Submitted on December 17, 2008
Revised on January 14, 2009
Accepted on January 28, 2009
The nitric oxide-mediated protection of endothelial cells from hydrogen peroxide is mediated by intracellular zinc and glutathione
Miriam Margherita Cortese-Krott1*, Christoph V Suschek2, Wiebke Wetzel3, Klaus-Dietrich Kröncke4, and Victoria Kolb-Bachofen5
1 University Hospital of the RWTH Aachen University
2 RWTH Aachen University
3 Heinrich Heine Universität Düsseldorf
4 Heinrich-Heine University of Düsseldorf
5 University of Dusseldorf
* To whom correspondence should be addressed. E-mail: mcortese{at}ukaachen.de.
Oxidative stress may cause endothelial dysfunction and vascular disease. It has been shown that NO protects endothelial cells (EC) against H2O2-induced toxicity. In addition, it is known that NO within cells induces a zinc release from proteins containing zinc sulfur complexes. The aim of this study was to investigate whether zinc released intracellularly by NO plays a signaling role in the NO-mediated protection against H2O2 in rat aortic EC. Our results show that the NO-mediated protection towards H2O2 depends on the activities of glutathione peroxidase and glutamate cysteine ligase (GCL), the rate-limiting enzyme of GSH de novo biosynthesis. Moreover, NO increases the synthesis of the antioxidant glutathione (GSH) via inducing the expression of the catalytic subunit of GCL. Chelating intracellular "free" zinc abrogates the NO-mediated increase of GCLC and of cellular GSH levels. As a consequence, the NO-mediated protection against H2O2-induced toxicity is impaired. We also show that under pro-inflammatory conditions both cellular NO synthesis as well as intracellular "free" zinc are required to maintain the cellular GSH levels. By using RNA interference and laser scanning microscopy we found that the NO-induced expression of GCLC depends on the activation of the transcription factor Nrf2, but not on the activity of the "zinc sensing" transcription factor MTF-1. These findings show that intracellular "free" zinc plays a signaling role in the protective activity of NO, and could explain why maintenance of an adequate zinc status in the endothelium is important to protect from oxidative stress and the development of vascular disease.
