Protein kinase C inhibition enhances platelet-activating factor-induced eicosanoid production in human eosinophils.
Previous investigations have suggested that protein kinase C (PKC) may regulate guinea pig eosinophil responses through a suppressive "negative feedback" mechanism. Using the selective PKC inhibitors bisindolylmaleimide I (Bis I, GF 109203X) and calphostin C, we examined the role of PKC in platelet-activating factor (PAF)-induced respiratory burst and generation of arachidonic acid metabolites in human peripheral blood eosinophils. Bis I inhibited PAF-induced generation of superoxide anion with substantially lower potency (geometric mean IC50 = 1.41 microM, 95% CI 0.94-2.11 microM) than it exhibited against responses to the phorbol esters 4-beta-phorbol 12-myristate 13-acetate (PMA; IC50 = 0.25 microM, 0.09-0.72 microM; P < 0.01) and 4-beta-phorbol 12,13-dibutyrate (IC50 = 0.48 microM, 0.20-1.14 microM; P < 0.05). The production of thromboxane (measured as TxB2) induced by 1 microM PAF was increased significantly by Bis I at concentrations of 1 microM (162 +/- 7.5% of control PAF response; P < 0.01) and 10 microM (194 +/- 17%; P < 0.001); TxB2 release induced by PMA was unaffected by concentrations of Bis I up to 1 microM and inhibited by 10 microM Bis I (48 +/- 11%; P < 0.05). Bis I (1 microM) significantly increased both thromboxane and leukotriene C4 (LTC4) production induced by 2 microM (P < 0.01 and P < 0.05, respectively) or 20 microM PAF (both P < 0.001). The actions of Bis I on PAF-stimulated thromboxane and leukotriene production were mimicked by a second PKC inhibitor, calphostin C, whereas the non-PKC-inhibitory analog, bisindolylmaleimide V, caused no enhancement of TxB2 or LTC4 production. The increase in intracellular free calcium induced by 1 microM PAF was heightened and prolonged in cells pre-treated with 1 microM Bis I or 1 microM calphostin C (peak increase, P < 0.05 for both drugs; level 60 s after addition of PAF, P < 0.001 and P < 0.05 for Bis I and calphostin C, respectively; time to return to 50% of peak, P < 0.05 for Bis I). We conclude that PKC inhibition causes augmentation of thromboxane and LTC4 production in PAF-stimulated human eosinophils despite suppressing respiratory burst activity, indicating that different signaling pathways predominate in these two responses and that PKC mediates a suppression of an early stage in an alternative pathway of activation.
Dent
G
G
Krankenhaus Grosshansdorf, Zentrum für Pneumologie und Thoraxchirurgie, LVA Hamberg, Germany. Gordon_Dent@compuserve.com
Muñoz
N M
NM
Rühlmann
E
E
Zhu
X
X
Leff
A R
AR
Magnussen
H
H
Rabe
K F
KF
D016718:Arachidonic Acid / Q000378:metabolism; D002118:Calcium / Q000378:metabolism; D002374:Catalase / Q000494:pharmacology; D015777:Eicosanoids / Q000096:biosynthesis*; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D004804:Eosinophils / Q000187:drug effects / Q000378:metabolism*; D005260:Female; D006801:Humans; D007211:Indoles / Q000494:pharmacology; D017997:Leukotriene C4 / Q000096:biosynthesis; D008297:Male; D008301:Maleimides / Q000494:pharmacology; D009281:Naphthalenes / Q000494:pharmacology; D015240:Phorbol 12,13-Dibutyrate / Q000494:pharmacology; D010972:Platelet Activating Factor / Q000494:pharmacology*; D011493:Protein Kinase C / Q000037:antagonists & inhibitors* / Q000502:physiology; D016897:Respiratory Burst / Q000187:drug effects; D013481:Superoxides / Q000378:metabolism; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D013929:Thromboxane B2 / Q000096:biosynthesis
D015777:Eicosanoids; D004791:Enzyme Inhibitors; D007211:Indoles; D008301:Maleimides; D009281:Naphthalenes; D010972:Platelet Activating Factor; D013481:Superoxides; D016718:Arachidonic Acid; D017997:Leukotriene C4; D015240:Phorbol 12,13-Dibutyrate; D013929:Thromboxane B2; D002374:Catalase; D011493:Protein Kinase C; C058819:calphostin C; C088060:bisindolylmaleimide; D013755:Tetradecanoylphorbol Acetate; D002118:Calcium