Injection Of T3 Research Articles

Effects of maternal injection of 3,5,3′-triiodo-l-thyronine (T3) on growth of newborn offspring of rockfish, Sebastes schlegeli

In the rockfish, Sebastes schlegeli, changes of thyroid hormone levels in plasma and egg of mature female broodstock and in whole body of newly born larvae were examined after maternal injection of 3,5,3′-triiodo-l-thyronine (T3) at a dose of 20 mg/kg body weight. Also, the growth and survival rate of offspring from T3-injected broodstock (T3 group) were compared for 30 days with those of vehicle-control (injected with DMSO: control group) and sham-control (needle inserted, but without any injection: sham group). Larvae were fed with rotifers, Artemia nauplii and commercial diet for flounder. Although there was no change of l-thyroxine (T4) level following maternal T3 injection, a significant increase of T3 level was found in maternal plasma, eggs and larvae. Growth of offspring in the T3 injection group was significantly faster than that of the two controls, and the condition factor was lower than that of controls. In addition, the survival rate of larvae from T3-injected broodstock was significantly higher than that of larvae from the two controls. We conclude that exogenous T3 in maternal circulation was transferred into oocytes and larvae. The transferred thyroid hormone appears to play some role in the early development of larvae and may confer a distinct advantage for the growth of offspring in rockfish, S. schlegeli.

Thyroid function and the development of endothermy in a marsupial, the Tasmanian bettong, Bettongia gaimardi (Demarest 1822)

The Tasmanian bettong ( Bettongia gaimardi) is a small rat-kangaroo (marsupial) found only in Tasmania, Australia. The duration of pouch life is 15 weeks. Adults and older young display non-shivering thermogenesis and this paper examines the role of thyroxine in the development of endothermy in pouch young. Free thyroxine (T4) concentrations varied throughout pouch life. The mean (± SE) concentration was 6.2 ± 1.9 pmol L −1 in week 7, increased and peaked at 19.2 ± 4.3 pmol L −1 in week 12, and declined to 5.6 ± 0.4 pmol L −1 by week 20. This was similar to adult levels (3.2 ± 3.8 pmol L −1). These concentrations showed significant differences. From pouch week 12 onwards, T4 injection raised oxygen consumption. Maximum levels of VO 2 after T4 injection occurred at weeks 14–15. Although adult levels were lower, the increase in adult oxygen consumption after T4 injection was about 50%. Peak free T4 levels and metabolic responses to nor-adrenalin occur at week 12 and we hypothesize that thyroid hormone may facilitate the development of adrenergic-receptors in this species. The data presented in the paper further attest to the likely important role of the thyroid gland in the development of endothermy in marsupial pouch young.

Thyroxine-induced cardiac hypertrophy: influence of adrenergic nervous system versus renin-angiotensin system on myocyte remodeling.

The present study assessed the possible involvement of the renin-angiotensin system (RAS) and the sympathetic nervous system (SNS) in thyroxine (T4)-induced cardiac hypertrophy. Hemodynamic parameters, heart weight (HW), ratio of HW to body weight (HW/BW), and myocyte width were evaluated in absence of thyroid hormone (hypothyroidism) and after T4 administration. Male Wistar rats were used. Some were subjected to thyroidectomies, whereas hyperthyroidism was induced in others via daily intraperitoneal injection of T4 (25 or 100 microg x 100 g BW(-1) x day(-1)) for 7 days. In some cases, T4 administration was combined with the angiotensin I-converting enzyme inhibitor enalapril (Ena), with the angiotensin type 1 (AT1) receptor blocker losartan (Los) or with the beta-adrenergic blocker propanolol (Prop). Hemodynamics and morphology were then evaluated. Systolic blood pressure (SBP) was not altered by administration of either T4 alone or T4 in combination with the specific inhibitors. However, SBP decreased significantly in hypothyroid rats. An increased heart rate was seen after administration of either T4 alone or T4 in combination with either Los or Ena. Although the higher dose of T4 significantly increased HW, HW/BW increased in both T4-treated groups. Ena and Prop inhibited the increase in HW or HW/BW in hyperthyroid rats. Morphologically, both T4 dose levels significantly increased myocyte width, an occurrence prevented by RAS or SNS blockers. There was a good correlation between changes in HW/BW and myocyte width. These results indicate that T4-induced cardiac hypertrophy is associated with both the SNS and the RAS.

