Effect of metabolic inhibitors on the MSU crystal-induced prostanoid synthesis and protein expression of COX-2 in human monocytes. Monocytes were incubated with diluent (C), or MSU crystals (M; 0.3 mg/mL) for 24 hours alone or in combination with a protein synthesis inhibitor, cycloheximide (CHX; 10 μg/mL); an inhibitor of transcription, actinomycin D (AD; 5 μg/mL), or a specific inhibitor of tyrosine kinases, herbimycin A (HA; 100 nmol/L). A p38 mitogen-activated protein kinase inhibitor, SB 203580 (SB; 10 μmol/L), as well as an inactive structural analog, SK&F 106978 (SKF; 10 μmol/L) were also used under similar conditions. (A) Cell-free supernatants were collected and analyzed for prostanoid synthesis as indicated in the Materials and Methods. Results are the mean ± SD from 3 separate experiments and are expressed as percent of MSU crystal-stimulated monocytes in the absence of metabolic inhibitor. (B) Corresponding cell samples were processed for evaluation of immunoreactive COX-2 by immunoblot as described in Materials and Methods. A representative immunoblot is shown.