Correction of stress-induced disorders of the ultrastructure of hypertrophied myocardium with thyroid hormones

In the experiment performed on 25 non-inbred male rats ultrastructural changes in cardiomyocytes of the hypertrophied heart have been studied under conditions of stress caused by immobilization and possibility to correct these changes by means of thyroid hormones. The stress intensifies destructive lesions in a number of organelles++, which develop at a prolonged hypertrophy, decreases essentially the ratio mitochondria/myofibrils in section area. Small doses of the thyroid hormones protect the hypertrophied heart from the damaging effect of the stress: prevent the stress-induced+ decrease in the ratio mitochondria/myofibrils, as well as stimulate development of the regenerative-adaptive processes (increase in size and number of mitochondria and their crists, elements of the sarcoplasmic reticulum, glycogen granules, increase in section areas of their nuclei and chromatin in them). The thyroid hormones restrict essentially decrease in correlation of organelles++, resulted from hypertrophy. Thus, the stress-induced disturbances in ultrastructure of the hypertrophied heart can be prevent by means of the thyroid hormones, administered in small doses.     

Myocardial damage during immobilization stress and the protective effect of thyroid hormones

In the experiments performed on 30 white inbred male rats, the stress produces an essential increase of dilated canals in the T-system and sarcoplasmic reticulum, lesions of sarcolemma and nucleolemma, fragmentation and homogenization of mitochondrial cristae; there are many pinocytic vesicles in the capillary walls. Preliminary injection of thyroid hormones in small doses, that do not result in any noticeable influence on the body mass, cardiac contraction rate and thyrotoxin concentration in blood serum, at the stress causes less pronounced changes in ultrastructure of cardiomyocytes, demonstrating as a poor swelling of the T-system elements, preservation of the sarcolemmic integrity at its larger length; besides, large mitochondria with compact arrangement of cristae appear, amount of the sarcoplasmic reticulum elements increases, chromatin margination becomes evident, i. e. a complex of adaptive processes develops. The data presented demonstrate a defensive role of physiological concentrations of the thyroid hormones under the stress.     

Stress-induced changes in the area ratio or the quantitative ratio of myocardial mitochondria and myofibrils and their correction with thyroid hormones

An experimental study performed on 30 male white rats has demonstrated that immobilization stress was associated with a decrease in mitochondrial to myofibril area ratio (Smch/Smf) in the left ventricle and interventricular septum by 34.6% and 46.9%, respectively. Pretreatment with small doses of thyroid hormones which did not influence body weight, heart rhythm or serum thyroxin level prevented the decrease and caused Smch/Smf increase in the left ventricle and interventricular septum by 68.7% and 45.8%, respectively. The doses of thyroid hormones applied caused a more marked increase of mitochondrial to myofibril area ratio in the control rats.     

Effect of the hyperbaric oxygenation of animals and man on mitochondrial function in their tissues (based on EPR study data)

The ration of differential intensities of EPR signal of free endogenous radicals of semiquinone type and iron-sulfur proteins recorded at the temperature of liquid nitrogen in tissues (R index) is proposed as a criterion for estimation of changes in the functional state of mitochondria in human and animal tissues treated with hyperbaric oxygenation (HBO). The increase in R value was observed in hearts of intact mice and rabbits treated with HBO in near-toxic doses. This indicates a shift in mitochondria redox state towards oxidation. The above effect was not observed in mitochondria from intact zone of myocardium in animals with experimental infarction, and the numbers of mitochondria in the tissue increased after HBO. HBO treatment of women with pathological pregnancy leads to the decrease in R value for placental mitochondria.     

The effect of irradiation by a He-Ne laser and phytohemagglutinin on lymphocyte mitochondria

Electron-microscopic morphometry has been applied to study mitochondria on ultrathin sections of lymphocytes from human peripheral blood. It has been shown that the stimulation of lymphocytes by the mitogen phytohemagglutinin (PHA) 1 h causes increases in the quantity of mitochondria per cellular section (17%) as well as in the total area of mitochondria per cell section (35%), i.e. an increase in mitochondrial mass. Taking into account known facts about growth and division of mitochondria in late phases of cellular cycle, one can suppose that described above changes in mitochondria during G0----G1 transition under action of PHA belong to an early phase of biogenesis of mitochondria. In the contrary, irradiation of lymphocytes with He-Ne-laser (lambda = 632.8 nm) in dose 56 J/m2 which does not cause the G0----C1 transition, results in the increase in the number of mitochondria per cellular section (20%) but not increase in the total area of mitochondria per cell section. The last finding indicates to some modification of space configuration of the mitochondria without any changes in their mass. The increase in the quantity of mitochondria per cellular section after the irradiation could be related with the increase in electrochemical proton gradient and in phosphorylating activity of mitochondria. He-Ne-laser radiation as well as mitogen PHA cause some deaggregation of mitochondria (this is more pronounced in case of PHA) which may be related to their functional activation.     

