Effect of ischemia–reperfusion on heart mitochondria from hyperthyroid rats (original) (raw)
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Cellular and Molecular Life Sciences, 2004
We investigated the role of nitric oxide (NO) in the mitochondrial derangement associated with the functional response to ischemia-reperfusion of hyperthyroid rat hearts. Mitochondria were isolated at 3000 g from hearts subjected to ischemia-reperfusion, with or without N w -nitro-L-arginine (L-NNA, an NO synthase inhibitor). During reperfusion, hyperthyroid hearts displayed tachycardia and low functional recovery. Their mitochondria exhibited O 2 consumption similar to euthyroid controls, while H 2 O 2 production, hydroperoxide, protein-bound carbonyl and nitrotyrosine levels, and susceptibility to swelling were higher. L-NNA blocked the reperfusion tachycardic response and increased inotropic recovery in hyperthyroid hearts. L-NNA decreased mitochondrial H 2 O 2 production and oxidative damage, and increased respiration and tolerance to swelling. Such effects were CMLS Cellular and Molecular Life Sciences higher in hyperthyroid preparations. These results confirm the role of mitochondria in ischemia-reperfusion damage, and strongly suggest that NO overproduction is involved in the high mitochondrial dysfunction and the low recovery of hyperthyroid hearts from ischemia-reperfusion. L-NNA also decreased protein content and cytochrome oxidase activity of a mitochondrial fraction isolated at 8000 g. This and previous results suggest that the above fraction contains, together with light mitochondria, damaged mitochondria coming from the heaviest fraction, which has the highest cytochrome oxidase activity and capacity to produce H 2 O 2 . Therefore, we propose that the high mitochondrial susceptibility to swelling, favoring mitochondrial population purification from H 2 O 2overproducing mitochondria, limits hyperthyroid heart oxidative stress.
Biochimica et Biophysica Acta (BBA) - Molecular Basis of Disease, 1994
In order to further investigate the mechanism regulating the control of mitochondrial respiration by thyroid hormones, the effect of the hyperthyroidism on the kinetic characteristics of cytocrome c oxidase in rat heart mitochondria was studied. Mitochondrial preparations from both control and hyperthyroid rats had equivalent g m values for cytochrome c, while the maximal activity of cytochrome oxidase was significantly increased (by around 30%) in mitochondrial preparation from hyperthyroid rats. This enhanced activity of cytochrome oxidase was associated to a parallel increase in mitochondrial State 3 respiration. The hormone treatment resulted in a decrease in the flux control coefficent of the oxidase. The enhanced activity of cytochrome oxidase in hyperthyroid rats does not appear to be dependent on an increase in the mass of this enzyme complex in that the heme aa 3 content was equivalent in both hyperthyroid and control preparations. The Arrhenius plot characteristics differ for cytochrome oxidase activity in mitochondria from hyperthyroid rats as compared with control rats in that the breakpoint of the biphasic plot is shifted to a lower temperature. Cardiolipin content was significantly increased in mitochondrial preparations from hyperthyroid rats, while there were no significant alterations in the fatty acid composition of cardiolipin of control and hyperthyroid preparations. The results support the conclusion that the enhanced cytochrome oxidase activity in heart mitochondrial preparations from hyperthyroid rats is due to a specific increase in the content of cardiolipin.
Free Radical Biology and Medicine, 1999
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.
