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Thyroid Function and Disorders

Writer and Curator: Larry H. Bernstein, MD, FCAP 

Normal thyroid function is maintained by endocrine interactions between the hypothalamus, anterior pituitary and thyroid gland [Matfin, 2009]. Iodide is transported across the basement membrane of the thyroid cells by an intrinsic membrane protein called the Na/I symporter (NIS). At the apical border, a second iodide transport protein called pendrin moves iodide into the colloid, where it is involved in hormono-genesis. Once inside the follicle, most of the iodide is oxidized by the enzyme thyroid peroxidase (TPO) in a reaction that facilitates combination with a tyrosine molecule to ultimately form thyroxine (T4) and triiodothyronine (T3). Thyroxine is the major thyroid hormone secreted into the circulation (90%, with T3 composing the other 10%). There is evidence that T3 is the active form of the hormone and that T4 is converted into T3 before it can act physiologically.

All of the major organs in the body are affected by altered levels of thyroid hormone. These actions are mainly mediated by T3. In the cell, T3 binds to a nuclear receptor, resulting in transcription of specific thyroid hormone response genes.

Maternal thyroid hormones are essential for neural development in zebrafish.

Marco A Campinho, João Saraiva, Claudia Florindo, Deborah M Power Molecular endocrinology (Baltimore, Md.) 05/2014;
http://dx.doi.org:/10.1210/me.2014-1032

ABSTRACT Teleost eggs contain an abundant store of maternal thyroid hormones (THs) and early in zebrafish embryonic development all the genes necessary for TH signalling are expressed. Nonetheless the function of THs in embryonic development remains elusive. To test the hypothesis that THs are fundamental for zebrafish embryonic development an MCT8 knockdown strategy was deployed to prevent maternal TH uptake. Absence of maternal THs did not affect early specification of the neural epithelia but profoundly modified later dorsal specification of the brain and spinal cord as well as specific neuron differentiation. Maternal THs acted upstream of pax2a, pax7 and pax8 genes but downstream of shha and fgf8a signalling. The lack of inhibitory spinal cord interneurons and increased motorneurons in the MCT8 morphants is consistent with their stiff axial body and impaired mobility. MCT8 mutations are associated with X-linked mental retardation in humans and the cellular and molecular consequences of MCT8 knockdown during embryonic development in zebrafish provides new insight into the potential role of THs in this condition.
Relationship between thyroid status and renal function in a general population of unselected outpatients

Giuseppe Lippi, Martina Montagnana, Giovanni Targher, Gian Luca Salvagno, Gian Cesare Guidi
Clin Biochem May 2008; 41(7–8): 625-627

When compared with euthyroid subjects, those with TSH < 0.2 mIU/L and > 2.5 mIU/L had increased and decreased estimated glomerular filtration rate (e-GFR), respectively. TSH levels were an independent predictor of e-GFR.

Serum Thyroid-Stimulating Hormone Measurement for Assessment of Thyroid Function and Disease

Douglas S. Ross
Endocr and Metab Clinics of N Am, Jun 2001; 30(2, 1): 245-264

Thyrotropin, or thyroid-stimulating hormone (TSH), is one of a family of glycoprotein hormones including luteinizing hormone (LH), follicle-stimulating hormone (FSH), and human chorionic gonadotropin (hCG) that share a common α-subunit and a unique β-subunit. Pituitary TSH regulates the secretion of the thyroid hormones T4 (thyroxine) and T3 (triiodothyronine). TSH secretion, in turn, is controlled through negative feedback by thyroid hormone on the pituitary thyrotrope. This relationship is negative log-linear. Small changes in serum free thyroid hormone concentrations result in large changes in serum TSH concentrations, and even subtle changes in thyroid hormone production are best assessed by measurement of serum TSH . Until the late 1980s, the detection limit of TSH assays was within the normal range, and these first-generation TSH assays were useful only for the detection of hypothyroidism. Free T4 measurements were primarily used for assessing thyroid function despite the technical difficulties in free thyroid hormone measurements owing to abnormal binding proteins, changes in binding protein concentrations, and the effects of drugs and illness on thyroid hormone binding. With the use of sensitive second- and third-generation TSH assays, TSH measurement has emerged as the single most useful test of thyroid function. It is widely and appropriately used as a screening test. Unfortunately, the trend has been to rely on TSH measurements alone for the assessment of complicated thyroid disease and patients undergoing treatment for thyroid dysfunction. This article focuses on the potential and real limitations of TSH measurement.
Correlation of creatinine with TSH levels in overt hypothyroidism — A requirement for monitoring of renal function in hypothyroid patients?

Vandana Saini, Amita Yadav, Megha Kataria Arora, Sarika Arora, Ritu Singh, Jayashree Bhattacharjee
Clin Biochem  Feb 2012; 45(3): 212-214

Highlights
► Increase serum creatinine levels in both subclinical and overt hypothyroidism. ► Creatinine levels progressively increase with increasing degree of hypothyroidism. ► Increase in creatinine correlated with TSH levels in overt hypothyroid subjects. ► Regular monitoring of renal function is required in hypothyroid patients.

Renal function is influenced by thyroid status. Therefore, this study was done to determine the relationship between renal function and different degrees of thyroid dysfunction.
Design and methods
Thyroid and kidney function tests were analyzed in 47 patients with overt (TSH ≥ 10.0 μIU/L) and 77 patients with subclinical hypothyroidism (TSH 6.0–9.9 μIU/L) in a cross-sectional study. These were compared with 120 age- and sex-matched euthyroid controls.
Results
Overt hypothyroid subjects showed significantly raised serum urea, creatinine and uric acid levels as compared to controls whereas subclinical hypothyroid patients showed significant increased levels of serum urea and creatinine levels. TSH showed significant positive correlation with creatinine and uric acid values and, fT4 had a negative correlation with uric acid in overt hypothyroidism.
Conclusion
Hypothyroid state is associated with significant derangement in biochemical parameters of renal function. Hence the renal function should be regularly monitored in hypothyroid patients.

1. Ability of Serum Thyroid-Stimulating Hormone Levels to Reflect Peripheral and Central Thyroid Hormone Action Appropriately

• Uncertainty Owing to Heterogeneity of T4 Deiodinases
• Uncertainty Owing to Heterogeneity of T3 Receptors
• Uncertainty Owing to Resetting of the Threshold for Negative Feedback

2. Clinical Utility of Thyroid-Stimulating Hormone Measurement
3. Screening for Thyroid Disease and Assessment of Patients Suspected of Having Thyroid Disease

• Limitations of Thyroid-Stimulating Hormone Testing in Patients with Known Thyroid Disease Central Hypothyroidism
• Thyrotoxicosis Owing to Inappropriate Thyroid-Stimulating Hormone Secretion
• Monitoring Thyroid Hormone Therapy
• Patients Treated for Hyperthyroidism

4. The Pituitary-Thyroid Axis in Nonthyroidal Illness

• Measurement of Thyroid-Stimulating Hormone
• Drugs that Affect Serum Thyroid-Stimulating Hormone Concentrations

Investigations into the etiology of elevated serum T3 levels in protein-malnourished rats

Robert C. Smallridge, Allan R. Glass, Leonard Wartofsky, Keith R. Latham, Kenneth D. Burman
Metabolism, V June 1982; 31(6): 538-542

Thyroid function studies and the peripheral metabolism of thyroid hormone were examined in rats fed a low protein diet (9% casein) for 4–8 wk. Compared to animals fed a normal protein diet ad libitum, both the low protein rats and a pair-fed control group weighed less at the end of the study. However, serum total T3 levels were significantly higher only in the protein deficient rats. The elevated serum T3 was not explainable by enhanced peripheral T4 to T3 conversion, as there was no evidence of any change in hepatic or renal 5′-deiodinase activity when homogenates were examined for conversion of T4 to T3, reverse T3 to 3,3′-diiodothyronine, or 3′,5′-diiodothyronine to 3′-monoiodothyronine. Neither was there an effect on hepatic T3 receptor maximal binding capacity (204 ± 24 versus 168 ± 15 fmol/mg DNA control) or binding affinity (2.07 ± 0.38 versus 2.49 ± 0.24 × 10−10 M control). In two separate experiments the dialyzable fraction of T3 was significantly lower in the low protein group while free T3 concentrations were unchanged or reduced. In contrast, serum total and free T4 were either normal or reduced and dialyzable T4 was unaffected by protein deficiency. We conclude that while serum total T3 is elevated in rats chronically fed a low protein diet, this elevation is not due to enhanced T4 to T3 conversion. Rather, the increased T3 levels can be accounted for by a striking alteration in protein binding to T3. Moreover, the failure to demonstrate similar changes in serum total and dialyzable T4 suggests that in the rat, protein deficiency has different effects on binding to the two major thyroid hormones. Dietary induced changes in serum thyroid hormone binding must be kept in mind in nutrition studies in the rat.

