Molecular basis of growth hormone daily mRNA and protein synthesis in rats.

AIMS: Daily and seasonal rhythms coordinate the endocrine and metabolic functions. The pituitary gland is the master regulator of several endocrine activities, and its function is classically regulated by endocrine signals from its target glands as well as from the hypothalamus. The growth hormone (GH) produced and secreted by the anterior pituitary presents a pulsatile secretion throughout the 24-hour cycle. However, the molecular mechanisms regulating the daily pattern of GH secretion are still unclear. Herein we investigated whether circadian GH mRNA and protein synthesis is modulated by acute adjustments in the stability and expression of GH mRNA. MAIN METHODS: GH mRNA and protein content were evaluated by real-time PCR and Western blotting, respectively, in pituitary gland of rats euthanized every 3 h during a 24-h period at the Zeitgeber times (ZT3 to ZT24). The GH mRNA poly(A) tail length was determined by RACE-PAT assay. KEY FINDINGS: We identified two main peaks of GH mRNA level in the pituitary gland of rats; one in the middle of the light-cycle and another in the middle of the dark-cycle. The latter was associated with an increase in pituitary GH protein content. Interestingly, an increment in the poly(A) tail length of the GH transcript was observed in association to reduced migration rate of the GH transcript and increased mRNA content in the dark-cycle period. SIGNIFICANCE: Our findings provide evidence that changes in the GH mRNA poly(A) length may underlie the circadian pattern of GH mRNA and protein levels in the pituitary gland of rats.

Novel aspects of T3 actions on GH and TSH synthesis and secretion: physiological implications.

Thyroid hormones (THs) classically regulate the gene expression by transcriptional mechanisms. In pituitary, the encoding genes for growth hormone (GH) and thyroid-stimulating hormone (TSH) are examples of genes regulated by triiodothyronine (T3) in a positive and negative way, respectively. Recent studies have shown a rapid adjustment of GH and TSH synthesis/secretion induced by T3 posttranscriptional actions. In somatotrophs, T3 promotes an increase in Gh mRNA content, poly(A) tail length and binding to the ribosome, associated with a rearrangement of actin cytoskeleton. In thyrotrophs, T3 reduces Tshb mRNA content, poly(A) tail length and its association with the ribosome. In parallel, it promotes a redistribution of TSH secretory granules to more distal regions of the cell periphery, indicating a rapid effect of T3 inhibition of TSH secretion. T3 was shown to affect the content of tubulin and the polymerization of actin and tubulin cytoskeletons in the whole anterior pituitary gland, and to increase intracellular alpha (CGA) content. This review summarizes genomic and non-genomic/posttranscriptional actions of TH on the regulation of several steps of GH and TSH synthesis and secretion. These distinct mechanisms induced by T3 can occur simultaneously, even though non-genomic effects are promptly elicited and precede the genomic actions, coexisting in a functional network within the cells.

3,5-diiodothyronine (3,5-T2) reduces blood glucose independently of insulin sensitization in obese mice.

AIM: Thyroid hormones regulate metabolic response. While triiodothyronine (T3) is usually considered to be the active form of thyroid hormone, one form of diiodothyronine (3,5-T2) exerts T3-like effects on energy consumption and lipid metabolism. 3,5-T2 also improves glucose tolerance in rats and 3,5-T2 levels correlate with fasting glucose in humans. Presently, however, little is known about mechanisms of 3,5-T2 effects on glucose metabolism. Here, we set out to compare effects of T3, 3,5-T2 and another form of T2 (3,3-T2) in a mouse model of diet-induced obesity and determined effects of T3 and 3,5-T2 on markers of classical insulin sensitization to understand how diiodothyronines influence blood glucose. METHODS: Cell- and protein-based assays of thyroid hormone action. Assays of metabolic parameters in mice. Analysis of transcript and protein levels in different tissues by qRT-PCR and Western blot. RESULTS: T3 and 3,5-T2 both reduce body weight, adiposity and body temperature despite increased food intake. 3,3'-T2 lacks these effects. T3 and 3,5-T2 reduce blood glucose levels, whereas 3,3'-T2 worsens glucose tolerance. Neither T3 nor 3,5-T2 affects markers of insulin sensitization in skeletal muscle or white adipose tissue (WAT), but both reduce hepatic GLUT2 glucose transporter levels and glucose output. T3 and 3,5-T2 also induce expression of mitochondrial uncoupling proteins (UCPs) 3 and 1 in skeletal muscle and WAT respectively. CONCLUSIONS: 3,5-T2 influences glucose metabolism in a manner that is distinct from insulin sensitization and involves reductions in hepatic glucose output and changes in energy utilization.

