Melatonin, a well-known antioxidant, has been shown to protect against ischemia-reperfusion myocardial damage. Mitochondrial permeability transition pore (MPTP) opening is an important event in ...cardiomyocyte cell death occurring during ischemia-reperfusion and therefore a possible target for cardioprotection. In the present study, we tested the hypothesis that melatonin could protect heart against ischemia-reperfusion injury by inhibiting MPTP opening. Isolated perfused rat hearts were subjected to global ischemia and reperfusion in the presence or absence of melatonin in a Langerdoff apparatus. Melatonin treatment significantly improves the functional recovery of Langerdoff hearts on reperfusion, reduces the infarct size, and decreases necrotic damage as shown by the reduced release of lactate dehydrogenase. Mitochondria isolated from melatonin-treated hearts are less sensitive than mitochondria from reperfused hearts to MPTP opening as demonstrated by their higher resistance to Ca(2+). Similar results were obtained following treatment of ischemic-reperfused rat heart with cyclosporine A, a known inhibitor of MPTP opening. In addition, melatonin prevents mitochondrial NAD(+) release and mitochondrial cytochrome c release and, as previously shown, cardiolipin oxidation associated with ischemia-reperfusion. Together, these results demonstrate that melatonin protects heart from reperfusion injury by inhibiting MPTP opening, probably via prevention of cardiolipin peroxidation.
ABSTRACT
Reactive oxygen species (ROS) are considered an important factor in ischemia‐reperfusion injury to cardiac myocites. We have examined the effects of ischemia (30 min) and ischemia followed ...by reperfusion (15 min) of rat hearts on the activity of complex III and on the cardiolipin content in isolated mitochondria. Mitochondrial production of H2O2 and lipid peroxidation was also measured. The capacity of mitochondria to produce both H2O2 and lipid peroxidation increased upon reperfusion. The activity of complex III was 22% and 46% lower in ischemic and reperfused rat heart mitochondria, respectively, than that of controls. These changes in complex III activity were associated to parallel changes in state 3 respiration. The mitochondrial content of cardiolipin, which is required for optimal activity of complex III, decreased by 28% and by 50% as a function of ischemia and reperfusion, respectively. The lower complex III activity in mitochondria from reperfused rat hearts could be completely restored to the level of normal hearts by exogenously added cardiolipin. It is proposed that the loss of complex III activity in reperfused rat hearts can be mainly ascribed to a loss of cardiolipin content, due to oxidative attack by oxygen free radicals.
ABSTRACT
Several recent works have shown that a brief ischemia applied during the onset of reperfusion (postconditioning) is cardioprotective in different animal models and that the early minutes of ...reperfusion are critical to its cardioprotection. This effect has been related to prevention of oxidative stress, but mechanisms have not been clearly demonstrated. The present study tested the hypothesis that mitochondria play a central role in peroxide production and oxidative stress during reperfusion and are responsible for the protective effect of postconditioning. Isolated perfused rat hearts were subjected to complete global ischemia for 45 min and reperfused for 40 min. Normoxic group was reperfused with a Krebs‐Henseleit solution with the preischemic pO2 level (600 mmHg); in the “hypoxic group”, normoxic reperfusion was preceded by 3 min with 150 mmHg pO2. Reperfusion was stopped at 3 and 40 min. The rate of hydroperoxide production, GSH, GSSG, and carbonyl protein levels were measured in mitochondria at 3 min and at the end of reperfusion. GSH and GSSG were also measured in tissue. Hemodinamic function was monitored during the experiment. LVEDp increased and LVDp decreased in the normoxic group but not in the hypoxic group. The rate of mitochondrial peroxide production was higher in normoxic than in the hypoxic group 3 min after reperfusion and at its conclusion. Accordingly, GSH was oxidized in normoxic but not in hypoxic hearts. Mitochondria carbonyl proteins were significantly higher in normoxic than in the hypoxic group at the end of reperfusion. In this model, 1) hypoxic reperfusion at the onset of reperfusion reduces myocardial injury; 2) the major rate of mitochondrial peroxide production is 3 min after the onset of reperfusion; 3) cardioprotection of postconditioning correlates with reduced mitochondria peroxide production and prevention of GSH oxidation.—Serviddio, G., Di Venosa, N., Federici, A., D'Agostino, D., Rollo, T., Prigigallo, F., Altomare, E., Fiore, T., Vendemiale, G. Brief hypoxia before normoxic reperfusion (postconditioning) protects the heart against ischemia‐reperfusion injury by preventing mitochondria peroxyde production and glutathione depletion. FASEB J. 19, 354–361 (2005)
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.
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.
Direct hemoperfusion using polymyxin B-immobilized fiber (PMX-DHP) is an established treatment method for septic shock caused by Gram-negative infections. We report one instance in which PMX-DHP ...therapy has been used successfully in a 33-year-old woman with septic shock from urosepsis. Although there is lack of recommendations in latest Surviving Sepsis Campaign Guidelines, evidence of PMX-DHP efficacy in this subset of patients is growing.
SUMMARY
1
One hour exposure to hyperoxia has been shown previously to limit a subsequent ischaemia–reperfusion injury in spontaneously breathing rats. We tested the cardioprotective effect of a ...shorter period of hyperoxia during mechanical ventilation and the possible contribution of reactive oxygen species (ROS) and mitochondrial ATP‐sensitive potassium (mitoKATP) channels.
2
Mechanically ventilated rats were exposed to normoxia (Fio2 = 0.3) or hyperoxia (Fio2 = 1.0) for 30 min and pH, Pco2, Po2, heart rate, airway and blood pressure were measured at baseline and after 30 min mechanical ventilation. Isolated hearts were subsequently subjected to 30 min ischaemia and 120 min reperfusion. Infarct size and left ventricular end‐diastolic pressure (LVEDP), developed pressure (LVDP) and coronary flow (CF) were measured. In order to investigate the role of ROS and KATP channels within the mechanism leading to cardioprotection, the free radical scavenger N‐acetylcysteine (NAC; 150 mg/kg) was infused in mechanically ventilated rats and the KATP channel blockers glibenclamide (200 mmol/L) or 5‐hydroxydecanoate (10 mmol/L) were infused in isolated hearts immediately before ischaemia.
3
No differences were detected in Pco2, pH, heart rate, airway and blood pressure between the groups. However, the Po2 in hyperoxic groups was significantly higher compared with that in normoxic groups (P < 0.01). After 30 min ischaemia, we found that hyperoxic preconditioning significantly improved CF (P < 0.01), LVDP (P < 0.01) and LVEDP (P < 0.01) and reduced the extent of infarct size in the reperfused heart compared with the normoxic group (P < 0.01). When rats were pretreated either with NAC before hyperoxic ventilation or with KATP channel blockers before ischaemia, myocardial protection was abolished.
4
Hyperoxic mechanical ventilation, prior to ischaemia, reduces myocardial reperfusion injury. This is likely to occur through the induction of oxidative stress, which leads to myocyte mitoKATP channel opening.
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