Role of thyrotropin in metabolism of thyroid hormones in nonthyroidal tissues

Udaya M. Kabadi
Metabolism, Jun 2006; 55(6): 748-750

T4 conversion into T3 in peripheral tissues is the major source of circulating T3. However, the exact mechanism of this process is ill defined. Several in vitro studies have demonstrated that thyrotropin facilitates deiodination of T4 into T3 in liver and kidneys. However, there is a paucity of in vitro studies confirming this activity of thyrotropin. Therefore, this study was conducted to examine the influence of thyrotropin on thyroid hormone metabolism in nonthyroidal tissues. We assessed T4, T3, reverse T3 (rT3), and T3 resin uptake (T3RU) responses up to 12 hours at intervals of 4 hours in 6 thyroidectomized female mongrel dogs rendered euthyroid with LT4 replacement therapy before and after subcutaneous (SC) administration of bovine thyrotropin (5 U) on one day and normal saline (0.5 mL) on another in a randomized sequence between 08:00 and 09:00 am. Euthyroid state after LT4 replacement was confirmed before thyrotropin administration. Serum T4, T3, rT3, and T3RU all remained unaltered after SC administration of normal saline. No significant alteration was noted in serum T3RU values on SC administration of thyrotropin. However, serum T3 rose progressively reaching a peak at 12 hours with simultaneous declines being noted in both serum T4 and rT3 concentrations (P < .05 vs prethyrotropin values for all determinations). The changes after SC administration were significantly different (P < .001) in comparison to those noted on SC administration of normal saline. Thyrotropin may promote both the conversion of T4 to T3 and metabolism of rT3 into T2 in nonthyroidal tissues via enhancement of the same monodeionase.

Effects of growth hormone administration on fuel oxidation and thyroid function in normal man

Jens Møller, Jens O.L. Jørgensen, Niels Møller, Jens S. Christiansen, Jørgen Weeke
Metabolism, Jul 1992;  41(7): 728-731

In a randomized, double-blind, placebo-controlled, cross-over study, we examined the effects of 14 days of growth hormone (GH) administration (12 IU/d subcutaneously) on energy expenditure (EE), respiratory exchange ratio (RER), and thyroid function in 14 normal adults of normal weight (eight men and six women). EE (kcal/24 h) was significantly elevated after GH administration (2,073 ± 392, [GH], 1,900 ± 310, [placebo], P = .01). RER was significantly lowered during GH administration (0.73 ± 0.04 v 0.78 ± 0.06, P = .02), reflecting increased oxidation of lipids. Total triiodothyronine (TT3) (nmol/L) and free T3 (FT3) (pmol/L) increased significantly during GH (TT3: 1.73 ± 0.06 [GH], 1.48 ± 0.08 [placebo], P = .01; FT3: 6.19 ± 0.56 [GH], 5.49 ± 0.56 [placebo], P = .01). Concomitantly, an insignificant decrease in reverse T3 (rT3) (nmol/L) was observed (0.07 ± 0.01 [GH], 0.15 ± 0.01 [placebo], P = .08). GH caused a highly significant increase in T3/thyroxine (T4 (×100) ratio (1.84 ± 0.12 [GH], 1.37 ± 0.06 [placebo]). Serum thyrotropin (TSH) was not significantly changed by GH. No changes in total thyroxine (TT4) (nmol/L) (98 ± 6 [GH], 111 ± 8 [placebo], P = .40) and free thyroxine (FT4) (pmol/L) (17.4 ± 1.3 [GH], 18.6 ± 1.1 [placebo], P = .37) after 14 days of GH administration were observed. In conclusion, 2 weeks of GH administration increases EE and lipidoxidation. This finding may partly be mediated by an increase in peripheral T4 to T3 conversion.

Studies on the deiodination of thyroid hormones in Xenopus laevis tadpoles

Helen Robinson, Valerie Anne Galton
Gen Compar Endocr, Sept 1976; 30(1): 83-90

Liver and tail tissues from Xenopus laevis tadpoles possess deiodinating systems capable of degrading both thyroxine (T4) and 3,5,3′-triiodothyronine (T3). Deiodinating activity in liver remains at a constant level throughout late development and metamorphosis with the exception of a transient increase at stage 59, the onset of metamorphosis. Tail activity remains constant during development but rises sharply during metamorphosis when the tail is undergoing regression. In contrast to these findings on spontaneously metamorphosing tadpoles, tail tips induced to regress in vitro do not exhibit any rise in deiodinating activity, even when the tail tips are undergoing extensive autolysis. These results indicate that, while a rise in deiodinating activity may coincide temporarily with hormone action during metamorphosis, the two phenomena may be separated. The deiodinating activity present in tadpole tissues appears to be enzymic and possesses properties characteristic of peroxidase activity. The reaction catalyzed by this mechanism does not appear to involve monodeiodination and hence cannot be considered a mechanism for the peripheral conversion of T4 to T3.

Mechanisms governing the relative proportions of thyroxine and 3,5,3′-triiodothyronine in thyroid secretion

Peter Laurberg
Metabolism, Apr 1984; 33(4): 379-392

In subjects with normal thyroid function only a minor part of circulating 3,5,3′-triiodothyronine (T3) originates directly from the thyroid; the majority is produced in the peripheral tissues by deiodination of thyroxine (T4). However, T3 of thyroidal origin constitutes a relatively high fraction of the total T3 produced in many patients with thyroid hyperfunction or hypofunction. Such a relatively high T3 content in the secretion of the thyroid could be caused by a low T4T3 ratio in thyroglobulin. Severe iodine deficiency is a well-known inducer of a low T4T3 ratio, but a low T4T3 ratio can also be produced independent of the iodine content. This is seen in in vitro studies of thyroglobulin iodination when small amounts of DIT are added to the incubation mixture and in vivo in TSH-treated animals and in patients with Graves’ disease. Another mechanism for high thyroidal secretion of T3 could be an enhanced fractional deiodination of T4 to T3 in the thyroid. In vitro thyroid perfusion studies have shown that the T3 content of thyroid secretions is higher than would be expected from the T4T3 ratio of thyroid hydrolysate and that the major mechanism is deiodination of T4 to T3. Thyroxine deiodinases are also present in the human thyroid, and the amount of T4 deiodinase is enhanced in the thyroids from patients with medically treated Graves’ disease and in the hyperstimulated thyroids of rats. Other factors of possible importance for the mixture of T3 and T4 secreted by the thyroid are a relatively faster liberation of T3 than of T4 from thyroglobulin during partial hydrolysis (this faster release of T3 is probably the mechanism behind the more “rapid” secretion of T3 than of T4), or some kind of thyroid heterogeneity leading to pinocytosis and hydrolysis of thyroglobulin with a lower T4T3 ratio than that of average thyroglobulin.

Starvation-induced alterations of circulating thyroid hormone concentrations in man

Thomas J. Merimee, E.S. Fineberg
Metabolism Jan 1976; 25(1): 79-83

Serum concentrations of triiodothyronine (T3), thyroxine (T4), and TSH were examined in seven men and seven women of normal weight during a 60-hr fast. Similar studies were conducted in two women who received daily for 1 mo before and during a similar fast, 0.4 mg and 0.5 mg of l-thyroxine.
The serum concentrations of T3 decreased in each of the untreated normal subjects (sign test of significance, p < 0.001). The mean control concentration of T3 in women was 152 ± 9 ng100 ml (X ± SEM); after 24 hr of fasting, 131 ± 31 ng100 ml; and at the termination of the fast, 90 ± 15 ng100 ml. The latter value differed from the control value with a p value of < 0.01. Similar changes of T3 concentration occurred in men (mean basal T = 160 ± 11 ng100 ml; mean at termination of fast = 87 ± 16 ng100 ml). The range of decrease for T3 in all subjects varied from 24% to 55%.
The mean T4 concentration at the beginning of the fast was  6.9 ± 0.9, and at the termination of the fast, 7.5 ± 0.6 (p = NS). TSH concentrations remained unchanged (Control, 3.8 ± 0.45 μU/ml; at 60 hr, 4.0 ± 0.26 μU/ml, p = NS).
Studies in two women who received, before and during a fast, T4, indicate that a decreased peripheral conversion of T4 to T3 is the most likely mechanism responsible for this change.