Thyroid hormone reduces inflammatory cytokines improving glycaemia control in alloxan-induced diabetic wistar rats.

AIM: This study aimed at evaluating whether thyroid hormone treatment could improve glycaemia and insulin response in alloxan-induced diabetic rats by altering cytokine expression in the skeletal muscle and epididymal white adipose tissue (eWAT) as well as altering inflammatory cell infiltration in eWAT. METHODS: Diabetes mellitus (DM) was induced in male Wistar rats by alloxan injection, and a subset of the diabetic rats was treated with T3 (1.5 µg per 100 g body weight) for a 28-day period (DT3 ). Cytokines were measured in serum (MILIplex assay kit) as well as in soleus and EDL skeletal muscles and eWAT by Western blotting. Thyroid function was evaluated by morphological, molecular and biochemical parameters. Cardiac function was assessed by measuring heart rate, blood pressure, maximal rate of pressure development (dp/dtmax ) and decline (dp/dtmin ) as well as the contractility index (CI). Sixty rats were used in the study. RESULTS: Diabetic rats exhibited decreased thyroid function and increased inflammatory cytokines in serum, soleus muscle and eWAT. T3 treatment decreased glycaemia and improved insulin sensitivity in diabetic animals. These alterations were accompanied by decreased TNF-alpha and IL-6 content in soleus muscle and eWAT, and inflammatory cell infiltration in eWAT. T3 treatment did not affect cardiac function of diabetic rats. CONCLUSIONS: The present data provide evidence that T3 treatment reduces glycaemia and improves insulin sensitivity in diabetic rats, and that at least part of this effect could result from its negative modulation of inflammatory cytokine expression.

The role of glucocorticoids in the stress-induced reduction of extrathyroidal 3,5,3'-triiodothyronine generation in rats.

Serum concentrations of T4, T3, and rT3 as well as liver and kidney 5'-deiodinase activity, have been examined in rats stressed by restraint. After immobilization, serum concentrations of T3 decreased significantly (6 hr, -33 +/- 1%; 8 h, -42 +/- 3%), while serum rT3 increased (6 h, +55 +/- 3%; 8 h, +75 +/- 5%). In the same or similarly treated animals, there was a time-dependent reduction in T4 5'-deiodinase activity in both liver (4 h, -23 +/- 2%; 8 h, -43 +/- 3%) and kidney (4 h, -18 +/- 1%; 8 h, -42 +/- 3%) homogenates. The reduction in hepatic and renal T3 production was due to reduced enzyme activity and not to reduced substrate availability. In spite of reductions in serum TSH (4 h, -9 +/- 1%; 8 h, -51 +/- 5%), the serum T4 concentration did not fall. The serum concentration of corticosterone reached 30 times the basal level after 8 h of restraint. Either adrenalectomy or metyrapone treatment, followed by replacement with nonstress doses of B, completely prevented the alterations of iodothyronine metabolism induced by restraint. These results indicate that the stress-induced elevation of plasma glucocorticoids plays a key role in the pathogenesis of the low T3 syndrome in this model. The reduction in serum T3 may be accounted for by a reduction in T3 production by liver and kidney, adding support to the concept that these organs are an important source of plasma T3 in the rat.

Thyroid hormone stimulates myoglobin expression in soleus and extensorum digitalis longus muscles of rats: concomitant alterations in the activities of Krebs cycle oxidative enzymes.

