Oxidative stress — an idiosyncrasy of nature

Danielle Feuvray
Physiologie Cellulaire, Université Paris-Sud XI, France
Correspondence: Professor Danielle Feuvray, Physiologie Cellulaire, CNRS UMR 8078,
Bâtiment 443, Université Paris XI, 91405 Orsay, France.
Tel: +33 1 69157898/40946732, e-mail: danielle.feuvray@ibaic.u-psud.fr

The idiosyncrasies of nature are such that molecular oxygen, which is essential for maintaining cell viability, may also act as a trigger for tissue injury. This was originally highlighted by the observation of David Hearse and colleagues of an “oxygen paradox.” The paradox was that, upon abrupt reoxygenation of an anoxic heart, myocardial injury was suddenly and markedly enhanced [1]. Since this initial observation, a large number of basic and clinical investigations have aimed at understanding the mechanisms involved and the functional consequences associated with oxidative stress. It has in particular become apparent that residual molecular oxygen in the ischemic myocardium, and more clearly the reintroduction of oxygen into the ischemic myocardium, results in the formation of oxygen-derived free radicals. Oxidative stress, which is usually associated with an increase in the formation of reactive oxygen species, modifies phospholipids and proteins, leading to lipid peroxidation and oxidation of thiol groups. These changes are thought to alter cell membrane permeability and configuration, in addition to producing functional modifications in various cellular proteins. This is summarized in two articles in this issue. Joël de Leiris provides a concise review of the biochemistry of free radicals, how they are produced or overproduced, and how they can be counteracted. It is the purpose of Sandeep Raha and Brian Robinson to focus mainly on how the mitochondria, a major source of cellular free radicals, contributes to the regulation of free radical metabolism.
Basic research is continually discovering new effects of excessive production of reactive oxygen species in the different aspects of cardiovascular disease. It is known that damage to mitochondria enhances the production of reactive oxygen species; and an increase in the formation of reactive oxygen species during ischemia-reperfusion has been reported using the electron paramagnetic resonance technique in both humans and animals [2, 3]. In addition, since investigations have reported a depletion of endogenous antioxidants in the ischemic heart upon reperfusion [4],it appears especially important to focus on antioxidant systems. Direct evidence, using genetically engineered animal models, has recently been presented to show the importance of several naturally occurring scavenging enzyme systems, such as catalase, superoxide dismutase, and glutathione peroxidase, in protecting the myocardium against ischemia-reperfusion injury [5–7]. During normal respiration, extensive oxidative damage is prevented by mitochondrial antioxidant enzyme systems [8]. Thus, manganese-containing mitochondrial superoxide dismutase, located in the mitochondrial matrix, eliminates superoxide radical °O2- by catalyzing dismutation to hydrogen peroxide H2O2. H2O2 is then inactivated by either catalase or by the glutathione redox system consisting of reduced glutathione as the cofactor for glutathione peroxidase and glutathione reductase. Although the relative contributions of catalase and glutathione peroxidase in H2O2 degradation remain unclear, these enzymes serve to minimize the accumulation of °O2- and H2O2, which in the presence of the redox-active transition metals, copper and iron, form the very reactive and damaging hydroxyl radical °OH, for which no antioxidant enzyme system exists [8]. Finally, when oxidative damage has occurred, mitochondria also possess enzyme systems that can repair. Phospholipid hydroperoxide glutathione peroxidase is a selenium-containing enzyme that directly reduces peroxidized acyl groups in phospholipids. Repair of oxidized protein sulfhydryl groups may also occur via thioredoxin and thioltransferase enzymes. However, these repairing enzyme systems are likely to be damaged or ineffective in most conditions of oxidative stress unless their reserve can be boosted by antioxidant enzyme mimics.
Another fascinating aspect of the consequences of oxidative stress has emerged from very recent studies. It has been suggested that oxidative stress may trigger apoptosis (ie, programmed cell death) during ischemia-reperfusion. However, reactive oxygen species generated during ischemic preconditioning can upregulate the anti-death gene Bcl-2 by activating a specific nuclear transcription factor (NFkB), which in turn reduces apoptosis [9, 10]. These results indicate an additional antioxidant pathway for the protection of myocardium by ischemic preconditioning. Hopefully, this should lead to exciting developments for new antioxidant therapy, taking into account important intracellular signaling pathways for treating oxidative stress-associated injury.