Effect of estrogens on thyroid function. II. Alterations in plasma thyroid hormone levels and their metabolism

Ramesh C. Sawhney, Indra Rastogi, Gopal K. Rastogi
Metabolism Mar 1978; 27(3): 279-288

The circulating levels of total triiodothyronine (TT3), thyroxine (TT4, and T4-bbinding globulin (TBG) and the kinetics of T3 and T4 were studied in five menstruating rhesus monkeys before, during, and after prolonged treatment with estradiol monobenzoate (E2B, 50 μg/kg body weight/day subcutaneously). A significant increase over pretreatment (p < 0.01) plasma TT3, TT4, and TBG was recorded on day 6 of E2B therapy. A further significant stepwise increase in these parameters was noted up to day 19 of E2B, when the levels plateaued for the rest of the period of E2B treatment. Two weeks after discontinuation of E2B, plasma TT3, TT4, and TBG had returned to the pretreatment range and remained so up to 40 days of observation. Although the percent free T3 and percent free T4 were significantly decreased (p < 0.01) during E2B therapy, the absolute concentrations of free T3 and free T4 were not altered. After prolonged E2B treatment the metabolic clearance rate, distribution space, and production rate (PR) of both T3 and T4 were decreased (p < 0.01). The extrathyroidal T4 pool (ETT4P) was significantly increased (p < 0.01), whereas ETT3P did not show any significant alterations (p > 0.05). The decreased PR of T4 might have been due to a direct inhibitory effect of E2B on the thyroid, whereas the decrease in PR of T3 might have been due to either decreased conversion of T4 to T3, to decreased secretion by the thyroid, or both.
Zebrafish as a model to study peripheral thyroid hormone metabolism in vertebrate development

Marjolein Heijlen, Anne M. Houbrechts, Veerle M. Darras
Gen Compar Endocr 1 Jul 2013; 188: 289-296

To unravel the role of thyroid hormones (THs) in vertebrate development it is important to have suitable animal models to study the mechanisms regulating TH availability and activity. Zebrafish (Danio rerio), with its rapidly and externally developing transparent embryo has been a widely used model in developmental biology for some time. To date many of the components of the zebrafish thyroid axis have been identified, including the TH transporters MCT8, MCT10 and OATP1C1, the deiodinases D1, D2 and D3, and the receptors TRα and TRβ. Their structure and function closely resemble those of higher vertebrates. Interestingly, due to a whole genome duplication in the early evolution of ray-finned fishes, zebrafish possess two genes for D3 (dio3 and dio3a) and for TRα (thraa and thrab). Transcripts of all identified genes are present during embryonic development and several of them show dynamic spatio-temporal distribution patterns. Transient morpholino-knockdown of D2, D3 or MCT8 expression clearly disturbs embryonic development, confirming the importance of each of these regulators during early life stages. The recently available tools for targeted stable gene knockout will further increase the value of zebrafish to study the role of peripheral TH metabolism in pre- and post-hatch/post-natal vertebrate development.

The consequences of inappropriate treatment because of failure to recognize the syndrome of pituitary and peripheral tissue resistance to thyroid hormone

Samuel Refetoff, Angel Salazar, Terry J. Smith, Neal H. Scherberg
Metabolism  Aug 1983; 32(8); 822-834

Since the description of the syndrome of global (peripheral tissues and pituitary) resistance to thyroid hormone, new cases are being recognized with increasing frequency. The patient described herein had a markedly elevated serum TSH concentration of 260 μU/mL at the time of diagnosis. Studies suggest that elevations of serum TSH levels in this and other patients with the syndrome are most likely iatrogenic in origin. The patient was 312 years old when a goiter and a high serum T4 concentration were detected. Despite subtotal thyroidectomy, antithyroid drugs were required to maintain her T4 level in the normal range. She was referred at age 1112 years because of recurrent goiter. Her parents and five older siblings had normal thyroid function. Off therapy, her serum T4 level was 14.9 μg/dL, FT4I was 17.0, T3 was 362 ng/dL, TSH was 260 μU/mL, and antibodies were negative. There were no signs of thyrotoxicosis, her bone age was 7 years, her growth was stunted (third percentile), her intellectual quotient (IQ) was 67, and there was a 30–50 dB sensorineural hearing loss. The presence of a pituitary adenoma was ruled out. Her TSH had normal bioreactivity and rose to 540 μU/mL in response to TRH. Triiodothyronine was given in incremental doses of 50, 100, 200, and 400 μg/d over 28 days. The log concentrations of serum TSH showed an inverse linear correlation with serum T3. While receiving the highest dose of T3, on which the level of serum T3 ranged from 1400 to 2500 ng/dL, the TSH response to TRH normalized (basal 4.2 and peak 20 μU/mL), as did the high levels of serum cholesterol, carotene, and T4. Her BMR rose from +5 to +22%, her IQ rose to 77, and she gained weight without an increase in caloric intake. Only minimal changes were observed in levels of urinary cAMP, hydroxyproline, magnesium, and nitrogen. All values, with the exception of the weight gain, returned to baseline 2 months after T3 treatment was discontinued. The TSH level was suppressed by l-dopa and by prednisone. Long-term therapy with equivalent doses of T4 (from 300 to 1000 μg/d) produced a growth of 3 cm during the initial 6 weeks, 10.5 cm over the ensuring year (above the 10th percentile), and regression of goiter without thyrotoxicosis. The patient exhibited resistance to thyroid hormone in pituitary and peripheral tissues. The optimal dose of T4 replacement could be predicted by studying tissue responses to incremental doses of T3. The marked elevation in serum TSH concentration, stunted growth, and laboratory evidence of hypothyroidism were due to the limited thyroidal reserve caused by thyroidectomy. All patients with an impaired ability to compensate for the defect as a result of inappropriate treatment should be given thyroid hormone in amounts short of producing catabolic effects. Such a dose is expected to normalize the basal serum TSH concentration and its response to TRH.

Solving the mystery of iodine uptake

Valda Vinson
Science 20 Jun 2014; 344(6190), p. 1355
http://dx.doi.org:/10.1126/science.344.6190.1355-a

The thyroid gland produces iodine-containing hormones that regulate metabolism. The cell membrane protein NIS (sodium/iodine symporter) transports iodine into thyroid cells, but because iodine concentrations outside of the cell are so low, how it does so is a mystery. The key? Moving two sodium ions along with the iodine ion, Nicola et al found. NIS also does not bind sodium very tightly, but the high concentrations of sodium outside the cell allow one sodium ion to bind. This binding increases the affinity of NIS for a second sodium ion and also for iodine. With the three ions bound, NIS changes its conformation so that it opens to the inside of the cell, where the sodium concentration is low enough for NIS to release its sodium ions. When the sodium goes away, so does NIS’s affinity for iodine, leading NIS to release it.

Unliganded Thyroid Hormone Receptor α Regulates Developmental Timing via Gene Repression in Xenopus tropicalis

Jinyoung Choi, Ken-ichi T. Suzuki, Tetsushi Sakuma, Leena Shewade, Takashi Yamamoto, and Daniel R. Buchholz
Endocr Feb 2015; 156(2): 735–744 http://dx.doi.org:/10.1210/en.2014-1554

Thyroid hormone (TH) receptor (TR) expression begins early in development in all vertebrates when circulating TH levels are absent or minimal, yet few developmental roles for unliganded TRs have been established. Unliganded TRs are expected to repress TH-response genes, increase tissue responsivity to TH, and regulate the timing of developmental events. Here we examined the role of unliganded TRα in gene repression and development in Xenopus tropicalis. We used transcription activator-like effector nuclease gene disruption technology to generate founder animals with mutations in the TRα gene and bred them to produce F1 offspring with a normal phenotype and a mutant phenotype, characterized by precocious hind limb development. Offspring with a normal phenotype had zero or one disrupted TRα alleles , and tadpoles with the mutant hind limb phenotype had two truncated TRα alleles with frame shift mutations between the two zinc fingers followed by 40–50 mutant amino acids and then an out-of-frame stop codon. We examined TH-response gene expression and early larval development with and without exogenous TH in F1 offspring. As hypothesized, mutant phenotype tadpoles had increased expression of TH-response genes in the absence of TH and impaired induction of these same genes after exogenous TH treatment, compared with normal phenotype animals. Also, mutant hind limb phenotype animals had reduced hind limb and gill responsivity to exogenous TH. Similar results in methimazole-treated tadpoles showed that increased TH-response gene expression and precocious development were not due to early production of TH. These results indicate that unliganded TRα delays developmental progression by repressing TH-response genes.
The discovery of thyroid replacement therapy. Part 2: The critical 19th century
Conceptualizing the link between the thyroid and myxoedema

