The Production of Thyroid Hormone Part Two

The Production of Thyroid Hormone Part Two

The thyroid has far reaching effects on health and fetal development.  Successful treatment of thyroid disorders requires a solid understanding of the pathophysiology of the production of thyroid hormone.

Header Image: Thyroid follicles microscopic view courtesy of wikimedia commons and Grays Anatomy.

What is Thyroid Hormone ?

The main job of the thyroid gland is to secrete thyroid hormone into the blood stream in a ratio of 80% T4 and 20 per cent T3.  In the periphery, at the cellular level,  Diodinase Enzyymes convert T4 to T3, the more biologically active form of the hormone. T4 is a pro-hormone, havingfour iodine atoms attached to two tyrosine rings coupled together. The De-Iodinase enzyme is intracellular, and removes an iodine from T4 making T3.

Chemical Structure of thyroid hormone: Above left image is T4 with four iodine atoms, and above right image is T3 with three Iodine atoms. Courtesy of de Castro, Guilherme Vieira, et al. “Synthesis of Analogues of Thyroid Hormones: Nuclear Receptor Modulators.” Orbital: The Electronic Journal of Chemistry 7.3 (2015): 282-291.(57)

In 2014, Dr Carla Portulano writes:

The significance of the thyroid gland for human health is difficult to overstate, given the wide-ranging effects of the thyroid hormones (THs) on prenatal and early development as well as on intermediary metabolism at all stages of life. (32)

Thyroid hormones have been described as “the major endocrine controllers of metabolic rate”, since increasing the thyroid hormone levels increases metabolic rate, primarily via effects on mitochondria, stimulating increased numbers and size of mitochondria (mitochondriogenesis) as well as respiratory chain components within the mitochondria. Left image: Electron Microscopy of mitochoindria courtesy of wikimedia commons.

In 2008, Dr. Mary Ellen Harper writes:

thyroid hormones stimulate mitochondriogenesis and thereby augment cellular oxidative capacity. Thyroid hormones induce substantial modifications in mitochondrial inner membrane protein and lipid compositions. Results are consistent with the idea that thyroid hormones activate the uncoupling of oxidative phosphorylation through various mechanisms involving inner membrane proteins and lipids. Increased uncoupling appears to be responsible for some of the hypermetabolic effects of thyroid hormones. (20)

In 2022, Dr Federica Cioffi writes:

almost all components of the respiratory chain [within mitochondria] are directly or indirectly affected by iodothyronines [thyroid hormones]. In some cases, the actions result in an activation of specific biochemical pathways while in others the effects would result into an increase in mRNA or protein levels of specific components of respiratory chain. (21)

Above image: light microsopy of thyroid follicles coutesy of wikimedia commons.

The Thyrocyte and Follicular Lumen:

The thyroid cell, or thyrocyte, is the workhorse of the thyroid gland, the first step is secretion of thyroglobulin into the central follicle.  Thyrocyte cells line the follicles, round storage filled with pink staining colloid, also called thyroglobulin. Microscopically, the thyroid gland demonstrates a pattern with thousands of small spherical follicles serving as storage chambers for thyroglobulin, the precursor protein later converted to thyroid hormone.

The thyrocytes are triangular shaped and have a specific orientation. The apex is nearest the lumen of the follicle and contains the villous apical membrane which does most of the work. The base of the thyrocyte is oriented away from the follicle lumen containing the NIS, the sodium iodine symporter, the active transport mechanism which takes up iodine from the blood stream and concentrates iodine in the thyrocyte. The NIS is imbedded is the basal membrane of the thyrocyte.

Five Steps of Thyroid Hormone Synthesis

In 2022, Dr. Muhammad Shahid discussed the five steps of thyroid synthesis.(1)

First Step: Synthesis of Thyroglobulin:

Thyrocytes in the thyroid follicles produce a protein called thyroglobulin (TG), the precursor to thyroid hormone which, initially, does not contain any iodine. TG is secreted by exocytosis and stored in the follicle.

Step Two: Uptake of Iodide, the Sodium Iodide Symporter:

TSH stimulation causes increased activity of active transport of Iodine into the thyrocyte.  This is accomplished by the Sodium Iodide Symporter (Na+-I-), located at the basolateral membrane of the thyrocyte which actively pumps iodide into the cell, maintaining a concentration 20 to 50 times higher than that of the blood stream. In Grave’s hyperthyroidism, this ratio is increased to greater than 100 times higher, due to massive TSH receptor stimulation by Grave’s disease auto-antibodies (TSI and TRAb). Even though the serum TSH will be quite low (suppressed), the thyroid gland in Graves’ disease is massively stimulated by the TSH receptor antibodies.  The thyroid gland will concentrate bromine as well as iodine. If so, bromine in the thyroid decreases the accumulation of iodine. If severe, this may lead to iodine deficiency and hypothyroidism.(58)

Iodide diffuses through the thyrocyte to the apical membrane where it is pumped into the follicular lumen by the pendrin transporter. Note: the role of pendrin is still matter for debate. (1-4)(15)

Step Three : Iodination of thyroglobulin:

Thyroglobulin provides the polypeptide backbone for synthesis and storage of thyroid hormones. Thyroglobulin is manufactured in the endoplasmic reticulum of the thyrocyte and secreted into the follicular lumen by exocytosis.  Thyroglobulin within the follicular lumen is called colloid.  The iodination of thyroglogulin, also called organification of iodine, is performed by the Thyroperoxidase (TPO) enzyme, a heme containing molecule with a porphyrin ring structure similar to hemoglobin. TPO is secreted into the follicular lumen at the thyrocyte apex villous membrane, and has three functions: 1) oxidation of iodine, 2) organification of iodine, and 3) coupling of Tyrosine residues. Once iodine is oxidized by TPO to molecular iodine (I2), it combines with ring-like Tyrosine residues on the thyroglobulin to form MIT and DIT (mono-iodo-tyrosine and di-iodotryrosine)

Oxidation of Iodide to Iodine: TPO uses hydrogen peroxide to oxidize iodide (I-) to iodine (I2). This hydrogen peroxide is generated by NADPH oxidases dual oxidases 1 and 2 (DUOX1 and DUOX2) located in the villous apex of the thyrocyte. Iodide, which is a negative ion, is oxidized to molecular iodine (I2) by the TPO thyroperoxidase enzyme using hydrogen peroxide as a substrate. This takes place inside the follicles on the micro-villous surface of the apical membrane of the thyrocytes, the same location as hydrogen peroxide produced by the DUOX enzyme system.

Organification: Iodine(I2) spontaneously attaches to tyrosine residues of thyroglobulin protein, generating monoiodotyrosine (MIT, one iodine) and diiodotyrosine (DIT, two iodines)

Coupling reaction: Iodinated Tyrosine residues are coupled by TPO making triiodothyronine (T3) and tetraiodothyronine (T4). T4 is made from 2 DIT’s. T3 is made from MIT and DIT.

