Negative regulation of human heme oxygenase in microvessel endothelial cells by dexamethasone

Heme oxygenase-l (HO-1) is a stress protein, and its induction has been sug gested to participate in defense mechanisms against agents that promote oxi dative injury such as endotoxins and heme, We have shown that the inflammat ory cytokines, interleukin-g (IL-6) and heme-induced HO-1 gene expression, were suppressed by dexamethasone (Dex) in a sustained manner, We examined t he mechanism by which the anti-inflammatory agent, Dex, inhibits IL-6 and h eme-induced HO-1 expression in rabbit coronary endothelial cells, Endotheli al cells treated with heme (10 mu M) and IL-6 (25 ng/ml), increased HO-1 mR NA 15- and 60-fold, respectively. The activity of HO was increased 3-fold a fter such treatment. Although Dex failed to inhibit heme-mediated HO-1 mRNA and HO activity, it was able to reverse IL-6-stimulated HO activity, Sever al human HO-1 promoter-drive chloramphenicol acetyltransferase (CAT) constr ucts were examined to analyze IL-6 and Dex-mediated modulation of the HO-1 gene in endothelial cells. CAT assays revealed that the HO-1 promoter regio n between -180 and -1500 might contain a Dex-mediated negative regulator, G el mobility shift assays using nuclear extracts from IL-6-treated endotheli al cells showed a binding to the synthetic 21 base pairs of the HO-1 sequen ce that contains the putative STAT3 sequence. STAT3-specific probe inhibite d nuclear binding protein to the putative HO-1-STAT3 sequence, This suggest s that IL-6 induction of human HO-1 is mediated via the JAK-STAT pathway an d that Dex inhibition of gene expression is carried out by activation of a transcriptional protein in competition with the STAT3 binding site.