Stefan Slater
R Soc Med 2011; 104: 59–63. http://dx.doi.org:/10.1258/jrsm.2010.10k051

Sir William Withey Gull (1816–1890)

Frederik Ruysch, anatomist in Leyden around 1690, adopted, according to Albrecht von Haller in 1766, the opinion that a peculiar fluid was elaborated in the gland and poured into the veins’. The 19th century thus began with thyroidology at best in embryo; but during that century endocrinology was born and the thyroid was its standard bearer. In 1836, Thomas Wilkinson King of Guys Hospital, regarded by some as the ‘Father of Endocrinology’, anticipated on the basis of observation and experiment the internal secretion of the thyroid. In a meticulous paper on its anatomy: he wrote of the thyroid gland that ‘its absorbent vessels carry its peculiar secretion to the great veins of the body’. This language is almost identical to that of Ruysch and Haller more than a century earlier. The idea was prompted by the thyroid’s disproportionately large vascular supply in the absence of any evident mechanical or other local function and also at what he described as its ‘peculiar’ fluid. King notes that his view ‘has been indirectly surmised by Morgagni [probably in 1761] and others’.
In 1850, at a meeting of the Royal Medical and Chirurgical Society of London, chaired by Thomas Addison, Thomas Blizzard Curling, surgeon at the London Hospital, provided a clear clinicopathological correlate in a paper entitled ‘Two cases of absence of the thyroid body and symmetrical swellings of fat tissue at the sides of the neck, connected with defective cerebral development’.  Postmortem examination in each revealed no trace of thyroid tissue and that the swellings consisted only of fat.  Curling’s important observation was not pursued until 1871 when, at another meeting of the Society, Curling himself then in the chair, Charles Hilton Fagge, a physician at Guy’s Hospital, presented a paper on sporadic cretinism. He described four living cases and noted that none of them had a goiter and that one had been well up to the age of eight and, although now physically cretinous at age 16, she remained very intelligent. He referred to Curling’s paper and reached the same conclusion that the ‘healthy thyroid body is capable of exerting a counteracting influence [on cretinism]’.
Two years later, in 1873, Fagge’s senior colleague at Guy’s, Sir William Withey Gull, presented before the Clinical Society of London two of the five cases he had seen of what he called ‘A Cretinoid State supervening in Adult Life in Women’. He described their cretin-like appearance, drawing particular attention to the broad and thick tongue and the guttural voice and its pronunciation ‘as if the tongue were too large for the mouth’. He acknowledged his remarks were tentative, hence, he said, his use of the word ‘cretinoid’, but he had no doubt this was a ‘substantive’ condition and not one of cardiac or renal origin.
Gull was an interesting personality with apparently a remarkable presence, resembling Napoleon in face, form and manner (Figure). In the 1970s, 80 years after his death in 1890, he was the subject of a theory, quickly discredited, that he had been ‘Jack the Ripper’, the killer in the still unsolved murders and mutilations of at least five Whitechapel prostitutes in 1888. He figured in the 1988 TV film series, Jack the Ripper, starring Michael Caine as the detective. Gull is credited with the first description of hypothyroidism in adults and his paper was important in defining a recognizable clinical syndrome.
Then, in 1877, William Miller Ord, read his paper before the Royal Medical and Chirurgical Society of London and proposed the term ‘myxoedema’ for the adult condition. He described the non-pitting, ‘mucous edema’.   He also presented an engaging theory to explain the lethargy, inertia and slow responses associated with the disease. He suggested that these might result from the sheathing and insulation of the body in a ‘jelly-like’, mucin-laden integument that interfered with sensory perceptions and stimulation. Six years later, he chaired the committee set up by the Clinical Society of London to investigate the whole matter. He also later undertook some of the earliest metabolic studies of the effects of treating myxoedema with thyroid extract, showing the rapid weight loss and rise in temperature and in urinary volume and nitrogen excretion that occurred.
The key papers, which advanced these English authors observations, were those of the Swiss surgeons, Jaques-Louis Reverdin in Geneva and Emil Theodor Kocher in Bern, Kocher later receiving the Nobel Prize for his work on the thyroid. How fitting it is that it should be two Swiss doctors whose practices unlocked an understanding of the importance of the thyroid. For they each identified the late effects of total ablation (extirpation) of goiters. they

noted the great similarity of Gull’s and Ord’s myxoedema cases with their affected postoperative patients, referring to the comparison as a ‘rapprochement complet’, clearly making the connection. They acknowledged Gull’s primacy in describing the clinical manifestations and Ord’s ‘christening’ the condition ‘myxoedema’, and proposed that surgical cases be known as ‘myxoedème opératoire’. In light of his findings in 1882, Reverdin thereafter sought to conserve a part of the gland during thyroidectomy for goiter, speculating that its complete removal may have been responsible for these late effects. He had noticed that no such problems followed a just unilateral lobectomy. Kocher called the disease picture in his affected cases ‘cachexia strumipriva’ – literally, a bad condition due to the removal of a struma (goiter) without reference to the earlier work of Reverdin. Halsted noted in his monumental review of goiter surgery: ‘It is interesting to follow the argumentation of a mind so exceptionally keen and sane as Kocher’s in its futile efforts to explain insufficiently illuminated phenomena’. In reading Kocher’s 1909 Nobel Prize Lecture (in English translation), one gets the impression that Kocher was aware in 1883 of Gull’s and Ord’s reports, despite not referring to them, and he dismisses Reverdin’s contribution.
There ensued a competition over the contribution to the thyroid discovery.  When post-thyroidectomy myxedema wsas brought to the attention of Kocher, he agreed it was analogous to his cases of cachexia strumipriva. It is also obvious that Kocher, like many surgeons of the time, cannot have engaged in routine postoperative outpatient follow-up, for otherwise the ensuing problems in his goiter-operated patients would have been detected years earlier. In respect of this key moment in the history of the thyroid, Reverdin could be said to hold the intellectual property. The thought has been expressed that perhaps he should have shared the 1909 Nobel Prize with Kocher.
The Emerging Roles of Thyroglobulin

Yuqian Luo, Yuko Ishido, Naoki Hiroi, Norihisa Ishii, and Koichi Suzuki
Adv in Endocr 2014, Article ID 189194, 7 pp http://dx.doi.org/10.1155/2014/189194

Thyroglobulin (Tg), the most important and abundant protein in thyroid follicles, is well known for its essential role in thyroid hormone synthesis. In addition to its conventional role as the precursor of thyroid hormones, we have uncovered a novel function of Tg as an endogenous regulator of follicular function over the past decade. The newly discovered negative feedback effect of Tg on follicular function observed in the rat and human thyroid provides an alternative explanation for the observation of follicle heterogeneity. Given the essential role of the regulatory effects of Tg, we consider that dysregulation of normal Tg function is associated with multiple human thyroid diseases including autoimmune thyroid disease and thyroid cancer. Additionally, extrathyroid Tg may serve a regulatory function in other organs. Further exploration of Tg action, especially at the molecular level, is needed to obtain a better understanding of both the physiological and pathological roles of Tg.