The responses of mitochondrial proteome in rat liver to the consumption of moderate ethanol: the possible roles of aldo-keto reductases

A large body of evidence supports the view that mitochondria are a primary target of alcohol stress. Changes in mitochondrial proteins due to moderate ethanol intake, however, have not been broadly and accurately estimated. For this study, rats were fed low doses of ethanol and the mitochondria were isolated from heart, kidney, and liver, using ultracentrifugation with Nycodenz density gradient. The mitochondrial proteins were well resolved upon two-dimensional electrophoresis (2DE), and the alcohol-responsive 2DE spots were identified by matrix-assisted laser desorption/ionization-time-of-flight mass spectrometry (MALDI-TOF/TOF MS). Compared with the control group, the proteins extracted from liver mitochondria of ethanol-fed rats exhibited the significant changes on 2DE images, whereas the 2DE images obtained from the kidney and the heart mitochondria remained almost unchanged by ethanol feeding. Significantly, over 50% of the alcohol-responsive proteins in liver mitochondria were members of aldo-keto reductase family (AKR), which were usually present in cytoplasm. The organelle distributions of AKR proteins in liver mitochondria were further confirmed by Western blot analysis as well as by confocal microscopy. In addition, translocations of AKR were examined in the CHANG cell line, which was cultured with and without ethanol. The results of Western blot strongly suggested that the abundances of AKR proteins in the mitochondria were greatly reduced by the presence of ethanol in culture medium. The results of this study show that, even with moderate ethanol feeding, the mitochondrial proteome in rat liver was more sensitive to alcohol stress than that of either the kidney or the heart. The translocation of AKR proteins may be involved in the detoxification of liver cells.     

Effect of thyroid state on susceptibility to oxidants and swelling of mitochondria from rat tissues

The effects of the thyroid state on oxidative damage, antioxidant capacity, susceptibility to in vitro oxidative stress and Ca(2+)-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 micro 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(2+)-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(2+)-induced swelling may have important implications for the thyroid hormone regulation of the turnover of proteins and whole mitochondria, respectively.     

Dependence of organism stability on chronic stress from thyroid status

In experiences on 108 male rats, the effect of acute (immobilization during 3 hrs) and chronic (immobilization for 3 hrs during 5 days) stresses on the organism general stability was studied as evaluated with changes of body weight, adrenal glands, spleen, thymus relative mass, gastric mucosa state, animals physical endurance. Chronic stress evoked more obvious decreasing of spleen and thymus relative mass than the acute one, as well as lesion of gastric mucosa accompanied with decrease of the rat resistance to physical loading. Thyroid function suppression by merkazolil (1.2 mg/100 g body weight during 14 days) promotes further the reduction of the organism stability in acute and, especially, in chronic stress, while physiological doses of thyroid hormones (5.0-8.0 mcg of thyroxin on kg of body weight during 28 days), on the contrary, increased it in both stress conditions. Existence of the organism stability dependence on thyroid status both in acute and chronic stress proves iodothyronine's important role in the organism antistress-system.     

Thyroid hormone effect on gene expression of the adenine nucleotide translocase in different rat tissues

The ADP/ATP transport across the mitochondrial membrane is achieved by the adenine nucleotide translocase (ANT), an integral inner mitochondrial membrane protein. As deduced from experiments in rat liver in vivo and in isolated rat liver mitochondria this ADP/ATP transport is accelerated by thyroid hormone application, thus explaining, at least to a considerable extent, the thyroid hormone mediated increase in mitochondrial metabolic activity. The present study investigates the effect of T3 on rat liver, heart, and kidney ANT gene expression. As shown by Northern blot analysis, a cDNA for beef heart ANT-mRNA showed cross-hybridization with the ANT-mRNA from rat heart, liver, and kidney. Hypo- and hyperthyroid rats showed no differences in size nor in amounts of heart, liver, and kidney ANT-mRNA. Measurement of heart ANT-protein level revealed no major differences among the various thyroid states. Thus, the long-term action of thyroid hormones on increasing the carrier-mediated ADP/ATP translocation cannot be ascribed to an effect of T3 on ANT gene expression. The mechanism by which T3 activates this transporter system remains to be identified but some possibilities are suggested.     

The mechanism of the increase in mitochondrial proton permeability induced by thyroid hormones.