Step 4: Storage: Thyroid hormones (T3 and T4) are stored in the follicular lumen bound to thyroglobulin as colloid, a combination of Free and Iodinated thyroglobulin.

Step 5: Release of thyroid hormone into circulation: Iodinated thyroglobulin is taken up at the apex of the thyrocytes as vesicles via endocytosis, travels back towards the basal membrane and while In transit, these vesicles are fused with lysosomes containing acid and proteolytic enzymes which digest the thyroglobulin, freeing the T3 and T4, subsequently released at the basolateral membrane into the capillary blood stream in ratio of 80% T4 and 20% T3.  Any extra iodine is salvaged and returned to the intracellular iodine pool.(1-4)

Hashimoto’s Thyroiditis:

In Hashimotos Disease, blood tests show elevated antibodies to the TPO (Thyroperoxidase) enzyme and to thyroglobulin.  Both are in close proximity to the hydrogen peroxide generating system called DUOX (dual oxidase) at the apical villous membrane. Damage caused by excess hydrogen peroxide to these proteins is thought to create antigenicity and autoimmunity. In both Graves’ and Hashimoto’s Disease, electron microscopy may show electron dense deposits along the basement membrane of the follicular cells which are immune complexes when visualized with immunoflourescent staining.(59-61)

Immunoflourescent staining in Hashimoto’s with high titers of TPO antibodies shows intense staining of the cytoplasm of thyrocytes without staining of colloid in follicles. In cases with high thyroglobulin antibody titers, there is diffuse staining of colloid within the follicles. (73)

Organification Defect in Hashimoto’s

Hashimotos’ disease results in organification defect, a decreased ability to organify iodine. This is called “inability to organify iodine”, and explains the low iodine content of the thyroid gland in Hashimoto’s patients, and the rapid washout of iodine with the perchlorate discharge test.

Similarly, methimazole, the most common thyroid blocking drug, works by irreversibly binding to and blocking function of the TPO enzyme, thus inhibiting organification of iodine. Methimazole also inhibits the DUOX enzyme, thus inhibiting hydrogen peroxide formation, a beneficial feature which prevents excess hydrogen peroxide damage to the thyrocytes.

A major difference in thyroid blocking function of methimazole vs. excess iodine or lithium is this: methimazole does not block release of iodine from the thyroid gland.  However, excess Iodine or lithium will block release of Iodine from the thyroid gland.

Loss of Auto-Regulation

The thyroid gland in Hashimoto’s patients has lost the autoregulation needed to escape from suppressive effects of iodine excess, rendering these patients more sensitive to “Iodine Blockade” described by Wolf as the suppressive effect of iodine excess on thyroid function. In normal thyroid glands, down regulation of the NIS symporter RNA by iodine as well as a direct effect on NIS within the basal membrane leads to eventual escape from the suppressive effects of excess iodine within two days.  However, this does not apply to thyroid blockade with methimazole or lithium, as there is no escape from their suppressive effects.(74-75)

As mentioned above, the second antibody in Hashimoto’s Disease is the anti-thyroglobulin antibody. Additional antibodies against the NIS (sodium iodine symporter), and against Pendrin have also been discovered.  However, lab tests for NIS and Pendrin are not yet available for clinicians.  Antibodies which stimulate the TSH receptor (TSI and TRAb) indicates Graves’ hyperthyroidism.  About 70 per cent of Graves’ disease patients will also have TPO and thyroglobulin antibodies found typically on Hashimotos’ hypothyroidism.  Indeed, some authors believe Graves and Hashimotos’ are two extremes of the same disease process.

Methimazole-antithyroid medication

Methimazole works by irreversibly inactivating the thyroid peroxidase enzyme (TPO), thus preventing organification, a necessary step for thyroid hormone synthesis.  In 1999, working in vitro, Dr Sugawara found that methimazole increases TPO mRNA, resulting in greater amounts of TPO enzyme in the thyroid gland. Thus, upon stopping the methimazole drug, there may be a rebound phenomenon as TPO activity is increased. Selenium analogs of Methimazole, still under development, may be the preferable form because the selenium protects the thyroid from oxidation. Until then, it would be prudent to supplement with selenium and magnesium to prevent thyroid damage from excess hydrogen peroxide. (8-10)(52-53)

Genetic Mutation in Thyroperoxidase

Failure to organify iodine can be found in those harboring genetic mutations in the thyroperoxidase enzyme. In these cases, iodide in the thyroid gland cannot be oxidized and/or bound to the thyroglobulin protein.(8)

The role of selenium and hydrogen peroxide

The thyroid gland has a high selenium content due to selenoproteins (glutathione peroxidases and deiodinases). The anti-oxidant selenoproteins protect the thyrocyte cells from oxidative damage associated with hydrogen peroxide production needed for thyroid hormone biosynthesis.

Iodination of thyroglobulin, also called organification, is the key step of thyroid hormone biosynthesis. It is catalyzed by thyroid peroxidase (TPO) and occurs within the follicular space at the apical plasma membrane. Hydrogen peroxide generating enzymes (DUOX) are also found at this same location, at the villous apical membrane of the thyrocyte just within the follicular lumen.  Both enzyme systems are needed for the organification of iodine to Tyrosine residues in the thyroglobulin. Various thyroid pathologies, including Hashimoto’s thyroiditis can be explained by overproduction and lack of degradation of hydrogen peroxide (H2O2) causing damage to the thyrocyte structures. (11-16)

Iodine Intake Inhibits H202 Generation-Wolf Chaikoff Effect

The “Wolff-Chaikoff” effect is the inhibition of H2O2 generation caused by iodine itself.  The intake of excess iodine limits the oxidation and binding of I2 (iodine) to thyroglobulin because of the reduced availability of hydrogen peroxide at the apical membrane. The Wolff-Chaikoff effect is the inhibition by iodide of its own organification. Since the organification of iodine is already reduced in Hashimoto’s thyroiditis, these patients are more sensitive to the inhibitory effect of iodine excess on thyroid function.  This will be discussed later in various studies which misinterpret the inhibitory effect of Iodine as a worsening in Hashimoto’s thyroiditis.

In 1988, Dr Bernard Corvilain Studied Dog Thyroid Slices

In 1988, Dr Bernard Corvilain studied the Wolff-Chaikoff effect in dog thyroid slices in vitro, finding elevated TSH stimulates hydrogen peroxide generation and organification. Excess iodide had the oppsoite effect, greatly inhibiting hydrogen peroxide generation as well as organification via reduction of intracellular signaling from TSH (The Wolf-Chaikoff Effect), writing:

In dog thyroid slices thyrotropin [TSH] and carbamylcholine greatly enhance protein iodination and H2O2 generation. The action of thyrotropin [TSH] is … mediated by cyclic AMP. This suggests that the effect of carbamylcholine is mediated by the two intracellular signals generated by the Ca++ phosphatidylinositol cascade: Ca++ and diacylglycerol. The Wolff-Chaikoff effect is the inhibition by iodide of its own organification…In dog thyroid slices, iodide greatly inhibited H2O2 generation stimulated by thyrotropin [TSH] and by carbamylcholine. Iodide decreased the production of intracellular signals induced by TSH.(17)

Note: carbamycholine is a common drug used in ophthalmology to dilate pupils. Emphasis Mine.

