Reactive oxygen species (ROS) are considered an important factor in ischemia/reperfusion injury to cardiac myocytes. Mitochondrial respiration, mainly at the level of complex I and III, is an ...important source of ROS generation and hence a potential contributor of cardiac reperfusion injury. Appropriate antioxidant strategies could be particularly useful to limit this ROS generation and associated mitochondrial dysfunction. Melatonin has been shown to effectively protect against ischemic-reperfusion myocardial damage. The mechanism by which melatonin exerts this cardioprotective effect is not well established. In the present study we examined the effects of melatonin on various parameters of mitochondrial bioenergetics in a Langerdoff isolated perfused rat heart model. After isolation of mitochondria from control, ischemic-reperfused and melatonin-treated ischemic-reperfused rat heart, various bioenergetic parameters were evaluated such as rates of mitochondrial oxygen consumption, complex I and complex III activity, H₂O₂ production as well as the degree of lipid peroxidation, cardiolipin content, and cardiolipin oxidation. We found that reperfusion significantly altered all these mitochondrial parameters, while melatonin treatment had strong protective effect attenuating these alterations. This effect appears to be due, at least in part, to the preservation, by ROS attack, of the content and integrity of cardiolipin molecules which play a pivotal role in mitochondrial bioenergetics. Protection of mitochondrial dysfunction was associated with an improvement of post-ischemic hemodynamic function of the heart. Melatonin had also strong protective effect against oxidative alterations to complex I and III as well as to cardiolipin in isolated mitochondria.--Petrosillo, G., Di Venosa, N., Pistolese, M., Casanova, G., Tiravanti, E., Colantuono, G., Federici, A., Paradies, G., Ruggiero, F. M. Protective effect of melatonin against mitochondrial dysfunction associated with cardiac ischemia-reperfusion: role of cardiolipin.
Reactive oxygen species (ROS) are considered an important factor in ischemia/reperfusion injury to cardiac myocites. Mitochondrial respiration is an important source of ROS generation and hence a ...potential contributor to cardiac reperfusion injury. Appropriate treatment strategy could be particularly useful to limit this ROS generation and associated mitochondrial dysfunction. In the present study, we examined the effect of lowering the oxygen tension, at the onset of the reperfusion, on various parameters of mitochondrial bioenergetics in rat heart tissue. After isolation of mitochondria from control, ischemic, normoxic and hypoxic reperfused rat heart, various bioenergetic parameters were evaluated such as rates of mitochondrial oxygen consumption, complex I and complex III activity, H
2O
2 production and in addition, the degree of lipid peroxidation, cardiolipin content and cardiolipin oxidation. We found that normoxic reperfusion significantly altered all these mitochondrial parameters, while hypoxic reperfusion had a protective effect attenuating these alterations. This effect appears to be due, at least in part, to a reduction of mitochondrial ROS generation with subsequent preservation of cardiolipin integrity, protection of mitochondrial function and improvement of post-ischemic hemodynamic function of the heart.
ABSTRACT—Reactive oxygen species (ROS) are considered an important factor in ischemia/reperfusion injury to cardiac myocytes. Mitochondrial respiration is an important source of ROS production and ...hence a potential contributor to cardiac reperfusion injury. In this study, we have examined the effect of ischemia and ischemia followed by reperfusion of rat hearts on various parameters related to mitochondrial function, such as complex I activity, oxygen consumption, ROS production, and cardiolipin content. The activity of complex I was reduced by 25% and 48% in mitochondria isolated from ischemic and reperfused rat heart, respectively, compared with the controls. These changes in complex I activity were associated with parallel changes in state 3 respiration. The capacity of mitochondria to produce H2O2 increased on reperfusion. The mitochondrial content of cardiolipin, which is required for optimal activity of complex I, decreased by 28% and 50% as function of ischemia and reperfusion, respectively. The lower complex I activity in mitochondria from reperfused rat heart could be completely restored to the level of normal heart by exogenous added cardiolipin. This effect of cardiolipin could not be replaced by other phospholipids nor by peroxidized cardiolipin. It is proposed that the defect in complex I activity in ischemic/reperfused rat heart could be ascribed to a ROS-induced cardiolipin damage. These findings may provide an explanation for some of the factors responsible for myocardial reperfusion injury.
