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.     

Alterations in thyroid hormone physiology induced by temperature and feeding in newly hatched chickens

In the chicken the transition of a poikilotherm to a homeotherm reaction upon cold exposure takes place in the perinatal period between pipping and hatching. However, newly hatched chicks cannot maintain their body temperature within narrow limits after cold exposure. The fact that relatively little attention was payed on the role of thyroid hormones in the thermoregulatory reaction to cold of young chicks was probably due to the hypothetically long latention time that was thought to be necessary to bring about changes in secretory activity by cold stimulation. However, more recently, rapid changes (within hours) of thyroid hormone concentrations upon cold exposure were described in the chickens and the quail. In this study, changes in circulating T3 and T4 concentrations upon cold exposure of young chicks during the first two weeks were followed, that means during the period wherein NST (non-shivering thermogenesis), if it exists at all, should be progressively replaced by ST (shivering thermogenesis). Because of the importance of feeding condition on thyroid hormone levels, the experiments were carried out with and without a preceeding fasting period. In all experiments a short-term cold exposure of young chickens (1-11 days) fed ad lib decreased T3 but increased T4 levels while a reversed picture was found after short cold exposure of the fasted animals. However, after prolonged cold stimulus (15 degrees C) of young chickens fed ad lib, plasma T3 was also significantly elevated over that of controls whereas T4 levels returned to normal values. A prolonged warm treatment (37 degrees C) of young chickens fed ad lib resulted in significantly lower T3 and higher T4 concentrations. After a prolonged cold treatment no differences in T4 or T3 response upon TRH were found whereas the warm treatment abolished these responses upon TRH. However, a cold treatment at the stage of incubation during which the hypothalamo-hypophyseal control of thyroid function is established (dag 10-14) enhanced the T4 response to TRH with a long lasting effect extending to the posthatch period. Since T3 is thought to be the active form of thyroid hormones with regard to thermopoiesis we have studied more specifically the effect of blocking peripheral conversion of T4 on thermoregulatory abilities in young chicks and the influence of temperature treatment on monodeiodination capacity. The lower rectal temperatures following the interference with the peripheral monodeiodination of T4, the effect being more pronounced at the lower ambient temperature, are indicative for a preponderant role of T3 on thermogenesis.(ABSTRACT TRUNCATED AT 400 WORDS)     

The mitochondrial protonic electrochemical potential difference as a point of hormone action. II. New proposals for the activity of glucagon

Many of the data concerning the action of glucagon upon liver mitochondria are assimilated in this communication. This leads to the novel proposal that glucagon acts to inhibit the conductance of the mitochondrial inner membrane. When the isolated mitochondria are incubated in the absence of ADP, this proposed action of glucagon provides an explanation for the increase in the aerobic protonic electrochemical potential difference across the inner membrane, without effect upon respiratory rate. However, when the organelles are incubated in the presence of ADP, a different action of glucagon is suggested to increase the rate of respiration. These proposals are compared with those recently put forward to explain the action of thyroid hormones upon mitochrondria. An analysis is also made of the likely respiratory state of mitochondria in vivo, and the possible physiological significance of the new proposals is discussed in this context.     

Circulating thyroid hormones in primates with mild or severe hepatitis following liver transplantation

In order to study the effects of acute immunologically mediated liver disease on circulating thyroid hormones, serum levels of thyroxine (T4, total and free) and triiodothyronine (T3) were measured in 8 baboons before and for 60 days after allogeneic liver transplantation. In 3 animals early rejection and jaundice developed; T4 levels declined as liver function deteriorated. In the 5 tolerant animals liver function was only temporarily deranged without jaundice and there was a consistent early rise in T4 (P less than 0.01) followed by a later fall. T3 concentrations were relatively normal in both groups. The T3 resin uptake test remained virtually unchanged in all animals. Serum T4 and T3 responses to exogenously administered bovine thyrotropin (TSH) were similar in the jaundiced and anicteric animals. We conclude that the early rise in T4 in the tolerant animals was caused by transient increases in thyroid binding globulin in (TBG) while the fall in thyroid hormones in these and in the jaundiced animals was related to a decline in TBG levels. Thyroid responsiveness to TSH is not disturbed by moderately deranged liver function.     