Toxic Effect of Acute Iodine Administration in Iodine Depleted Animal

In 2000, Dr. Bernard Corvilain again studied the effect of iodine administration on H202 production in animal thyroid slices, finding iodine depletion is a condition in which acute iodine administration stimulates H202 production, rather than inhibits it.  When iodine is acutely given to an iodine deficient animal, there is stimulation of H202, presumably to promote efficient oxidation and organification of the iodine into thyroid hormone.  This stimulation of H202 explains the toxic effect of acute iodine administration in an iodine depleted animal. Dr. Bernard Corvilain writes:

In comparison with conditions in which an inhibitory effect of iodide on H2O2 generation is observed, the stimulating effect was observed for lower concentrations and for a shorter incubation time with iodide. Such a dual control of H2O2 generation by iodide has the physiological interest of promoting an efficient oxidation of iodide when the substrate is provided to a deficient gland [iodine deficient] and of avoiding excessive oxidation of iodide and thus synthesis of thyroid hormones when it is in excess. The activation of H2O2 generation may also explain the well described toxic effect of acute administration of iodide on iodine-depleted thyroids. (17B)  Emphasis Mine

RESUME HERE ZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZZ

Thyroid Hormone Synthesis Courtesy of Szanto 2019 Fig 3.  Szanto, Ildiko, Marc Pusztaszeri, and Maria Mavromati. “H2O2 metabolism in normal thyroid cells and in thyroid tumorigenesis: focus on NADPH oxidases.” Antioxidants 8.5 (2019): 126.

Hydrogen Peroxide Metabolism

in 2019, Dr Ildiko Szanto reviewed the role of hydrogen peroxide in  normal thyroid metabolism in organification of iodine and production of thyroid hormone.  However, excessive hydrogen peroxide generation is carcinogenic, and simulates autoimmunity, writing:

The synthesis of thyroid hormones… utilize hydrogen peroxide (H2O2) as an oxidative agent. Hydrogen peroxide is contained within the lumen of the thyroid follicles which are considered as the functional units of the thyroid gland where hormone synthesis, storage and release take place. The follicles are formed by a monolayer of polarized epithelial cells, termed thyrocytes, that surround the central lumen of the follicle. The lumen is delimited by the apical surface of thyrocytes that are connected to each other by tight junctions preventing “leaking” of the lumen content into the extra-follicular space. The outer side of the follicle is sealed by the basolateral plasma membranes of thyrocytes. The lumen is filled by a protein-rich substance termed “colloid”, mainly consisting of thyroglobulin (TG), a 660kDa thyroid-specific protein. TG accounts for approximatively 50% of the protein content of the thyroid gland and serves as the precursor of thyroid hormones. Thyroid hormone synthesis requires the oxidative iodination of specific tyrosine residues of TG, a process termed “iodide organification”. Oxidative iodination is catalyzed by the enzyme thyroid peroxidase (TPO). This process is a key step in thyroid hormonogenesis requiring an appropriate amount of H2O2 for oxidation…The major source of H2O2 in the thyroid follicle is the isoform DUOX2 [5,6]. In contrast to DUOX2-generated physiological reactive oxygen species (ROS) production, pathologically elevated H2O2 levels are linked to thyroid carcinogenesis resulting from enhanced mitogenic receptor signals or oncogene activation . (22)

Regulation of H202 generation by TSH and Iodine

The Hydrogen peroxide generating system is called DUOX2, and as one might expect is upregulated by TSH in two ways. 1) messenger RNA is increased. This increases production of the DUOX2 protein. 2) TSH increases DUOX2 enzymatic activity.

Excess Iodine inhibits DUOX2 mediated production of Hydrogen peroxide.  This is the Wolf Chaikoff Effect first described in 1948, i.e. Iodine inhibits its own organization. This is a form of thyroid autoregulation which is mediated by Iodolactones and iodoaldehydes formed when molecular iodine reacts with lipids. Iodolactones not only mediate thyroid autoregulation, they also play an important role as anti-cancer agents in prevention and treatment of breast cancer.  See the chapter on prevention and treatment of breast cancer with iodine. Dr Ildiko Szanto writes:

DUOX2-mediated H2O2 production is inhibited by excess iodine leading to decreased TPO activity and reduced incorporation of iodine into TG [thyroglobulin]. This inhibitory effect of iodine on its own organification was already described in 1948 and was named the “Wolff-Chaikoff effect” after the authors of the original paper.(22)(62-63)

In Graves’ Disease, elevated antibodies to the TSH receptor (TSI and TRAb) stimulate the thyroid gland to “go into overdrive”, producing massive quantities of thyroid hormones.  The elevated T3 and T4 suppress the TSH which will be quite low in Graves’ disease. One might assume this same TSH receptor stimulation would increase H202 generation. This would be an incorrect assumption. While TSH hormone stimulates H202 generation, TSH receptor antibodies in Graves’ disease do not stimulate H202 production.(64)

This may explain why the difference in appearance of the thyroid gland in classical Graves disease versus Hashimotos’.  In Classical Graves disease the thyroid gland is smoothly enlarged without nodularity or fibrosis and other inflammatory changes associated with excess hydrogen peroxide generation.  Of course, over time, the thyroid gland in Graves’ Disease may develop irregular nodularity after repeated bouts of thyroiditis, and may change its imaging appearance to that of multinodular goiter.

How can we explain the undulating course of Graves disease with cycles of relapse and remission.  During remission obrained by using methimazole, a thyroid blocking drug, the TSH may go too high, stimulating excess H202 production which may then cause thyroiditis, the release of excess thyroid and relapse of hyperthyroidism.

Block and Replace Treatment Strategy

During treatment of Grave’s Disease with thyroid blocking drugs such as Methimazole, Iodine excess, or Lithium, treatment may suppress thyroid hormone production enough to cause TSH elevation, which can then stimulate hydrogen peroxide production and damage the thyroid gland. In this scenario, “Block and Replace” would be useful to decrease the TSH and prevent thyroid stimulation and damage from excess H202.  Remember, Methimazole not only irreversibly blocks the TPO enzyme, it also inhibits hydrogen peroxide generation.

