PYRROLIDINE DITHIOCARBAMATE INHIBITS INTERCELLULAR-ADHESION MOLECULE-1 BIOSYNTHESIS INDUCED BY CYTOKINES IN HUMAN FIBROBLASTS

Intercellular adhesion molecule-1 (ICAM-1), the ligand of lymphocyte function-associated antigen-1, plays an important role in the interactions of a variety of hemopoietic and nonhemopoietic cells, including leukocytes, fibroblasts, and endothelial cells. ICAM-1 is known to be involved in the onset of several diseases such as inflammation, allograft rejection, and so on. In this report, we investigated the effects of dexamethasone, cyclosporin A, FK506, and pyrrolidine dithiocarbamate (PDTC) on the induction of the ICAM-1 gene by cytokines in fibroblasts. PDTC, a potent inhibitor of NF-kappa B, was shown by ELISA and FAGS analysis to prevent dramatically the expression of the ICAM-1 gene stimulated by IL-1 alpha, IFN-gamma, and PMA, although the other reagents inhibited it only slightly. Ribonuclease protection assay revealed that PDTC blocked the expression of the ICAM-1 gene at the mRNA level. To elucidate the mechanism of this inhibition, we constructed a series of ICAM-1 promoter deletion mutants linked to the chloramphenicol acetyl transferase gene and analyzed the effect of PDTC on their activities. Transient transfection analysis indicated that the critical region for inhibition by PDTC is an NF-kappa B binding site-like motif (GGGAGGATTCC, ICAM-1 kappa B) that is located at position -540. Electrophoresis mobility shift assay revealed that PDTC actually inhibits the binding of NF-kappa B (or NF-kappa B-like) protein to the ICAM-1 kappa B site. These findings suggest that PDTC inhibits ICAM-1 gene expression by inhibiting the association of NF-kappa B (or NF-kappa B-like) protein with the ICAM-1 kappa B site.