Electron paramagnetic resonance study of lipid and protein membrane components of erythrocytes oxidized with hydrogen peroxide
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Data
2012-05
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Resumo
Electron paramagnetic resonance (EPR) spectroscopy of spin labels was used to monitor membrane dynamic changes in
erythrocytes subjected to oxidative stress with hydrogen peroxide (H2O2). The lipid spin label, 5-doxyl stearic acid, responded
to dramatic reductions in membrane fluidity, which was correlated with increases in the protein content of the membrane.
Membrane rigidity, associated with the binding of hemoglobin (Hb) to the erythrocyte membrane, was also indicated by a spinlabeled
maleimide, 5-MSL, covalently bound to the sulfhydryl groups of membrane proteins. At 2% hematocrit, these alterations
in membrane occurred at very low concentrations of H2O2 (50 μM) after only 5 min of incubation at 37°C in azide phosphate
buffer, pH 7.4. Lipid peroxidation, suggested by oxidative hemolysis and malondialdehyde formation, started at 300 μM H2O2
(for incubation of 3 h), which is a concentration about six times higher than those detected with the probes. Ascorbic acid and
α-tocopherol protected the membrane against lipoperoxidation, but did not prevent the binding of proteins to the erythrocyte
membrane. Moreover, the antioxidant (+)-catechin, which also failed to prevent the cross-linking of cytoskeletal proteins with
Hb, was very effective in protecting erythrocyte ghosts from lipid peroxidation induced by the Fenton reaction. This study also
showed that EPR spectroscopy can be useful to assess the molecular dynamics of red blood cell membranes in both the lipid
and protein domains and examine oxidation processes in a system that is so vulnerable to oxidation.
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Electron paramagnetic resonance, Spin label, Erythrocyte, Membrane fluidity, Protein oxidation, Membrane oxidation
Citação
MENDANHA, S. A.; ANJOS, J. L. V.; SILVA, A. H. M.; ALONSO, A. Electron paramagnetic resonance study of lipid and protein membrane components of erythrocytes oxidized with hydrogen peroxide. Brazilian Journal of Medical and Biological Research, Ribeirão Preto, v. 45, n. 6, p. 473-481, May 2012.