Figure 1 Thyroid hormone biosynthesis, secretion and major signaling pathways in thyrocytes. AC, adenyl cyclase; cAMP, cyclic adenosine monophosphate; DAG, diacylglycerol; DEHAL1, dehalogenase 1; DIT, diiodotyrosine; DUOX, dual oxidase; DUOXA, dual oxidase maturation factor; IP3, inositol trisphosphate; MIT, monoiodotyrosine; NIS, sodium-iodide symporter; PDS, pendrin; PKA, protein kinase A; PKC, protein kinase C; PLC, phospholipase C; T3, triiodothyronine; T4, thyroxine; TG, thyroglobulin; TPO, thyroid peroxidase; TSH, thyrotropin; TSHR, thyrotropin receptor . Courtesy of (Ohye, 2010)

Dr. Hidemi Ohye Explains Production of Thyroid Hormone

In 2010, Dr. Hidemi Ohye explained the production of thyroids hormone as follows:

Iodide is actively transported into thyrocytes by a sodium/iodide symporter (NIS) on the basolateral membrane and to the follicular lumen by, in part, pendrin (PDS/SLC26A4) at the apical membrane. Iodide is rapidly oxidized by TPO in the presence of H2O2 resulting in covalent binding to the tyrosyl residues of thyroglobulin (Tg) on the luminal side of the apical membrane. This step produces monoiodotyrosine (MIT) and diiodotyrosine (DIT). Then only properly spaced MIT and DIT in Tg participate in the coupling reactions to form thyroxine (T4) and triiodothyronine (T3); this reaction is also catalyzed by TPO with H2O2. The source of thyroid H2O2 is DUOX2 expressed in the apical plasma membrane coordinated with DUOXA2. Thyroid hormones are released into the circulation after digestion of Tg. MIT and DIT are deiodinated by an iodotyrosine dehalogenase 1 (DEHAL1).23 Thyroid hormone formation is predominantly regulated by thyrotropin (TSH). The binding of TSH to the TSH receptor activates both Gs and Gq proteins. The former activates the growth regulation, differentiation and thyroid hormone secretion, whereas the latter activates H2O2 generation and iodide binding to protein through the phospholipase C-dependent inositol phosphate Ca2þ/diacylglycerol pathway…. Methimazole and propylthiouracil inhibit NADPH oxidase activity… Thyroid-stimulating antibody found in patients with Graves’ disease does not appear to stimulate H2O2 generation. (16)(64) Emphasis Mine.

Both TSH and Iodide Control H202 Levels

Note in the above quote, Dr. Hidemi Ohye  explains how TSH controls thyroid gland growth regulation, differentiation and thyroid hormone secretion, as well as H2O2 generation and iodide binding to protein (organification).  Higher TSH stimulates all these steps in thyroid hormone synthesis as well as thyroglobulin, sodium Iodide Symporter and Thyroid Peroxidase protein expression.

Dr. Hidemi Ohye goes on to explain how Iodide controls H202 levels.  While TSH stimulates greater H202 production, Iodide excess reduces H202 production. This is done through post-transcription change in the DUOX molecule. Note DUOX is the Dual Oxidase system which generates hydrogen peroxide.  Dr. Hidemi Ohye writes:

Iodide controls H2O2 generation in thyroid cells. Morand et al. have studied the effect of KI on H2O2 generation in porcine thyroid follicles, the most physiological thyroid culture system. They exposed follicles to 1 mmol/L KI for two days under cAMP stimulation and showed reduction in H2O2 production without affecting DUOX mRNA levels. Post-transcriptional change of the DUOX molecule by KI appears to be responsible for the decreased H2O2 generation. Quote from : Ohye, Hidemi, and Masahiro Sugawara. “Dual oxidase, hydrogen peroxide and thyroid diseases.” Experimental biology and medicine 235.4 (2010): 424-433.

Wolff–Chaikoff effect Protects Thyrocytes from Iodine Excess

In the normal thyroid gland, the Wolff–Chaikoff effect protects thyrocytes from Iodine excess by inhibiting iodide organification, leading to discharge of iodide which cannot be organified. For example, when a 65 mg potassium iodide capsule is distributed to the population surrounding a nuclear accident, the Wolff–Chaikoff effect prevents entry of radioactive iodine into the thyroid gland, protecting the population from increased risk of thyroid cancer. However, the auto-immune thyroid patient may have a defective Wolff–Chaikoff effect. instead of reducing H202, the excess iodine increases H202 production. This creates a damaging effect on thyroglobulin which then serves as antigen for auto-immune attack.  Methimazole is a TPO enzyme inhibitor. However, methimazole also inhibits H202 generation by inhibiting the NAD oxidase. (Freitas Ferreira 2003)

Dr. Hidemi Ohye writes:

The animal experiments suggest that susceptible hosts have defective Wolff–Chaikoff effect allowing them to generate H2O2 in response to increased iodide influx, which ordinarily should not happen as seen in the non-susceptible mouse.67 Thus, abnormality of thyroid H2O2 generation in response to high iodide may play a role in the development of Hashimoto’s thyroiditis in susceptible individuals. Whether iodide-mediated H2O2 generation is driven by activated DUOX or NOX4 or defective antioxidants has yet to be studied.(16)(64)

Graves Antibodies DO NOT Stimulate hydrogen Peroxide !

As mentioned avbove, TSH stimulates all steps in thyroid hormone synthesis including generation of hydrogen peroxide (H202). In 1991, Dr Eric Laurent studied human thyroid slices in vitro finding, unlike TSH (thyrotropin), Graves Antibidies do not stimulate hydrogen peroxide generation.  This makes sense to me since if Graves antibodies did stimulate excess hydrogen peroxide, this would lead to severe thyroid damage, chronic thyroiditis and ealry complete destruction of the thyroid gland similar to the myedematous cretinsim described in Zaire, Africa. This would lead to immune cell infiltration and rapid complete destruction of the thyroid gland. Instead, in early untreated Graves disease, the gland is smoothly enlarged without evidence of chronic changes. Dr Eric Laurent writes:

The effects of thyroid-stimulating antibodies (TSAb) and of thyrotropin (TSH) were compared…The patterns of the response curves of TSAb and TSH on cyclic AMP accumulation were different, suggesting that different mechanisms may be involved. In addition, unlike TSH, TSAb were not able to stimulate H2O2 generation, which in human tissue mainly depends on the activation of the phosphatidylinositol-Ca2+ cascade. … In conclusion, our results show that TSAb do not share all the metabolic actions of TSH on human thyroid tissue. (64)

As mentioned above, Hydrogen peroxide generation is inhibited by excess iodine, and partially inhibited by Methimazole and PTU (propylthiouracil). (65-69)

Dr. Guy Abraham Explains Thyroid Hormone Production- Iodine Deficiency Causes Hashimotos Thyroiditis