REFERENCES

 
Networking antioxidants in the isolated rat heart are selectively depleted by ischemia-reperfusion.

Haramaki N, Stewart DB, Aggarwal S, Ikeda H, Reznick AZ, Packer L.

The Department of Internal Medicine III, Kurume University School of Medicine, Japan. haramaki@med.kurume-u.ac.jp

Although cardiac endogenous antioxidants have been reported to be oxidized and decreased by ischemia-reperfusion, little is known whether the changes in these antioxidants are correlated with each other in a systematic relationship. In this study, isolated rat hearts were subjected to various periods of ischemia-reperfusion using the Langendorff method, and the content and/or redox status of tissue antioxidants were analyzed. Significant losses in the tissue hydrophilic antioxidants, ascorbate, and glutathione were observed. These losses were dependent on the duration of the reperfusion period (between 0-40 min) but not of ischemia (20-60 min). Marked increases of dehydroascorbate and glutathione disulfide, the oxidized forms of ascorbate and glutathione, respectively, were found during reperfusion, but these changes were not observed during ischemia. These findings indicate that the tissue hydrophilic antioxidants are easily oxidized and may be the first line of antioxidant defenses during reperfusion. Lipophilic antioxidants, like ubiquinol 9 and vitamin E, were not decreased during ischemia-reperfusion using regular buffer; however, if oxidative stress was induced by addition of H2O2 to the buffer solution during reperfusion after 20 min of ischemia, decreases in both the hydrophilic and hydrophobic antioxidants were noticeable. With 100 microM H2O2, the tissue antioxidant decreases were ubiquinol 9 (39%), vitamin E (3%), glutathione (44%) and ascorbate (58%). Only with 500 microM H2O2 treatment were marked decreases in tissue vitamin E (65%) observed; this was associated with almost complete depletion of tissue ubiquinol 9 (95%). These results suggest that prior to the consumption of vitamin E, other antioxidants are depleted and that vitamin E may serve as the ultimate antioxidant, protecting the integrity of cellular membranes. Thus, in this work, cardiac antioxidants were demonstrated to change in a systematically organized relationship under ischemia-reperfusion. This graded utilization of antioxidants supports the redox based antioxidant network concept, found to be present in other biological systems.

PMID: 9680179 [PubMed - indexed for MEDLINE]

 
Overexpression of MnSOD protects against myocardial ischemia/reperfusion injury in transgenic mice.

Chen Z, Siu B, Ho YS, Vincent R, Chua CC, Hamdy RC, Chua BH.

Cecile Cox Quillen Laboratory of Geriatrics, James H. Quillen School of Medicine, East Tennessee State University, James H. Quillen Veterans Affairs Medical Center, Johnson City 37614, USA.

Generation of free radicals upon reperfusion has been cited as one of the major causes of ischaemia/reperfusion injury. The following series of experiments was designed to study the effect of manganese superoxide dismutase (MnSOD) overexpression in transgenic mice on ischemia/reperfusion injury. A species of 1.4 kb human MnSOD mRNA was expressed, and a 325% increase in MnSOD activity was detected in the hearts of transgenic mice with no changes in the other antioxidant enzymes or heat shock proteins. Immunocytochemical study indicated an increased labeling of MnSOD mainly in the heart mitochondria of the transgenic mice. When these hearts were perfused as Langendorff preparations for 45 min after 35 min of global ischemia, the functional recovery of the hearts, expressed as heart rate x left ventricular developed pressure, was 52 +/- 4% in the transgenic hearts as compared to 31 +/- 4% in the non-transgenic hearts. This protection was accompanied by a significant decrease in lactate dehydrogenase release from the transgenic hearts. Overexpression of MnSOD limited the infarct size in vivo in a left coronary artery ligation model. Our results demonstrate that overexpression of MnSOD renders the heart more resistant to ischemia/reperfusion injury.

