ROLE OF NF-KAPPA-B IN THE REGULATION OF INDUCIBLE NITRIC-OXIDE SYNTHASE IN AN MTAL CELL-LINE

The effects of cytokines, lipopolysaccharide (LPS), 8-bromoadenosine 3 ',5'-cyclic monophosphate (8-BrcAMP), and pyrrolidine dithiocarbamate (PDTC), an inhibitor of nuclear factor kappa B (NF-kappa B) activation , on inducible nitric oxide synthase (iNOS) expression were studied in the medullary thick ascending limb of Henle's loop cell line ST-1. LP S + interferon-gamma (IF-gamma) promoted a time-dependent increase in nitrite (a NO metabolite) and iNOS mRNA and the appearance of NF-kappa B p50 and p65 in nuclear protein extracts. Actinomycin D but not cycl oheximide prevented the LPS + IF-gamma induction of iNOS mRNA and NO s ynthesis, indicating that iNOS transcriptional activation by LPS + IF- gamma does not require newly synthesized proteins. PDTC inhibited the LPS + IF-gamma induction of NO, iNOS mRNA, and the appearance of NF-ka ppa B in nuclear protein extracts, suggesting that NF-kappa B mobiliza tion and trans-activation of the iNOS gene mediates this induction. In contrast to other cell types, cycloheximide did not alter iNOS mRNA s tability, and 8-BrcAMP did not alter basal or LPS + IF-gamma induced N O production in ST-1 cells.