Dr. Guy Abraham, the inventor of Iodoral, a popular iodine supplement, explains how iodine deficiency causes increased TSH. This stimulates excess hydrogen peroxide which damages TPO and thyroglobulin causing Hashimotos thyroiditis.  The mechanism of excess H2O2 production is caused by combined iodine and magnesium deficiencies resulting in  low levels of iodinated lipids and high cytosolic calcium, respectively. The excess hydrogen peroxide damages structures in closest proximity, TPO and the substrate thyroglobulin (Tg). These damaged proteins stimulate autoimmunity, thus explaining the origin of Hashimotos Disease from iodine and magnesium deficiency. In Dr. Abraham’s opinion, supplementing with iodine (12.5 mg/day or above) and magnesium (1200 mg per day) should reverse auto-immune thyroiditis, both Hashimotos and Graves’ disease. Dr. Guy Abraham writes:

We would like to propose a mechanism for the oxidative damage caused by low levels of iodide combined with antithyroid drugs: inadequate iodide supply to the thyroid gland, aggravated by goitrogens [a substance that blocks iodine uptake, worsening the iodine deficiency], activates the thyroid peroxydase (TPO) system through elevated TSH, low levels of iodinated lipids, and high cytosolic free calcium, resulting in excess production of H2O2. …This H2O2 production is above normal due to a deficient feedback system caused by high cytosolic calcium resulting from magnesium deficiency and low levels of iodinated lipids which requires for their synthesis iodide levels two orders of magnitude greater than the RDA for iodine. Once the low iodide supply is depleted, TPO in the presence of H2O2 and organic substrate reverts to its peroxydase function which is the primary function of  haloperoxydases, causing oxidative damage to molecules nearest to the site of action: TPO and the substrate thyroglobulin (Tg). Oxydized TPO and Tg elicit an autoimmune reaction with production of antibodies against these altered proteins with subsequent damage to the apical membrane of the thyroid cells, resulting in the lymphocytic infiltration and in the clinical manifestations of Hashimoto’s thyroiditis.

In laboratory animals prone to autoimmune thyroiditis, the genetic defect may be in the production of H2O2 in excess of what is needed. The iodination of tyrosine residues by TPO requires the presence of Tg, H2O2, and iodide. The supply of H2O2 comes from the NADPH oxydase system. This system is inhibited by certain iodinated lipids and is enhanced by cytosolic free calcium Ca++.89 The equation for organification of iodide by TPO is displayed in Figure 1, together with the feedback system controlling the production of H2O2. The logical deduction from this equation is that increased cytosolic free calcium will cause an excess of H2O2. Increased levels of iodinated lipids, on the other hand, would limit the production of H2O2. How much iodide is required for the production of iodinated lipids? In 1976, Rabinovitch, et al reported their results regarding the effect of three levels of iodide supplementation on the production of iodinated lipids in the thyroid glands of dogs: low iodide diet, normal iodide diet, and high iodine diet. The dogs were kept on those diets for
six weeks. Iodinated lipids in the plasma membrane and in the cell total lipids were observed only in the dogs receiving the high iodide diet. What about human subjects? In 1994, Dugrillon, et al reported for the first time the presence of 5-hydroxy-6-iodo- 8,11,14-eicosatrienoic delta lactone (delta iodolactone) in a human thyroid, following the ingestion of 15 mg iodide/day for 10 days in the host. It was the first time this biologically active iodolipid was isolated from human thyroid glands. The amount of iodide the host received was 100 times the RDA, but it is the amount of iodine/iodide we recommended for orthoiodosupplementation 30 Dugrillon, et al stated, “These results demonstrate for the first time that delta-iodo-lactone is present in iodide-treated human thyroid.” Magnesium deficiency, which is prevalent in the US population, results in increased levels of cytosolic free calcium.98 Intracellular free calcium levels above the normal range are cytotoxic causing calcification of mitochondria
and cell death. The cell membrane possesses an ATP-dependant calcium pump that keeps intracellular levels of free ionized calcium within narrow limits. This calcium pump is magnesium-dependent for normal function. Magnesium deficiency results in a defective calcium pump and intracellular accumulation of ionized calcium. Inadequate iodine/iodide intake below orthoiodosupplementation results in decreased levels of
delta-iodo-lactone. Combined magnesium and iodine/
iodide deficiency based on the concept of  orthoiodosupplementation are the basic factors involved in the
oxidative damage caused by excess H2O2 and reactive
oxygen species. If this proposed mechanism is valid, orthoiodosupplementation,30 combined with magnesium intake between 800-1,200 mg/day, a daily amount this author recommended 21 years ago98 for magnesium sufficiency, should reverse autoimmune thyroiditis. This nutritional approach is also effective in Graves’ autoimmune thyroiditis as previously discussed.(70-72)

Various Thyroid Pathologies

In 2008, Dr Song explains that various thyroid pathologies are caused by either overproduction of hydrogen peroxide, or its lack of degradation due to selenium deprivation and consequent GSH [glutathione] peroxidase depletion, thus explaing the etiology of auto-immune thyroid disease, goiter, thyroid nodules, thyroid cancer etc.

Dr. Song reminds us excess H2O2 in thyroid cells is a carcinogen and “a killer”. Dr. Song then discusses the etiology of myxedematous endemic cretinism in Zaire Africa, a form of thyroid destruction after birth asociated with iodine deficiency (which stimulates elevated TSH and consequently H2O2 generation), combined with selenium deficiency (causing decreased GSH peroxidase and thioredoxin reductase activity), combined with dietary thiocyanate.  Dr Song proposes a similar mechanism for patholophysiology of thyroiditis, and proposes selenium dietary supplementation for prevention and treatment of thyroiditis. Dr. Song writes:

In thyrocytes of most species, including humans, TSH and its receptor activates…H2O2 generation, iodide binding to proteins, and thyroid hormone formation, and secretion…In all species studied, iodide at high concentrations presumably through an iodinated lipid, iodohexadecanal [6-iodo-lactone], inhibits H2O2 generation (the Wolff-Chaikoff effect) and adenylate cyclase…

H2O2 in various cell types, and presumably in thyroid
cells, is a signal, a mitogen, a mutagen, a carcinogen, and a killer…It is proposed that various pathologies can be explained, at least in part, by overproduction and lack of degradation of H2O2 (tumorigenesis, myxedematous cretinism, and thyroiditis) and by failure of the H2O2 generation or its positive control system (congenital hypothyroidism)……We have repeatedly suggested that the important generation of H2O2 in thyroid cells might account for mutagenesis and the important generation of nodules in the thyroid. This would also explain in part why more nodules are found in iodine-deficient areas.

Myxedematous endemic cretinism [Zaire Afreica], caused by thyroid destruction after birth, has been linked to low iodine supply in early life, leading to intense stimulation [by elevated TSH]  and presumably H2O2 generation, to passage from low O2 to high O2 at birth, to selenium deficiency, and thus to decreases in GSH peroxidase and thioredoxin reductase activity and to dietary thiocyanate…Interestingly, a similar scenario has been proposed for the physiopathology of thyroiditis. Selenium dietary supplementation has therefore been proposed for prevention and treatment of thyroiditis and has indeed alleviated it. (4) Emphasis Mine.

