Recent observations have led to the hypothesis that significant mitochondrial damage occurs during ischemia due to the production of reactive oxygen species (ROS) under the hypoxic and other conditions that prevail. Because of the difficulty of studying reactions at controlled PO2, few studies of mitochondrial ROS production during hypoxia exist. This absence of information leaves an unexplained conundrum: how does ROS production increase as the substrate 02, decreases. Defects identified in elderly heart mitochondria are hypothesized to augment ROS production during ischemia, resulting in increased oxidative damage during ischemic periods in the elderly heart. The first two specific aims test two alternative mechanisms that could individually or in combination account for the enhanced production of ROS during ischemia. The first hypothesis is that the carriers in the electron transport chain are more reduced, resulting in sites of increased leakage to a low but nonzero 02 concentration, to generate . O2. The second mechanism is that the enhanced ROS production comes from changes in the cellular environment induced as a response to ischemia and/or hypoxia. Among these cellular responses are the release of Fe, the influx of Ca ++, a decrease in pH, and the production of NO. A novel apparatus has been developed that permits the production of ROS to be monitored, concurrent with continuous respiratory utilization of 02 present at controlled variable partial pressures of 2-15 torr. Using submitochondrial particles, the variation in PO2 has been shown to alter both H202 and superoxide generation. The production of these and other ROS will be monitored as a function of the presence of Fe(II), Ca 2+, decreased pH and NO, all at concentrations mimicking physiological changes observed during ischemia. The response of both adult and elderly and both subsarcolemmal and interfibrillar mitochondria will be compared. The site of ROS production, under conditions that lead to the greatest ROS generation, will be investigated by varying the reducing substrate in the presence of specific inhibitors as well as comparing results obtained with submitochondrial particles, mitoplasts and mitochondria. In the third specific aim the chemical damage inflicted by the ROS on proteins and cardiolipin will be investigated by mass spectrometric methods. Methods of isolating the individual electron transport complexes for MS analyses have been developed so that the location of the most significant functional and structural damage can now be compared. In the fourth specific aim, the variations in mitochondria detected or engineered by the other three projects will be mimicked in our system so that their effects on the direct production of mitochondrial ROS can be characterized. Additionally, therapeutic interventions suggested by the PPG studies will be tested.

Agency
National Institute of Health (NIH)
Institute
National Institute on Aging (NIA)
Type
Research Program Projects (P01)
Project #
5P01AG015885-09
Application #
7651107
Study Section
Special Emphasis Panel (ZAG1)
Project Start
Project End
Budget Start
2008-08-01
Budget End
2009-07-31
Support Year
9
Fiscal Year
2008
Total Cost
$193,905
Indirect Cost
Name
Case Western Reserve University
Department
Type
DUNS #
077758407
City
Cleveland
State
OH
Country
United States
Zip Code
44106
Kerner, Janos; Yohannes, Elizabeth; Lee, Kwangwon et al. (2015) Acetyl-L-carnitine increases mitochondrial protein acetylation in the aged rat heart. Mech Ageing Dev 145:39-50
Minkler, Paul E; Stoll, Maria S K; Ingalls, Stephen T et al. (2015) Quantitative acylcarnitine determination by UHPLC-MS/MS--Going beyond tandem MS acylcarnitine ""profiles"". Mol Genet Metab 116:231-41
Xu, Aijun; Szczepanek, Karol; Maceyka, Michael W et al. (2014) Transient complex I inhibition at the onset of reperfusion by extracellular acidification decreases cardiac injury. Am J Physiol Cell Physiol 306:C1142-53
Dadabayev, Alisher R; Yin, Guotian; Latchoumycandane, Calivarathan et al. (2014) Apolipoprotein A1 regulates coenzyme Q10 absorption, mitochondrial function, and infarct size in a mouse model of myocardial infarction. J Nutr 144:1030-6
Kerner, Janos; Minkler, Paul E; Lesnefsky, Edward J et al. (2014) Fatty acid chain elongation in palmitate-perfused working rat heart: mitochondrial acetyl-CoA is the source of two-carbon units for chain elongation. J Biol Chem 289:10223-34
Solinas, Paola; Fujioka, Hisashi; Radivoyevitch, Tomas et al. (2014) Aging effects on oxidative phosphorylation in rat adrenocortical mitochondria. Mech Ageing Dev 138:10-4
Gao, Xing-Huang; Qanungo, Suparna; Pai, Harish V et al. (2013) Aging-dependent changes in rat heart mitochondrial glutaredoxins--Implications for redox regulation. Redox Biol 1:586-98
Kim, Junhwan; Hoppel, Charles L (2013) Comprehensive approach to the quantitative analysis of mitochondrial phospholipids by HPLC-MS. J Chromatogr B Analyt Technol Biomed Life Sci 912:105-14
Solinas, Paola; Fujioka, Hisashi; Tandler, Bernard et al. (2012) Isolation of rat adrenocortical mitochondria. Biochem Biophys Res Commun 427:96-9
Xi, Lei; Zhu, Shu-Guang; Das, Anindita et al. (2012) Dietary inorganic nitrate alleviates doxorubicin cardiotoxicity: mechanisms and implications. Nitric Oxide 26:274-84

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