Cardiovascular Research Advance Access first published online on April 7, 2009
This version [Corrected Proof] published online on April 29, 2009
Cardiovascular Research, doi:10.1093/cvr/cvp107
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Nrf2-dependent upregulation of antioxidative enzymes: a novel pathway for proteasome inhibitor-mediated cardioprotection
Medizinische Klinik für Kardiologie und Angiologie, Campus Mitte, Charité—Universitätsmedizin Berlin, Charitéplatz 1, D-10117 Berlin, Germany
* Corresponding author. Tel: +49 30 450513075; fax: +49 30 450513932. E-mail address: karl.stangl{at}charite.de
Aims: We have shown previously that non-toxic inhibition of the ubiquitin–proteasome system upregulates antioxidative defence mechanisms and protects endothelial cells from oxidative stress. Here, we have addressed the question whether the induction of antioxidative enzymes contributes to cardioprotection by non-toxic proteasome inhibition.
Methods and results: Treatment with 0.5 µmol/L MG132 for 48 h proved to be non-toxic and protected neonatal rat cardiac myocytes against H2O2-mediated oxidative stress in lactate dehydrogenase assays. This correlated with reduced levels of intracellular reactive oxygen species as determined by loading myocytes with dichlorofluorescein. Immunoblots showed significant upregulation of superoxide dismutase 1 (SOD1), haem oxygenase 1, and catalase upon proteasome inhibition. Luciferase assays using a reporter driven by the SOD1 promoter revealed proteasome inhibitor-mediated induction of luciferase activity. Deletion and mutation analyses identified an antioxidant response element (ARE) in the SOD1 promoter to be not only essential but also sufficient for transcriptional upregulation by proteasome inhibition. An essential role for the antioxidative transcription factor NF-E2-related factor 2 (Nrf2)—which was stabilized by proteasome inhibition—in ARE-mediated transcriptional activation was revealed in cardiac myocytes from Nrf2 wild-type and knockout mice: proteasome inhibition upregulated antioxidative enzymes and conferred protection against H2O2-mediated oxidative stress in Nrf2 wild-type cells. In contrast, the induction of antioxidative enzymes and cytoprotection were completely abolished in cardiac myocytes from Nrf2 knockout mice.
Conclusion: Non-toxic proteasome inhibition upregulates antioxidative enzymes via an Nrf2-dependent transcriptional activation of AREs and confers cardioprotection.
KEYWORDS Oxidative stress; Preconditioning; Cardiac myocytes; Proteasome inhibitor; Nrf2; Antioxidant response element
Time for primary review: 10 days
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  H. Dreger, K. Westphal, N. Wilck, G. Baumann, V. Stangl, K. Stangl, and S. Meiners Protection of vascular cells from oxidative stress by proteasome inhibition depends on Nrf2 Cardiovasc Res, September 4, 2009; (2009) cvp279v2. [Abstract] [Full Text] [PDF]
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