Thyroid hormone enhances aggrecanase-2/ADAM-TS5 expression and proteoglycan degradation in growth plate cartilage.

Effects of thyroid hormone on proteoglycan degradation in various regions of cartilage were investigated. In propylthiouracil-treated rats with hypothyroidism, proteoglycan degradation in epiphyseal cartilage during endochondral ossification was markedly suppressed. However, injections of T(4) reversed this effect of propylthiouracil on proteoglycan degradation. In pig growth plate explants, T(3) also induced breakdown of proteoglycan. T(3) increased the release of aggrecan monomer and core protein from the explants into the medium. Accordingly, the level of aggrecan monomer remaining in the tissue decreased after T(3) treatment, and the monomer lost hyaluronic acid-binding capacity, suggesting that the cleavage site is in the interglobular domain. The aggrecan fragment released from the T(3)-exposed explants underwent cleavage at Glu(373)-Ala(374), the major aggrecanase-cleavage site. The stimulation of proteoglycan degradation by T(3) was less prominent in resting cartilage explants than in growth plate explants and was barely detectable in articular cartilage explants. Using rabbit growth plate chondrocyte cultures, we explored proteases that may be involved in T(3)-induced aggrecan degradation and found that T(3) enhanced the expression of aggrecanase-2/ADAM-TS5 (a disintegrin and a metalloproteinase domain with thrombospondin type I domains) mRNA, whereas we could not detect any enhancement of stromelysin, gelatinase, or collagenase activities or any aggrecanase-1/ADAM-TS4 mRNA expression. We also found that the aggrecanse-2 mRNA level, but not aggrecanase-1, increased at the hypertrophic stage during endochondral ossification. These findings suggest that aggrecanse-2/ADAM-TS5 is involved in aggrecan breakdown during endochondral ossification, and that thyroid hormone stimulates the aggrecan breakdown partly via the enhancement of aggrecanase-2/ADAM-TS5.

Experimentally induced hypo- and hyper-thyroidism influence on the antioxidant defence system in adult rat testis.

The objective of the present experiment was to study the effect of thyroid hormone on the antioxidant defence system of rat testis. Hypothyroidism induced in rats by 6-n propyl 2-thiouracil (PTU) treatment resulted in a reduction in body weight, seminal vesicle and ventral prostate gland. A further decrease in the weight of seminal vesicle was recorded following administration of T3 to hypothyroid rats. The oxidative stress parameters such as hydrogen peroxide and protein carbonyl content increased in the crude homogenate of testis of hypothyroid rats. T3 administration to hypothyroid rats resulted in no further change in the hydrogen peroxide level but the protein carbonyl content further elevated in the crude homogenate of testis. No significant change was observed in the endogenous lipid peroxidation level of the crude homogenate of testis whereas the FeSO4/ascorbic acid induced lipid peroxidation level decreased in hypothyroid rats and did not change further by T3 administration. Although the reduced glutathione level in the crude homogenate of testis did not change following hypothyroidism, oxidized glutathione level increased. The reduced and oxidized glutathione level decreased and increased, respectively following T3 administration to hypothyroid rats in comparison with PTU-treated rats. Activities of superoxide dismutase and catalase decreased in the post-mitochondrial fraction (PMF) of testis of hypothyroid rats. T3 injection to PTU-treated rats resulted in an elevation in the level of catalase activity only. The activity of glutathione peroxidase in the PMF of testis elevated in the hypothyroid rats and reduced following T3 treatment to hypothyroid rats. The results of the present study suggest that any alteration in the thyroid hormone level in the body affects the antioxidant defence system of testis of adult rats and, thereby, may affect the physiology of testis through oxidative stress.

Effect of thyroid hormone on the development and gene expression of hormone receptors in rat testes in vivo