Myoglobin (Mb) gene expression, Citrate Synthase (CS) and Succinate Dehydrogenase (SDH) activities of Soleus (S) and Extensorum Digitalis Longus (EDL) muscles were studied in intact, thyroidectomized and T3-treated (25 microg/100g, BW, ip, 15 days) rats. The fiber type composition of S muscle was also evaluated and used as control of the T3-induced effects. In the S muscle, the T3 treatment increased the Mb mRNA and protein expression, as well as the CS and SDH activity. These changes occurred parallel to the expected increase in type II (fast) and decrease in type I (slow)-fibers in S muscle. In the hypothyroid state, the Mb mRNA was decreased, while the Mb expression and CS activity tended to decrease. In contrast the SDH activity was increased, probably due to the enhanced motor activity that occurs as a short-term response to the hypothermia induced by hypothyroidism. In the EDL, the alterations were milder than those in S muscle in both thyroid states. These findings show that Mb gene expression is induced by T3. This is concomitant with the enhancement of Krebs Cycle enzyme activities and provides additional evidence that thyroid hormone increases the aerobic potential of skeletal muscles, as well as the speed of muscle contraction.

Role of thyroid hormone in the control of growth hormone gene expression.

Growth hormone (GH) gene expression was examined in male Wistar rats (200 g) subjected to different manipulations of thyroid status. Thyroidectomy followed by 10 days of treatment with 0.03% methimazole added to drinking water caused a marked decrease in GH mRNA levels estimated by Northern Blot analysis. T3 administration (100 micrograms/100 g body weight, ip, twice daily) to euthyroid rats for one week caused a substantial increase in GH mRNA levels. In another set of experiments, thyroidectomized methimazole-treated rats were killed at different times after a single T3 injection (100 micrograms/100 g body weight, ip). T3 induced a prompt response in GH gene expression by 15 min that reached a maximum after 1 h, remaining so up to 4 h. We conclude that in the rat, GH gene expression is highly dependent on thyroid hormones. Because of the rapidity of the response, the effect is probably mediated by a transcriptional mechanism.

Hyperthyroid-like metabolic changes induced by chronic administration of compound BW 48/80.

The present study examines the metabolic changes caused by chronic mast cell degranulation in rats. Three groups of 5 adult rats each were used: 1) rats treated with increasing doses of BW 48/80 (2 to 6 mg kg-1 day-1, ip, for 4 days); 2) a control group receiving daily saline injections, and 3) rats treated with disodium chromoglycate (DCG) (40 mg kg-1 day-1,ip, for 4 days). The third group was included as a control since DCG blocks mast cell degranulation. Saline- and DCG-treated rats showed little differences except for an increase in spleen wet weight and liver glycogen content in the latter. Chronic treatment with compound BW 48/80 resulted in statistically significant changes such as body weight loss, reduction in thymus and spleen wet weights, decrease in spleen protein content, liver glycogen and blood insulin concentrations, increase in plasma free fatty acids concentration and adrenal wet weight compared to control rats. These metabolic changes are similar to those reported for hyperthyroidism and support previous findings on the possible role of mast cells in the control of thyroid function.

Extrathyroidal conversion of thyroxine to 3,5,3'-triiodothyronine in cold-acclimated thyroxine-maintained thyroidectomized rats.

Chronic exposure of rats to cold (4 degrees C) leads to thyroid gland hyperactivity as a compensatory mechanism for activating body heat production. There is increased extrathyroidal production of T3 from T4 in parallel to thyroid hormone hypersecretion. Since the 5'-deiodination (5'-D) of T4 can be modulated by thyroid hormones, it has been suggested that the increased thyroid hormone secretion may activate the 5'-D enzymatic pathway leading to increased extrathyroidal T3 production. In an attempt to explore this possibility, T4 to T3 conversion was studied in liver and kidney homogenates of thyroidectomized rats which received T4 (0.5 to 50 micrograms/100 g body weight per day) for 10 days. Tissue homogenates were incubated with T4 (5 micrograms) for 2 h and the T3 generated was measured by RIA as an index of the activity of the 5'-D pathway. A direct relationship between T4 dose and the production of T3 by the homogenate was observed. 5'-D activity was significantly decreased in hypothyroid rats and greatly increased in hyperthyroid rats. Thyroidectomized rats treated with a replacement dose of T4 (1 microgram/100 g body weight/day) were exposed to 4 degrees C for 60 days. Despite the absence of the thyroid gland, increased 5'-D activity was observed in both liver and kidney homogenates compared to both intact and T4-treated thyroidectomized rats maintained at 25 degrees C. We conclude that chronic cold exposure of rats stimulates 5'-D activity which is independent of the concomitant thyroid gland hyperactivity.

Facilitatory role of serotonin (5-HT) in the control of thyrotropin releasing hormone/thyrotropin (TRH/TSH) secretion in rats.