Thyroid state affects H2O2 removal by rat heart mitochondria
Archives of Biochemistry and Biophysics, 2018
We investigated the effects of thyroid state on the mechanisms underlying rat heart mitochondrial capacity to remove H 2 O 2 produced by an exogenous source. The removal rates were higher in the presence of respiratory substrates independently from thyroid state and were higher in hyperthyroid than in hypothyroid preparations. The thyroid state-linked changes in H 2 O 2 removal rates, mirrored those in H 2 O 2 release rates, showing that endogenous and exogenous H 2 O 2 do not compete for the removing system. Mitochondrial content of coenzyme Q9 and Q10 was lower in hypothyroidism and higher in hyperthyroidism suggesting that the thyroid state-linked changes in the rates of H 2 O 2 production are due to changes in the ubiquinone mitochondrial content. The rates of H 2 O 2 removal in the presence of antioxidant enzyme inhibitors indicated that the contribution of each antioxidant is dependent on the thyroid state. This was supported by enzymatic activity measurements. Pharmacological inhibition also showed that the overall percentage contribution of the enzymatic processes, as well as that of non-enzymatic processes, is not affected by thyroid state. Cytochrome levels, inferred by light emission measurements, and western blot determination of cytochrome c, were lower in hypothyroid and higher in hyperthyroid preparations supporting the idea that the levels of reducing compounds were modified in opposite way by the changes in thyroid state. Further support was obtained showing that the whole antioxidant capacity, which provides an evaluation of capacity of the systems, different from cytochromes, assigned to H 2 O 2 scavenging, was lower in hyperthyroid than in hypothyroid state.
Life Sciences, 2000
Hypet-thyroidism has been reported to decrease heart antioxidant capacity and increase its susceptibility to in vitro oxidative stress. This may affect the heart response to &hernia-reperfusion, a condition that increases free radical production. We compared the functional recovery from in vitro ischemia-repermsion (Langendorff) of hearts from euthyroid (E), hyperthyroid (H, ten daily intraperitoneal injections of T,, 10 ug/lOOg body weight), vitamin E-treated (VE, ten daily intramuscular injections, 20 mg/lOOg body weight) and hyperthyroid vitamin Etreated (HVE) rats. We also determined lipid peroxidation, tissue antioxidant capacity and the tissue capability to face an oxidative stress in vitro. A significant tachycardia was displayed during reperfusion following 20 min ischemia by the hyperthyroid hearts, together with a low recovery of left ventricular developed pressure (LVDP) and left ventricular dPid$,,,. When H hearts were paced at 300 beats/min, the functional recovery (LVDP and dPidt,,_J was close to 100% and significantly higher than in E paced hearts. At the end of the ischemia-reperfusion protocol, myocardium antioxidant capacity was significantly lower, whereas lipid peroxidation and the susceptibility to in vitro oxidative stress were higher in the T, treated (H) than in euthyroid rats. The in vifro tachycardic response, the reduction in the antioxidant capacity and the increase in lipid peroxidation were prevented by treatment of hyperthyroid rats with vitamin E (HVE). These results suggest that the tachycardic response to reperfusion following chronic T, pretreatment was associated with the reduced capability of the heart to face oxidative stresses in hyperthyroidism. Key K&K hyperthyroidism, heart ischemia-reperfision, vitamin E, antioxidant capacities Hyperthyroidism has been associated with a hyperdynamic state of the heart characterized by tachycardia (1, 2) and increased contractile performance (3). These changes in the electrical and mechanical activity of the heart result in an increased total cardiac output and work which match the increased oxygen demand by peripheral tissues owing to the thyroid hormone-induced increase in their metabolism. These changes in heart function and tissue metabolism may represent an integrate, suitable response to physiological increases in thyroid state, such as those elicited by seasonal
Molecular and Cellular Endocrinology, 2000
Mitochondria seem to be involved in oxygen radical damage and aging. However, the possible relationships between oxygen consumption and oxygen radical production by functional mitochondria, and oxidative DNA damage, have not been studied previously. In order to analyze these relationships, male Wistar rats of 12 weeks of age were rendered hyper-and hypothyroid by chronic T 3 and 6-n-propyl-2-thiouracil treatments, respectively. Hypothyroidism decreased heart mitochondrial H 2 O 2 production in States 4 (to 51% of controls; P B0.05) and 3 (to 21% of controls; PB 0.05). In agreement with this, 8-oxo-7,8-dihydro-2%-deoxyguanosine (8-oxodG) decreased in the heart genomic DNA of hypothyroid animals to 40% of controls (PB0.001). Studies with respiratory inhibitors showed that the decrease in oxygen radical generation observed in hypothyroidism occurred at Complex III (mainly) and at Complex I; that decrease was due to the presence of a lower free radical leak in the respiratory chain (P B 0.05). Hyperthyroidism did not significantly change heart mitochondrial H 2 O 2 production since the increase in State 4 oxygen consumption in comparison with control and hypothyroid animals (PB 0.05) was compensated by a decrease in the free radical leak in relation to control animals (PB 0.05). In agreement with this, heart 8-oxodG was not changed in hyperthyroid animals. The lack of increase in H 2 O 2 production per unit of mitochondrial protein will protect mitochondria themselves against self-inflicted damage during hyperthyroidism.