Effect of propranolol on the metabolism of thyroid hormones

The effect of propranolol on thyroxine (T4) and 3,5',3'-triiodothyronine (T3) plasma concentrations, fractional disappearance rates (k), metabolic clearance rates (MCR) and catabolic rates has been investigated in young pigs. The animals were examined in the period 18-24 h after feeding. Plasma concentrations of T3 were lower after treatment with propranolol, but T4 was not significantly affected. The k value of T4 was decreased by propranolol but that for T3 was unaffected. The MCR of T4 and the catabolic rates of both hormones were reduced by propranolol. The reduction in metabolic rate after propranolol is thus probably related to its action on thyroid hormone metabolism.     

The nature of thyroid hormone receptors. Intracellular functions of thyroxine-binding prealbumin

The effect of tyroxin-binding prealbumin (TBPA) of blood serum on the template activity of chromatin was studied. It was found that the values of binding constants of TBPA for T3 and T4 are 2 X 10(-11) M and 5 X 10(-10) M, respectively. The receptors isolated from 0.4 M KCl extract of chromatin and mitochondria as well as hormone-bound TBPA cause similar effects on the template activity of chromatin. Based on experimental results and the previously published comparative data on the structure of TBPA, nuclear, cytoplasmic and mitochondrial receptors of thyroid hormones as well as on translocation across the plasma membrane and intracellular transport of TBPA, a conclusion was drawn, which suggested that TBPA is the "core" of the true thyroid hormone receptor. It was shown that T3-bound TBPA caused histone H1-dependent conformational changes in chromatin. Based on the studies with the interaction of the TBPA-T3 complex with spin-labeled chromatin, a scheme of functioning of the thyroid hormone nuclear receptor was proposed.     

Effect of thyroid hormones and their analogues on the mitochondrial calcium transport activity.

In this paper the authors studied the effects of thyroid hormones and their structural analogues on the mitochondrial calcium transport activities. The thyroid hormones, 3,5,3' L-triiodothyronine (LT3) and 3,5,3'5' L-tetraiodothyronine (LT4) at physiological intracellular concentrations between 7.2 and 9 nM, decouple total Ca++ transport, as well as inhibit the passive transport of Ca++, either due to oxidation of pyruvate, malate or succinate or after inhibition with rotenone. The optical isomers 3,5,3' D-triiodothyronine (DT3) and 3,5,3',5' D-tetraiodothyronine (DT4) are less effective at all the used concentrations. Furthermore the structural analogues 3,3',5' L-triiodothyronine (LrT3), 3,5-dicloro, 3',5' L-diiodothyronine (LDiClT2) and 3,5 L-diiodothyronine (LT2) furnished even less effects on the same activities. The effect of the thyroid hormones and of their structural analogues has revealed that the mitochondrial calcium transport may be influenced both by a stereospecific interaction between hormones and protein ligands and by a lipophilic chaotropic action on the mitochondrial membranes lipids. In this context it is interesting to consider that both thyroid hormones and Ca++ transport activity are interacting with the energetic metabolism by means of phosphorylation and substrate oxidation mechanism.     

Evaluation of the antioxidant properties of thyroid hormones and propylthiouracil in the brain-homogenate autoxidation system and in the free radical-mediated oxidation of erythrocyte membranes

The antioxidant capacity of thyroid hormones and the antithyroid drug propylthiouracil was studied in three model systems, namely, autoxidation of rat brain homogenates and oxidation of rat erythrocyte plasma membranes (EPM) induced by either 2,2'-azobis-(2-amidinopropane) (AAP) thermolysis or by gamma irradiation. Thyroid hormones significantly inhibited the development of lipid peroxidation in these systems at micromolar concentrations, as assessed either by visible light emission, thiobarbituric acid reactive substances accumulation or oxygen uptake. This behaviour was not observed when L-3,3',5-triiodothyronine (T3) and L-thyroxine (T4) were assayed at nanomolar concentrations. In EPM exposed to AAP or gamma irradiation, propylthiouracil inhibited the induced lipid peroxidation, with Q1/2 values of 112-150 microM. It is concluded that the antioxidant capacity of thyroid hormones found in vitro may not be of relevance in physiological conditions, which exhibit variations of T3 and T4 levels in the nanomolar range. On the other hand, the behaviour of propylthiouracil as an inhibitor of EPM lipid peroxidation is observed at concentrations close to the therapeutic levels, thus representing a possible complementary action to its antithyroid activity.     

