Differential effects of heme oxygenase isoforms on heme mediation of endothelial intracellular adhesion molecule 1 expression

Heme oxygenase (HO), by catabolizing heme to bile pigments, down-regulates cellular hemoprotein, hemoglobin, and heme; the latter generates pro-oxidan t products, including free radicals. Two HO isozymes, the products of disti nct genes, have been described; HO-1 is the inducible isoform, whereas HO-2 is suggested to be constitutively expressed. We studied the inducing effec t of several metal compounds (CoCl2, stannic mesoporphyrin, and heme) on HO activity. Additionally, we studied HO-1 expression in experimental models of adhesion molecule expression produced by heme in endothelial cells, and the relationship of HO-1 expression to the induced adhesion molecules. Flow cytometry analysis showed that heme induces intracellular adhesion molecul e 1 (ICAM-1) expression in a concentration (10-100 mu M)- and time (1-24 h) - dependent fashion in human umbilical vein endothelial cells. Pretreatment with stannic mesoporphyrin, an inhibitor of HO activity, caused a 2-fold i ncrease in heme-induced ICAM-1 expression. In contrast, HO induction by CoC l2 decreased heme-induced ICAM-1 expression by 33%. To examine the contribu tion of HO-1 and HO-2 to endothelial HO activity, specific antisense oligon ucleotides (ODNs) of each isoform were tested for their specificity to inhi bit HO activity in cells exposed to heme. Endothelial cells exposed to heme elicited increased HO activity, which was prevented (70%) by HO-1 antisens e ODNs. HO-2 antisense ODN inhibited heme-induced HO activity by 21%. Addit ion of HO-1 antisense ODNs prevented heme degradation and resulted in eleva tion of microsomal heme. Western blot analysis showed that HO-1 antisense O DNs selectively inhibited HO-1 protein and failed to inhibit HO-2 protein. Incubation of endothelial cells with HO-1 antisense enhanced heme-dependent increase of ICAM-1. In contrast, addition of HO-2 antisense to endothelial cells failed to increase adhesion molecules. The role of glutathione, an i mportant antioxidant, was examined on heme-induced ICAM-1 expression. Endot helial cells pretreated with a glutathione precursor, N-acetylcysteine, or glutathione ester, showed a decrease in heme-induced ICAM-1 expression of 3 7 and 44%, respectively, suggesting that the mechanism of ICAM-1 induction by heme may be partly dependent on the levels of antioxidant. It is possibl e that amelioration of the heme-induced oxidative stress and expression of ICAM-1 is due, in part, to the induction of HO-1 activity. Regulation of HO activity in this manner may have clinical applications.