Release of cytochrome c from mitochondria is considered a critical, early event in the induction of an apoptosis cascade that ultimately leads to programmed cell death. Mitochondrial Ca(2+) loading ...is a trigger for the release of cytochrome c, although the molecular mechanism underlying this effect is not fully clarified. This study tested the hypothesis that distinct Ca(2+) thresholds may induce cytochrome c release from rat liver mitochondria by membrane permeability transition (MPT)-dependent and independent mechanisms. The involvement of reactive oxygen species (ROS) and cardiolipin in the Ca(2+)-induced cytochrome c release was also investigated. Cytochrome c was quantitated by a new, very sensitive, and rapid reverse-phase high performance liquid chromatography method with a detection limit of 0.1 pmol/sample. We found that a low extramitochondrial Ca(2+) level (2 microM) promoted the release of approximately 13% of the total alamethicin releasable pool of cytochrome c from mitochondria. This release was not depending of MPT; it was mediated by Ca(2+)-induced ROS production and cardiolipin peroxidation and appears to involve the voltage-dependent anion channel. High extramitochondrial Ca(2+) level (20 microM) promoted approximately 45% of the total releasable pool of cytochrome c. This process was MPT-dependent and was also mediated by ROS and cardiolipin. It is suggested that distinct Ca(2+) levels may determine the mode and the amount of cytochrome c release from rat liver mitochondria. The data may help to clarify the molecular mechanism underlying the Ca(2+)-induced release of cytochrome c from rat liver mitochondria and the role played by ROS and cardiolipin in this process.
The aim of this study was to investigate the influence of reactive oxygen species (ROS) on the activity of complex I and on the cardiolipin content in bovine heart submitochondrial particles (SMP). ...ROS were generated through the use of xanthine/xanthine oxidase (X/XO) system. Treatment of SMP with X/XO resulted in a large production of superoxide anion, detected by acetylated cytochrome c method, which was blocked by superoxide dismutase (SOD). Exposure of SMP to ROS generation resulted in a marked loss of complex I activity and to parallel loss of mitochondrial cardiolipin content. Both these effects were completely abolished by SOD+catalase. Exogenous added cardiolipin was able to almost completely restore the ROS-induced loss of complex I activity. No restoration was obtained with other major phospholipid components of the mitochondrial membrane such as phosphatidylcholine and phosphatidylethanolamine, nor with peroxidized cardiolipin. These results demonstrate that ROS affect the mitochondrial complex I activity via oxidative damage of cardiolipin which is required for the functioning of this multisubunit enzyme complex. These results may prove useful in probing molecular mechanisms of ROS-induced peroxidative damage to mitochondria, which have been proposed to contribute to those pathophysiological conditions characterized by an increase in the basal production of reactive oxygen species such as aging, ischemia/reperfusion and chronic degenerative diseases.
Cytochrome
c release from mitochondria is a critical event in the apoptosis induction. Dissociation of cytochrome
c from the mitochondrial inner membrane (IMM) is a necessary first step for ...cytochrome
c release. In the present study, the effect of reactive oxygen species (ROS) on the dissociation of cytochrome
c from beef-heart submitochondrial particles (SMP) and on the cardiolipin content was investigated. Exposure of SMP to mitochondrial-mediated ROS generation resulted in a large dissociation of cytochrome
c from SMP and in a parallel loss of cardiolipin. Both these effects were directly and significantly correlated and also abolished by superoxide dismutase+catalase. These results demonstrate that ROS generation induces the dissociation of cytochrome
c from IMM via cardiolipin peroxidation. The data may prove useful in clarifying the molecular mechanism underlying the release of cytochrome
c from the mitochondria to the cytosol.
ATLAS data are distributed centrally to Tier-1 and Tier-2 sites. The first stages of data selection and analysis take place mainly at Tier-2 centres, with the final, iterative and interactive, stages ...taking place mostly at Tier-3 clusters. The Italian ATLAS cloud consists of a Tier-1, four Tier-2s, and Tier-3 sites at each institute. Tier-3s that are grid-enabled are used to test code that will then be run on a larger scale at Tier-2s. All Tier-3s offer interactive data access to their users and the possibility to run PROOF. This paper describes the hardware and software infrastructure choices taken, the operational experience after 10 months of LHC data, and discusses site performances.