Three possible mechanisms by which different levels of thyroid hormones in rats might cause the observed sevenfold change in the apparent proton permeability of the inner membrane of isolated liver mitochondria were investigated. (a) Cytochrome c oxidase was isolated from the livers of hypothyroid, euthyroid and hyperthyroid rats and incorporated into liposomes made with soya phospholipids. There was no difference between the proton current/voltage curves of the three types of vesicles. The hormonal effects, therefore, were not an inherent property of the enzymes, and were not due to different coupling of electron flow through the enzyme to proton transport. (b) The surface area of the mitochondrial inner membrane was shown by three different assays to be greater by a factor of between two and three in mitochondria from hyperthyroid animals than in mitochondria from hypothyroid animals; euthyroid controls were intermediate. This difference in surface area of the inner membrane explains less than half of the difference in apparent proton permeability. (c) The proton permeability of liposomes prepared from phospholipids extracted from mitochondrial inner membranes of hyperthyroid rats was three times greater than the proton permeability of those from hypothyroid rats; euthyroid controls were intermediate. This suggests, first, that the proton permeability of the phospholipid bilayer is an important component of the proton permeability in intact mitochondria and, second, thyroid hormone-induced changes in the bilayer are a major part of the mechanism of increased proton permeability. Such changes may be due to the known differences in fatty acid composition of mitochondrial phospholipids in different thyroid states. Thus we have identified two mechanisms by which thyroid hormone levels in rats change proton flux/mass protein in isolated liver mitochondria: a change in the area of the inner membrane/mass protein and a change in the intrinsic permeability of the phospholipid bilayer.     

Increase of probability of cell loss in the endothelium of blood vessel capillaries in progeny of rats after a low-dose irradiation of one of the parents (electron microscopy study)

The total gamma-irradiation of Wistar rats at a dose of 0.25 Gy as well as at higher doses (0.5, 2 and 4.5 Gy) produces in the capillary endothelial cells of myocardium and lung a pronounced, dose-independent increase of the yield of necrotized cells. Similar changes were revealed in the animals, of which one of the parents (a male one day, a female seven days prior to copulation) was irradiated at doses of 0.25 and 0.5 Gy. This effect was observed in all studied descendants. The massive induction of the changes already by low radiation actions and their dose-independence allow considering the revealed effects as a manifestation of peculiar cellular reactions that presumably have epigenetic nature.     

Ultrastructural-morphometric characterization of adaptive and compensatory myocardial changes using heart enlargement as an example

A morphometric analysis of the heart muscle cell shows significant differences in the reaction of different cellular structures in different experimental stress models of the myocardium (swimming exercise and ligature infarction) in the rat. For instance, the number of mitochondria is 44% higher only 2 days after infarction than after 45 h of swimming exercise over a period of 4 weeks. The mitochondria/myofibrils ratio is higher than in the case of training. But later on in the course of the experiment the number of mitochondria and their membrane surface density diminishes compared with training. The differences in the rate of formation and in the kind and intensity of the changes are probably attributable to different adaptive mechanisms by means of which the heart reacts to acute and chronic loads.     

Mitochondria as a target of cardioprotection in models of preconditioning

Over the recent years the view on mitochondria in the heart as a cellular powerhouse providing ATP supply needed to sustain contractile function, basal metabolic processes, and ionic homeostasis has changed radically. At present it is known that dysfunctions of these organelles are essential in the development of a large number of diseases, including cardiovascular diseases. Moreover, mitochondria are considered to be a very promising target of endogenous strategies that are essential in the protection of the myocardium from acute ischemia/reperfusion injury. These strategies including ischemic preconditioning, remote ischemic preconditioning as well as the acute phase of streptozotocin-induced diabetes mellitus, provide a similar effect of protection. Alterations observed in the functional and structural properties of heart mitochondria caused by short-term pathological impulses are associated with endogenous cardioprotective processes. It seems that the extent of mitochondrial membrane fluidization could be an active response mechanism to injury with a subtle effect on membrane-associated processes which further affect the environment of the whole organelle, thus inducing metabolic changes in the heart. In this review article, we provide an overview of endogenous protective mechanisms induced by hypoxic, pseudohypoxic and ischemic conditions with special consideration of the role of heart mitochondria in these processes.     

Changes in hexokinase isoenzymes in regions of rat brain during thyroid deficiency

In this work, activities of hexokinase isoenzymes Type I and Type II were measured in the soluble and particulate fractions from the brain regions (cerebral hemispheres (cerebrum), cerebellum and brain stem) of the thyroidectomized adult rats as well as of the thyroidectomized rats administered with triiodothyronine. Thyroidectomy generally decreased the hexokinase activity associated with particulate and soluble fractions. Hexokinase Type II isoenzyme was more affected than the Type I isoenzyme. Administration of triiodothyronine to the hypothyroid rats abolished the effect of thyroidectomy. Adult brain enzymes have been generally considered not be affected by thyroid hormones. The data obtained in this work are suggestive of an effect of thyroid hormones on hexokinase in the adult brain. Since the effects of thyroidectomy on the energy metabolism of the heart tissue are well known, the heart tissue was also studied for comparison.     