Thyroid hormone is known to play a pivotal role in the regulation of prepuberal rat testes development and function with specific influence on the differentiation of Sertoli cells, the only cell type that expresses thyroid hormone receptors in testes. To explore in vivo effects of thyroid hormone on testes development and the regulation of testicular gene expression, the hyper- and hypothyroid rat models were established by T3 injection to pups (ip 100 microg/kg bw) and by oral administration of 6-N-propyl-2-thiouracil (PTU) to the lactating mother from days 1 to 21 post-delivery. Half of the rats from each group were sacrificed at 21 days of age, and the other half were allowed to recover with discontinued treatments from day 22 to day 50. At 21 days of age, a significantly elevated serum T3 level was observed in hyperthyroid rats (179.5 ng/dl) vs controls (97.5 ng/dl), and in hypothyroid rats a significantly lower level of T3 was detected (26.1 ng/dl). However, serum T4 concentration was significantly lower in both hyper- (0.105 microg/dl) and hypothyroid (0.058 microg/dl) rats compared to the controls (2.48 microg/dl). In recovered rats in which the serum T3 and T4 were restored to normal, the serum T levels remained remarkably lower in both hyper- and hypothyroid rats. The significantly decreased body and testes weights observed in both hyper- and hypothyroid rats at 21 days of age were not restored by the time they were 50 days old. Histological analyses of testes of 21-day-old hypothyroid rats revealed smaller-sized seminiferous tubules, incomplete lumen formation and delayed germ cell differentiation and in hyperthyroid rats an increased number of early stage spermatocytes was found. Testicular mRNA levels of follicle-stimulating hormone receptor (FSH-R), luteinizing hormone receptor (LH-R) and androgen binding protein (ABP) were studied by Northern blot hybridization. At 21 days of age data showed that FSH-R mRNA levels were significantly higher in both hyper- and hypothyroid rat testes compared to controls, but no differences were detected in recovered 50-day-old rats. Significantly decreased ABP mRNA levels were detected only in hypothyroid rat testes compared to those in both the hyperthyroid and control groups at 21 days of age, but no significant change was observed in recovered 50-day-old rats. To further evaluate the effect of thyroid hormone on the Leydig cell function, the 2.3/2.6 kb specific LH-R hybridization bands were detected with rat LH-R cRNA probe. Significant suppression of LH-R mRNA levels was only observed in the hypothyroid rat testes at 50 days of age. The testicular thyroid hormone receptors (TRs) and the regulation of TR by thyroid hormone were investigated using semi-quantitative reverse transcription-polymerase chain reaction (RT-PCR) assays. Both TRalpha and TRbeta mRNAs were identified in the testes from 21- and/or 50-day-old rats. TRalpha mRNA levels were significantly increased in hypothyroid rat testes and were suppressed in hyperthyroid rats at 21 days of age and no changes of TRalpha mRNA were found in recovered animals. Our in vivo data strongly suggest that the thyroid hormone directly affects the development of prepuberal testes and the regulation of FSH-R and ABP gene expression in Sertoli cells, as well as the LH-R mRNA levels in Leydig cells, which may lead to further modulating the effect of gonadotropins on testes function.

Triiodothyronine-mediated myosin heavy chain gene transcription in the heart.

We developed an RT-PCR assay to study both the time course and the mechanism for the triiodothyronine (T(3))-induced transcription of the alpha- and beta-myosin heavy chain (MHC) genes in vivo on the basis of the quantity of specific heterogeneous nuclear RNA (hnRNA). The temporal relationship of changes in transcriptional activity to the amount of alpha-MHC mRNA and the coordinated regulation of transcription of more than one gene in response to T(3) are demonstrated here for the first time. Quantitation of alpha-MHC hnRNA demonstrated that T(3) induced alpha-MHC transcription in hypothyroid rats within 30 min of a single injection of T(3) (0.5 microg/100 g body wt). Maximal transcription rates (135% +/- 15.8 of euthyroid values) occurred 6 h after injection and subsequently declined in parallel with serum T(3) levels. The transcription of beta-MHC was reduced to 86% of peak hypothyroid levels 6 h after a single T(3) injection and reached a nadir of 59% of hypothyroid levels at 36 h. Analysis of the time course of T(3)-mediated induction of alpha-MHC hnRNA and repression of beta-MHC hnRNA indicates that separate molecular mechanisms are involved in the coordinated regulation of these genes.

Thyroid hormone stimulates hepatic IGF-I mRNA expression in a bony fish, the tilapia Oreochromis mossambicus, in vitro and in vivo