The effect of reactive oxygen species (ROS), produced by the mitochondrial respiratory chain, on the activity of cytochrome
c oxidase and on the cardiolipin content in bovine heart submitochondrial ...particles (SMP) was studied. ROS were produced by treatment of succinate-respiring SMP with antimycin A. This treatment resulted in a large production of superoxide anion, measured by epinephrine method, which was blocked by superoxide dismutase (SOD). Exposure of SMP to mitochondrial mediated ROS generation, led to a marked loss of cytochrome
c oxidase activity and to a parallel loss of cardiolipin content. Both these effects were completely abolished by SOD+catalase. Added cardiolipin was able to almost completely restore the ROS-induced loss of cytochrome
c oxidase activity. No restoration was obtained with peroxidized cardiolipin. These results demonstrate that mitochondrial mediated ROS generation affects the activity of cytochrome
c oxidase via peroxidation of cardiolipin which is needed for the optimal functioning of this enzyme complex. These results may prove useful in probing molecular mechanism of ROS-induced peroxidative damage to mitochondria which have been proposed to contribute to aging, ischemia/reperfusion and chronic degenerative diseases.
In the framework of WLCG, Tier-1s need to manage large volumes of data ranging in the PB scale. Moreover they need to be able to transfer data, from CERN and with the other centres (both Tier-1s and ...Tier-2s) with a sustained throughput of the order of hundreds of MB/s over the WAN offering at the same time a fast and reliable access also to the computing farm. In order to cope with these challenging requirements, at INFN Tier-1 we have adopted a storage model based on StoRM/GPFS/TSM for the D1T0 and D1T1 Storage Classes and on CASTOR for the D0T1. In this paper we present the results of the commissioning tests of this system for the ATLAS experiment reproducing the real production case with a full matrix transfer from the Tier-0 and with all the other involved centres. Noticeably also the new approach of direct file access from farm to data is covered showing positive results. GPFS/StoRM has also been sucessfully deployed, configured and commissioned as storage solution for an ATLAS INFN Tier-2, specifically the one of Milano. The results are shown and discussed in this paper together with the ones obtained for the Tier-1.
Ischemia-reperfusion injury to cardiac myocytes involves membrane damage mediated by oxygen free radicals. Lipid peroxidation is considered a major mechanism of oxygen free radical toxicity in ...reperfused heart. Mitochondrial respiration is an important source of these reactive oxygen species and hence a potential contributor to reperfusion injury. We have examined the effects of ischemia (30 min) and ischemia followed by reperfusion (15 min) of rat hearts, on the kinetic parameters of cytochrome
c oxidase, on the respiratory activities and on the phospholipid composition in isolated mitochondria. Mitochondrial content of malonyldialdheyde (MDA), an index of lipid peroxidation, was also measured. Reperfusion was accompanied by a significant increase in MDA production. Mitochondrial preparations from control, ischemic and reperfused rat heart had equivalent
K
m values for cytochrome
c, although the maximal activity of the oxidase was 25 and 51% less in ischemic and reperfused mitochondria than that of controls. These changes in the cytochrome
c oxidase activity were associated to parallel changes in state 3 mitochondrial respiration. The cytochrome aa
3 content was practically the same in these three types of mitochondria. Alterations were found in the mitochondrial content of the major phospholipid classes, the most pronounced change occurring in the cardiolipin, the level that decreased by 28 and by 50% as function of ischemia and reperfusion, respectively. The lower cytochrome
c oxidase activity in mitochondria from reperfused rat hearts could be almost completely restored to the level of control hearts by exogenously added cardiolipin, but not by other phospholipids nor by peroxidized cardiolipin. It is proposed that the reperfusion-induced decline in the mitochondrial cytochrome
c oxidase activity can be ascribed, at least in part, to a loss of cardiolipin content, due to peroxidative attack of its unsaturated fatty acids by oxygen free radicals. These findings may provide an explanation for some of the factors that lead to myocardial reperfusion injury.
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