Cyclosporine A-induced nitration of tyrosine 34 MnSOD in endothelial cells: role of mitochondrial superoxide
| Autor: | Santiago Lamas, Eduardo Rial, Natalia Romero, Mariano Redondo-Horcajo, Pablo Martínez-Acedo, Antonio Martínez-Ruiz, Cátia F. Lourenço, José Antonio Enríquez, Jesús Vázquez, Nieves Movilla, Celia Quijano, Rafael Radi, Fernando Rodríguez-Pascual |
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| Rok vydání: | 2010 |
| Předmět: |
Nitric Oxide Synthase Type III
Physiology Cell Respiration Mitochondrion Nitric Oxide Transfection Mass Spectrometry Peroxynitrite Nitroxidative stress Nitric oxide Superoxide dismutase Mice chemistry.chemical_compound Superoxides Calcineurin inhibitors Peroxynitrous Acid Physiology (medical) Animals Cells Cultured Reactive nitrogen species Aconitate Hydratase Membrane Potential Mitochondrial Mice Knockout biology Superoxide Dismutase Superoxide Electron Spin Resonance Spectroscopy Endothelial Cells Recombinant Proteins Mitochondria Cell biology Mice Inbred C57BL Endothelial stem cell Peroxynitrous acid Biochemistry chemistry Cyclosporine biology.protein Tyrosine Cattle Vascular toxicity Cardiology and Cardiovascular Medicine Immunosuppressive Agents |
| Zdroj: | Digital.CSIC. Repositorio Institucional del CSIC instname |
| ISSN: | 1755-3245 0008-6363 |
| Popis: | 10 páginas, 5 figuras -- PAGS nros. 356-365 Aims Cyclosporine A (CsA) has represented a fundamental therapeutic weapon in immunosupression for the past three decades. However, its clinical use is not devoid of side effects, among which hypertension and vascular injury represent a major drawback. Endothelial cells are able to generate reactive oxygen and nitrogen species upon exposure to CsA, including formation of peroxynitrite. This may result in endothelial cell toxicity and increased tyrosine nitration. We have now studied the subcellular origin of superoxide formation in endothelial cells treated with CsA and the biochemical consequences for the function of mitochondrial enzymes. Methods and results By using electron spin resonance and endothelial cells lacking functional mitochondria, we showed that superoxide anion is generated in mitochondria. This was associated with an effect of CsA on bioenergetic parameters: increased mitochondrial membrane potential and inhibition of cellular respiration. In addition, CsA inhibited the activity of the mitochondrial enzymes aconitase and manganese superoxide dismutase (MnSOD). The use of murine lung endothelial cells deficient in endothelial nitric oxide synthase (eNOS) and NOS/peroxynitrite inhibitors allowed us to establish that the presence of eNOS and concomitant NO synthesis and peroxynitrite formation were essential for CsA induced nitration and inhibition of MnSOD activity. As the latter has been shown to become inactivated by nitration, we sought to identify this modification by mass spectrometry analysis. We found that CsA induced specific MnSOD tyrosine 34 nitration both in the recombinant protein and in endothelial cells overexpressing MnSOD. Conclusion We propose that CsA induced endothelial damage may be related to increased mitochondrial superoxide formation and subsequent peroxynitrite-dependent nitroxidative damage, specifically targeting MnSOD. The inactivation of this key antioxidant enzyme by tyrosine nitration represents a pathophysiological cellular mechanism contributing to self-perpetuation and amplification of CsA-related vascular toxicity This work was supported by grants [SAF 2006-02410] and Consolider [CSD-2007-0020]. Network of Excellence for the Research on Oxidative Stress in Spain, Reactive Oxygen Species and Systems (ROSAS) from Plan Nacional de I + D + I; CARDIOVREP from Comunidad de Madrid; an Accion Integrada [2007UY0018 CSIC], Spain- Universidad de la Republica, Uruguay; a grant from the Sociedad Española de Nefrología and Fundación Renal Iñigo Alvarez de Toledo to S.L.; a ‘Miguel Servet’ grant [CP07/00143] from Instituto de Salud Carlos III (Spain) to A.M.-R.; a grant from Fundación de Investigación Médica Mutua Madrileña to F.R.-P.; by grants [BIO2006-10085, GR/SAL/0141/2004 (CAM), CAM BIO/0194/2006], from the Fondo de Investigaciones Sanitarias (Ministerio de Sanidad y Consumo, Instituto Salud Carlos III, RECAVA) and by an institutional grant by Fundación Ramón Areces to CBMSO to J.V. N.R. received grant support from Fondo Clemente Estable/Agencia Nacional de Investigación e Innovación-Uruguay and R.R. from the Howard Hughes Medical Institute and the International Centre for Genetic Engineering and Biotechnology |
| Databáze: | OpenAIRE |
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