To gain more knowledge about the physiological regulation of hepatic insulin-like growth factor-I (IGF-I) production in bony fish, we examined the potential influence of thyroid hormone (T3, 3,5,3 ′-triiodothyronine) on the expression of IGF-I in the liver of the tilapia Oreochromis mossambicus, using in vitro and in vivo methods. The in vitro experiments were performed using a recently established primary hepatocyte cell culture and IGF-I expression was determined by means of semiquantitative RT-PCR. T3 (100 nM) significantly enhanced the synthesis rate of IGF-I mRNA in short (>8 h) and long (>42 h) time courses. The stimulating effect of T3 was detected already after 1 h. After 4 h, the IGF-I mRNA expression was more than 150% of the starting amount. In long time courses, after 6 h the IGF-I mRNA value was about 170% of that in untreated cells and at the end of the experiment, it was still three times higher than in the control. In addition, the increase in IGF-I mRNA expression evoked by T3 (1 nM to 1 μM) was dose-dependent. In the in vivo approach, 10 individuals of tilapia received 4 daily intraperitoneal injections of T3 (6 μg/g body weight). IGF-I mRNA was assessed using dot blot technique with a tilapia specific IGF-I cRNA probe. The T3 treatment led to an increase of the IGF-I mRNA level up to 45% in the liver compared to the untreated animals. In conclusion, our results show that T3 directly stimulates the hepatic production of IGF-I in the tilapia in vitro and in vivo and indicate that in tilapia liver regulatory mechanisms seem to exist, as they are discussed for mammals.

The expression of mRNA of cytokines and of extracellular matrix proteins in triiodothyronine-treated rat hearts.

In various models of cardiac hypertrophy, e.g. treatment of rats with norepinephrine infusion or pressure overload, increased expression of cytokines together with increase in extracellular matrix proteins (ECMP) was reported. In this study the effect of triiodothyronine (T3) on the expression of mRNA for cytokines and ECMP was investigated. Female Sprague-Dawley rats were treated daily with T3 in a dose of 0.2 mg x kg(-1) of body weight s.c. Changes in the left (LV) and right (RV) ventricular function were measured 6, 24, 48, 72 h and 7 and 14 days after the first T3-injection using Millar ultraminiature pressure catheter transducers. RNA was isolated from LV and RV tissue, and the expression of cytokines and ECMP was measured using the ribonuclease protection assay. T3-treatment induced a significant increase in LV dP/dtmax and RV dP/dtmax, (p < 0.05) 24 h after the first injection of T3 together with an increase in heart rate (p < 0.01). The RV systolic pressure increased 48 h after the first T3 injection, whereas the LV systolic pressure remained unchanged. After 48 h the heart weight to body weight ratio was increased (p < 0.01). Hypertrophy of the RV was more prominent than that of the LV (155.9 vs. 137.7%). In all groups the expression of mRNA for interleukins (IL) IL-6, IL-1beta, IL-1alpha and tumour necrosis factor (TNF)-alpha in both ventricles did not change (p > 0.05). There was a significant increase in the mRNA for colligin 24 h after the T3 injection in both LV (p < 0.01) and RV (p < 0.05). This was followed by an increase in the mRNA for collagen I and III 72 h after the first T3-dose (p < 0.05 in RV; p < 0.01 in LV). At this point, the mRNA for tissue inhibitor of matrix metalloproteinases-2 (TIMP-2) was increased (p < 0.01) in the LV only. Moreover, after 7 days also the mRNA for matrix metalloproteinase (MMP)-2 increased (p < 0.01) in the LV. Both, TIMP-2 and MMP-2 were increased in the RV only after 14 days (p < 0.05). The gelatinase activity of MMP-2, however, was unchanged in both ventricles. The T3-induced cardiac hypertrophy was not accompanied by fibrosis as measured by the Sirius red staining after 14-days of T3-treatment. The moderate increase in mRNA for ECMP and MMP may be attributed more to the increasing mass of the ventricles with the accompanying remodelling of the ECM than to increased fibrosis.

Effects of Melatonin on Skeletal Muscle of Rats with Experimental Hyperthyroidism

The aim of this study was to investigate structural changes that occurred in the skeletal muscle of rats with experimental hyperthyroidism and the effect of melatonin on these changes. Groups of animals were designated as controls, 3,3′,5‐triiodothyronine (T3) injected and T3 + melatonin injected group. At the end of the study the tissue specimens were harvested and their structure examined. In the skeletal muscle of T3 injected rats a decrease was observed in muscle fiber diameter, splitting of fiber, collections of adipose tissue in perimysium, and gathering of nuclei in central compared to the control. Electron microscopic examination showed that mitochondria were dilated and the I band was less clear. In the T3 + melatonin injected group, the structure of fibers was similar to control. In conclusion, this study showed that T3 injection caused structural changes in the skeletal muscle and that melatonin had a positive effect on these changes.

