Characterization of glucocorticoid inhibition of antigen-induced inositolphosphate formation by rat basophilic leukemia cells: possible involvement of phosphatases

Abstract The suppress effect of glucocortocoids (GC) upon antigen-induced phosphatidylinositol phospholipase C (PI-PLC) activity and inositol phosphate formation by rat basophilic leukemia cells (RBL-2H3) has been characterized. Addition of antigen for a period of 1–30 min enhanced production of [ 3 H ]inositol monophosphate (IP 1 ), inositol 1,4-biphosphate (IP 2 ) and inositol 1,4,5-triphosphate (IP 3 ) by about 5–10-fold. Pretreatment with hydrocortisone (HC) reduced formation of the various inositol phosphates (IPs) and degradation of phosphatidylinositol 4,5-bisphosphate (PIP 2 ) by an average of 50%. Maximal inhibition of hydrolysis of PIP 2 and reduction in stimulation of IP 3 formation was reached after 4 h of preincubation with 2·10 −6 M of HC. Cycloheximide and RU486, a GC receptor antagonist, completely prevented the inhibitory effect of HC on IP formation. Other GC, dexamethasone (DEX) and triamcinolone (each at 2·10 −7 M) markedly suppressed antigen induced IP 3 production, while aldosterone and sex steroids such as estradiol and progesterone (each at 2·10 −6 M) were virtually inactive. Antigen-stimulated phosphorylation of a 18 kDa and other proteins was inhibited by about 60% following pretreatment with the GC. This inhibition was in turn prevented by cycloheximide. DEX also doubled the activity of cellular acid phosphatase activity. The results suggest that the inhibitory effect of GC is specific, receptor-mediated, dependent on protein synthesis and possibly mediated by protein phosphatase activity.