A matter of life, death and diseases: mitochondria from a proteomic perspective.

Warda, Mohamad,Kim, Hyoung Kyu,Kim, Nari,Ko, Kyung Soo,Rhee, Byoung Doo,Han, Jin Future Drugs Ltd 2013 Expert review of proteomics Vol.10 No.1

<P>Mitochondria are highly ordered, integrated organelles that energize cellular activities and contribute to programmed death by initiating disciplined apoptotic cascades. This review seeks to clarify our understanding of mitochondrial structural-functional integrity beyond the resolved nuclear genome by unraveling the dynamic mitochondrial proteome and elucidating proteome/genome interplay. The roles of mechanochemical coupling between mitoskeleton and cytoskeleton and crosstalk with other organelles in orchestrating cellular outcomes are explained. The authors also review the modulation of mitochondrial-related oxidative stress on apoptosis and cancer development and the context is applied to interpret pathogenetic events in neurodegenerative disorders and cardiovascular diseases. The accumulated proteomics evidence is used to describe the integral role that mitochondria play and how they influence other intracellular organelles. Possible mitochondrial-targeted therapeutic interventions are also discussed.</P>

Simulated hyperglycemia in rat cardiomyocytes: A proteomics approach for improved analysis of cellular alterations

Warda, Mohamad,Kim, Hyoung Kyu,Kim, Nari,Youm, Jae Boum,Kang, Sung Hyun,Park, Won Sun,Khoa, Tran Min,Kim, Young Hwan,Han, Jin WILEY-VCH Verlag 2007 Proteomics Vol.7 No.15

<P>Diabetic hyperglycemia can lead to stress-related cellular apoptosis of cardiac tissue. However, the mechanism by which hyperglycemia inflicts this damage on the structure and function of the heart is unclear. In this study, we examined the relationship between proteome alterations, mitochondrial function, and major biochemical and electrophysiological changes affecting cardiac performance during simulated short-term hyperglycemia. Two-dimensional comparative proteomics analysis of rat hearts perfused with glucose at high (30 mM) or control (5.5 mM) levels revealed that glucose loading alters cardiomyocyte proteomes. It increased expression levels of initial enzymes of the tricarboxylic acid cycle, and of enzymes of fatty acid β-oxidation, with consequent up-regulation of enzymes of mitochondrial electron transport. It also markedly decreased expression of enzymes of glycolysis and the final steps of the tricarboxylic acid cycle. Glucose loading increased the rate of Bax-independent apoptosis. High glucose increased the duration of the action potential and elevated level of intracellular cytoplasmic calcium. Surprisingly, glucose loading did not influence levels of nitric oxide or mitochondrial superoxide in isolated cardiomyocytes. In summary, short-term simulated hyperglycemia attenuated expression of many anti-apoptotic proteins. This effect was apparently mediated via alterations in multiple biochemical pathways that collectively increased apoptotic susceptibility.</P>

Toward a better understanding of preeclampsia: Comparative proteomic analysis of preeclamptic placentas

Kim, Young Nam,Kim, Hyoung Kyu,Warda, Mohamad,Kim, Nari,Park, Won Sun,Prince, Ab del Bary,Jeong, Dae Hoon,Lee, Dae Shim,Kim, Ki Tae,Han, Jin Wiley (John WileySons) 2007 Proteomics. Clinical applications Vol.1 No.12

<P>Preeclampsia (PE), a pregnancy-specific syndrome of hypertension, proteinuria, and other systemic disturbances, is a state of widespread endothelial dysfunction secondary to defective placentation. Morphologically, the current data displayed degenerative and apoptotic changes in the mitochondria and villous trophoblasts of preeclamptic placenta. To reveal the superimposing alterations in placental proteins that might explain the pathophysiology of PE, we performed 2-DE MALDI-TOF MS/MS proteomics analysis of differentially expressed placental proteins with placenta from eight normal and eight preeclamptic pregnancies. The identified proteins were confirmed by Western blot analysis. We also performed morphologic evaluation of preeclamptic placentas under both electron and light microscopy. The results disclosed the marked overexpression of chaperonin 60, GST, VDAC, ERp29, and cathepsin D in PE. These proteomics findings clearly suggest the possible cellular battle against mitochondria-originated oxidative stress during PE that either end up with recovery or apoptosis. These results provide a better understanding of proteomic alterations and may help in clarification of stress-related changes in preeclamptic placentas.</P>

Nitric oxide-cGMP-protein kinase G signaling pathway induces anoxic preconditioning through activation of ATP-sensitive K+ channels in rat hearts