Myocardial ischemia and in vitro mitochondrial metabolic efficiency
Molecular and Cellular Biochemistry, 1996
The purpose of this study was to evaluate the oxidative capacities and the rate of energy synthesis in isolated mitochondria extracted from normal and post-ischemic myocardium. Isolated rat hearts were perfused according to the working mode with a Krebs Heinseleit buffer containing glucose (11 mM), insulin (10 IU/I) and caprylic acid (25 ~tM). After a 15 min perfusion in normoxic conditions, the hearts were subjected to a 20 min local zero-flow ischemia followed by a 20 min reperfusion. During the perfusion, the aortic and coronary flows, the aortic pressure and the electrocardiogram were monitored. At the end of the reperfusion period, the non-ischemic and ischemic zones (NIZ and IZ, respectively) were separated and the mitochondria were harvested from each zone. The oxygen uptake and the rate of energy production of the NIZ and IZ mitochondria were then assessed with palmitoylcarnitine as substrate in 2 buffers differing in their free calcium concentration (0.041 and 0.150/aM). Ischemia provoked a 50% reduction of coronary and aortic flows. The reperfusion of the IZ allowed the partial recovery of coronary flow, but the aortic flow decreased beneath its ischemic value because of the occurrence of severe arrhythmias, stunning and probably hibernation. The IZ mitochondria displayed a lower rate of oxygen consumption, whatever the buffer free calcium concentration. Conversely, their rate of energy production was increased, indicating that their metabolic efficiency was improved as compared to NIZ mitochondria. This might be due to the mitochondrial calcium overload persisting during reperfusion, to the activation of the inner membrane Na÷/Ca 2+ exchange and to a significant mitochondrial swelling. On the other hand, the presence of an elevated free calcium concentration in the respiration buffer provoked some energy wasting characterized by a constant AMP production. This was attributed to some accumulation of acetate and the activation of the energy-consuming acetylCoA synthetase. In conclusion, ischemia and reperfusion did not alter the membrane integrity of the mitochondria but improved their metabolic efficiency. Nevertheless, these in vitro results can not reflect the mitochondrial function in the reperfused myocardium. The mitochondrial calcium overload reported to last during reperfusion in the cardiomyocytes might mimic the free calcium-induced reduction of metabolic efficiency observed in vitro in the present study. The resulting energy wasting might be responsible for the contractile abnormalities noticed in the reperfused myocardium.
Hyperthyroidism causes cardiac dysfunction by mitochondrial impairment and energy depletion
Journal of Endocrinology, 2013
This study elucidates the role of metabolic remodeling in cardiac dysfunction induced by hyperthyroidism. Cardiac hypertrophy, structural remodeling, and expression of the genes associated with fatty acid metabolism were examined in rats treated with triiodothyronine (T3) alone (8 μg/100 g body weight (BW), i.p.) for 15 days or along with a peroxisome proliferator-activated receptor alpha agonist bezafibrate (Bzf; 30 μg/100 g BW, oral) and were found to improve in the Bzf co-treated condition. Ultrastructure of mitochondria was damaged in T3-treated rat heart, which was prevented by Bzf co-administration. Hyperthyroidism-induced oxidative stress, reduction in cytochromecoxidase activity, and myocardial ATP concentration were also significantly checked by Bzf. Heart function studied at different time points during the course of T3treatment shows an initial improvement and then a gradual but progressive decline with time, which is prevented by Bzf co-treatment. In summary, the results...