The features of mitochondria of cardiomyocytes from rats with chronic heart failure

Electron-microscopy study of rat myocardium 2 weeks after a heart attack revealed significant alterations in the ultrastructure of cardiomyocytes than for the control. The location of myofibrils was less regular than for normal cells. The population of interfibrillar mitochondria decreased. Mitochondrial cristae were located less densely and formed cellated structures. Swollen mitochondria were observed in the periinfarction and intact areas, indicating the development of ischemia in the myocardium as a whole. Six months after the occlusion of coronary vessels alterations in the location of myofibrils and mitochondria were mainly observed in the peri-infarction area. Mitochondria also formed cellated structures. A 30% decrease in the density of the arrangement of the inner membranes of mitochondria on an area unit was found in the periinfarction zone. The ratio between the relative volumes of mitochondria and myofibrils in the cardiomyocytes of the peri-infarction area was increased by 20%. The area of mitochondria in the intact zone of the left ventricle was 30% greater than for the control. A study of isolated living cardiomyocytes revealed that the mitochondrial- membrane potential in the rats subjected to myocardial infarction half a year ago previously was significantly lower than for the mitochondrial-membrane potential in the control rats. Thus, cardiomyocytes that were similar to healthy cardiomyocytes in their morphology exhibited lower total mitochondrial-membrane potential, indicating their decreased energy state.     

Thyroid hormones and mitochondria

Because of their central role in the regulation of energy-transduction, mitochondria, the major site of oxidative processes within the cell, are considered a likely subcellular target for the action that thyroid hormones exert on energy metabolism. However, the mechanism underlying the regulation of basal metabolic rate (BMR) by thyroid hormones still remains unclear. It has been suggested that these hormones might uncouple substrate oxidation from ATP synthesis, but there are no clear-cut data to support this idea. Two iodothyronines have been identified as effectors of the actions of thyroid hormones on energy metabolism: 3',3,5-triiodo-L-thyronine (T3) and 3,5-diiodo-L-thyronine (T2). Both have significant effects on BMR, but their mechanisms of action are not identical. T3 acts on the nucleus to influence the expression of genes involved in the regulation of cellular metabolism and mitochondria function; 3,5-T2, on the other hand, acts by directly influencing the mitochondrial energy-transduction apparatus. A molecular determinant of the effects of T3 could be uncoupling protein-3 (UCP-3), while the cytochrome-c oxidase complex is a possible target for 3,5-T2. In conclusion, it is likely that iodothyronines regulate energy metabolism by both short-term and long-term mechanisms, and that they act in more than one way in affecting mitochondrial functions.     

Restriction of stress-induced activation of lipid peroxidation by small doses of thyroid hormones]

The experiments on 57 female rats demonstrated that small doses of thyroid hormones (thyroidin) significantly (55-118%) restrict stress induced increase in the concentration of initial and terminal products of lipid peroxidation (LP) in the myocardium and in the blood plasma. After hormone injection stress decreases the activity of key antioxidant enzyme, superoxide dismutase of erythrocytes (SOD), to a lesser degree and increases the rate of malonyldialdehyde (MDA) production induced by Fe2+ in homogenates of the myocardium to the same degree as well in comparison with rats that had not been injected thyroidin. In normal rates thyroidin does not influence the concentration of products of LP, increases the activity of SOD and decreases increment of MDA induced by Fe2+ in homogenates of the myocardium. Thus, small doses of thyroid hormones restrict significantly stress induced activity of LP membranes, increasing the power of antioxidant systems both in the myocardium and in the organism.     

Regulation of vernal migration in Gambel's white-crowned sparrows: Role of thyroxine and triiodothyronine

Appropriate timing of migratory behavior is critical for migrant species. For many temperate zone birds in the spring, lengthening photoperiod is the initial cue leading to morphological, physiological and behavior changes that are necessary for vernal migration and breeding. Strong evidence has emerged in recent years linking thyroid hormone signaling to the photoinduction of breeding in birds while more limited information suggest a potential role in the regulation of vernal migration in photoperiodic songbirds. Here we investigate the development and expression of the vernal migratory life history stage in captive Gambel's white-crowned sparrows (Zonotrichia leucophrys gambelii) in a hypothyroidic state, induced by chemical inhibition of thyroid hormone production. To explore possible variations in the effects of the two thyroid hormones, triiodothyronine and thyroxine, we subsequently performed a thyroid inhibition coupled with replacement therapy. We found that chemical inhibition of thyroid hormones resulted in complete abolishment of mass gain, fattening, and muscle hypertrophy associated with migratory preparation as well as resulting in failure to display nocturnal restlessness behavior. Replacement of thyroxine rescued all of these elements to near control levels while triiodothyronine replacement displayed partial or delayed rescue. Our findings support thyroid hormones as being necessary for the expression of changes in morphology and physiology associated with migration as well as migratory behavior itself.