Effects of triiodothyronine and imipramine on basal 5-HT levels and 5-HT 1 autoreceptor activity in rat cortex

Clinical studies have shown that triiodothyronine (T3) both augments and accelerates the therapeutic response to antidepressant drugs, particularly tricyclics. There is evidence that this effect is mediated by the serotonergic system. We show here that T3 administered daily for 7 days over the range 0.02–0.5 mg/kg increases basal serotonin (5-hydroxytryptamine, 5-HT) levels, as measured by in vivo microdialysis in rat cortex, in a dose-dependent fashion. All the doses of T3 examined reduced 5-HT 1A autoreceptor activity, as measured by the effect of 8-hydroxy-2-(di- n-propylamino)tetralin (8-OH-DPAT, 0.05 mg/kg s.c.) to decrease 5-HT levels in frontal cortex. T3 administered daily for 14 days at 0.02 mg/kg also reduced 5-HT 1B autoreceptor activity, as measured by the effect of locally administered 3-(1,2,5,6-tetrahydropyrid-4-yl)pyrrolo[3,2-b]pyrid-5-one (CP 93129, 10 μM) to decrease 5-HT levels. In animals administered imipramine (10 mg/kg/day by osmotic minipump) concurrently with T3 injections, no further changes in either 5-HT 1A or 5-HT 1B autoreceptor activity were seen. We suggest that the effect of T3 to accelerate the therapeutic actions of antidepressant drugs may be due to a combination of the actions of T3 at autoreceptors and the actions of the drugs at postsynaptic 5-HT 1A receptors.

Effect of ischemia-reperfusion on heart mitochondria from hyperthyroid rats.

We investigated the effect of hyperthyroidism on the functional response of mitochondria to ischemia-reperfusion and its relationship with changes in mitochondrial susceptibility to stress conditions. Hyperthyroidism was elicited by ten daily intraperitoneal injections of T3 (10 microg/100 g body weight). Mitochondria were isolated at 3000xg (M3) from homogenates of hearts perfused by the Langendorff technique after either 25 min reperfusion following 20 min ischemia or 45 min perfusion (controls). Rates of O2 consumption and H2O2 release with complex II-linked substrate, capacity to remove H2O2, extent of oxidative damage, levels of liposoluble antioxidants, such as ubiquinols and vitamin E, and susceptibility to Ca2+ -induced swelling were determined. During reperfusion, hyperthyroid hearts displayed a significant tachycardia together with a low functional recovery. In comparison to the respective controls, mitochondria from both euthyroid and hyperthyroid hearts subjected to ischemia-reperfusion protocol exhibited decreases in the rate of O2 consumption, capacity to remove H2O2, and concentration of antioxidants, and increases in the rate of H2O2 release, concentration of hydroperoxides and protein-bound carbonyls, and susceptibility to Ca2+ -induced swelling. Such changes were higher in mitochondria from hyperthyroid hearts. The increase in the protein percent content and cytochrome oxidase activity of a mitochondrial fraction isolated at 8000xg (M8) from hyperthyroid hearts after reperfusion, suggests that the decline of mitochondrial respiration of M3 fraction could be due to the degradation of the oldest, mature mitochondria endowed of high oxidative capacity, but low antioxidant capacity, which would be lost by heavy mitochondrial fraction and recovered in the light fraction. The higher susceptibility to ischemia-reperfusion of the heart from hyperthyroid animals is associated with a significant increase in mitochondrial dysfunction.

Thyroid hormone regulates the expression of NeuroD/BHF1 during the development of rat cerebellum

During the postnatal development of the rat cerebellum, there is an extensive proliferation of granular neurones in the external layer, followed by their migration and differentiation in the internal layer. These processes are impaired by neonatal hypothyroidism and can be restored by thyroid hormone therapy. They are also abolished in transgenic mice in which the neuroD gene is not expressed. This gene encodes a basic helix-loop-helix (bHLH) transcription factor (NeuroD), which induces the differentiation of neuronal precursors. We studied the expression of neuroD/BHF1-A mRNA during the postnatal development of euthyroid and hypothyroid rats, and compared it with that of neurotrophin-3 (NT-3), a marker of granular neurone differentiation. In euthyroid animals, the neuroD/BHF1-A mRNA increases 6-fold between days 4 and 15 after birth, and then decreases to 50% of this level in the adult. NT-3 mRNA expression followed a similar pattern, although it was increased only 3-fold. Hypothyroidism reduced both mRNA levels by 35–45%, depending on the postnatal stage. In hypothyroid pups, the injection of triiodothyronine (T3) restored normal levels of both mRNAs within 6 h. In 15-day old hypothyroid rats, the amount of NeuroD protein was reduced by about 35%. It increased about 2-fold 24 h after T3 injection. In conclusion, our results indicate that thyroid hormones (TH) regulate the expression of NeuroD during the ‘critical period’ of cerebellum development. This regulation may constitute an early event in the control of differentiation of the cerebellar granular neurones by TH.