Above image courtesy of Dr Robyn Murphy, (2016) (76)

Dr. Robyn Murphy Clears Everything Up

In 2016, Dr. Robyn Murphy agrees with Dr. Guy Abraham and Dr. Song suggesting iodine deficiency as a causative factor in patients with auto immune thyroid disease. Firstly, Dr. Murphy points out the observed increase in auto-immune thyroiditis after introduction of salt iodination programs. Then Dr. Murphy details how chronic iodine deficeincy affects the mechanism of thyroid hormone production in a way that gives a satidfactory explaination of the above observation.

Chronic iodine deficiency creates a loss of autoregulation in the thyroid leading to excess hydrogen peroxide generation when iodine is re-introduced. This loss of autoregulation relates to high TSH levels which stimulate NIS (sodium iodide transporter), the active transport of iodine into the thyrocyte. In some patients, namely those with aiuto immune thyroid disease, the NIS can not be turned by reintrodcution of iodine, leading to loss of autoregulation. This loss of autoregulation also relates to absence of Iodolactones the main feedback mechanism for turning off excess hydrogen peroxide production.  Intracellular calcium excess is also a factor in excess hydrogen peroxide production, hence the benefits of magnesium supplementation. Dr. Murphy writes:

In iodine deficiency, the loss of negative feedback on H2O2 production has been implicated in thyroid dysfunction and as a possible mechanism in the generation of autoantibodies.(76)

Dr. Dr. Robyn Murphy goes on to describe the train of events in the production of thyroid hormones, writing:

The sodium-iodide symporter (NIS) transports iodide (I) into the thyrocyte to be organified to iodine (I2) and bound to thyroglobulin (TG) by thyroid peroxidase (TPO) and hydrogen peroxide (H2O2).  The iodine–thyroglobulin complexes then combine to form thyroid hormones. The nicotinamide adenine dinucleotide phosphate (NADPH) oxidase system is upregulated by intracellular calcium to generate H2O2, a reactive oxygen species (ROS). To prevent excess H2O2, iodolactones negatively inhibit NADPH oxidase, and glutathione peroxidase degrades H2O2Iodinated lipids provide negative feedback on NADPH oxidase to reduce H2O2 and oxidative damage to the thyrocyte…Studies suggest that when iodine intake is above 1.5 mg daily, TPO synthesizes iodolactones. TPO facilitates the iodination of arachidonic acid and other polyunsaturated fatty acids to produce iodolactones. Research shows that measurable concentrations of δ-iodolactone are found in the thyroid tissue, prevent excess iodide uptake and the generation of H2O2 by NADPH oxidase, and regulate thyroid function. However, in iodine deficiency, iodolactones are obsolete, and the thyroid gland is susceptible to oxidative damage and loss of cell-cycle control…In chronic iodine deficiency, compensatory mechanisms fail to maintain iodine concentration, and the thyroid is susceptible to damage. As iodine concentrations decrease, the pituitary gland secretes thyroid-stimulating hormone (TSH) to induce NIS expression. TSH remains high, and the NIS fails to shut down, predisposing the gland to excess iodine during supplementation, a phenomenon found in individuals with preexisting autoimmune thyroiditis (AIT). In addition, thyroid activity remains high, leading to hyperplasia and goiter. Researchers in cross-sectional studies reported that patients with nodular goiters often have an increase in thyroid antibodies, which is more common in iodine deficiency. These observations suggest that immune stimulation to thyroid proteins occurs during iodine deficiency and may be a causative factor in the development of thyroid autoimmunity…In iodine deficiency, the thyroid is susceptible to oxidative damage. As TSH stimulates NIS and TPO activity, low levels of iodinated lipids with high cytosolic free calcium allow for excess H2O2 to be produced. Iodine alone fails to initiate immune activation; rather, experimental studies show that iodine in the presence of inflammatory cytokines augments immune function. In an in vitro study, researchers observed increased ICAM-1 mRNA expression in the presence of potassium iodide only when low-dose IFN-γ was added. In the presence of excess H2O2, oxidative damage to TPO and TG cells leads to antigen presentation and lymphocytic infiltration, which facilitates the production of anti-TPO and anti-TG antibodies. This mechanism may account for the prevalence of AIT upon iodine supplementation through USI programs…The lowest concentrations of iodine are found in individuals with thyroid autoimmunity. In 13 patients with overt symptoms of hypothyroidism and AIT, an average of 2.3 mg of iodine was found in the thyroid, as compared with 10 mg in healthy subjects. On the basis of data derived from the National Health and Nutrition Examination Survey, although iodine intake in the United States continues to decrease, the prevalence of AIT continues to increase. This correlation suggests that iodine deficiency may be a causative factor in patients with AIT. (76)

TSH Stimulates Conversion of Intrathyroidal T4 to T3

In 1990, Dr. Kohrle studied the TSH control of T4 to T3 conversion in thyrocytes finding TSH controls the activity of D1 deiodinase in thyrocytes, which converts T4 to T3. This high TSH stimulation increases the peripheral T3/T4 ratio. Low TSH reduced this ration.  Thus high Free T3 blood level is an indicator of more intense thyroid stimulation by TSH receptor antibodies in Graves’ hyperthyroidism. He writes:

TSH, the major signalling factor for the thyroid follicles, controls thyrocyte function…TSH is involved in the regulation of thyroidal uptake of small molecules and nutrients, intracellular transport of thyrocyte specific proteins, and in most of the steps of thyroid hormone synthesis, storage and release… Thyrocytes also express a highly active Type I iodothyronine 5′ deiodinase which is controlled by TSH stimulated cAMP production. The thyrocyte specific 5′ deiodinase isozyme has marked influence on the amount of T3 secreted by the thyroid. This 5′ deiodinase isozyme shows most of the characteristics of the type I 5′ deiodinase found in liver and kidney and is also blocked by PTU, other 5′ deiodinase inhibitors [methimazole and potassium iodide], and iodinated X-ray contrast agents such as iopanoic acid, which are occasionally used in thyrotoxicosis to inhibit thyroidal T3-production by this enzyme. (  Köhrle, 1990    ) Emphasis mine

Köhrle, J. “Thyrotropin (TSH) action on thyroid hormone deiodination and secretion: one aspect of thyrotropin regulation of thyroid cell biology.” Hormone and Metabolic research. Supplement Series 23 (1990): 18-28.

The Role of the NIS Sodium/ Iodide Symporter

Dohan, Orsolya, et al. “The sodium/iodide symporter (NIS): characterization, regulation, and medical significance.” Endocrine reviews 24.1 (2003): 48-77.

