cytochrome-c-t and Hyperthyroidism

We investigated the effects of thyroid state on the mechanisms underlying rat heart mitochondrial capacity to remove H

Topics: Animals; Cytochromes c; Hydrogen Peroxide; Hyperthyroidism; Male; Mitochondria, Heart; Oxidative Stress; Rats; Rats, Wistar; Thyroid Gland; Triiodothyronine

We investigated thyroid state effect on capacity of rat liver mitochondria to remove exogenously produced H2O2, determining their ability to decrease fluorescence generated by an H2O2 detector system. The rate of H2O2 removal by both non respiring and respiring mitochondria was increased by hyperthyroidism and decreased by hypothyroidism. However, the rate was higher in the presence of respiratory substrates, in particular pyruvate/malate, indicating a respiration-dependent process. Generally, the changes in H2O2 removal rates mirrored those in H2O2 release rates excluding the possibility that endogenous and exogenous H2O2 competed for the removing system. Pharmacological inhibition revealed thyroid state-linked differences in antioxidant enzyme contribution to H2O2 removal which were consistent with those in antioxidant system activities. The H2O2 removal was only in part due to enzymatic systems and that imputable to non-enzymatic processes was higher in hyperthyroid and lower in hypothyroid mitochondria. The levels of cytochrome c and the light emissions, due to luminol oxidation catalyzed by cytochrome/H2O2, exhibited similar changes with thyroid state supporting the idea that non-enzymatic scavenging was mainly due to hemoprotein action, which produces hydroxyl radicals. Further support was obtained showing that the whole antioxidant capacity, which provides an evaluation of capacity of the systems, different from cytochromes, assigned to H2O2 scavenging, was lower in hyperthyroid than in hypothyroid state. In conclusion, our results show that mitochondria from hyperthyroid liver have a high capacity for H2O2 removal, which, however, leading in great part to more reactive oxygen species, results harmful for such organelles.

Topics: Animals; Cell Fractionation; Cytochromes c; Glutathione Peroxidase; Glutathione Reductase; Hepatocytes; Hydrogen Peroxide; Hydroxyl Radical; Hyperthyroidism; Hypothyroidism; Liver; Malates; Male; Mitochondria, Liver; Oxidative Phosphorylation; Oxidative Stress; Oxygen Consumption; Pyruvic Acid; Rats; Rats, Wistar; Thyroid Gland

Hyperthyroidism, known to have deleterious effects on heart function, and is associated with an enhanced metabolic state, implying an increased production of reactive oxygen species. Tamoxifen is a selective antagonist of estrogen receptors. These receptors make the hyperthyroid heart more susceptible to ischemia/reperfusion. Tamoxifen is also well-known as an antioxidant. The aim of the present study was to explore the possible protective effect of tamoxifen on heart function in hyperthyroid rats. Rats were injected daily with 3,5,3'-triiodothyronine at 2mg/kg body weight during 5 days to induce hyperthyroidism. One group was treated with 10mg/kg tamoxifen and another was not. The protective effect of the drug on heart rhythm was analyzed after 5 min of coronary occlusion followed by 5 min reperfusion. In hyperthyroid rats not treated with tamoxifen, ECG tracings showed post-reperfusion arrhythmias, and heart mitochondria isolated from the ventricular free wall lost the ability to accumulate and retain matrix Ca(2+) and to form a high electric gradient. Both of these adverse effects were avoided with tamoxifen treatment. Hyperthyroidism-induced oxidative stress caused inhibition of cis-aconitase and disruption of mitochondrial DNA, effects which were also avoided by tamoxifen treatment. The current results support the idea that tamoxifen inhibits the hypersensitivity of hyperthyroid rat myocardium to reperfusion damage, probably because its antioxidant activity inhibits the mitochondrial permeability transition.