Are the effects of T3 on resting metabolic rate in euthyroid rats entirely caused by T3 itself?

Because we previously reported that T3 and 3,5-diiodo-L-thyronine (3,5-T2) both increase resting metabolic rate (RMR), 3,5-T2 could be another thyroidal regulator of energy metabolism. This effect of 3,5-T2 is evident in rats made hypothyroid by propylthiouracil and iopanoic acid, not in normal euthyroid (N) rats. Possibly, under euthyroid conditions, active 3,5-T2 may need to be formed intracellularly from a precursor such as T3. We tested this hypothesis by giving a single injection of T3 to N rats and comparing the time course of the variations in RMR with those of the changes in the serum and hepatic levels of 3,5-T2. Acute injection had an evident effect on RMR, 25 h earlier, in N rats than in rats made hypothyroid by propylthiouracil and iopanoic acid, maximal values (+40%) being reached in the former at 24-26 h. In N rats, the simultaneous injection of actinomycin D with the T3 inhibited the late part of the effect (after 24 h) more strongly than the early part (14-24 h). In serum and liver, 3,5-T2 levels were increased significantly at 12-24 h after T3 injection into N rats, a time at which RMR was rising rapidly to peak. These results seem to indicate that when T3 is injected into N animals, not all the effects on RMR are attributable to T3 itself, the early effect presumably being largely because of its in vivo deiodination to 3,5-T2. Because the effects of T3 and 3,5-T2 are additive, in N rats, the two iodothyronines probably cooperate in vivo to determine the total metabolic rate.

Drug-induced and postnatal hypothyroidism impairs the accumulation of diacylglycerol in liver and liver cell plasma membranes.

Thyroid hormones are well known modulators of signal transduction. The effect of hyper- and hypo-thyroidism on diacylglycerol/protein kinase C (DAG/PKC) signaling in cardiomiocytes has been determined. Triiodothyronine (T3) has been shown to prevent the alpha1-adrenoreceptor-mediated activation of PKC but does not alter the stimulation of enzyme and hepatic metabolism by phorbol ethers. It has been suggested that the elevation of endogenous DAG in senescent or hypothyroid cells changes the PKC-dependent response of cells to phorbol esters and hormones. In the present study, was examined the formation of DAG and activation of PKC in liver cells from rats of different thyroid status. The results obtained provide the first demonstration of DAG accumulation in liver and cell plasma membranes at age- and drug-dependent thyroid gland malfunction. The experiments were performed in either the [14C]CH3COOH-labeled rat liver, liver slices or hepatocytes labeled by [14C] oleic acid and [3H]arachidonic acid or [14C]palmitic acid as well as in the isolated liver cell plasma membranes of 90- and 720-day-old rats of different thyroid status. The decrease of T4 and T3 levels in blood serum of 720-day-old rats and mercazolil-treated animals was associated with increases of both the DAG mass in liver and liver cell plasma membranes and newly synthesized [14C]DAG level in liver and isolated hepatocytes. Hypothyroidism decreased PKC activity in both membrane and cytosol as well as phospholipid and triacylglycerol synthesis in liver. These hypothyroidism effects were restored in liver by injection of T4. T4 administration to the intact animals of different ages decreased the DAG level in liver and isolated plasma membranes and the content of newly synthesized DAG in liver. The reduction of DAG level in liver was not associated with increasing free fatty acid level. DAG labeling ratio 14C/3H in liver slices of rats of different thyroid state sharply differed from PL. DAG was relatively enriched in [14C]oleic acid whereas PL were enriched in [3H]arachidonic acid. The above data have indicated that thyroid hormones are important physiological modulators of DAG level in rat liver and cell plasma membranes. Age- and drug-induced malfunction of thyroid gland resulted in a prominent decrease of glycerolipid synthesis which may promote DAG accumulation in liver.

