Thyroid hormone profile in breast cancer patients in postmenopause

Objective: The aim of this study was to determine thyroid hormone (TH) profile in postmenopausal patients with breast cancer (BC). Subjects and methods: 12 CaM patients stages I or II, without interventions that could interfere with tumor progression were selected, as well as and a control group with 18 postmenopausal women without CaM. We measured serum anti-thyroperoxidase antibody (TPOAB), thyroid-stimulating hormone (TSH), free thyroxine (T4L), estradiol (E2), follicle-stimulating hormone (FSH), and luteinizing hormone (LH), before and after surgery, besides immunohistochemistry for estrogen (ER) and progesterone (PR) receptors. Results: Four patients with CaM showed changes in thyroid hormone profile: two had hyperthyroidism, one hypothyroidism, and one was positive for TPO-AB. All of them positive for ER and PR.TSH levels in breast cancer patients were not different from levels found in the control group (1.89 +/- 1.56 vs. 2.86 +/- 3.12 mIU/mL), but the levels of T4L in patients with CaM were statistically higher than those of the control group (1.83 +/- 0.57 vs. 1.10 +/- 0.20 ng/dL). Conclusion: These results reinforce the need for assessment of thyroid status in CaM patients, since in the absence of E2, changes in clinical HTs can act in E2-controlled processes. Arq Bras Endocrinol Metab. 2012;56(4):238-43

MiRNA-208a and miRNA-208b are triggered in thyroid hormone-induced cardiac hypertrophy - role of type 1 Angiotensin II receptor (AT1R) on miRNA-208a/α-MHC modulation

Hyperthyroidism promotes cardiac hypertrophy and the Angiotensin type 1 receptor (AT1R) has been demonstrated to mediate part of this response. Recent studies have uncovered a potentially important role for the microRNAs (miRNAs) in the control of diverse aspects of cardiac function. Then, the objective of the present study was to investigate the action promoted by hyperthyroidism on β-MHC/miR-208b expression and on α-MHC/miR-208a expression, as well as the possible contribution of the AT1R in this event. The findings of this study confirmed that AT1R is a key mediator of the cardiac hypertrophy induced by hyperthyroidism. Additionally, we demonstrated that like β-MHC, miR-208b was down-regulated in the hyperthyroid group. Similarly, like the expression of its host gene, α-MHC, miR-208a expression was up-regulated in response to hyperthyroidism. Finally, our data suggest for the first time that AT1R mediates the hyperthyroidism-induced increase on cardiac miRNA-208a/α-MHC levels, while does not influence on the reduction of miRNA-208b/β-MHC levels.

Identification of thyroid hormone response elements in the human fatty acid synthase promoter

To investigate the regulation of the human fatty acid synthase gene by the thyroid hormone triiodothyronine, various constructs of the human fatty acid synthase promoter and the luciferase reporter gene were transfected in combination with plasmids expressing the thyroid hormone and the retinoid X receptors in HepG2 cells. The reporter gene was activated 25-fold by the thyroid hormone in the presence of the thyroid hormone receptor. When both the thyroid hormone and the retinoid X receptors were expressed in HepG2 cells, there was about a 100-fold increase in reporter gene expression. 5′-Deletion analysis disclosed two thyroid hormone response elements, TRE1 (nucleotides −870 to −650) and TRE2 (nucleotides −272 to −40), in the human fatty acid synthase promoter. The presence of thyroid hormone response elements in these two regions of the promoter was confirmed by cloning various fragments of these two regions in the minimal thymidine kinase promoter−luciferase reporter gene plasmid construct and determining reporter gene expression. The results of this cloning procedure and those of electrophoretic mobility shift assays indicated that the sequence GGGTTAcgtcCGGTCA (nucleotides −716 to −731) represents TRE1 and that the sequence GGGTCC (nucleotides −117 to −112) represents TRE2. The sequence of TRE1 is very similar to the consensus sequence of the thyroid hormone response element, whereas the sequence of TRE2 contains only a half-site of the thyroid hormone response element consensus motif because it lacks the direct repeat. The sequences on either side of TRE2 seem to influence its response to the thyroid hormone and retinoid X receptors.