Effect of thyroid state on susceptibility to oxidants and swelling of mitochondria from rat tissues
Free Radical Biology and Medicine, 2003
The effects of the thyroid state on oxidative damage, antioxidant capacity, susceptibility to in vitro oxidative stress and Ca 2ϩ -induced permeabilization of mitochondria from rat tissues (liver, heart, and gastrocnemious muscle) were examined. Hypothyroidism was induced by administering methimazole in drinking water for 15 d. Hyperthyroidism was elicited by a 10 d treatment of hypothyroid rats with triiodothyronine (10 g/100 g body weight). Mitochondrial levels of hydroperoxides and protein-bound carbonyls significantly decreased in hypothyroid tissues and were reported above euthroid values in hypothyroid rats after T 3 treatment. Mitochondrial vitamin E levels were not affected by changes of animal thyroid state. Mitochondrial Coenzyme Q9 levels decreased in liver and heart from hypothyroid rats and increased in all hyperthyroid tissues, while Coenzyme Q10 levels decreased in hypothyroid liver and increased in all hyperthyroid tissues. The antioxidant capacity of mitochondria was not significantly different in hypothyroid and euthyroid tissues, whereas it decreased in the hyperthyroid ones. Susceptibility to in vitro oxidative challenge decreased in mitochondria from hypothyroid tissues and increased in mitochondria from hyperthyroid tissues, while susceptibility to Ca 2ϩ -induced swelling decreased only in hypothyroid liver mitochondria and increased in mitochondria from all hyperthyroid tissues. The tissue-dependence of the mitochondrial susceptibility to stressful conditions in altered thyroid states can be explained by different thyroid hormone-induced changes in mitochondrial ROS production and relative amounts of mitochondrial hemoproteins and antioxidants. We suggest that susceptibilities to oxidants and Ca 2ϩ -induced swelling may have important implications for the thyroid hormone regulation of the turnover of proteins and whole mitochondria, respectively.
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It is widely accepted that mitochondria contribute to their own damage in the settings of experimental ischemia/reperfusion injury. The aim of the present work was to investigate the degree of mitochondrial dysfunction elicited by an in vivo rat heart model of regional ischemia/reperfusion injury. Mitochondria were isolated from ischemic and non-ischemic zones rat hearts subjected to 30 min of regional ischemia by coronary ligation and 15 min of reperfusion. Oxygen consumption was assessed by polarographic oxymetry in a Clark-type electrode in the presence of complex I substrates (glutamate/malate) and complex II (succinate). State 3 respiration rate and the respiratory control ratio for glutamate/malate were significantly decreased in mitochondria isolated from the ischemic area, suggesting an early alteration of complex I supported respiration (p < 0.01 vs. non-ischemic area). In another set of experiments mitochondrial calcium retention capacity was evaluated by luminescence spectroscopy using the calcium-sensitive probe Calcium Green 5N and mitochondrial swelling was monitored as the decrease in light scattering at 520 nm. In the presence of complex IIdependent substrate, less calcium was required to zone induce permeability transition, suggesting the pre-existence of calcium overload in mitochondria isolated from ischemic zone vs. non-ischemic zone. Mitochondria isolated from the ischemic (vs. non-ischemic) zone presented an increased rate of swelling in the case of complex I supported respiration. These results are suggestive for an early damage to the phosphorylation apparatus at complex I in mitochondria isolated at reperfusion after regional ischemia in the in vivo rat hearts.