Increases in Tumor Necrosis Factor-α in Response to Thyroid Hormone-induced Liver Oxidative Stress in the Rat

Thyroid hormone-induced calorigenesis contributes to liver oxidative stress and promotes an increased respiratory burst activity in Kupffer cells, which could conceivably increase the expression of redox-sensitive genes, including those coding for cytokines. Our aim was to test the hypothesis that l -3,3',5-triiodothyronine (T3)-induced liver oxidative stress would markedly increase the production of TNF- α by Kupffer cells and its release into the circulation. Sprague-Dawley rats received a single dose of 0.1 mg T3/kg or vehicle (controls) and determinations of liver O2 consumption, serum TNF-α, rectal temperature, and serum T3 levels, were carried out at different times after treatment. Hepatic content of total reduced glutathione (GSH) and biliary glutathione disulfide (GSSG) efflux were measured as indices of oxidative stress. In some studies, prior to T3 injection animals were administered either (i) the Kupffer cell inactivator gadolinium chloride (GdCl3), (ii) the antioxidants α-tocopherol and N-acetyl-L-cysteine (NAC), or (iii) an antisense oligonucleotide against TNF-α (ASO TJU-2755). T3 elicited an 80-fold increase in the serum levels of TNF-α at 22h after treatment, which coincided with the onset of thyroid calorigenesis. Pretreatment with GdCl3 , α-tocopherol, NAC, and ASO TJU-2755 virtually abolished this effect and markedly reduced T3-induced liver GSH depletion and the increases in biliary GSSG efflux. It is concluded that the hyperthyroid state in the rat increases the circulating levels of TNF-α by actions exerted at the Kupffer cell level and these are related to the oxidative stress status established in the liver by thyroid calorigenesis.

Prevention of auditory dysfunction in hypothyroid Tshr mutant mice by thyroxin treatment during development.

Based on previous work, it is clear that genetically hypothyroid Tshr(hyt) mutant mice are congenitally deaf [O'Malley et al. (1995) Hear. Res. 88: 181-189, Sprinkle et al. 2001b, J. Assoc. Res. Otolaryngol. DOI: 10.1007/s101620010077]. However, the extent to which auditory development is dependent on the availability of thyroxin (T4) during specific developmental stages is unknown. The aim of this study was to determine the relative importance of prenatal and postnatal thyroxin on the ontogeny of hearing in the hyt mouse. Experimental hypothyroid subjects were offspring of hyt/hyt breeders implanted with T4 or placebo controlled-release pellets 14 days prior to mating. Pups received T4 or saline placebo injections from birth through postnatal day 14 (P14) or the time of testing on P28. In the absence of exogenous T4 replacement, very high stimulus levels (>80 dB SPL) were required to elicit responses. Remarkably, T4 treatment confined to the postnatal period failed to significantly improve auditory function relative to untreated animals, while response thresholds, latencies, and amplitudes of mice born to dams that received T4 during pregnancy were significantly improved relative to both of the untreated groups. Response thresholds were improved somewhat when maternal T4 replacement was followed by treatment during the first 14 days of life, and animals treated throughout prenatal and postnatal life were comparable to those of age-matched euthyroid individuals. Findings from this study show that treatment of hyt/hyt mice with exogenous T4 significantly attenuates hypothyroid-induced otopathology in a develop-mental-stage-dependent manner. In addition, we demonstrate that postnatal development is critically dependent on prenatal exposure to thyroxin and that the critical window of T4 dependence extends throughout development.

The type 2 iodothyronine deiodinase is essential for adaptive thermogenesis in brown adipose tissue.

Type 2 iodothyronine deiodinase (D2) is a selenoenzyme, the product of the recently cloned cAMP-dependent Dio2 gene, which increases 10- to 50-fold during cold stress only in brown adipose tissue (BAT). Here we report that despite a normal plasma 3,5,3'-triiodothyronine (T3) concentration, cold-exposed mice with targeted disruption of the Dio2 gene (Dio2(-/-)) become hypothermic due to impaired BAT thermogenesis and survive by compensatory shivering with consequent acute weight loss. This occurs despite normal basal mitochondrial uncoupling protein 1 (UCP1) concentration. In Dio2(-/-) brown adipocytes, the acute norepinephrine-, CL316,243-, or forskolin-induced increases in lipolysis, UCP1 mRNA, and O(2) consumption are all reduced due to impaired cAMP generation. These hypothyroid-like abnormalities are completely reversed by a single injection of T3 14 hours earlier. Recent studies suggest that UCP1 is primarily dependent on thyroid hormone receptor beta (TR beta) while the normal sympathetic response of brown adipocytes requires TR alpha. Intracellularly generated T3 may be required to saturate the TR alpha, which has an approximately fourfold lower T3-binding affinity than does TR beta. Thus, D2 is an essential component in the thyroid-sympathetic synergism required for thermal homeostasis in small mammals.

