In activated macrophages, large amounts of nitric oxide (NO) are generated by inducible nitric oxide synthase (iNOS). This is an important mechanism in macrophage-induced cytotoxicity and inflammation. In the present study, a synthetic carbazole compound, 9-(2-chlorobenzyl)-9H-carbazole-3-carbaldehyde (LCY-2-CHO), was found to have an inhibitory effect on lipopolysaccharide (LPS)stimulated NO generation in RAW 264.7 macrophages (IC50 value of 1.3 +/- 0.4 muM). LCY-2-CHO did not induce cytotoxicity and had a negligible effect on iNOS activity. To explore the mechanism of inhibition of NO generation by LCY-2-CHO, the expression of the iNOS gene was examined. LCY-2-CHO abolished the LPS-induced expression of both iNOS protein and mRNA in a parallel concentration-dependent manner with IC50 values similar to those required for inhibition of NO generation. LCY-2-CHO did not enhance the degradation of iNOS mRNA. In cells transiently transfected with an iNOS promoter-chloramphenicol acetyltransferase (CAT) reporter construct, LCY-2-CHO attenuated the LPS-induced iNOS promoter activity. However, LCY-2-CHO had no effect on the degradation of IkappaB-alpha or IkappaB-beta, DNA binding activity, or transcriptional activity of nuclear factor-kappaB (NF-kappaB). These results indicate that LCY-2-CHO inhibits NO generation via a decrease in the transcription of iNOS mRNA through a signaling pathway that does not involve NF-kappaB activation. (c) 2002 Elsevier Science Inc. All rights reserved.