An unliganded thyroid hormone receptor causes severe neurological dysfunction

Congenital hypothyroidism and the thyroid hormone (T3) resistance syndrome are associated with severe central nervous system (CNS) dysfunction. Because thyroid hormones are thought to act principally by binding to their nuclear receptors (TRs), it is unexplained why TR knock-out animals are reported to have normal CNS structure and function. To investigate this discrepancy further, a T3 binding mutation was introduced into the mouse TR-β locus by homologous recombination. Because of this T3 binding defect, the mutant TR constitutively interacts with corepressor proteins and mimics the hypothyroid state, regardless of the circulating thyroid hormone concentrations. Severe abnormalities in cerebellar development and function and abnormal hippocampal gene expression and learning were found. These findings demonstrate the specific and deleterious action of unliganded TR in the brain and suggest the importance of corepressors bound to TR in the pathogenesis of hypothyroidism.

Thyroid hormone and estrogen interact to regulate behavior.

Environmental perturbations that increase plasma thyroid hormone (T3) concentrations also profoundly affect female reproductive behavior and physiology. We explored whether these effects were mediated by interactions between T3 receptor (TR) and estrogen receptor (ER). This hypothesis was of interest because the half-site of a consensus T3 response element DNA sequence is identical to an ER response element (ERE), and TRs bind to a consensus ERE. Molecular data presented in the accompanying paper [Zhu, Y.-S., Yen, P.M., Chin, W.W.& Pfaff, D.W. (1996) Proc. Natl. Acad. Sci. USA 93, 12587-12592] demonstrate that TRs and ERs are both present in rat hypothalamic nuclear extracts and that both can bind to the promoter the hypothalamic gene preproenkephalin and that interations between liganded TRs and ERs affect preproenkephalin transcription. In this paper, we show that molecular interactions between TRs and ERs are sufficient to mediate environmental effects on estrogen-controlled reproductive behavior. Ovariectomized (OVX) rats treated with high doses of T3 showed significantly lower levels of lordosis behavior in response to estradiol benzoate (EB) compared with OVX females treated with EB alone. Conversely, thyroidectomized/OVX females treated with EB showed significantly greater levels of lordosis behavior compared with OVX females treated with EB, showing the effect of endogenous T3. Thyroid hormone interference with EB-induced behavior could not be explained by a reduction in plasma E2 concentrations or by a general reduction in responsiveness of EB-sensitive tissues. Moreover, numbers of hypothalamic ER-immunoreactive cells increased dramatically following T3 treatment. These data suggest that T3 may reduce EB-dependent sexual behavior through interactions between TR and ER in the nuclei of behaviorally relevant hypothalamic neurons, envisioning for the first time a functional consequence of interactions between two nuclear hormone receptors in brain. These results also open up the possibility of molecular interactions on DNA encoding environmental signals, a new field for the study of neuronal integration.

Ligand induction of a transcriptionally active thyroid hormone receptor coactivator complex.

Transcriptional regulation by nuclear hormone receptors is thought to involve interactions with putative cofactors that may potentiate receptor function. Here we show that human thyroid hormone receptor alpha purified from HeLa cells grown in the presence of thyroid hormone (T3) is associated with a group of distinct nuclear proteins termed thyroid hormone receptor-associated proteins (TRAPs). In an in vitro system reconstituted with general initiation factors and cofactors (and in the absence of added T3), the "liganded" thyroid hormone receptor (TR)/TRAP complex markedly activates transcription from a promoter template containing T3-response elements. Moreover, whereas the retinoid X receptor is not detected in the TR/TRAP complex, its presence is required for the function of the complex. In contrast, human thyroid hormone receptor alpha purified from cells grown in the absence of T3 lacks the TRAPs and effects only a low level of activation that is dependent on added ligand. These findings demonstrate the ligand-dependent in vivo formation of a transcriptionally active TR-multisubunit protein complex and suggest a role for TRAPs as positive coactivators for gene-specific transcriptional activation.

Simian virus 40 late gene expression is regulated by members of the steroid/thyroid hormone receptor superfamily.

Transcription of the late genes of simian virus 40 (SV40) is repressed during the early phase of the lytic cycle of infection of binding of cellular factors, called IBP-s, to the SV40 late promoter; repression is relieved after the onset of viral DNA replication by titration of these repressors. Preliminary data indicated that one of the major components of IBP-s was human estrogen-related receptor 1 (hERR1). We show here that several members of the steroid/thyroid hormone receptor superfamily, including testis receptor 2, thyroid receptor alpha 1 in combination with retinoid X receptor alpha, chicken ovalbumin upstream promoter transcription factors 1 and 2 (COUP-TF1 and COUP-TF2), as well as hERR1, possess the properties of IBP-s. These receptors bind specifically to hormone receptor binding sites present in the SV40 major late promoter. Recombinant COUP-TF1 specifically represses transcription from the SV40 major late promoter in a cell-free transcription system. Expression of COUP-TF1, COUP-TF2, or hERR1 in monkey cells results in repression of the SV40 late promoter, but not the early promoter, in the absence of the virally encoded large tumor antigen. Overexpression of COUP-TF1 leads to a delay in the early-to-late switch in SV40 gene expression during the lytic cycle of infection. Thus, members of this superfamily can play major direct roles in regulating expression of SV40. Possibly, natural or synthetic ligands to these receptors can serve as antiviral drugs. Our findings also provide the basis for the development of assays to screen for the ligands to testis receptor 2 and hERR1.