In order to investigate the role of serotonin in the regulation of thyrotropin (TSH) secretion, control and propylthiouracil (PTU)-treated male Wistar rats weighing approximately 250 g were subjected to ip injections of methysergide (MET, 10 micrograms/100 g body weight), a serotonergic receptor blocker, and killed 60 min later by decapitation. Serum and pituitary concentrations of TSH were measured by radioimmunoassay. An addition, the pituitary release of TSH was estimated in an in vitro system in which pituitary glands were incubated with hypothalamic extracts. MET treatment led to a decrease in pituitary (94.12 +/- 18.55 vs 199.30 +/- 31.47 micrograms/mg, N = 20), and serum (1.95 +/- 0.92 vs 4.26 +/- 1.40 ng/ml, N = 20) TSH concentration (P < 0.001) and also to a decreased in vitro pituitary response to control hypothalamic extracts (55 +/- 8 vs 78 +/- 7%, N = 5, P < 0.005). In addition, hypothalamic extracts of MET-treated rats significantly facilitated in vitro pituitary TSH secretion, suggesting an enhanced hypothalamic thyrotropin releasing hormone (TRH) activity (347 +/- 62 vs 78 +/- 7%, N = 5, P < 0.001). These results suggest that serotonin participates in the physiological control of TRH/TSH secretion, probably by increasing TRH production/secretion, and/or by facilitating the pituitary TSH response to TRH.

GLUT4 protein expression in obese and lean 12-month-old rats: insights from different types of data analysis.

GLUT4 protein expression in white adipose tissue (WAT) and skeletal muscle (SM) was investigated in 2-month-old, 12-month-old spontaneously obese or 12-month-old calorie-restricted lean Wistar rats, by considering different parameters of analysis, such as tissue and body weight, and total protein yield of the tissue. In WAT, an approximately 70% decrease was observed in plasma membrane and microsomal GLUT4 protein, expressed as microg protein or g tissue, in both 12-month-old obese and 12-month-old lean rats compared to 2-month-old rats. However, when plasma membrane and microsomal GLUT4 tissue contents were expressed as g body weight, they were the same. In SM, GLUT4 protein content, expressed as microg protein, was similar in 2-month-old and 12-month-old obese rats, whereas it was reduced in 12-month-old obese rats, when expressed as g tissue or g body weight, which may play an important role in insulin resistance. Weight loss did not change the SM GLUT4 content. These results show that altered insulin sensitivity is accompanied by modulation of GLUT4 protein expression. However, the true role of WAT and SM GLUT4 contents in whole-body or tissue insulin sensitivity should be determined considering not only GLUT4 protein expression, but also the strong morphostructural changes in these tissues, which require different types of data analysis.

[Neonates born to drug-addicted mothers]. / Recém-nascidos filhos de mães toxicodependentes.

A test was carried out involving 94 newborn babies whose birth occurred in Alfredo da Costa Maternity between November 1991 and February 1994. The aims of this study were to find out the frequency of social and obstetric risks, to assess the neonatal, mobility, namely as regards the withdrawal syndrome, and also the repercussion of drug-addiction in the children's development. The frequency was 1 newborn baby of a drug-addicted mother for 106 pregnant women. In 67 out of 94 cases there was pre-natal follow-up. In such cases, the frequency of prematurity, of the withdrawal syndrome, and the possibility of preventive treatment of sexually contagious illnesses (Syphilis, hepatitis B, and aids) decreased. 8.5% of the 94 newborn babies were handed over to relatives or for adoption. Only in a small number of cases was it possible to keep the babies under regular observation. However, all of them showed hypertony and bulimia, which disappeared between the 8th and 9th months.

Neonatal hyper- and hypothyroidism alter the myoglobin gene expression program in adulthood.