Ravera, Silvia, et al. “The sodium/iodide symporter (NIS): molecular physiology and preclinical and clinical applications.” Annual review of physiology 79 (2017): 261. TSH is the primary regulator of NIS in the thyroid at both the transcriptional and post-transcriptional levels (25, 31–35) (see 36 for a recent review). Another regulator of NIS function is I− itself. Wolff & Chaikoff (37) showed that when I− reaches a critical high concentration in the plasma, TH biosynthesis decreases (a.k.a. the Wolff-Chaikoff effect). However, there is an “escape” from this acute Wolff-Chaikoff effect that restores normal TH biosynthesis even in the continued presence of high plasma I− concentrations (38) (see 14 for a recent review). At the molecular level, excess I− may have a deleterious effect on the thyroid by modifying NIS mRNA stability and increasing the production of reactive oxygen species (39, 40).

Riesco-Eizaguirre, Garcilaso, Pilar Santisteban, and Antonio de la Vieja. “The complex regulation of NIS expression and activity in thyroid and extrathyroidal tissues.” (2021).

Martín, Mariano, et al. “Implications of Na+/I-symporter transport to the plasma membrane for thyroid hormonogenesis and radioiodide therapy.” Journal of the Endocrine Society 3.1 (2019): 222-234.

Eleftheriadou, Anna-Maria, et al. “Re-visiting autoimmunity to sodium-iodide symporter and pendrin in thyroid disease.” European Journal of Endocrinology 183.6 (2020): 571-580.

Micali, Salvatore, et al. “Sodium iodide symporter (NIS) in extrathyroidal malignancies: focus on breast and urological cancer.” BMC cancer 14.1 (2014): 1-12.

Shiozaki, Atsushi, et al. “Functional analysis and clinical significance of sodium iodide symporter expression in gastric cancer.” Gastric Cancer 22.3 (2019): 473-485.

Dwyer, R. M., et al. “Sodium iodide symporter-mediated radioiodide imaging and therapy of ovarian tumor xenografts in mice.” Gene Therapy 13.1 (2006): 60-66.

Nicola, Juan Pablo, et al. “The Na+/I− symporter mediates active iodide uptake in the intestine.” American Journal of Physiology-Cell Physiology 296.4 (2009): C654-C662.

Spitzweg, Christine, et al. “Expression of the sodium iodide symporter in human kidney.” Kidney international 59.3 (2001): 1013-1023.

Dr Margaret Rayman Explains Importance of Iron

TPO is a Heme Protein with Central Iron

As mentioned above, TPO, thyroperoxidase is directly responsible for organification of iodine into thyroglobulin. TPO is a heme enzyme containing a porphyrin ring with a central iron atom. Thus, thyroid hormone production is dependent on adequate iron stores.  Iron supplementation to a ferritin level of 100 μg/l is suggested by Dr Margaret Rayman who writes in 2019:

It is important to recognise that low iron stores may contribute to symptom persistence in patients treated for hypothyroidism…An example is afforded by a small study in twenty-five Finnish women with persistent symptoms of hypothyroidism, despite appropriate L-T4 [Levothyroxine] therapy, who became symptom-free when treated with oral iron supplements for 6–12 months… all had serum ferritin <60 μg/l. Restoration of serum ferritin above 100 μg/l ameliorated the symptoms in two-thirds of the women. At least 30–50 % of hypothyroid patients with persisting symptoms despite adequate L-T4 therapy may, in fact, have covert ID [iron deficiency]…Patients with AITD or hypothyroidism should be routinely screened for ID [iron deficiency]. If either ID or serum ferritin below 70 μg/l is found, coeliac disease or autoimmune gastritis may be the cause and should be treated.(Rayman, 2019)

Rayman, Margaret P. “Multiple nutritional factors and thyroid disease, with particular reference to autoimmune thyroid disease.” Proceedings of the Nutrition Society 78.1 (2019): 34-44.

Fayadat, Laurence, et al. “Role of heme in intracellular trafficking of thyroperoxidase and involvement of H2O2 generated at the apical surface of thyroid cells in autocatalytic covalent heme binding.” Journal of Biological Chemistry 274.15 (1999): 10533-10538.

Conclusion: The Nuclear Reactor Analogy:

The thyroid gland can be compared to a nuclear reactor.  Under normal working conditions the nuclear reactor generates energy safery. However when the cooling system fails, the reactor may go into catastrophic melt down. a “nuclear accident”  with leakage of radioactivity.  Similarly when the cooling system of the thyroid, the selenium based glutathione system is dysfunctional or when H202 generation is excessive, from high TSH stimulation, the thyroid gland undergoes “nuclear meltdown”, causing various thyroid pathologies, auto-immmune thyroid disease, thyroiditis, goiter, nodules, and thyroid cancer.

References

1) Shahid, Muhammad A., Muhammad A. Ashraf, and Sandeep Sharma. “Physiology, thyroid hormone.” StatPearls [Internet]. StatPearls Publishing, 2022.

The five steps of thyroid synthesis

Synthesis of Thyroglobulin: Thyrocytes in the thyroid follicles produce a protein called thyroglobulin (TG). TG does not contain any iodine, and it is a precursor protein stored in the lumen of follicles. It is produced in the rough endoplasmic reticulum. Golgi apparatus pack it into the vesicles, and then it enters the follicular lumen through exocytosis.

Iodide uptake: Protein kinase A phosphorylation causes increased activity of basolateral Na+-I- symporters, driven by Na+-K+-ATPase, to bring iodide from the circulation into the thyrocytes. Iodide then diffuses from the basolateral side to the apex of the cell, where it is transported into the colloid through the pendrin transporter.

Iodination of thyroglobulin: Protein kinase A also phosphorylates and activates the enzyme thyroid peroxidase (TPO). TPO has three functions: oxidation, organification, and coupling reaction.

Oxidation: TPO uses hydrogen peroxide to oxidize iodide (I-) to iodine (I2). NADPH-oxidase, an apical enzyme, generates hydrogen peroxide for TPO.

Organification: TPO links tyrosine residues of thyroglobulin protein with I2. It generates monoiodotyrosine (MIT) and diiodotyrosine (DIT). MIT has a single tyrosine residue with iodine, and DIT has two tyrosine residues with iodine.

Coupling reaction: TPO combines iodinated tyrosine residues to make triiodothyronine (T3) and tetraiodothyronine (T4). MIT and DIT join to form T3, and two DIT molecules form T4.

Storage: thyroid hormones are bound to thyroglobulin for stored in the follicular lumen.

Release: thyroid hormones are released into the fenestrated capillary network by thyrocytes in the following steps:

Thyrocytes uptake iodinated thyroglobulin via endocytosis

Lysosome fuse with the endosome containing iodinated thyroglobulin

Proteolytic enzymes in the endolysosome cleave thyroglobulin into MIT, DIT, T3, and T4.

T3 (20%) and T4 (80%) are released into the fenestrated capillaries via MCT8 transporter.[5]

Deiodinase enzymes remove iodine molecules from DIT and MIT. Iodine can be salvaged and redistributed to an intracellular iodide pool.[1][6][4]

 

2) Rousset, Bernard, et al. “Thyroid hormone synthesis and secretion, Chapter 2.” Endotext [Internet] (2015).

In this chapter “iodine” refers to the element in general, and

“molecular iodine” refers to I2.

