title: Reversible blockade of complex I or inhibition of PKCβ reduces activation and mitochondria translocation of p66Shc to preserve cardiac function after ischemia.
abstract: OBJECTIVE
  Excess mitochondrial reactive oxygen species (mROS) play a vital role in cardiac ischemia reperfusion (IR) injury. P66Shc, a splice variant of the ShcA adaptor protein family, enhances mROS production by oxidizing reduced cytochrome c to yield H2O2. Ablation of p66Shc protects against IR injury, but it is unknown if and when p66Shc is activated during cardiac ischemia and/or reperfusion and if attenuating complex I electron transfer or deactivating PKCβ alters p66Shc activation during IR is associated with cardioprotection.
  
  
  METHODS
  Isolated guinea pig hearts were perfused and subjected to increasing periods of ischemia and reperfusion with or without amobarbital, a complex I blocker, or hispidin, a PKCβ inhibitor. Phosphorylation of p66Shc at serine 36 and levels of p66Shc in mitochondria and cytosol were measured. Cardiac functional variables and redox states were monitored online before, during and after ischemia. Infarct size was assessed in some hearts after 120 min reperfusion.
  
  
  RESULTS
  Phosphorylation of p66Shc and its translocation into mitochondria increased during reperfusion after 20 and 30 min ischemia, but not during ischemia only, or during 5 or 10 min ischemia followed by 20 min reperfusion. Correspondingly, cytosolic p66Shc levels decreased during these ischemia and reperfusion periods. Amobarbital or hispidin reduced phosphorylation of p66Shc and its mitochondrial translocation induced by 30 min ischemia and 20 min reperfusion. Decreased phosphorylation of p66Shc by amobarbital or hispidin led to better functional recovery and less infarction during reperfusion.
  
  
  CONCLUSIONS
  Our results show that IR activates p66Shc and that reversible blockade of electron transfer from complex I, or inhibition of PKCβ activation, decreases p66Shc activation and translocation and reduces IR damage. These observations support a novel potential therapeutic intervention against cardiac IR injury.
authors.affiliation: Department of Anesthesiology, Medical College of Wisconsin, Milwaukee, WI, United States of America.
authors.forename: Meiying
authors.initials: M
authors.lastname: Yang
authors.affiliation: Department of Anesthesiology, Medical College of Wisconsin, Milwaukee, WI, United States of America; Department of Physiology, Medical College of Wisconsin, Milwaukee, WI, United States of America; Cardiovascular Research Center, Medical College of Wisconsin, Milwaukee, WI, United States of America; Research Service, Zablocki VA Medical Center, Milwaukee, WI, United States of America; Department of Biomedical Engineering, Marquette University, Milwaukee, WI, United States of America.
authors.forename: David F
authors.initials: DF
authors.lastname: Stowe
authors.affiliation: Department of Anesthesiology, Medical College of Wisconsin, Milwaukee, WI, United States of America.
authors.forename: Kenechukwu B
authors.initials: KB
authors.lastname: Udoh
authors.affiliation: Department of Anesthesiology, Medical College of Wisconsin, Milwaukee, WI, United States of America.
authors.forename: James S
authors.initials: JS
authors.lastname: Heisner
authors.affiliation: Department of Anesthesiology, Medical College of Wisconsin, Milwaukee, WI, United States of America; Cardiovascular Research Center, Medical College of Wisconsin, Milwaukee, WI, United States of America.
authors.forename: Amadou K S
authors.initials: AK
authors.lastname: Camara
mesh_terms: D000654:Amobarbital / Q000494:pharmacology; D000818:Animals; D004579:Electron Transport / Q000187:drug effects; D042967:Electron Transport Complex I / Q000037:antagonists & inhibitors*; D004789:Enzyme Activation / Q000187:drug effects; D006168:Guinea Pigs; D006321:Heart / Q000187:drug effects / Q000503:physiopathology*; D008928:Mitochondria / Q000187:drug effects / Q000378:metabolism*; D017202:Myocardial Ischemia / Q000378:metabolism / Q000473:pathology / Q000503:physiopathology; D064546:Protein Kinase C beta / Q000037:antagonists & inhibitors* / Q000378:metabolism; D047428:Protein Kinase Inhibitors / Q000494:pharmacology*; D021381:Protein Transport / Q000187:drug effects; D015427:Reperfusion Injury / Q000378:metabolism / Q000473:pathology / Q000503:physiopathology*; D055765:Shc Signaling Adaptor Proteins / Q000378:metabolism*
chemical_list: D047428:Protein Kinase Inhibitors; D055765:Shc Signaling Adaptor Proteins; D064546:Protein Kinase C beta; D042967:Electron Transport Complex I; D000654:Amobarbital