Skeletal muscle and nuclear hormone receptors: Implications for cardiovascular and metabolic disease

Skeletal muscle is a major mass peripheral tissue that accounts for similar to 40% of the total body mass and a major player in energy balance. It accounts for > 30% of energy expenditure, is the primary tissue of insulin stimulated glucose uptake, disposal, and storage. Furthermore, it influences metabolism via modulation of circulating and stored lipid (and cholesterol) flux. Lipid catabolism supplies up to 70% of the energy requirements for resting muscle. However, initial aerobic exercise utilizes stored muscle glycogen but as exercise continues, glucose and stored muscle triglycerides become important energy substrates. Endurance exercise increasingly depends on fatty acid oxidation (and lipid mobilization from other tissues). This underscores the importance of lipid and glucose utilization as an energy source in muscle. Consequently skeletal muscle has a significant role in insulin sensitivity, the blood lipid profile, and obesity. Moreover, caloric excess, obesity and physical inactivity lead to skeletal muscle insulin resistance, a risk factor for the development of type II diabetes. In this context skeletal muscle is an important therapeutic target in the battle against cardiovascular disease, the worlds most serious public health threat. Major risk factors for cardiovascular disease include dyslipidemia, hypertension, obesity, sedentary lifestyle, and diabetes. These risk factors are directly influenced by diet, metabolism and physical activity. Metabolism is largely regulated by nuclear hormone receptors which function as hormone regulated transcription factors that bind DNA and mediate the pathophysiological regulation of gene expression. Metabolism and activity, which directly influence cardiovascular disease risk factors, are primarily driven by skeletal muscle. Recently, many nuclear receptors expressed in skeletal muscle have been shown to improve glucose tolerance, insulin resistance, and dyslipidernia. Skeletal muscle and nuclear receptors are rapidly emerging as critical targets in the battle against cardiovascular disease risk factors. Understanding the function of nuclear receptors in skeletal muscle has enormous pharmacological utility for the treatment of cardiovascular disease. This review focuses on the molecular regulation of metabolism by nuclear receptors in skeletal muscle in the context of dyslipidemia and cardiovascular disease. (c) 2005 Published by Elsevier Ltd.

Two Novel Mutations In The Thyroid Hormone Receptor β In Patients With Resistance To Thyroid Hormone (rth β): Clinical, Biochemical, And Molecular Data.

The syndrome of resistance to thyroid hormone (RTH β) is an inherited disorder characterized by variable tissue hyposensitivity to 3,5,30-l-triiodothyronine (T3), with persistent elevation of free-circulating T3 (FT3) and free thyroxine (FT4) levels in association with nonsuppressed serum thyrotropin (TSH). Clinical presentation is variable and the molecular analysis of THRB gene provides a short cut diagnosis. Here, we describe 2 cases in which RTH β was suspected on the basis of laboratory findings. The diagnosis was confirmed by direct THRB sequencing that revealed 2 novel mutations: the heterozygous p.Ala317Ser in subject 1 and the heterozygous p.Arg438Pro in subject 2. Both mutations were shown to be deleterious by SIFT, PolyPhen, and Align GV-GD predictive methods.

Development and validation of a fast RP-HPLC method to determine the analogue of the thyroid hormone, 3,5,3 '-triiodothyroacetic acid (TRIAC), in polymeric nanoparticles

The objective of this work was to develop and validate a rapid Reversed-Phase High-Performance Liquid Chromatography method for the quantification of 3,5,3 '-triiodothyroacetic acid (TRIAC) in nanoparticles delivery system prepared in different polymeric matrices. Special attention was given to developing a reliable reproductive technique for the pretreatment of the samples. Chromatographic runs were performed on an Agilent 1200 Series HPLC with a RP Phenomenex (R) Gemini C18 (150 x 4, 6 mm i.d., 5 mu m) column using acetonitrile and triethylamine buffer 0.1% (TEA) (40 : 60 v/v) as a mobile phase in an isocratic elution, pH 5.6 at a flow rate of 1 ml min(-1). TRIAC was detected at a wavelength of 220 nm. The injection volume was 20 mu l and the column temperature was maintained at 35 degrees C. The validation characteristics included accuracy, precision, specificity, linearity, recovery, and robustness. The standard curve was found to have a linear relationship (r(2) - 0.9996) over the analytical range of 5-100 mu g ml(-1) . The detection and quantitation limits were 1.3 and 3.8 mu g ml(-1), respectively. The recovery and loaded TRIAC in colloidal system delivery was nearly 100% and 98%, respectively. The method was successfully applied in polycaprolactone, polyhydroxybutyrate, and polymethylmethacrylate nanoparticles.