The Surgical Management of Thyroid Cancer

Sara A. Morrison, Hyunsuk Suh, and Richard A. Hodin
Rambam Maimonides Med J 2014; 5(2):e0008. http://dx.doi.org:/10.5041/RMMJ.10142

There are approximately 63,000 reported cases of thyroid carcinoma annually in the United States, representing roughly 4% of all documented malignancies.1 Diagnosis typically stems from work-up of a thyroid nodule. Data from the Framingham study suggests that palpable thyroid nodules are present in 4% of the US population,2 but non-palpable nodules may exist in up to 67% of the population. Such nodules are often found incidentally secondary to the rising use of imaging modalities in medical settings. The large majority of thyroid nodules are benign, with an overall reported risk of malignancy from 5% to 15%.
Thyroid cancer has been increasing in incidence, with the number of reported cases in the US rising by 25% over the last 3 years. With growing technological advances in the field and improved contributions of diagnostics, surgical decision-making and operative planning have taken on new challenges. Herein, we review the current clinical practice recommendations and active areas of surgical controversy, reflective of the most recently published professional consensus guidelines and a systematic review of the literature.
The use of FNA in current clinical practice has resulted in post-surgical pathology findings of malignancy in over 50% of specimens.7 The Bethesda System for Reporting Thyroid Cytopathology (TBSRTC) was developed in order to allow pathologists among varying institutions to communicate results to clinical care-takers with widely under-stood descriptors. Results of FNA biopsies are broken down into the following categories with the corresponding risks of malignancy: non-diagnostic or unsatisfactory (1%–4%), benign (0%–3%), atypia of undetermined significance or follicular lesion of undetermined significance (AUS/FLUS; 5%–15%), follicular neoplasm or suspicious for a follicular neoplasm (FN/sFN; 15%–30%), suspicious for malignancy (60%–75%), and malignant (97%–99%).
Mutational Panels.
AsuragenmiR Inform (Austin, TX, USA) mutation analysis assay and Thyroid Cancer Mutation Panel by Quest Diagnostics (Madison, NJ, USA) are the two main commercially available mutational tests which test for known genetic alterations such as BRAF, RAS, RET/PTC, and PAX8/PPAR. These mutational panels are highly specific for malignancy; however, due to the low overall frequency of these mutations in thyroid cancers, negative results do not rule out cancer. Therefore, mutational panel tests are considered a “rule-in” test. If a preoperative mutational test is positive, the nodule should be considered malignant, and total thyroidectomy should be recommended.
Gene Expression Profiling.
The most widely known gene expression profiling test is Afirma Gene Expression Classifier (Veracyte, San Francisco, CA, USA), and, with its recent clinical validation by Alexander et al., Afirma is already being utilized in many clinical settings. The Afirma Gene Expression Classifier (GEC) is an RNA-based assay that utilizes FNA samples to evaluate 167 molecular genes associated with benign nodules based on their proprietary algorithm. Unlike the mutational panel testing, Afirma testing is considered a “rule-out” test since the test has a high negative predictive value in distinguishing benign nodules. However, a positive result reported as “suspicious” carries only 38% risk of malignancy.
In all, these molecular tests should be utilized judiciously and should be considered as a complementary diagnostic tool in the management of thyroid nodules. In the future, molecular testing could become more cost-effective and accurate as a diagnostic tool while providing prognostic and therapeutic information.
Papillary Thyroid Cancer.
Total thyroidectomy is the gold standard for patients with a preoperative diagnosis of papillary thyroid cancer when the nodule is greater than 1 cm in size. Completion thyroidectomy is indicated in patients who have undergone prior lobectomy and are found on final pathology to have papillary thyroid cancer that is larger than 1 cm. The completion thyroidectomy should generally be performed within 6 months of the original procedure in order to minimize the risk of lymph node metastasis.
Involvement of cervical lymph nodes in papillary thyroid cancer is frequent, reported to occur in up to 50% of patients. The role of neck dissection at the time of total thyroidectomy is somewhat controversial, however, since most of the nodal involvement is microscopic and does not affect overall survival. It is generally agreed upon that a therapeutic neck dissection should be pursued in the setting of well-differentiated thyroid cancer patients with clinically positive lymph nodes, whether in the central or lateral neck compartments. Prophylactic neck dissection is not done for follicular thyroid cancer, as the rates of lymph node metastasis are typically less than 10%.
Medullary thyroid cancer (MTC) comprises 4% of all thyroid malignancies. The majority of cases are sporadic in nature; approximately 20%–25% represent familiar/hereditary syndromes. Diagnosis is commonly made by FNA biopsy with specific staining for the presence of calcitonin in the tissue specimen. All patients with a diagnosis of medullary thyroid cancer must be evaluated for multiple endocrine neoplasia (MEN) 2 and be ruled out for the synchronous presence of pheochromocytoma prior to scheduling thyroid surgery.
Effects of Dose Level of Anti-thyroid Drug Carbimazole on Thermoregulation and Blood Constituents in Male Rabbits (Oryctolagus cuniculus)

Intisar H. Saeed, Abdalla M. Abdelatif and Mohamed E. Elnageeb
Adv in Research 2014; 2(3): 129-144. Article no. AIR.2014.002

Carbimazole (CBZ) is an anti-thyroid drug commonly used in the treatment of hyperthyroidism. The objective of this study was to evaluate the effects of dose level of CBZ on thermoregulation and blood constituents in mature male rabbits. Twenty animals were assigned to 4 groups (A, B, C, D) of 5 each. Group A served as control and treated animals in groups B,C,D, received daily orally CBZ doses of 10, 15 and 20 mg/animal for 3 weeks, respectively.
The values of rectal temperature (Tr,), respiration rate (RR) and heart rate (HR) decreased in treated rabbits and the mean values of HR decreased with increase in the dose level of CBZ. The packed cell volume (PCV),  Hb concentration and total leukocyte count (TLC) were lower in CBZ treated rabbits. Serum levels of total protein and globulins increased and serum albumin level decreased in treated groups of rabbits. Serum urea level was lower in CBZ treated groups and there was an increase in serum urea level with increase in CBZ dose level. Serum cholesterol level was higher in treated groups and there was an increase in serum cholesterol level with increase in CBZ dose level. Plasma glucose level decreased significantly in CBZ treated groups compared with the control and the mean values decreased with increase in the dose level of CBZ. The results indicate that the responses of basic physiological parameters were almost dose dependent in the range adopted in this study.
Phosphatase Inhibitor Calyculin A Activates TRPC2 Channels in Thyroid FRTL-5 Cells

Pramod Sukumaran, MY Asghar, C Löf, T Viitanen, and Kid Törnquist
Calcium Signaling Jun 2014; 1(2)  http://www.researchpub.org/journal/cs/cs.html

We have previously shown that rat thyroid FRTL-5 cells express a calcium entry pathway regulated by a phosphatase. The nature of the calcium entry pathway is presently unknown. We have also shown that FRTL-5 cells express only the TRPC2 channel of the TRPC family of cation channels. In the present investigation we show, using pharmacological inhibitors, the measurement of sodium and calcium entry, stable TRPC2 knock-down cells, and transfection with a non-conducting form of TRPC2, that the calcium entry pathway regulated by a phosphatase is, in fact, the TRPC2 channel. Our data thus point to a novel mechanism by which the TRPC2 channels can be regulated.

Thyroxine Uptake by Perfused Rat Liver
No Evidence for Facilitation by Five Different Thyroxine-binding Proteins

Carl M. Mendel and Richard A. Weisiger
J. Clin. Invest.  1990; 86: 1840-1847

For each of the five protein-hormone complexes studied, the rate of hepatic uptake of T4 (measured under conditions expected to result in dissociation-limited uptake) closely approximated the rate of spontaneous dissociation of the protein-hormone complex within the hepatic sinusoids. These findings indicate an absence of special cellular mechanisms that facilitate the hepatic uptake of T4 from its plasma binding proteins, and support the view that uptake occurs from the free T4 pool after spontaneous dissociation of T4 from its binding proteins.
Thyroxine Transport and Distribution in Nagase Analbuminemic Rats

Carl M. Mendel, RR Cavalieri, LA Gavin, T Pettersson, and M Inoue
J. Clin. Invest. 1989; 83: 143-148

The postulate that thyroxine (T4) in plasma enters tissues by protein-mediated transport or enhanced dissociation from plasma-binding proteins leads to the conclusion that almost all T4 uptake by tissues in the rat occurs via the pool of albumin bound T4 (Pardridge, W. M., B. N. Premachandra, and G. Fierer. 1985. Am. J. Physiol. 248:G545-G550).
To directly test this postulate, and to test more generally whether albumin might play a special role in T4 transport in the rat, we performed in vivo kinetics studies in six Nagase analbuminemic rats and in six control rats, all of whom had similar serum T4 concentrations and percent free T4 values.
Evaluation of the plasma disappearance curves of simultaneously injected ¹²⁵I-T4 and ^I31I-albumin indicated that the flux of T4 from the extracellular compartment into the rapidly exchangeable intracellular compartment was similar in the analbuminemic rats (51±21 ng/min, mean±SD) and in the control rats (54±15 ng/min), as was the size of the rapidly exchangeable intracellular pool of T4 (1.13±0.53 vs. 1.22±036 Mg). This latter finding was confirmed by direct analysis of tissue samples (liver, kidney, and brain). We also performed in vitro kinetics studies using the isolated perfused rat liver. The single-pass fractional extraction by normal rat liver of T4 in pooled analbuminemic rat serum was indistinguishable from that of T4 in pooled control rat serum (10.9±3.3%, n = 3, vs. 11.4±3.4%). When > 98% of the albumin was removed from normal rat serum by chromatography with Affi-Gel blue, the single-pass fractional extraction of T4 (measured by a bolus injection method) did not change (16.3±2.1%, n = 5, vs. 15.2±2.5%). These data provide the first valid experimental test of the enhanced

dissociation hypothesis and indicate that there is no special, substantive role for albumin in T4 transport in the rat.
Influence of thyroid receptors on breast cancer cell proliferation

