Cyclooxygenase-2 (COX-2), an inducible isozyme of cyclooxygenase, is selectively expressed in response to lipopolysaccharide (LPS) and its expression is suppressed by the glucocorticoid dexamethasone (DEX) in the monocytic differentiated U937 cells. However, COX-2 mRNA was not detected nor induced by LPS before the cells differentiated. To study the transcriptional role of the NF-kappa B site (nucleotides -223 to -214) in the COX-2 gene, the luciferase reporter vector driven by the COX-2 promoter region (nucleotides -327 to +59) mutated at both the cAMP response element and the NF-IL6 site was stably transfected into U937 cells. The substantial luciferase activity observed in the undifferentiated cells was not induced by LPS. However, after the cells had differentiated, luciferase activity was induced by LPS and its induction was suppressed by DEX. Moreover, a protein tyrosine kinase inhibitor herbimycin A suppressed both the expression of COX-2 mRNA and the luciferase activity induced by LPS. These results suggest that the NF-kappa B site is involved in both the LPS-induced expression of the COX-2 gene and its suppression by DEX and herbimycin A in a differentiation-dependent manner.