A TR beta-selective agonist confers resistance to diet-induced obesity

Thyroid hormone receptor beta (TR beta also listed as THRB oil the MGI Database)-selective agonists activate brown adipose tissue (BAT) thermogenesis, while only minimally affecting cardiac activity or lean body mass. Here, we tested the hypothesis that daily administration of the TR beta agonist GC-24 prevents the metabolic alterations associated with a hypercaloric diet. Rats were placed on a high-fat diet and after a month exhibited increased body weight (BW) and adiposity, fasting hyperglycemia and glucose intolerance, increased plasma levels of triglycerides, cholesterol, nonesterified Fatty acids and interleukin-6. While GC-24 administration to these animals did not affect food ingestion or modified the progression of BW gain, it did increase energy, g the increase in adiposity Without expenditure, eliminating causing cardiac hypertrophy Fasting hyperglycemia remained unchanged, but treatment with GC-24 improved glucose I tolerance by increasing insulin Sensitivity and also normalized plasma triglyceride levels. plasma cholesterol levels were only Partially normalized and liver cholesterol content remained high in the GC-24-treated animals. Gene expression in liver, skeletal muscle, and white adipose tissue was only minimally affected by treatment with GC-24, with the main target being BAT In conclusion, during high-fat feeding treatment with the TR beta-selective agonist, GC-24 only partially improves metabolic control probably as a result Of accelerating the resting metabolic rate. Journal of Endocrinology (2009) 203, 291-299

Synthesis of an analog of the thyroid hormone-binding protein transthyretin via regioselective chemical ligation

Transthyretin is an essential protein responsible for the transport of thyroid hormones and retinol in human serum and is also implicated in the amyloid diseases familial amyloidotic polyneuropathy and senile systemic amyloidosis. Its folding properties and stabilization by ligands are of current interest due to their importance in understanding and combating these diseases, Here we report the solid phase synthesis of the monomeric unit of a transthyretin analog (equivalent to 127 amino acids) using t-Boc chemistry and peptide ligation and its folding to form a functional 54-kDa tetramer, The monomeric unit of the protein was chemically synthesized in three parts (positions 1-51, 54-99, and 102-127) and ligated using a chemoselective thioether ligation chemistry. The synthetic protein was folded and assembled to a tetrameric structure in the presence of transthyretin's native ligand, thyroxine, as shown by gel filtration chromatography, native gel electrophoresis, transthyretin antibody recognition, and thyroid hormone binding. Other folding products included a high molecular weight aggregate as well as a transient dimeric species. This represents one of the largest macromolecules chemically synthesized to date and demonstrates the potential of protein chemical synthesis for investigations of protein-ligand interactions.

Regulation of the mouse Nas1 promoter by vitamin D and thyroid hormone

The renal sodium-sulfate cotransporter, NaSi-1, a protein implicated to control serum sulfate levels, has been shown to be regulated in vivo by 1,25-dihydroxyvitamin D-3 (1,25-(OH)(2)D-3) and tri-iodothyronine (T-3). Recently, we cloned the mouse NaSi-1 gene (Nas1) and in the present study identified a 1,25-(OH)(2)D-3- and T-3-responsive element located within the Nas1 promoter. Mutational analysis of the Nas1 promoter resulted in identification of a direct repeat 6-type vitamin-D-responsive element (DR6 VDRE) at -525 to -508 and an imperfect inverted repeat 0-type T-3-responsive element (IR0 T3RE) at -436 to -425 which conferred 1,25(OH)(2)D-3 and T3 responsiveness, respectively. In summary, we have identified responsive elements that mediate the enhanced transcription of Nas1 by 1,25-(OH)(2)D-3 and T-3, and these mechanisms may provide important clues to the physiological control of sulfate homeostasis.

Thyroid hormone modulation of early neocortical network development

Magdeburg, Univ., Fak. für Naturwiss., Diss., 2012