10. Conde, R. Paniagua, J. Zamora, M. J. Blanquez, B. Fraile, A. Ruiz & M. I. Arenas
    Ann Oncol 2005; http://dx.doi.org:/10.1093/annonc/mdj040

Background: The involvement of thyroid hormones in the development and differentiation of normal breast tissue has been established. However, the association between breast cancer and these hormones is controversial. Therefore, the objective of the present study was to determine the protein expression pattern of thyroid hormone receptors in different human breast pathologies and to evaluate their possible relationship with cellular proliferation.
Patients and methods: The presence of thyroid hormone receptors was evaluated by immunohistochemistry and western blot analysis in 84 breast samples that included 12 cases of benign proliferative diseases, 20 carcinomas in situ and 52 infiltrative carcinomas.
Results: TR-α was detected in the nuclei of epithelial cells from normal breast ducts and acini, while in any pathological type this receptor was located in the cytoplasm. However, TR-b presented a nuclear location in benign proliferative diseases and carcinomas in situ and a cytoplasmatic location in normal breast and infiltrative carcinomas. The highest proliferation index was observed in carcinomas in situ, although in infiltrative carcinomas an inverse correlation between this index and the TR-α expression was encountered.
Conclusions: The results of this study reveal substantial changes in the expression profile of thyroid hormone.
Zebrafish as a model for monocarboxyl transporter 8-deficiency

GD Vatine, D Zada, T Lerer-Goldshtein, A Tovin, G Malkinson, K Yaniv and L Appelbaum
J Biol Chem Nov 2012; Manuscript M112.413831
http://dx.doi.org:/10.1074/jbc.M112.413831

Background: Mutations in the thyroid hormone transporter MCT8 are associated with psychomotor retardation AHDS.
Results: In zebrafish, as in humans, mct8 is expressed primarily in the nervous system. Elimination of MCT8 causes severe neural impairment.
Conclusion: MCT8 is a crucial regulator during zebrafish embryonic development. Significance: Establishment of the first vertebrate model for MCT8-deficiency, which exhibits a neurological phenotype.
Unusual Ratio between Free Thyroxine and Free Triiodothyronine in a Long-Lived Mole-Rat Species with Bimodal Ageing

Yoshiyuki Henning, Christiane Vole, Sabine Begall, Martin Bens, et al.
PlusOne Nov 2014; 9(11),e113698. http://dx.doi.org:/10.1371/journal.pone.0113698

Ansell’s mole-rats (Fukomys anselli) are subterranean, long-lived rodents, which live in eusocial families, where the maximum lifespan of breeders is twice as long as that of non-breeders. Their metabolic rate is significantly lower than expected based on allometry, and their retinae show a high density of S-cone opsins. Both features may indicate naturally low thyroid hormone levels.
In the present study, we sequenced several major components of the thyroid hormone pathways and analyzed free and total thyroxine and triiodothyronine in serum samples of breeding and non-breeding F. anselli to examine whether
a) their thyroid hormone system shows any peculiarities on the genetic level,
b) these animals have lower hormone levels compared to euthyroid rodents (rats and guinea pigs), and
c) reproductive status, lifespan and free hormone levels are correlated.
Genetic analyses confirmed that Ansell’s mole-rats have a conserved thyroid hormone system as known from other mammalian species. Interspecific comparisons revealed that free thyroxine levels of F. anselli were about ten times lower than of guinea pigs and rats, whereas the free triiodothyronine levels, the main biologically active form, did not differ significantly amongst species. The resulting fT4:fT3 ratio is unusual for a mammal and potentially represents a case of natural hypothyroxinemia.
Comparisons with total thyroxine levels suggest that mole-rats seem to possess two distinct mechanisms that work hand in hand to downregulate fT4 levels reliably. We could not find any correlation between free hormone levels and reproductive status, gender or weight. Free thyroxine may slightly increase with age, based on subsignificant evidence. Hence, thyroid hormones do not seem to explain the different ageing rates of breeders and nonbreeders. Further research is required to investigate the regulatory mechanisms responsible for the unusual proportion of free thyroxine and free triiodothyronine.
Transthyretin Regulates Thyroid Hormone Levels in the Choroid Plexus, But Not in  the Brain Parenchyma: Study in a Transthyretin-Null Mouse Model

JA Palha, R Fernandes, GM De Escobar, V Episkopou, M Gottesman, and MJ Saraiva
Endocr 2000; 141(9): 3267–3272.

Transthyretin (TTR) is the major T4-binding protein in rodents. Using a TTR-null mouse model we asked the following questions.
1) Do other T4 binding moieties replace TTR in the cerebrospinal fluid (CSF)?
2) Are the low whole brain total T4 levels found in this mouse model associated with hypothyroidism, e.g. increased 59-deiodinase type 2 (D2) activity and RC3-neurogranin messenger RNA levels?
3) Which brain regions account for the decreased total whole brain T4 levels?
4) Are there changes in T3 levels in the brain?
Our results show the following.
1) No other T4-binding protein replaces TTR in the CSF of the TTR-null mice.
2) D2 activity is normal in the cortex, cerebellum, and hippocampus, and total brain RC3-neurogranin messenger RNA levels are not altered.
3) T4 levels measured in the cortex, cerebellum, and hippocampus are normal. However T4 and T3 levels in the choroid plexus are only 14% and 48% of the normal values, respectively.
4) T3 levels are normal in the brain parenchyma.
The data presented here suggest that TTR influences thyroid hormone levels in the choroid plexus, but not in the brain. Interference with the blood-choroid-plexus-CSF-TTR-mediated route of T4 entry into the brain caused by the absence of TTR does not produce measurable features of hypothyroidism. It thus appears that TTR is not required for T4 entry or for maintenance of the euthyroid state in the mouse brain.
Identification of monocarboxylate transporter 8 as a specific thyroid hormone transporter

E.C.H. Friesema, S Ganguly, A. Abdalla, J.E.M. Fox, AP. Halestrap, and TJ. Visser
J Biol Chem 2003; Manuscript M300909200
http://dx.doi.org/10.1074/jbc.M300909200

Transport of thyroid hormone across the cell membrane is required for its action and

metabolism. Recently, a T-type amino acid transporter was cloned which transports aromatic amino acids but not iodothyronines. This transporter belongs to the monocarboxylate transporter (MCT) family, and is most homologous with MCT8 (SLC16A2). Therefore, we cloned rat MCT8, and tested it for thyroid hormone transport in Xenopus laevis oocytes. Oocytes were injected with rat MCT8 cRNA, and after 3 days immunofluorescence microscopy demonstrated expression of the protein at the plasma membrane. MCT8 cRNA induced a ~10-fold increase in uptake of 10 nM ¹²⁵I-labeled thyroxine (T4), 3,3′,5-triiodothyronine (T3), 3,3′,5′-triiodothyronine (rT3) and 3,3′-diiodothyronine. Due to the rapid uptake of the ligands, transport was only linear with time for <4 min. MCT8 did not transport Leu, Phe, Trp or Tyr. [¹²⁵I]T4 transport was strongly inhibited by L-T4, D-T4, L-T3, D-T3, 3,3’,5-triiodothyroacetic acid, N-bromoacetyl-T3, and bromosulfophthalein. T3 transport was less affected by these inhibitors. Iodothyronine uptake in uninjected oocytes was reduced by albumin but the stimulation induced by MCT8 was markedly increased. Saturation analysis provided apparent Km values of 2-5 μM for T4, T3 and rT3. Immunohistochemistry showed high expression in liver, kidney, brain and heart. In conclusion, we have identified MCT8 as a very active and specific thyroid hormone transporter.
Thyroid hormones,T3 andT4, in the brain
Amy C. Schroeder and Martin L. Privalsky
Front Endocr Mar 2014; 5 article 40.  http://dx.doi.org:/10.3389/fendo.2014.00040