Topics: Animals; Arrhythmias, Cardiac; Cytochromes c; Estrogen Antagonists; Female; Hyperthyroidism; Mitochondria, Heart; Myocardial Reperfusion Injury; Oxidative Stress; Rats; Rats, Wistar; Tamoxifen; Thiobarbituric Acid Reactive Substances

Digitonin solubilizes mitochondrial membrane, breaks the integrity of the respiratory chain and releases two mobile redox-active components: coenzyme Q (CoQ) and cytochrome c (cyt c). In the present study we report the inhibition of glycerol-3-phosphate- and succinate-dependent oxygen consumption rates by digitonin treatment. Our results show that the inhibition of oxygen consumption rates is recovered by the addition of exogenous synthetic analog of CoQ idebenone (hydroxydecyl-ubiquinone; IDB) and cyt c. Glycerol-3-phosphate oxidation rate is recovered to 148 % of control values, whereas succinate-dependent oxidation rate only to 68 %. We find a similar effect on the activities of glycerol-3-phosphate and succinate cytochrome c oxidoreductase. Our results also indicate that succinate-dependent oxidation is less sensitive to digitonin treatment and less activated by IDB in comparison with glycerol-3-phosphate-dependent oxidation. These findings might indicate the different mechanism of the electron transfer from two flavoprotein-dependent dehydrogenases (glycerol-3-phosphate dehydrogenase and succinate dehydrogenase) localized on the outer and inner face of the inner mitochondrial membrane, respectively.

Topics: Animals; Cytochromes c; Digitonin; Disease Models, Animal; Dose-Response Relationship, Drug; Glycerolphosphate Dehydrogenase; Glycerophosphates; Hyperthyroidism; Kinetics; Male; Mitochondria, Liver; Mitochondrial Membranes; Oxidation-Reduction; Oxygen Consumption; Rats; Rats, Wistar; Recovery of Function; Succinate Cytochrome c Oxidoreductase; Succinic Acid; Ubiquinone

Thyrotoxicosis may be associated with a variety of abnormalities of liver function. The pathogenesis of hepatic dysfunction in thyrotoxicosis is unknown, but has been attributed to mitochondrial dysfunction. We studied the effect of altered thyroid function on the apoptotic index in rat liver. Extensive DNA fragmentation and significantly increased caspase-3 activity (P <.001) and caspase-9 activation (P <.005) were observed in hyperthyroid rat liver; cell death by apoptosis was confirmed. In hyperthyroid rat liver, 60% of mitochondria exhibited disruption of their outer membranes and a decrease in the number of cristae. These findings, along with significant translocation of cytochrome c and second mitochondria-derived activator of caspases to cytosol (P <.005), suggest activation of a mitochondrial-mediated pathway. However, no change in the expression levels of Bcl-2, Bax, and Bcl-x(L) were found in hyperthyroidism. For in vitro experiments, rat liver mitochondria were isolated and purified in sucrose density gradients and were treated with triiodothyronine (T3; 2-8 microM). T3 treatment resulted in an abrupt increase in mitochondrial permeability transition. Using a cell-free apoptosis system, the apoptogenic nature of proteins released from mitochondria was confirmed by observing changes in nuclear morphologic features and DNA fragmentation. Proteins released by 6 microM T3 contained significantly increased amounts of cytochrome c (P <.01) and induced apoptotic changes in 67% of nuclei. In conclusion, using in vivo and in vitro approaches, we provide evidence that excess T3 causes liver dysfunction by inducing apoptosis, as a result of activation of a mitochondria-dependent pathway. Thus, the results of this study provide an explanation for liver dysfunction associated with hyperthyroidism.

Topics: Alanine Transaminase; Animals; Aspartate Aminotransferases; Caspase 3; Caspase 9; Caspases; Cytochromes c; Cytosol; DNA Fragmentation; Hyperthyroidism; Hypothyroidism; Liver; Male; Mitochondria; Proto-Oncogene Proteins c-bcl-2; Rats; Rats, Sprague-Dawley; Severity of Illness Index; Triiodothyronine