Myoglobin acts as an oxygen store and a reactive oxygen species acceptor in muscles. We examined myoglobin mRNA in rat cardiac ventricle and skeletal muscles during the first 42 days of life and the impact of transient neonatal hypo- and hyperthyroidism on the myoglobin gene expression pattern. Cardiac ventricle and skeletal muscles of Wistar rats at 7-42 days of life were quickly removed, and myoglobin mRNA was determined by Northern blot analysis. Rats were treated with propylthiouracil (5-10 mg/100 g) and triiodothyronine (0.5-50 µg/100 g) for 5, 15, or 30 days after birth to induce hypo- and hyperthyroidism and euthanized either just after treatment or at 90 days. During postnatal (P) days 7-28, the ventricle myoglobin mRNA remained unchanged, but it gradually increased in skeletal muscle (12-fold). Triiodothyronine treatment, from days P0-P5, increased the skeletal muscle myoglobin mRNA 1.5- to 4.5-fold; a 2.5-fold increase was observed in ventricle muscle, but only when triiodothyronine treatment was extended to day P15. Conversely, hypothyroidism at P5 markedly decreased (60%) ventricular myoglobin mRNA. Moreover, transient hyperthyroidism in the neonatal period increased ventricle myoglobin mRNA (2-fold), and decreased heart rate (5%), fast muscle myoglobin mRNA (30%) and body weight (20%) in adulthood. Transient hypothyroidism in the neonatal period also permanently decreased fast muscle myoglobin mRNA (30%) and body weight (14%). These results indicated that changes in triiodothyronine supply in the neonatal period alter the myoglobin expression program in ventricle and skeletal muscle, leading to specific physiological repercussions and alterations in other parameters in adulthood.

Beta hydroxy beta methylbutyrate supplementation impairs peripheral insulin sensitivity in healthy sedentary Wistar rats.

AIM: Investigate, in healthy sedentary rats, the potential mechanisms involved on the effects of beta hydroxy beta methylbutyrate (HMB) supplementation upon the glycaemic homeostasis, by evaluating the insulin sensitivity in liver, skeletal muscle, and white adipose tissue. METHODS: Rats were supplemented with either beta hydroxy beta methylbutyrate (320 mg kg(-1)  BW) or saline by gavage for 4 weeks. After the experimental period, the animals were subjected to the glucose tolerance test (GTT) and plasma non-esterified fatty acids (NEFA) concentration measurements. The soleus skeletal muscle, liver and white adipose tissue were removed for molecular (western blotting and RT-PCR) and histological analysis. RESULTS: The beta hydroxy beta methylbutyrate supplemented rats presented: (i) higher ratio between the area under the curve (AUC) of insulinaemia and glycaemia during glucose tolerance test; (ii) impairment of insulin sensitivity on liver and soleus skeletal muscle after insulin overload; (iii) reduction of glucose transporter 4 (GLUT 4) total and plasma membrane content on soleus; (iv) increased hormone-sensitive lipase (HSL) mRNA and protein expression on white adipose tissue and plasma NEFA levels and (v) reduction of fibre cross-sectional area of soleus muscle. CONCLUSION: The data altogether indicate that beta hydroxy beta methylbutyrate supplementation impairs insulin sensitivity in healthy sedentary rats, which, in the long-term, could lead to an increased risk of developing type 2 diabetes.

Erythropoietin reduces the expression of myostatin in mdx dystrophic mice.

Erythropoietin (EPO) has been well characterized as a renal glycoprotein hormone regulating red blood cell production by inhibiting apoptosis of erythrocyte progenitors in hematopoietic tissues. EPO exerts regulatory effects in cardiac and skeletal muscles. Duchenne muscular dystrophy is a lethal degenerative disorder of skeletal and cardiac muscle. In this study, we tested the possible therapeutic beneficial effect of recombinant EPO (rhEPO) in dystrophic muscles in mdx mice. Total strength was measured using a force transducer coupled to a computer. Gene expression for myostatin, transforming growth factor-ß1 (TGF-ß1), and tumor necrosis factor-α (TNF-α) was determined by quantitative real time polymerase chain reaction. Myostatin expression was significantly decreased in quadriceps from mdx mice treated with rhEPO (rhEPO = 0.60 ± 0.11, control = 1.07 ± 0.11). On the other hand, rhEPO had no significant effect on the expression of TGF-ß1 (rhEPO = 0.95 ± 0.14, control = 1.05 ± 0.16) and TNF-α (rhEPO = 0.73 ± 0.20, control = 1.01 ± 0.09). These results may help to clarify some of the direct actions of EPO on skeletal muscle.

Triiodothyronine (T3) induces proinsulin gene expression by activating PI3K: possible roles for GSK-3ß and the transcriptional factor PDX-1.