Mitochondrial Interaction with Helminthosporium maydis Race T Toxin: Blocking by Dicyclohexylcarbodiimide

Treatment of mitochondria isolated from Texas male sterile cytoplasmcorn (T mitochondria) with high concentrations of dicyclohexylcarbodiimide(DCCD) (140 nmol DCCD mg^–1 mitochondrial protein) completelyand immediately inhibited T mitochondrial swelling by Helminthosporiummaydis Race T toxin (HmT toxin). In order to obtain a specificinteraction between DCCD and the ATPase complex T mitochondriawere incubated with lower DCCD concentrations (1–5 nmolDCCD mg^–1 mitochondrial protein) for up to 8 h at 4 °C.After 8 h incubation in the presence of 3.75 nmol DCCD mg^–1mitochondrial protein, toxin-induced swelling was decreasedby 69%. Specificity of DCCD action upon the ATPase complex wasconfirmed by (1) SDS gel electrophoresis and fluorographic analysesof proteins from [¹⁴C]-DCCD-treated T mitochondria and immunoprecipitatesand (2) physiological experiments showing that DCCD exertednone of its other documented effects. These data suggest thatHmT toxin interacts with the ATPase complex of T mitochondriaeither at or near the DCCD-binding protein within the membranesector of the complex. Key words: Zea mays L., Helminthosporium maydis, Mitochondria     

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 effects of age, season and geographic region on thyroid hormones in Steller sea lions (Eumetopias jubatus)

The purpose of this study was to investigate thyroid hormone concentrations, thyroxine (T4) and triiodothyronine (T3), in order to determine basal levels in Steller sea lions of different ages and over seasons. Serum concentrations of total T4 were highest in Steller sea lions followed by total T3 concentrations. Concentrations of free T4 and free T3 were three to four orders of magnitude lower. Concentrations for all four thyroid hormone measurements tended to a lower level as animals matured beyond the neonatal stage. When thyroid hormones from captive sea lions were evaluated across seasons, all thyroid hormones were highest in the July to September period. When compared across the geographic range, animals in southeast Alaska tended to have lower thyroid hormone levels, while the Steller sea lions west of Prince William Sound and animals from the Russian Far East had significantly higher concentrations. Significant inter-annual differences in concentrations were also observed across the geographic range. With an understanding of the basic changes in thyroid hormone concentrations, changes in plane of nutrition or life history states (i.e. fasting, pregnancy or lactation) can now be evaluated for their effect on the overall health of this endangered species.     

A novel interaction between thyroid hormones and 1,25(OH)(2)D(3) in osteoclast formation

Thyroid hormones enhance osteoclast formation and their excess is an important cause of secondary osteoporosis. 3,5,3' -Triiodo-L-thyronine (T3) induced the mRNA expression of receptor activator of nuclear factor-kappa B ligand (RANKL), which is a key molecule in osteoclast formation, in primary osteoblastic cells (POB). This effect was amplified in the copresence of 1 alpha,25-dihydroxyvitamin D(3) (1,25(OH)(2)D(3)). Although T3 alone did not induce octeoclasts in coculture of bone marrow cells with POB, T3 enhanced 1,25(OH)(2)D(3)-induced osteoclast formation. Thyroxine (T4) also enhanced 1,25(OH)(2)D(3)-induced osteoclast formation. These data suggested that T4 was locally metabolized to T3 for its action, since T4 is a prohormone with little hormonal activity. The mRNA expression of type-2 iodothyronine deiodinase (D2), which is responsible for maintaining local T3 concentration, was induced by 1,25(OH)(2)D(3) dose- and time-dependently. Our data would facilitate our understanding of the mechanism of osteoclast formation by thyroid hormones and suggest a novel interaction between thyroid hormones and 1,25(OH)(2)D(3).     

