanine nucleotide-binding (G) protein-dependent pathway in porcine alveolar macrophages.

DESIGN

Randomized complete block design was used with 2 or 3 replicates/block.

ANIMALS

Porcine alveolar macrophages were obtained by lavage of excised lungs from Yorkshire-type pigs (mean +/- SEM, 21 +/- 2 kg).

PROCEDURE

Phospholipase C activation was assessed, using anion exchange chromatography to measure accumulation of inositol phosphates in [3H]myo-inositol-labeled alveolar macrophages. Macrophages were incubated with saline solution, pertussis toxin (4.75 nM), or B-oligomer (4.75 nM) for 2 hours. Cells then were washed and incubated for 5 minutes with PAF (0, 0.1, 1.0, or 10 microM; n = 15). Results were expressed as total inositol phosphates (inositol monophosphate, bisphosphate, trisphosphate, and tetrakisphosphate).

RESULTS

Concentrations of total inositol phosphates were significantly (P < 0.05) increased to 162 +/- 7, 172 +/- 4, and 194 +/- 9% of control in response to 0.1, 1.0, and 10 microM PAF, respectively. Pertussis toxin attenuated the PAF-induced increase in total inositol phosphates by approximately 50% (P < 0.05). The B-oligomer of pertussis toxin failed to modify PAF-induced increases in total inositol phosphates. The specific PAF receptor antagonist WEB 2086 markedly attenuated PAF-induced. (10 microM) increase in inositol phosphates.

CONCLUSIONS

We conclude that PAF stimulates accumulation of inositol phosphates through a specific receptor and that a pertussis toxin-sensitive G protein is involved in the signal transduction process leading to activation of phospholipase C in porcine alveolar macrophages.