Thyroid hormones (THs) are essential for fetal and post-natal nervous system development and also play an important role in the maintenance of adult brain function. Of the two major THs, T4 (3,5,30,50-tetraiodo-l-thyronine) is classically viewed as an pro-hormone that must be converted toT3 (3,5,30-tri-iodo-l-thyronine) via tissue-level deiodinases for biological activity. THs primarily mediate their effects by binding to thyroid hormone receptor (TR) isoforms, predominantly TRα1 and TRβ1, which are expressed in different tissues and exhibit distinctive roles in endocrinology. Notably, the ability to respond toT4 and toT3 differs for the two TR isoforms, with TRα1 generally more responsive to T4 than TRβ1. TRα1 is also the most abundantly expressed TR isoform in the brain, encompassing 70–80% of all TR expression in this tissue. Conversion of T4 into T3 via deiodinase 2 in astrocytes has been classically viewed as critical for generating local T3 for neurons. However, deiodinase-deficient mice do not exhibit obvious defectives in brain development or function. Considering that TRα1 is well-established as the predominant isoform in brain, and that TRα1 responds to both T3 and T4, we suggest T4 may play a more active role in brain physiology than has been previously accepted.
Thyroid hormone action: astrocyte–neuron communication

Beatriz Morte and Juan Bernal
Front Endocr May 2014; 5, Article 82 http://dx.doi.org:/10.3389/fendo.2014.00082

Thyroid hormone (TH) action is exerted mainly through regulation of gene expression by binding of T3 to the nuclear receptors.T4 plays an important role as a source of intracellular T3 in the central nervous system via the action of the type 2 deiodinase (D2), expressed in the astrocytes. A model of T3 availability to neural cells has been proposed and validated. The model contemplates that brain T3 has a double origin: a fraction is available directly from the circulation, and another is produced locally from T4 in the astrocytes by D2. The fetal brain depends almost entirely on theT3 generated locally. The contribution of systemic T3 increases subsequently during development to account for approximately 50% of total brain T3 in the late postnatal and adult stages. In this article, we review the experimental data in support of this model, and how the factors affectingT3 availability in the brain, such as deiodinases and transporters, play a decisive role in modulating local TH action during development.
The Significance of Thyroid Hormone Transporters in the Brain

Juan Bernal
Endocr Apr 2005; 146(4):1698–1700. http://dx.doi.org:/10.1210/en.2005-0134

The MCT family comprises up to 14 members, some of which are involved in the transport of important substrates for the brain such as lactate and pyruvate. MCT8 has been shown to act as a specific transporter for T4 and T3 and displays slightly higher affinity for T3. Heuer et al. have also studied the regional expression of MCT8 mRNA. In addition to high expression levels in the choroid plexus, they found that MCT8 is expressed in neurons of the neocortex, hippocampus, basal ganglia, amygdala, hypothalamus, and the Purkinje cells of the cerebellum, all regions known to be sensitive to thyroid hormones. Expression of MCT8 in neurons suggests that neuronal uptake of the T3 produced in astrocytes is facilitated by this transporter.
The physiological significance ofMCT8 as a transporter for thyroid hormone is supported by the finding of mutations in humans by Dumitrescu et al. and Friesema et al.  The syndrome affects children from an early age and consists of severe developmental delay and neurological damage together with an unusually altered pattern of thyroid hormone levels in blood. The patients presented low total and free T4, high total and free T3, and low rT3. TSH was moderately elevated in two of the patients and normal or slightly elevated in the other five. Inactivating mutations of the MCT8 transporter could result in the altered thyroid hormone levels. In vitro uptake of T4 and T3 by fibroblasts isolated from affected males was strongly reduced, and intracellular D2 was increased 6- to 8-fold. It is thus hypothesized that the resulting increase in intracellularly generated T3 accumulates in blood because of its poor reuptake into cells.
The second trimester is also the period when thyroid hormone receptors increase in concentration in the brain. If MCT8 is needed at this stage of development for T3 entry into neurons, mutations of the transporter could interfere with T3-dependent developmental processes. Knowledge of the ontogenetic patterns of MCT8 in the human fetal brain would certainly be helpful. On the other hand, there is also the possibility that MCT8 mutations interfere with transport of other substrates for brain metabolism that could be even more important than T3 in determining the severity and outcome of the syndrome. Other members of the family transport metabolic substrates such as pyruvate and lactate, but MCT8 so far appears to be specific for iodothyronines

Peripheral markers of thyroid function: The effect of T4 monotherapy versus T4/T3 combination therapy in hypothyroid subjects: A randomized cross-over study

Ulla Schmidt, B Nygaard, EW Jensen, J Kvetny, A Jarløv, and Jens Faber
Endocrine Connections Jan 10, 2013 http://dx.doi.org:/10.1530/EC-12-0

Background: A recent randomized controlled trial suggests that hypothyroid subjects may find L-T4 and L-T3 combination therapy to be

superior to L-T4 monotherapy in terms of quality of life, suggesting that the brain registered increased T3 availability during the

combination therapy.

Hypothesis: Peripheral tissue might also be stimulated during T4/T3 combination therapy compared to T4 monotherapy.
Methods: Serum levels of Sex Hormone-Binding Globulin (SHBG), pro-collagen-1-N-terminal peptide (PINP), and N-terminal pro-brain natriuretic peptide (NT-proBNP) (representing hepatocyte, osteoblast, and cardiomyocyte stimulation, respectively) were measured in 26 hypothyroid subjects in a double blind, randomized, cross-over trial, which compared the replacement therapy with T4/T3 in combination (50 Fg T4 was substituted with 20 Fg T3) to T4 alone (once daily regimens). This was performed to obtain unaltered serum thyroid stimulating hormone (TSH) levels during the trial and between the two treatment groups. Blood sampling was performed 24 hours after the last intake of thyroid hormone medication.
Results: TSH remained unaltered between the groups ((median) 0.83 vs. 1.18 mU/l in T4/T3 combination and T4 mono-therapy, respectively; p=0.534). SHBG increased from (median) 75 nmol/l at baseline to 83 nmol/l in the T4/T3 group (p=0.015), but remained unaltered in the T4 group (67 nmol/l); thus, it was higher in the T4/T3 vs. T4 group (p=0.041). PINP levels were higher in the T4/T3 therapy (48 vs. 40 Fg/l (p<0.001)). NT-proBNP did not differ between the groups. Conclusions: T4/T3 combination therapy in hypothyroidism seems to have more metabolic effects than the T4 monotherapy.
Stimulatory effects of thyroid hormone on brain angiogenesis in vivo and in vitro

Liqun Zhang, CM Cooper-Kuhn, U Nannmark, K Blomgren and HG Kuhn
J Cereb Blood Flow & Metab 2010; 30:323–335. http://dx.doi.org:/10.1038/jcbfm.2009.216

Thyroid hormone is critical for the proper development of the central nervous system. However, the specific role of thyroid hormone on brain angiogenesis remains poorly understood. Treatment of rats from birth to postnatal day 21 (P21) with propylthiouracil (PTU), a reversible blocker of triiodothyronine (T3) synthesis, resulted in decreased brain angiogenesis, as indicated by reduced complexity and density of microvessels. However, when PTU was withdrawn at P22, these parameters were fully recovered by P90. These changes were paralleled by an  altered expression of vascular endothelial growth factor A (Vegfa) and basic fibroblast growth factor (Fgf2). Physiologic concentrations of T3 and thyroxine (T4) stimulated proliferation and tubulogenesis of rat brain derived endothelial (RBE4) cells in vitro. Protein and mRNA levels of VEGF-A and FGF-2 increased after T3 stimulation of RBE4 cells. The thyroid hormone receptor blocker NH-3 abolished T3-induced Fgf2 and Vegfα upregulation, indicating a receptor-mediated effect. Thyroid hormone inhibited the apoptosis in RBE4 cells and altered mRNA levels of apoptosis-related genes, namely Bcl2 and Bad. The present results show that thyroid hormone has a substantial impact on vasculature development in the brain. Pathologically altered vascularization could, therefore, be a contributing factor to the neurologic deficits induced by thyroid hormone deficiency.

