We used proteomics to detect regional differences in protein expression levels from mitochondrial fractions of control, ischemia-reperfusion (IR), and ischemic preconditioned (IPC) rabbit hearts. Using 2-DE, we identified 25 mitochondrial proteins that were differentially expressed in the IR heart compared with the control and IPC hearts. For three of the spots, the expression patterns were confirmed by Western blotting analysis. These proteins included 3-hydroxybutyrate dehydrogenase, prohibitin, 2-oxoglutarate dehydrogenase, adenosine triphosphate synthases, the reduced form of nicotinamide adenine dinucleotide (NADH) oxidoreductase, translation elongation factor, actin alpha, malate dehydrogenase, NADH dehydrogenase, pyruvate dehydrogenase and the voltage-dependent anion channel. Interestingly, most of these proteins are associated with the mitochondrial respiratory chain and energy metabolism. The successful use of multiple techniques, including 2-DE, MALDI-TOF-MS and Western blotting analysis demonstrates that proteomic analysis provides appropriate means for identifying cardiac markers for detection of ischemia-induced cardiac injury.

No abstract

Heparan sulfate (HS), is a proteoglycan (PG) found both in the extracellular matrix and on cell surface. It may represent one of the most biologically important glycoconjugates, playing an essential role in a variety of different events at molecular level. The publication of the mouse genome, and the intensive investigations aimed at understanding the proteome it encodes, has motivated us to initiate studies in mouse glycomics focused on HS. The current study is aimed at determining the quantitative and qualitative organ distribution of HS in mice. HS from brain, eyes, heart, lung, liver, kidney, spleen, intestine and skin was purified from 6-8 week old male and female mice. The recovered yield of HS from these organs is compared with the recovered whole body yield of HS. Structural characterization of the resulting HS relied on disaccharide analysis and 1 H-NMR spectroscopy. Different organs revealed a characteristic HS structure. These data begin to provide a structural understanding of the role of HS in cell-cell interactions, cell signaling

. Nitric oxidecGMP-protein kinase G signaling pathway induces anoxic preconditioning through activation of ATP-sensitive K ϩ channels in rat hearts. Am J Physiol Heart Circ Physiol 290: H1808 -H1817, 2006. First published December 9, 2005 doi:10.1152/ajpheart.00772.2005.-Nitric oxide (NO) plays an important role in anoxic preconditioning to protect the heart against ischemia-reperfusion injuries. The present work was performed to study better the NO-cGMP-protein kinase G (PKG) signaling pathway in the activation of both sarcolemmal and mitochondrial ATP-sensitive K ϩ (KATP) channels during anoxic preconditioning (APC) and final influence on reducing anoxia-reperfusion (A/R)-induced cardiac damage in rat hearts. The upstream regulating elements controlling NO-cGMP-PKG signal-induced KATP channel opening that leads to cardioprotection were investigated. The involvement of both inducible and endothelial NO synthases (iNOS and eNOS) in the progression of this signaling pathway was followed. Final cellular outcomes of ischemia-induced injury after different preconditioning in the form of lactate dehydrogenase release, DNA strand breaks, and malondialdehyde formation as indexes of cell injury and lipid peroxidation, respectively, were investigated. The lactate dehydrogenase and malondialdehyde values decreased in the groups that underwent preconditioning periods with specific mitochondrial KATP channels opener diazoxide (100 M), nonspecific mitochondrial KATP channels opener pinacidil (50 M), S-nitroso-Nacetylpenicillamine (SNAP, 300 M), or ␤-phenyl-1,N 2 -etheno-8-bromoguanosine-3Ј,5Ј-cyclicmonophosphorothioate, Sp-isomer (10 M) before the A/R period. Preconditioning with SNAP significantly reduced the DNA damage. The effect was blocked by glibenclamide (50 M), 5-hydroxydecanoate (100 M), N G -nitro-L-arginine methyl ester (200 M), and ␤-phenyl-1,N 2 -etheno-8-bromoguanosine-3Ј,5Ј-cyclic monophosphorothioate, Rp-isomer (1 M). The results suggest iNOS, rather than eNOS, as the major contributing NO synthase during APC treatment. Moreover, the PKG shows priority over NO as the upstream regulator of NO-cGMP-PKG signal-induced KATP channel opening that leads to cardioprotection during APC treatment.guanosine 3Ј,5Ј-cyclic monophosphate; adenosine 5Ј-triphosphate; oxidative damage ISCHEMIC PRECONDITIONING, in which short-term occlusion and reperfusion of a coronary artery are followed by long-term occlusion, can reduce subsequent ischemia-induced injury to the heart (42). Nitric oxide (NO), protein kinase G (PKG), and ATP-sensitive K ϩ (K ATP ) channels (both the sarcolemmal and mitochondrial subtypes) can mimic the effects of ischemic preconditioning in the heart, and mitochondrial K ATP channels appear to be the end effectors (19,20,25). The activation of these channels may improve the recovery of regional contractility of myocardium by shortening the duration of action potentials and by attenuating membrane depolarization, both of which would decrease myocardial contractility and reduce energy expenditure during ischem...

We compared mitochondrial function, morphology, and proteome in the rat normal gastric cell line RGM-1 and the human gastric cancer cell line AGS. Total numbers and cross-sectional sizes of mitochondria were smaller in AGS cells. Mitochondria in AGS cells were deformed and consumed less oxygen. Confocal microscopy indicated that the mitochondrial inner membrane potential was hyperpolarized and the mitochondrial Ca 2ϩ concentration was elevated in AGS cells. Interestingly, two-dimensional electrophoresis proteomics on the mitochondria-enriched fraction revealed high expression of four mitochondrial proteins in AGS cells: ubiquinol-cytochrome c reductase, mitochondrial short-chain enoyl-coenzyme A hydratase-1, heat shock protein 60, and mitochondria elongation factor Tu. The results provide clues as to the mechanism of the mitochondrial changes in cancer at the protein level and may serve as potential cancer biomarkers in mitochondria.