Catecholamine and thyroid hormone metabolism in a case of anorexia nervosa

Alterations in catecholamine (CA) and thyroid hormone metabolism were examined in a 12-year-old girl with anorexia nervosa during 3 months of treatment. According to her body weight change, the observation period was divided into 3 stages: initial emaciation (stage 1), stable maintenance of the -30% level of the previous weight (stage 2) and convalescent stage (stage 3). Stage 1 was characterized by relatively high urinary norepinephrine (NE) and epinephrine (E) but low dopamine (DA) excretion, elevated plasma DA-beta-hydroxylase (DBH) activity and reduced serum thyroid hormones, especially the triiodothyronine (T3) level. In stage 2, urinary CAs were markedly suppressed, while serum thyroid hormones gradually increase. In stage 3, a great increase in DA excretion, a fall in plasma DBH activity and normalization of thyroid hormones were observed. In the low T3 state below 60 ng/dl, urinary NE + E/DA ratios were elevated and widely fluctuated (0.58 +/- 0.30, SD), but were gradually decreased and completely stabilized in the normal T3 state (0.07 +/- 0.02, P less than 0.001). These results indicate that (1) although total CA production was depressed in anorexia nervosa, a change from an adrenergic-dominant to a dopaminergic-dominant state occurs in accordance with body weight gain, and (2) this shift in the CA profile is associated with concomitant recovery of reduced thyroid hormone concentrations. Thus, as for the energy expenditure, compensatory changes were observed in CAs and thyroid hormones in relation to caloric restriction.     

Mitochondrial respiration in muscle and liver from cold-acclimated hypothyroid rats.

The effects of long-term cold exposure on muscle and liver mitochondrial oxygen consumption in hypothyroid and normal rats were examined. Thyroid ablation was performed after 8-wk acclimation to 4 degrees C. Hypothyroid and normal controls remained in the cold for an additional 8 wk. At the end of 16-wk cold exposure, all hypothyroid rats were alive and normothermic and had normal body weight. At ambient temperature (24 degrees C), thyroid ablation induced a 65% fall in muscle mitochondrial oxygen consumption, which was reversed by thyroxine but not by norepinephrine administration. After cold acclimation was reached, suppression of thyroid function reduced muscle mitochondrial respiration by 30%, but the hypothyroid values remained about threefold higher than those in hypothyroid muscle in the warm. Blockade of beta- and alpha1-adrenergic receptors in both hypothyroid and normal rats produced hypothermia in vivo and a fall in muscle, liver, and brown adipose tissue mitochondria respiration in vitro. In normal rats, cold acclimation enhanced muscle respiration by 35%, in liver 18%, and in brown adipose tissue 450% over values in the warm. The results demonstrate that thyroid hormones, in the presence of norepinephrine, are major determinants of thermogenic activity in muscle and liver of cold-acclimated rats. After thyroid ablation, cold-induced nonshivering thermogenesis replaced 3,5,3'-triiodothyronine-induced thermogenesis, and normal body temperature was maintained.     

beta-2-Aminobicyclo-(2.2.1)-heptane-2-carboxylic acid. A new activator of glutaminase in intact rat liver mitochondria

beta-(+/-)-2-Aminobicyclo-(2.2.1)-heptane-2-carboxylic acid (BCH) stimulated, in a concentration-dependent manner, the formation of glutamate by mitochondria isolated from rat liver and incubated with 20 mM glutamine. Maximum enhancement was seen with 10 mM BCH while 5 mM leucine was without effect. The initial lag in the rate of glutamate formation was not eliminated by BCH. Preincubation of the mitochondria without glutamine also did not abolish the lag period; to the contrary, it resulted in a progressive deactivation of the glutaminase. The decrease in enzyme activity during the preincubation without glutamine was partially reversed by the addition of either 10 mM BCH or 1.4 mM NH4Cl and was essentially abolished by their combined action. The apparently sigmoid rise in the activity of glutaminase with increasing concentration of glutamine became hyperbolic in the presence of 1.4 mM NH4Cl. BCH stimulated the NH4Cl-activated glutaminase in the entire range of glutamine concentrations studied (2-40 mM) without changing the S50 value. In mitochondria disrupted by repeated cycles of freezing and thawing, the enzymatic activity was maximal even in the absence of BCH. It is postulated that BCH is a potent activator of mitochondrial glutaminase and that manifestation of its action requires intact organelle structure. In addition, it is concluded that BCH-induced stimulation of glutamine catabolism in isolated hepatocytes (Zaleski, J., Wilson, D. F., and Erecinska, M. (1986) J. Biol. Chem. 261, 14082-14090) is the consequence of activation of the mitochondrial glutaminase.     