Molecules important for thyroid hormone

synthesis and action – known facts and future perspectives

Klaudia Brix, Dagmar Führer, Heike Biebermann
Thyroid Research 2011, 4(Suppl 1):S9 http://www.thyroidresearchjournal.com/content/4/S1/S9

Thyroid hormones are of crucial importance for the functioning of nearly every organ. Remarkably, disturbances of thyroid hormone synthesis and function are among the most common endocrine disorders affecting approximately one third of the working German population. Over the last ten years our understanding of biosynthesis and functioning of these hormones has increased tremendously. This includes the identification of proteins involved in thyroid hormone biosynthesis like Thox2 and Dehal where mutations in these genes are responsible for certain degrees of hypothyroidism. One of the most important findings was the identification of a specific transporter for triiodothyronine (T3), the monocarboxylate transporter 8 (MCT8) responsible for directed transport of T3 into target cells and for export of thyroid hormones out of thyroid epithelial cells. Genetic disturbances of MCT8 in patients result in a biochemical constellation of high T3 levels in combination with low or normal TSH and thyroxine levels leading to a new syndrome of severe X-linked mental retardation. Importantly mice lacking MCT8 presented only with a mild phenotype, indicating that compensatory mechanisms exist in mice. Moreover, it has become clear that not only genomic actions of T3 exist. T3 is also capable to activate adhesion receptors and it signals via activation of PI3K and MAPK pathways. Most recently, thyroid hormone derivatives were identified, the thyronamines which are decarboxylated thyroid hormones initiating physiological actions like lowering body temperature and heart rate, thereby acting in opposite direction to the classical thyroid hormones. So far it is believed that thyronamines function via the activation of a G-protein coupled receptor, TAAR1. The objective of this review is to summarize the recent findings in thyroid hormone synthesis and action and to discuss their implications for diagnosis of thyroid disease and for treatment of patients.

Retinoic Acid Induces Expression of the Thyroid Hormone Transporter, Monocarboxylate Transporter 8 (Mct8)

T Kogai, Yan-Yun Liu, LL Richter, K Mody, H Kagechika, and GA Brent
J Biol Chem Jun 2010. Manuscript M110.123158
http://www.jbc.org/cgi/doi/10.1074/jbc.M110.123158

Retinoic acid (RA) and thyroid hormone are critical for differentiation and organogenesis in the embryo. The monocarboxylate transporter-8 (Mct8), expressed predominantly in brain and placenta, mediates thyroid hormone uptake from the circulation and is required for normal neural development. RA induces differentiation of F9 mouse teratocarcinoma cells towards neurons as well as extraembryonal endoderm. We hypothesized that Mct8 is functionally expressed in F9 cells and induced by RA.  All trans RA (tRA), and other RA receptor (RAR) agonists, dramatically (> 300-fold) induced Mct8. tRA treatment significantly increased uptake of triiodothyronine and thyroxine (4.1 fold and 4.3 fold, respectively), which was abolished by a selective Mct8 inhibitor, bromosulfophthalein. Sequence inspection of the Mct8 promoter region and
5′-rapid amplification of cDNA ends (5’-RACE) PCR analysis in F9 cells identified
11 transcription start sites and a proximal Sp1 site, but no TATA-box.  tRA significantly enhanced Mct8 promoter activity through a consensus RA responsive element located 6.6 kilobases upstream of the coding region. Chromatin immunoprecipitation assay demonstrated binding of RAR and retinoid-X receptor (RXR) to the RA response element. The promotion of thyroid hormone uptake through the transcriptional up-regulation of Mct8 by RAR is likely to be important for extraembryonic endoderm development and neural differentiation. This finding demonstrates crosstalk between RA signaling and thyroid hormone signaling in early development at the level of the thyroid hormone transporter.
Abnormal thyroid hormone metabolism in mice lacking the monocarboxylate transporter 8

Marija Trajkovic, Theo J. Visser, Jens Mittag, Sigrun Horn, et al.
J. Clin. Invest.  2007; 117:627–635. http://dx.doi.org:/10.1172/JCI28253

In humans, inactivating mutations in the gene of the thyroid hormone transporter monocarboxylate transporter 8 (MCT8; SLC16A2) lead to severe forms of psychomotor retardation combined with imbalanced thyroid hormone serum levels. The MCT8-null mice described here, however, developed without overt deficits but also exhibited distorted 3,5,3′-triiodothyronine (T3) and thyroxine (T4) serum levels, resulting in increased hepatic activity of type 1 deiodinase (D1). In the mutants’ brains, entry of T4 was not affected, but uptake of T3 was diminished. Moreover, the T4 and T3 content in the brain of MCT8-null mice was decreased, the activity of D2 was increased, and D3 activity was decreased, indicating the hypothyroid state of this tissue. In the CNS, analysis of T3 target genes revealed that in the mutants, the neuronal T3 uptake was impaired in an area-specific manner, with strongly elevated thyrotropin-releasing hormone transcript levels in the hypothalamic paraventricular nucleus and slightly decreased RC3 mRNA expression in striatal neurons; however, cerebellar Purkinje cells appeared unaffected, since they did not exhibit dendritic outgrowth defects and responded normally to T3 treatment in vitro.
In conclusion, the circulating thyroid hormone levels of MCT8-null mice closely resemble those of humans with MCT8 mutations, yet in the mice, CNS development is only partially affected.
3-Monoiodothyronamine: the rationale for its action as an endogenous adrenergic-blocking neuromodulator

HS Gompf, JH Greenberg, G Aston-Jones, A Ianculescu, TS Scanlan, and MB Dratman
Brain Res. 2010 Sep 10; 1351: 130–140. http://dx.doi.org:/10.1016/j.brainres.2010.06.067

The investigations reported here were designed to gain insights into the role of
3-monoiodothyronamine (T1AM) in the brain, where the amine was originally identified and characterized.
Extensive deiodinase studies indicated that T1AM was derived from the T4 metabolite, reverse triiodothyronine (revT3), while functional studies provided well-confirmed evidence that T1AM has strong adrenergic blocking effects. Because a state of adrenergic overactivity prevails when triiodothyronine (T3) concentrations becomes excessive, the possibility that T3’s metabolic partner, revT3, might give rise to an antagonist of those T3 actions was thought to be reasonable.
All T1AM studies thus far have required use of pharmacological doses.
Therefore we considered that choosing a physiological site of action was a priority and focused on the locus coeruleus (LC), the major noradrenergic control center in the brain. Site-directed injections of T1AM into the LC elicited a significant, dose-dependent neuronal firing rate change in a subset of adrenergic neurons with an EC50=2.7 μM, a dose well within the physiological range. Further evidence for its physiological actions came from autoradiographic images obtained following intravenous carrier-free 125I-labeled T1AM injection. These showed that the amine bound with high affinity to the LC and to other selected brain nuclei, each of which is both an LC target and a known T3 binding site. This new evidence points to a physiological role for T1AM as an endogenous adrenergic-blocking neuromodulator in the central noradrenergic system.

Thyroid hormones are transported through the blood-brain barrier

Thyroid hormones are transported through the blood-brain barrier (OATP) or the blood-CSF barrier (OATP and MCT8). In the astrocytes and tanycytes T4 is converted to T3 which then enters the neurons through MCT8. In the neurons both T4 and T3 are degraded by D3. T3 from the tanycytes may reach the portal vessels in the median eminence. Other transporters may be present on the astrocyte or tanycyte membranes. In most cases the transport could be bidirectional, although only one direction is shown.
Juan Bernal – Instituto de Investigaciones Biomedicas – 28029 Madrid, Spain

the interactions of maternal, placental and fetal thyroid

Old and new concepts of thyroid hormone action.

A: Old concept of thyroid hormone action. In former times it was assumed that thyroid hormones are able to pass the plasma membrane by passive transport. Once in the cytosol T4 is deiodinated to T3 which exerts genomic effects by binding to the thyroid hormone receptor (TR). After hetero-dimerization with other nuclear receptors like retinoic X receptor (RXR), transcriptional regulation is initiated resulting in activation or inactivation of target genes.
B: New concepts of thyroid hormone action. Thyroid hormones enter a target cell via specific transporters, e.g. T3 uses the monocarboxylate transporter MCT8 while T4 entry is mediated by Lat2 or Oatp14. Moreover, T3 can interact with avb3 integrins to induce ERK1/2 signalling. Cytosolic T3 exerts genomic effects but can additionally also act by non-genomic means after TR binding and activation of down-stream PI-3 kinase. Likewise, the naturally occurring iodothyronine T2 is believed to stimulate metabolic rates via mitochondrial pathways, thereby bypassing genomic regulation. Besides thyroid hormones, derivatives like the thyronamines T1AM or T0AM, modulate the action of T3, e.g. counter-acting its effects in certain target cells. Thyronamines (TAMs) bind to and activate G-protein coupled receptors (GPCRs) of the trace amine associated receptor (TAAR) family. So far, it is only known that TAAR1 is activated by TAMs and signals via adenylylcyclase (AC) activation with subsequent rise of cAMP levels. However other GPCRs are likely targets for thyroid hormone derivatives

Brix et al.: Molecules important for thyroid hormone synthesis and action – known facts and future perspectives. Thyroid Research 2011 4(Suppl 1):S9.
http://dx.doi.org:/10.1186/1756-6614-4-S1-S9

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