Fetal Monkey Surfactants After Intra-amniotic or Maternal Administration of Betamethasone and Thyroid Hormone

In Brief OBJECTIVE To compare direct intra-amniotic injection of betamethasone and thyroxine (T4) with maternal treatment and controls for accelerating pulmonary surfactant production. METHODS Twelve pregnant monkeys (Macaca mulatta) on gestational day 125 (term 165 ± 10 days) had surfactant protein A and B concentrations measured in amniotic fluid. In four controls, normal saline was injected into the amniotic fluid; four others (intra-amniotic) received intra-amniotic betamethasone (1 mg) and T4 (60 μg); and in four others (maternal), the dam was given betamethasone (12 mg) intramuscularly, repeated in 24 hours, plus TRH (400 μg) intravenously, repeated every 6 hours for 24 hours. Seventy-two hours after the initial amniocentesis, a hysterotomy was performed and fetal tissue and amniotic fluid harvested for determination of surfactant protein A and B concentrations and immunohistochemical staining for surfactant protein A. RESULTS Amniotic fluid surfactant protein A was higher with intra-amniotic injection than with maternal treatment (P < .04) or controls (P = .07). Amniotic fluid surfactant protein B was higher in the intra-amniotic group than in controls (P = .06). Immunohistochemical staining for surfactant protein A in the lung tissue was increased in the intra-amniotic group compared with controls (0.145 ± 0.01 versus 0.097 ± 0.001, percent positive staining for surfactant protein A cells per lung tissue cells; P < .03). Birth weight was greater in the intra-amniotic group compared with the maternal group (P < .03) although not different from the controls. Finally, gut motility and the presence of formed meconium were increased in the intra-amniotic group compared with the other groups (P < .05). CONCLUSION Intra-amniotic injection of betamethasone and T4 enhanced lung (and possibly intestinal) maturation of the preterm rhesus fetal monkey compared with maternal injections. Intra-amniotic injections of betamethasone and thyroxine are more effective than maternal administration of betamethasone and TRH in increasing surfactants in preterm rhesus monkeys.

Fetal monkey surfactants after intra-amniotic or maternal administration of betamethasone and thyroid hormone

OBJECTIVE: To compare direct intra-amniotic injection of betamethasone and thyroxine (T4) with maternal treatment and controls for accelerating pulmonary surfactant production. METHODS: Twelve pregnant monkeys ( Macaca mulatta) on gestational day 125 (term 165 ± 10 days) had surfactant protein A and B concentrations measured in amniotic fluid. In four controls, normal saline was injected into the amniotic fluid; four others (intra-amniotic) received intra-amniotic betamethasone (1 mg) and T4 (60 μg); and in four others (maternal), the dam was given betamethasone (12 mg) intramuscularly, repeated in 24 hours, plus TRH (400 μg) intravenously, repeated every 6 hours for 24 hours. Seventy-two hours after the initial amniocentesis, a hysterotomy was performed and fetal tissue and amniotic fluid harvested for determination of surfactant protein A and B concentrations and immunohistochemical staining for surfactant protein A. RESULTS: Amniotic fluid surfactant protein A was higher with intra-amniotic injection than with maternal treatment ( P < .04) or controls ( P = .07). Amniotic fluid surfactant protein B was higher in the intra-amniotic group than in controls ( P = .06). Immunohistochemical staining for surfactant protein A in the lung tissue was increased in the intra-amniotic group compared with controls (0.145 ± 0.01 versus 0.097 ± 0.001, percent positive staining for surfactant protein A cells per lung tissue cells; P < .03). Birth weight was greater in the intra-amniotic group compared with the maternal group ( P < .03) although not different from the controls. Finally, gut motility and the presence of formed meconium were increased in the intra-amniotic group compared with the other groups ( P < .05). CONCLUSION: Intra-amniotic injection of betamethasone and T4 enhanced lung (and possibly intestinal) maturation of the preterm rhesus fetal monkey compared with maternal injections.