Dual effects of cyclosporine A on arachidonate metabolism by peritoneal macrophages. Phospholipase activation and partial thromboxane-synthase blockage.
Because the oxygenated metabolites of arachidonate synthesized by macrophages, particularly prostaglandins (PG)I2 and E2, thromboxane (TX)A2 and 12-hydroxyeicosatetraenoic acid (12-HETE) have been shown to modulate the immune response of T-cells, we tested the effect of cyclosporine A (CsA), a potent immunosuppressor agent, on arachidonate (AA) metabolism in cultured peritoneal rat macrophages. Endogenous AA release and 12-HETE synthesis were estimated by radiometric HPLC after prelabelling of macrophages with [3H]AA whereas PG were determined either by radiometric HPLC or by direct radioimmunoassay in the culture mediums. Exposure of prelabelled cells for 16 hr to CsA led to a large increase in the release of AA itself and of its oxygenated metabolites, PG and 12-HETE, indicating stimulation of phospholipase activity. This effect was time- and dose-dependent at concentrations of CsA between 2 and 50 microM. There was also a marked increase in the ratio PGI2/TX, suggesting, in addition to activation of phospholipase, a partial blockade of TX synthase. When macrophages were triggered by A 23187 calcium ionophore (2 microM) or opsonized zymosan (1 mg/ml), the only detectable effect of CsA was a strong and specific inhibition (50%) of TX synthesis. Addition of an excess of exogenous AA (5 micrograms/ml) to cells treated by CsA confirmed the fact that CsA acted by specifically blocking the transformation of AA into TX without affecting PGI2 or 12-HETE synthesis. These results demonstrate that CsA acts at two different levels: it promotes phospholipase activation on resting cells but simultaneously induces a partial blockade of TX-synthase. This latter effect predominates when cells are stimulated. The resulting change in the ratio PGI2/TX promotes immunosuppression to the expense of immunostimulation. This may represent one of the factors underlying the potent immunosuppressive role of CsA.
Sraer
J
J
INSERM 64, Hôpital Tenon, Paris, France.
Bens
M
M
Ardaillou
R
R
D015121:6-Ketoprostaglandin F1 alpha / Q000096:biosynthesis; D000818:Animals; D001095:Arachidonic Acids / Q000378:metabolism*; D002851:Chromatography, High Pressure Liquid; D003524:Cyclosporins / Q000494:pharmacology*; D004305:Dose-Response Relationship, Drug; D004789:Enzyme Activation; D006893:Hydroxyeicosatetraenoic Acids / Q000096:biosynthesis; D008264:Macrophages / Q000201:enzymology / Q000378:metabolism*; D010529:Peritoneal Cavity / Q000166:cytology; D010740:Phospholipases / Q000378:metabolism*; D010741:Phospholipases A / Q000378:metabolism*; D011453:Prostaglandins / Q000096:biosynthesis; D011863:Radioimmunoassay; D051381:Rats; D011919:Rats, Inbred Strains; D013930:Thromboxane-A Synthase / Q000037:antagonists & inhibitors*
D001095:Arachidonic Acids; D003524:Cyclosporins; D006893:Hydroxyeicosatetraenoic Acids; D011453:Prostaglandins; D015121:6-Ketoprostaglandin F1 alpha; D010740:Phospholipases; D010741:Phospholipases A; D013930:Thromboxane-A Synthase