Thyroid hormone (TH) activates PI3K and Akt, leading to glucose uptake in rat skeletal muscle cells and proliferation of insulinoma cells, respectively. However, TH actions on pancreatic beta cells have been little explored, which lead us to evaluate the TH eff ects on proinsulin gene expression, and the involvement of PI3K/Akt/GSK-3ß signaling pathway, and a transcriptional factor for insulin (PDX-1). INS-1E cells were sorted into 3 groups: control and TH-depleted treated or not with T3 for 30 min. Cells were also previously treated with actinomycin D (ActD), cycloheximide (CHX), wortmannin or Akt inhibitor. Proinsulin mRNA expression was evaluated by real time PCR, and pGSK-3ß and PDX-1 protein content was analyzed by Western blotting. TH depletion decreased proinsulin mRNA content, which was restored after acute T3 treatment. ActD, CHX and wortmannin, but not Akt inhibitor, prevented the rapid stimulatory eff ect of T3 on proinsulin mRNA expression. TH depletion did not affect the phosphorylated GSK-3ß and PDX-1 protein content; but T3 treatment led to an increase in the content of these proteins. These data indicate that T3 acutely increases proinsulin mRNA expression, by mechanisms which depends on the activation of PI3K, but not of Akt, and may involve the inactivation of GSK-3ß by phosphorylation. Since GSK-3ß enhances PDX-1 degradation rate, the GSK-3ß inactivation could explain the increase of PDX-1 content in T3-treated cells. Considering that PDX-1 is one of the most important transcriptional factors for proinsulin gene expression, its enhancement may underlie the increased proinsulin mRNA content acutely induced by T3.

Arginine induces GH gene expression by activating NOS/NO signaling in rat isolated hemi-pituitaries.

The amino acid arginine (Arg) is a recognized secretagogue of growth hormone (GH), and has been shown to induce GH gene expression. Arg is the natural precursor of nitric oxide (NO), which is known to mediate many of the effects of Arg, such as GH secretion. Arg was also shown to increase calcium influx in pituitary cells, which might contribute to its effects on GH secretion. Although the mechanisms involved in the effects of Arg on GH secretion are well established, little is known about them regarding the control of GH gene expression. We investigated whether the NO pathway and/or calcium are involved in the effects of Arg on GH gene expression in rat isolated pituitaries. To this end, pituitaries from approximately 170 male Wistar rats (~250 g) were removed, divided into two halves, pooled (three hemi-pituitaries) and incubated or not with Arg, as well as with different pharmacological agents. Arg (71 mM), the NO donor sodium nitroprusside (SNP, 1 and 0.1 mM) and a cyclic guanosine monophosphate (cGMP) analogue (8-Br-cGMP, 1 mM) increased GH mRNA expression 60 min later. The NO acceptor hemoglobin (0.3 µM) blunted the effect of SNP, and the combined treatment with Arg and L-NAME (a NO synthase (NOS) inhibitor, 55 mM) abolished the stimulatory effect of Arg on GH gene expression. The calcium channel inhibitor nifedipine (3 µM) also abolished Arg-induced GH gene expression. The present study shows that Arg directly induces GH gene expression in hemi-pituitaries isolated from rats, excluding interference from somatostatinergic neurons, which are supposed to be inhibited by Arg. Moreover, the data demonstrate that the NOS/NO signaling pathway and calcium mediate the Arg effects on GH gene expression.

Hypothyroidism decreases proinsulin gene expression and the attachment of its mRNA and eEF1A protein to the actin cytoskeleton of INS-1E cells.