Destabilization of the cytosolic calcium level and the death of cardiomyocytes in the presence of derivatives of long-chain fatty acids

By means of fluorescent microscopy, long-chain fatty acid derivatives, myristoylcarnitine and palmitoylcarnitine, were shown to exert the most toxic effect on rat ventricular cardiomyocytes. The addition of 20–50 μM acylcarnitines increased calcium concentration in cytoplasm ([Ca²⁺]_i) and caused cell death after a lag-period of 4–8 min. This effect was independent of extracellular calcium level and Ca²⁺ inhibitors of L-type channels. Free myristic and palmitic acids at concentrations of 300–500 μM had little effect on [Ca²⁺]_i within 30 min. We suggest that the toxic effect is due to the activation of calcium channels of sarcoplasmic reticulum by acylcarnitines and/or arising acyl-CoA. Mitochondria play a role of calcium-buffer system under these conditions. The calcium capacity of the buffer determines the duration of the lag-period. Phosphate increases the calcium capacity of mitochondria and the lag-period. In the presence of rotenone and oligomycin, the elevation of [Ca²⁺]_i after the addition of acylcarnitines occurs without the lag-period. The exhaustion of the mitochondrial calcium-buffer capacity or significant depolarization of mitochondria leads to a rapid release of calcium from mitochondria and cell death. Thus, the activation of reticular calcium channels is the main reason of the toxicity of myristoylcarnitine and palmitoylcarnitine.     

Piperine lowers the serum concentrations of thyroid hormones, glucose and hepatic 5'D activity in adult male mice.

Piperine, the main alkaloid of Piper nigrum fruits, was evaluated for its thyroid hormone and glucose regulatory efficacy in adult male Swiss albino mice. Its daily oral administration (2.50 mg/kg) for 15 days lowered the serum levels of both the thyroid hormones, thyroxin (T (4)) and triiodothyronine (T (3)) as well as glucose concentrations with a concomitant decrease in hepatic 5'D enzyme and glucose-6-phospatase (G-6-Pase) activity. However, no significant alterations were observed in animals treated with 0.25 mg/kg of piperine in any of the activities studied except an inhibition in serum T (3) concentration. The decrease in T (4), T (3) concentrations and in G-6-Pase were comparable to that of a standard antithyroid drug, Proylthiouracil (PTU). The hepatic lipid-peroxidation (LPO) and the activity of endogenous antioxidants, superoxide dismutase (SOD), and catalase (CAT) were not significantly altered in either of the doses. It appears that the action of P. nigrum on thyroid functions is mediated through its active alkaloid, piperine. We also suggest that a higher dose of piperine may inhibit thyroid function and serum glucose concentration in euthyroid individuals.     

Thyroid hormone kinetics during late pregnancy in the ovine fetus

The factors responsible for the changes in the plasma concentrations of thyroid hormones in the ovine fetus in late pregnancy were investigated by making serial measurements of the concentrations, metabolic clearance rates and production rates of T3 and T4 in 17 fetuses. The concentrations of T3 in fetuses of 135-145 days gestational age were four times higher than in those of 110-125 days but the concentrations of T4 were unchanged. The metabolic clearance rate of T3 halved over this period whereas that of T4 rose slightly. The production rate of T3 more than doubled and of T4 increased slightly but not significantly. We conclude that the concentration of T4 shows little change with increasing gestational age because the trends in metabolic clearance rates and production rates are weak and in the same direction. The sharp rise in the concentration of T3 is attributable to a fall in metabolic clearance rate coupled with a rise in production rate.     

Cellular mechanism of action of thyroid hormones

It has emerged in the last decade that the molecular mechanism of action of thyroid hormones resembles that of steroids; thyroid hormones indeed exert their effects mainly by directly regulating gene expression, on association with specific chromatin-bound receptors. Of the two thyroid hormones, thyroxine (T4) appears to be a sort of prohormone, whereas triiodothyronine (T3) seems to be the active form; in this respect, T4-deiodination, which occurs at the level of the target tissues, may be crucial in the local homeostasis of T3. Moreover, many cellular compartments, other than the nucleus, can bind thyroid hormone, and at least some of these further sites might play some role in modulating T3 supply to the nucleus. The binding of the T3-receptor complex to chromatin is likely to regulate the structural organization of specific genes and, in some instances, of the chromatin as a whole.