PMID: 9925365 [PubMed - indexed for MEDLINE]

 
Transgenic mice overexpressing glutathione peroxidase are resistant to myocardial ischemia reperfusion injury.

Yoshida T, Watanabe M, Engelman DT, Engelman RM, Schley JA, Maulik N, Ho YS, Oberley TD, Das DK.

Department of Surgery, University of Connecticut School of Medicine, Farmington 06030, USA.

To test the authors' hypothesis that cellular antioxidant enzymes constitute a cellular defense against acute stress, myocardial ischemia reperfusion injury in transgenic mice overexpressing the cellular glutathione peroxidase (GSHPx-1) was studied. Transgenic mice were generated using the entire mouse GSHPx-1 gene including approximately 2.0 kb 5'flanking sequence. A 400% increase of GSHPx activity was found in the hearts of transgenic mice compared with non-transgenic controls. Isolated perfused hearts were prepared from two groups of mice: transgenic overexpressed; non-transgenic controls. Hearts were perfused by Langendorff mode, and after 10 min of stabilization subjected to 30 min of ischemia followed by 20 min of reperfusion. In addition, a group of hearts were perfused for 50 min without subjecting them to ischemia and reperfusion to demonstrate the stability of heart preparation. Transgenic mouse hearts demonstrated significantly improved recovery of contractile force and the rate of contraction, compared to non-transgenic control mouse hearts. The infarct size was also lower in transgenic mouse hearts compared to those of non-transgenic controls. In concert, following ischemia, release of creatine kinase from the transgenic hearts was significantly lower than the control group. The results of this study indicate that increased GSHPx-1 expression renders the heart more resistant to myocardial ischemia reperfusion injury.

PMID: 8877785 [PubMed - indexed for MEDLINE]

 
Mitochondrial dysfunction in cardiac disease: ischemia--reperfusion, aging, and heart failure.

Lesnefsky EJ, Moghaddas S, Tandler B, Kerner J, Hoppel CL.

Division of Cardiology, Case Western Reserve University and Geriatric Research, Education and Clinical Center, Louis Stokes Veterans Affairs Medical Center, Cleveland, Ohio 44106, USA. EXL9@po.cwru.edu

Mitochondria contribute to cardiac dysfunction and myocyte injury via a loss of metabolic capacity and by the production and release of toxic products. This article discusses aspects of mitochondrial structure and metabolism that are pertinent to the role of mitochondria in cardiac disease. Generalized mechanisms of mitochondrial-derived myocyte injury are also discussed, as are the strengths and weaknesses of experimental models used to study the contribution of mitochondria to cardiac injury. Finally, the involvement of mitochondria in the pathogenesis of specific cardiac disease states (ischemia, reperfusion, aging, ischemic preconditioning, and cardiomyopathy) is addressed. Copyright 2001 Academic Press.

Publication Types:

• Review
• Review, Academic

PMID: 11444914 [PubMed - indexed for MEDLINE]

 
Ischemic preconditioning reduces apoptosis by upregulating anti-death gene Bcl-2.

Maulik N, Engelman RM, Rousou JA, Flack JE 3rd, Deaton D, Das DK.

Department of Surgery, University of Connecticut School of Medicine, Farmington, CT 06030-1110, USA. nmaulik@panda.uchc.edu