Cuong, Dang Van,Kim, Nari,Youm, Jae Boum,Joo, Hyun,Warda, Mohamad,Lee, Jae-Wha,Park, Won Sun,Kim, Taeho,Kang, Sunghyun,Kim, Hyungkyu,Han, Jin American Physiological Society 2006 American journal of physiology, Heart and circulat Vol.290 No.5

<P> 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<SUP>+</SUP> (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 μM), nonspecific mitochondrial KATP channels opener pinacidil (50 μM), S-nitroso- N-acetylpenicillamine (SNAP, 300 μM), or β-phenyl-1, N<SUP>2</SUP>-etheno-8-bromoguanosine-3′,5′-cyclicmonophosphorothioate, Sp-isomer (10 μM) before the A/R period. Preconditioning with SNAP significantly reduced the DNA damage. The effect was blocked by glibenclamide (50 μM), 5-hydroxydecanoate (100 μM), N<SUP>G</SUP>-nitro-l-arginine methyl ester (200 μM), and β-phenyl-1, N<SUP>2</SUP>-etheno-8-bromoguanosine-3′,5′-cyclic monophosphorothioate, Rp-isomer (1 μM). 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. </P>

Cardiac Mitochondrial Inner Membrane Anion Channel during Ischemic-Reperfusion Injury

Kang, Sunghyun,Kim, Hyunggyu,Ko, Jaehong,Park, Won Sun,Mohamad Warda,Kim, Nari,Youm, Jae Boum,Han, Jin 이화여자대학교 세포신호전달연구센터 2008 고사리 세포신호전달 심포지움 Vol. No.10

Mitochondria are involved in oxidative phosphorylation, thermogenesis, reactive oxygen species. production, and intracellular Ca^(2+) homeostasis. Mitochondrial dysfunction during metabolic inhibition causes energy depletion and loss of cellular function and integrity, leading to cell death. Mitochondria possess a highly permeable outer membrane and an inner membrane that was originally thought to be relatively impermeable to ions to prevent dissipation of the electrochemical gradient for protons. Ion channels on the mitochondrial inner membrane influence membrane potential(ΔΨ_(m)) and cell function in specific ways that can be detrimental to cell survival. Mitochondrial K^(+) channels(mitoK_(Ca) and mitoK_(ATP)) are important determinants of resistance to ischemic damage and apoptosis, and may be clinically recruitable to prevent cardiac ischemic injury. In contrast, inner membrane anion channel(IMAC) initiates oscillations of mitochondrial redox and membrane potential, and thus might influence the overall function of the cell. In this study we recorded IMAC in cardiac mitochondrial inner membrane using nano patch-clamp techniques, and we tested whether IMAC confers protection against ischemic-reperfusion injury or not. We demonstrated that metabolic inhibition can trigger synchronized oscillations in ΔΨ_(m), ROS production, and mitochondrial redox potential. In the open-channel current-voltage curve, single channel currents with a full unitary conductance of 107 pS were often observed. DIDS, 4'-chlorodiazepham and PK11195 decreased the channel activity and prevented metabolic inhibition-induced ΔΨ_(m) loss. They also protected cardiac myocytes against ischemic-reperfusion injury. We are now trying to confirm the nature and molecular identity of the channel components. Our results suggest that IMAC is present in cardiac mitochondria and plays a role as key arbiters of cell life and death. Our studies may contribute to understanding the close relationship between mitochondrial ion channels, membrane potential, and the overall function of the cell. The ion channels should be present in all cell types containing mitochondria and the implications for normal as well as for pathophysiological cell function are universal.

Mitochondrial alterations in human gastric carcinoma cell line

Kim, Hyoung Kyu,Park, Won Sun,Kang, Sung Hyun,Warda, Mohamad,Kim, Nari,Ko, Jae-Hong,Prince, Abd El-bary,Han, Jin American Physiological Society 2007 American journal of physiology. Cell physiology Vol.293 No.2

<P>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<SUP>2+</SUP>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.</P>

Regional Differences in Mitochondrial Anti-oxidant State during Ischemic Preconditioning in Rat Heart

Vu Thi Thu,Dang Van Cuong,김나리,염재범,Mohamad Warda,박원선,고재홍,김의용,한진 대한약리학회 2007 The Korean Journal of Physiology & Pharmacology Vol.11 No.2

