Heme oxygenase-1 inhibition of Nox oxidase activation is a microvascular endothelial antioxidant effect of NO.

Astudy by Jiang et al 1 in this issue of Hypertension reports evidence suggesting that NO has a novel antioxidant-type effect in cultured human microvascular endothelial cells associated with an inhibition of Nox oxidase activation, which seems to be mediated by increasing the expression and activity of heme oxygenase-1 (HO-1). The induction of HO-1 was observed to occur when the cultured endothelium was exposed to a long-acting NO donor for 6 hours. Because freshly isolated vascular tissue from normal animals is generally thought to contain low levels of HO-1, the levels of NO seen in endothelium under physiological conditions do not seem to be a stimulus for maintaining an elevated level of this enzyme. Physiological stresses associated with oxidant production were initially observed to increase HO-1 expression in multiple tissue preparations.2 The formation of reactive NO-derived species (NOx) from the reaction of NO with superoxide, such as peroxynitrite, were initially observed to promote HO-1 expression in endothelial cells exposed to NO donors in a manner that was modulated by free thiol availability.3 Although the precise mechanism involved in how NOx induces HO-1 expression is currently not known, the gene for HO-1 has many sites in its promoter region for redox regulation by intracellular and nuclear signaling events that lead to transcriptional activation.2 Thus, increases in HO-1 expression by elevated levels of NO are likely to occur when endothelium is exposed to physiological stress or pathophysiological conditions associated with vascular disease processes, such as hypertension. The decreased detection of superoxide associated with induction of HO-1 by NO in endothelium was investigated to assess whether it was related to alterations in the expression of Nox oxidase subunits or Cu,Zn-superoxide dismutase (SOD) or a pattern of effects that could be equated with the increased activity of HO-1.1 Inhibition of HO activity or prevention of increased HO-1 expression with small interfering RNA attenuated the effects of exposure to NO, suggesting that the catalytic activity of HO-1 and not an alternative effect of NO on superoxide scavenging is responsible for the antioxidant effects of NO that were detected. The heme-depleting activity resulting from increased HO-1 expression was observed in a previous study4 to decrease NADPH oxidase activity in macrophages through processes thought to involve impairment of protein maturation and increased degradation in its heme-containing catalytic component, the gp91 (Nox2) subunit. However, in the present study it was observed that cellular levels of the key heme-containing Nox subunits present in endothelium, other subunits thought to regulate the Nox oxidases investigated, and Cu,Zn-SOD were not altered by the increased HO-1 expression elicited by exposure to NO.1 These data on the absence of detectible changes in Nox oxidase expression also rule out another recently reported mechanism5 involving NO donors inhibiting the expression of mRNA for Nox1. Thus, the superoxide-depressing effects of exposure of endothelium to NO seemed to be associated with an increased HO-1 activity and not with an alternative antioxidant action of NO or a change in the expression of Cu,Zn-SOD or key components of Nox oxidases present in endothelium. The product of the metabolism of heme by the HO-1 reaction, bilirubin, was observed to have an action potentially associated with decreasing Nox oxidase activation by a p47-dependent mechanism,1 which could contribute to the antioxidant effects of increased HO-1 expression by NO. An early observation made in studies characterizing modulation of the activation of Nox oxidase in neutrophils was that bilirubin, a key metabolic product of increased heme metabolism by HO, interacted with a cytosolic component in a manner that inhibited the activation of superoxide production by this oxidase.6 Bilirubin has also been observed to inhibit what seems to be a growth factor activation of Nox oxidases in airway smooth muscle cells.7 Although the mechanism through which bilirubin inhibits Nox oxidase activation is not known, data in the study of Jiang et al1 suggest that this agent may interfere with the membrane binding of p47 associated with Nox oxidase activation or possibly the protein kinase C-mediated regulation of this pathway. Regulation by protein kinase C–p47 is likely to activate Nox oxidases containing Nox1 or Nox2 but not the Nox4 subunits8 of the oxidases, which are observed to be present in the endothelial cells studied.1 Overall, the results of the study of Jiang et al1 support a mechanism shown in the Figure according to which prolonged exposure to NO elicits increased expression of HO-1, and the generation of bilirubin, resulting from the metabolism of heme by this enzyme, attenuates Nox oxidase–derived superoxide production by preventing oxidase activation by p47dependent pathways. Based on previous work in the literature demonstrating the potential importance of NOx in increasing the expression of HO-1 and the redox stress-associated regulation of transcriptional activation of this gene,2,3 NOx derived from the reaction of superoxide with NO or the oxidation of NO are likely to mediate the increase in HO-1 expression caused by prolonged exposure of endothelium to NO. The opinions expressed in this editorial are not necessarily those of the editors or of the American Heart Association. From the Departments of Physiology (M.S.W.) and Pharmacology (N.G.A.), New York Medical College, Valhalla, NY. Correspondence to Michael S. Wolin, Department of Physiology, Basic Science Bldg, Room 604, New York Medical College, Valhalla, NY 10595. E-mail [email protected] (Hypertension. 2006;48:1-2.) © 2006 American Heart Association, Inc.