Redox activation of aldose reductase in the ischemic heart.
Aldose reductase (AR) reduces cytotoxic aldehydes and glutathione conjugates of aldehydes derived from lipid peroxidation. Its inhibition has been shown to increase oxidative injury and abolish the late phase of ischemic preconditioning. However, the mechanisms by which ischemia regulates AR activity remain unclear. Herein, we report that rat hearts subjected to ischemia, in situ or ex vivo, display a 2-4-fold increase in AR activity. The AR activity was not further enhanced by reperfusion. Activation increased Vmax of the enzyme without affecting the Km and decreased the sensitivity of the enzyme to inhibition by sorbinil. Enzyme activation could be prevented by pretreating the hearts with the radical scavenging thiol, N-(2-mercaptoproprionyl)glycine or the superoxide dismutase mimetic, Tiron, or by treating homogenates with dithiothreitol. In vitro, the recombinant enzyme was activated upon treatment with H2O2 and the activated, but not the native enzyme, formed a covalent adduct with the sulfenic acid-specific reagent dimedone. The enzyme activity in the ischemic, but not the nonischemic heart homogenates was inhibited by dimedone. Separation of proteins from hearts subjected to coronary occlusion by two-dimensional electrophoresis and subsequent matrix-assisted laser desorption ionization time-of-flight/mass spectrometry analysis revealed the formation of sulfenic acids at Cys-298 and Cys-303. These data indicate that reactive oxygen species formed in the ischemic heart activate AR by modifying its cysteine residues to sulfenic acids.
Institute of Molecular Cardiology, Department of Physiology and Biophysics, University of Louisville, Louisville, Kentucky 40202, USA.
Karin
K
Kaiserova
Sanjay
S
Srivastava
Joseph D
JD
Hoetker
Sunday O
SO
Awe
Xian-Liang
XL
Tang
Jian
J
Cai
Aruni
A
Bhatnagar
D000449:Aldehyde Reductase / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D000818:Animals; D003545:Cysteine / Q000737:chemistry / Q000378:metabolism; D015180:Electrophoresis, Gel, Two-Dimensional; D004789:Enzyme Activation / Q000187:drug effects; D016166:Free Radical Scavengers / Q000494:pharmacology; D006861:Hydrogen Peroxide / Q000494:pharmacology; D066298:In Vitro Techniques; D007700:Kinetics; D008297:Male; D017202:Myocardial Ischemia / Q000201:enzymology*; D009206:Myocardium / Q000201:enzymology*; D010084:Oxidation-Reduction; D010477:Perfusion; D051381:Rats; D017207:Rats, Sprague-Dawley; D017382:Reactive Oxygen Species / Q000378:metabolism; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D013434:Sulfenic Acids / Q000737:chemistry / Q000378:metabolism
D016166:Free Radical Scavengers; D017382:Reactive Oxygen Species; D013434:Sulfenic Acids; D006861:Hydrogen Peroxide; D000449:Aldehyde Reductase; D003545:Cysteine