Ischemic preconditioning (IPC) is known to protect the heart against ischemia/reperfusion (IR)-induced injuries, and regional differences in the mitochondrial antioxidant state during IR or IPC may promote the death or survival of viable and infarcted cardiac tissues under oxidative stress. To date, however, the interplay between the mitochondrial antioxidant enzyme system and the level of reactive oxygen species (ROS) in the body has not yet been resolved. In the present study, we examined the effects of IR- and IPC-induced oxidative stresses on mitochondrial function in viable and infarcted cardiac tissues. Our results showed that the mitochondria from viable areas in the IR-induced group were swollen and fused, whereas those in the infarcted area were heavily damaged. IPC protected the mitochondria, thus reducing cardiac injury. We also found that the activity of the mitochondrial antioxidant enzyme system, which includes manganese superoxide dismutase (Mn-SOD), was enhanced in the viable areas compared to the infarcted areas in proportion with decreasing levels of ROS and mitochondrial DNA (mtDNA) damage. These changes were also present between the IPC and IR groups. Regional differences in Mn-SOD expression were shown to be related to a reduction in mtDNA damage as well as to the release of mitochondrial cytochrome c (Cyt c). To the best of our knowledge, this might be the first study to explore the regional mitochondrial changes during IPC. The present findings are expected to help elucidate the molecular mechanism involved in IPC and helpful in the development of new clinical strategies against ischemic heart disease.

Regional Differences in Mitochondrial Anti-oxidant State during Ischemic Preconditioning in Rat Heart

Thu, Vu Thi,Cuong, Dang Van,Kim, Na-Ri,Youm, Jae-Boum,Warda, Mohamad,Park, Won-Sun,Ko, Jae-Hong,Kim, Eui-Yong,Han, Jin The Korean Society of Pharmacology 2007 The Korean Journal of Physiology & Pharmacology Vol.11 No.2

Ischemic preconditioning (IPC) is known to protect the heart against ischemia/reperfusion (IR)-induced injuries, and regional differences in the mitochondrial antioxidant state during IR or IPC may promote the death or survival of viable and infarcted cardiac tissues under oxidative stress. To date, however, the interplay between the mitochondrial antioxidant enzyme system and the level of reactive oxygen species (ROS) in the body has not yet been resolved. In the present study, we examined the effects of IR- and IPC-induced oxidative stresses on mitochondrial function in viable and infarcted cardiac tissues. Our results showed that the mitochondria from viable areas in the IR-induced group were swollen and fused, whereas those in the infarcted area were heavily damaged. IPC protected the mitochondria, thus reducing cardiac injury. We also found that the activity of the mitochondrial antioxidant enzyme system, which includes manganese superoxide dismutase (Mn-SOD), was enhanced in the viable areas compared to the infarcted areas in proportion with decreasing levels of ROS and mitochondrial DNA (mtDNA) damage. These changes were also present between the IPC and IR groups. Regional differences in Mn-SOD expression were shown to be related to a reduction in mtDNA damage as well as to the release of mitochondrial cytochrome c (Cyt c). To the best of our knowledge, this might be the first study to explore the regional mitochondrial changes during IPC. The present findings are expected to help elucidate the molecular mechanism involved in IPC and helpful in the development of new clinical strategies against ischemic heart disease.

Increased Inhibition of Inward Rectifier K ⁺ Channels by Angiotensin II in Small-Diameter Coronary Artery of Isoproterenol-Induced Hypertrophied Model

Park, Won Sun,Ko, Jae-Hong,Kim, Nari,Son, Youn Kyoung,Kang, Sung Hyun,Warda, Mohamad,Jung, In Duk,Park, Yeong-Min,Han, Jin Ovid Technologies Wolters Kluwer -American Heart A 2007 Arteriosclerosis, thrombosis, and vascular biology Vol.27 No.8

Tissue Microarrays in Biomedical Research

Chung, Joon-Yong,Kim, Nari,Joo, Hyun,Youm, Jae-Boum,Park, Won-Sun,Lee, Sang-Kyoung,Warda, Mohamad,Han, Jin Korean Society for Bioinformatics and Systems Biol 2006 Bioinformatics and Biosystems Vol.1 No.1

Recent studies in molecular biology and proteomics have identified a significant number of novel diagnostic, prognostic, and therapeutic disease markers. However, validation of these markers in clinical specimens with traditional histopathological techniques involves low throughput and is time consuming and labor intensive. Tissue microarrays (TMAs) offer a means of combining tens to hundreds of specimens of tissue onto a single slide for simultaneous analysis. This capability is particularly pertinent in the field of cancer for target verification of data obtained from cDNA micro arrays and protein expression profiling of tissues, as well as in epidemiology-based investigations using histochemical/immunohistochemical staining or in situ hybridization. In combination with automated image analysis, TMA technology can be used in the global cellular network analysis of tissues. In particular, this potential has generated much excitement in cardiovascular disease research. The following review discusses recent advances in the construction and application of TMAs and the opportunity for developing novel, highly sensitive diagnostic tools for the early detection of cardiovascular disease.