The actions of thyroid hormone (TH) on pancreatic beta cells have not been thoroughly explored, with current knowledge being limited to the modulation of insulin secretion in response to glucose, and beta cell viability by regulation of pro-mitotic and pro-apoptotic factors. Therefore, the effects of TH on proinsulin gene expression are not known. This led us to measure: a) proinsulin mRNA expression, b) proinsulin transcripts and eEF1A protein binding to the actin cytoskeleton, c) actin cytoskeleton arrangement, and d) proinsulin mRNA poly(A) tail length modulation in INS-1E cells cultured in different media containing: i) normal fetal bovine serum - FBS (control); ii) normal FBS plus 1 µM or 10 nM T3, for 12 h, and iii) FBS depleted of TH for 24 h (Tx). A decrease in proinsulin mRNA content and attachment to the cytoskeleton were observed in hypothyroid (Tx) beta cells. The amount of eEF1A protein anchored to the cytoskeleton was also reduced in hypothyroidism, and it is worth mentioning that eEF1A is essential to attach transcripts to the cytoskeleton, which might modulate their stability and rate of translation. Proinsulin poly(A) tail length and cytoskeleton arrangement remained unchanged in hypothyroidism. T3 treatment of control cells for 12 h did not induce any changes in the parameters studied. The data indicate that TH is important for proinsulin mRNA expression and translation, since its total amount and attachment to the cytoskeleton are decreased in hypothyroid beta cells, providing evidence that effects of TH on carbohydrate metabolism also include the control of proinsulin gene expression.

Chronic supplementation of beta-hydroxy-beta methylbutyrate (HMß) increases the activity of the GH/IGF-I axis and induces hyperinsulinemia in rats.

OBJECTIVE: Beta-hydroxy-beta-methylbutyrate (HMß) is a metabolite of leucine widely used for improving sports performance. Although HMß is recognized to promote anabolic or anti-catabolic effects on protein metabolism, the impact of its long-term use on skeletal muscle and/or genes that control the skeletal protein balance is not fully known. This study aimed to investigate whether chronic HMß treatment affects the activity of GH/IGF-I axis and skeletal muscle IGF-I and myostatin mRNA expression. DESIGN: Rats were treated with HMß (320mg/kg BW) or vehicle, by gavage, for 4 weeks, and killed by decapitation. Blood was collected for evaluation of serum insulin, glucose and IGF-I concentrations. Samples of pituitary, liver, extensor digitorum longus (EDL) and soleus muscles were collected for total RNA or protein extraction to evaluate the expression of pituitary growth hormone (GH) gene (mRNA and protein), hepatic insulin-like growth factor I (IGF-I) mRNA, skeletal muscle IGF-I and myostatin mRNA by Northern blotting/real time-PCR, or Western blotting. RESULTS: Chronic HMß treatment increased the content of pituitary GH mRNA and GH, hepatic IGF-I mRNA and serum IGF-I concentration. No changes were detected on skeletal muscle IGF-I and myostatin mRNA expression. However, the HMß-treated rats although normoglycemic, exhibited hyperinsulinemia. CONCLUSIONS: The data presented herein extend the body of evidence on the potential role of HMß-treatment in stimulating GH/IGF-I axis activity. In spite of this effect, HMß supplementation also induces an apparent insulin resistance state which might limit the beneficial aspects of the former results, at least in rats under normal nutritional status and health conditions.

Effect of triiodothyronine on the maxilla and masseter muscles of the rat stomatognathic system.

The maxilla and masseter muscles are components of the stomatognathic system involved in chewing, which is frequently affected by physical forces such as gravity, and by dental, orthodontic and orthopedic procedures. Thyroid hormones (TH) are known to regulate the expression of genes that control bone mass and the oxidative properties of muscles; however, little is known about the effects of TH on the stomatognathic system. This study investigated this issue by evaluating: i) osteoprotegerin (OPG) and osteopontine (OPN) mRNA expression in the maxilla and ii) myoglobin (Mb) mRNA and protein expression, as well as fiber composition of the masseter. Male Wistar rats (~250 g) were divided into thyroidectomized (Tx) and sham-operated (SO) groups (N = 24/group) treated with T3 or saline (0.9%) for 15 days. Thyroidectomy increased OPG (~40%) and OPN (~75%) mRNA expression, while T3 treatment reduced OPG (~40%) and OPN (~75%) in Tx, and both (~50%) in SO rats. Masseter Mb mRNA expression and fiber type composition remained unchanged, despite the induction of hypo- and hyperthyroidism. However, Mb content was decreased in Tx rats even after T3 treatment. Since OPG and OPN are key proteins involved in the osteoclastogenesis inhibition and bone mineralization, respectively, and that Mb functions as a muscle store of O2 allowing muscles to be more resistant to fatigue, the present data indicate that TH also interfere with maxilla remodeling and the oxidative properties of the masseter, influencing the function of the stomatognathic system, which may require attention during dental, orthodontic and orthopedic procedures in patients with thyroid diseases.