BACKGROUND: Reperfusion of ischemic myocardium causes cardiomyocyte apoptosis in concert with downregulation of Bcl-2 gene. Ischemic preconditioning (PC) mediated by cyclic episodes of short-term ischemia and reperfusion reduces apoptotic cell death. PC also triggers a signaling pathway by potentiating tyrosine kinase phosphorylation leading to the activation of p38 MAP kinase and MAPKAP kinase 2. The nuclear transcription factor, NFkappaB, plays a crucial role in this signaling process. Because NFkappaB is a target of oxygen free radicals and Bcl-2 is an antioxidant gene, we hypothesized that reactive oxygen species might play a role in the signaling process. METHODS AND RESULTS: Isolated rat hearts were perfused in the absence or presence of either dimethyl thiourea (DMTU), a hydroxyl radical scavenger, or SN50 peptide, a NFkappaB blocker. Hearts were then subjected to PC by 4 repeated episodes of 5-minute ischemia, each followed by 10 minutes reperfusion. All hearts were then made globally ischemic at normothermia for 30 minutes followed by 2 hours of normothermic reperfusion. Creatine kinase release and malonaldehyde formation were determined in the coronary effluent collected during reperfusion. At the end of each experiment, hearts were processed for infarct size determination and analyses of apoptosis, DNA fragmentation, NFkappaB, and Bcl-2. Myocardial infarction was reduced by PC. DMTU and SN50 abolished this cardioprotective effect of PC. PC resulted upregulation of Bcl-2 gene which was partially prevented by DMTU and SN50. Both ischemia/reperfusion and PC caused nuclear translocation and activation of NFkappaB, which was blocked by both DMTU and SN50. PC reduced cardiomyocyte apoptosis which was partially inhibited by DMTU and SN50. A substantial number of apoptotic cardiomyocytes were identified in the hearts subjected to 30 minutes ischemia and 2-hour reperfusion. PC significantly inhibited the extent of cardiomyocyte apoptosis and DMTU and SN50 reversed it only minimally. CONCLUSIONS: The results demonstrate that reactive oxygen species play a crucial role in signal transduction mediated by PC. This signaling process appears to involve NFkappaB. NFkappaB becomes translocated and activated by both ischemia/reperfusion, which induces apoptosis and PC which reduces apoptosis. However, the amount of NFkappaB binding activity is significantly higher in the PC hearts compared with ischemic reperfused hearts. The upregulation of the antioxidant gene, Bcl-2, is inversely correlated with the reduction of cardiomyocyte apoptosis associated with PC.

PMID: 10567332 [PubMed - indexed for MEDLINE]

 
An essential role of the antioxidant gene Bcl-2 in myocardial adaptation to ischemia: an insight with antisense Bcl-2 therapy.

Hattori R, Hernandez TE, Zhu L, Maulik N, Otani H, Kaneda Y, Das DK.

Department of Surgery, University of Connecticut School of Medicine, Farmington 06030-1110, USA.

Reperfusion of ischemic myocardium results in apoptotic cell death, which can be blocked by adapting the heart to ischemic stress induced by cyclic episodes of brief periods of ischemia and reperfusion. In concert, the antiapoptotic gene bcl-2 is decreased by ischemia/reperfusion, but increased in the ischemically adapted myocardium. To examine if bcl-2 plays a crucial role in cardioprotection, adaptive cardioprotection was further examined in the hearts treated with antisense bcl-2 oligodeoxynucleotides (ODN). Isolated Langendorff-perfused rat hearts were divided into three groups: control (perfused with Krebs-Henseleit bicarbonate buffer for 210 min); 30-min ischemia followed by 2-h reperfusion; ischemic adaptation followed by 30-min ischemia and 2-h reperfusion. The last (adapted heart) group was subdivided into another two groups: one was transfected 48 h earlier with antisense bcl-2 ODN, whereas the other group was transfected with sense bcl-2 ODN. Cardioprotection was examined by determining cardiomyocyte death due to necrosis and apoptosis. Antisense gene therapy almost completely abolished bcl-2 protein expression in the hearts. Bcl-2 mRNA was down-regulated in the ischemic/reperfused heart, but up-regulated in the adapted myocardium. Adapted myocardium showed decreased infarct size and reduced number of apoptotic cardiomyocytes. Ischemia/reperfusion resulted in increased oxidative stress as evidenced by increased malonaldehyde formation. Adapted myocardium had a reduced amount of malonaldehyde. Antisense bcl-2 ODN completely abolished the cardioprotective effects of adaptation by eliminating the antideath signal of bcl-2. In concert, reduced oxidative stress in the adapted myocardium no longer persisted. The results suggest an antioxidant role of bcl-2 that appeared to be essential for the cardioprotection achieved by ischemic adaptation.

PMID: 11491653 [PubMed - indexed for MEDLINE]