“Iodide” refers specifically to the ion I-.

The normal thyroid maintains a concentration of free iodide 20 to 50 times higher than that of plasma, This concentration gradient may be more than 100:1 in the hyperactive thyroid of patients with Graves’ disease. The thyroid can also concentrate other ions, including bromide, astatide, pertechnetate, rhenate, and chlorate, but not fluoride (25;26).

Functional studies clearly show that NIS is responsible for most of the events previously described for iodide concentration by the thyroid. TSH stimulates NIS expression (39;40) and iodide transport (31;32). TSH exerts its regulatory action at the level of transcription through a thyroid-specific far-upstream enhancer denominated NUE (NIS Upstream Enhancer) that contains binding sites for the transcription factor Pax8 and a cAMP response element-like sequence. This original demonstration made on the rat NIS gene (41) has now been extended to human (42) and mouse (43) NIS genes. It has been suggested that TSH could also regulate NIS expression at post-transcriptional level (44). Data from TSH receptor-null mice (44-46) clearly show that TSH is required for expression of NIS.

Moderate doses of iodide in the TSH-stimulated dog thyroid inhibit expression of the mRNAs for NIS and TPO, while not affecting that for Tg and TSH receptor (40) . The decrease in thyroid iodide transport resulting from excess iodide administration (escape from the Wolff-Chaikoff effect, see further) is related to a decrease in NIS expression (40;47). Both NIS mRNA and NIS protein are suppressed by TGFb, which also inhibits iodide uptake (48;49). Reviews focus on NIS and its functional importance (50;51)

Iodide that enters the thyroid remains in the free state only briefly before it is further metabolized and bound to tyrosyl residues in Tg. A significant proportion of intrathyroidal iodide is free for about 10-20 minutes after administration of a radioactive tracer (81), but in the steady state, iodide contributes less than 1% of the thyroid total iodine. A major fraction of the intrathyroidal free iodide pool comes from deiodination of MIT and DIT; this iodide is either recycled within the thyroid or leaked into the circulation. Some data suggest that iodide entering the gland by active transport segregates from that generated by deiodination of Tg within the gland (82;83). Once in the thyroid, iodide is organically bound at a rate of 50 to 100% of the pool each minute (24;84). The proportion of an iodide load that is bound varies little, despite wide shifts in daily intake. In contrast, NIS activity is sensitive to both iodine availability and TSH stimulation, and transport rather than intrathyroidal binding is the controlling factor in making iodide available for hormonogenesis.

Extrathyroid NIS

The thyroid is not the only organ to concentrate iodine; the others endowed with this capacity are salivary glands, gastric mucosa, mammary glands, and choroid plexus. Ductal cells of the salivary glands express NIS (67) . The plasma membrane of the mammary gland epithelium contains a NIS protein with a molecular mass different from that of thyroid NIS (~75 kDa vs ~90 kDa). In the mammary gland, NIS is processed differently after translation and subjected to regulation by lactogenic stimuli (68). It has been reported that over 80% of human breast cancer samples express this symporter. As it is absent in normal non-lactating tissue, NIS may represent a marker for breast malignancy and even a possible target for radioiodine therapy (69). The thyroid, salivary glands, and gastric mucosa share a common embryologic derivation from the primitive alimentary tract and, in each of these tissues; iodide transport is inhibited by thiocyanate, perchlorate, and cardiac glycosides. TSH stimulates transport only in the thyroid. An active transport for iodide in the gastric mucosa has an obvious value because it provides iodine to the circulation for use in the thyroid. Active concentration by the breast helps transfer iodide to milk. Iodide concentration by the choroid plexus and salivary glands does not have any obvious physiologic benefit, but needs to be remembered for possible insights into pathways as yet undiscovered.

Thyroperoxidase oxidizes iodide in the presence of H2O2. TPO synthesized on polysomes is inserted in the membrane of the endoplasmic reticulum and undergoes core glycosylation. TPO is then transported to the Golgi where it is subjected to terminal glycosylation and packaged into transport vesicles along with Tg (112) (Fig. 2-6). These vesicles fuse with the apical plasma membrane in a process stimulated by TSH. TPO delivered at the apical pole of thyrocytes exposes its catalytic site with the attached heme in the thyroid follicular lumen (113). TPO activity is restricted to the apical membrane, but most of the thyroid TPO is intracellular, being located in the perinuclear part of the endoplasmic reticulum (114;115). Most of this intracellular protein is incompletely or improperly folded; it contains only high mannose-type carbohydrate units, while the membrane TPO has complex carbohydrate units. Glycosylation is essential for enzymatic activity (115). Chronic TSH stimulation increases the amount of TPO and its targetting at the apical membrane (116).

Thyroglobulin is the most abundant protein in the thyroid gland; its concentration within the follicular lumen can reach 200-300 g/L. Its main function is to provide the polypeptide backbone for synthesis and storage of thyroid hormones (180). It also offers a convenient depot for iodine storage and retrieval when external iodine availability is scarce or uneven. Neosynthesised Tg polypeptide chains entering the lumen of the rough endoplasmic reticulum (RER) are subjected to core glycosylation, dimerise and are transferred to the Golgi where they undergo terminal glycosylation (Fig. 2-7). Iodination and hormone formation of Tg occur at the apical plasma membrane-lumen boundary and the mature hormone-containing molecules are stored in the follicular lumen, where they make up the bulk of the thyroid follicle colloid content.

The step preliminary to thyroid hormone formation is the attachment of iodine to tyrosyl residues in Tg to produce MIT and DIT. This process occurs at the apical plasma membrane-follicle lumen boundary and involves H2O2, iodide, TPO, and glycosylated Tg. All rendezvous at the apical membrane to achieve Tg iodination (Fig. 2-8). First, iodide must be oxidized to an iodinating form. The final step in hormone synthesis is the coupling of two neighbouring iodotyrosyl residues to form iodothyronine

 

These findings suggested that iodination of lipids impairs H2O2 production and, therefore, decreases further Tg iodination. This is the most probable mechanism for the Wolff-Chaikoff effect (128).

The most important controlling factors are iodine availability and TSH. Inadequate amounts of iodine lead to inadequate thyroid hormone production, increased TSH secretion and thyroid stimulation, and goiter in an attempt to compensate. Excess iodide acutely inhibits thyroid hormone synthesis, the Wolff-Chaikoff effect (243), apparently by inhibiting H2O2 generation, and therefore, blocking Tg iodination (127). A proposed mechanism is that the excess iodide leads to the formation of 2-iodohexadecanal (255), which is endowed with an inhibitory action on H2O2 generation.

To summarize, TSH stimulates the expression of NIS, TPO, Tg and the generation of H2O2 , increases formation of T3 relative to T4, alters the priority of iodination and hormonogenesis among tyrosyls and promotes the rapid internalization of Tg by thyrocytes. These several steps are interrelated and have the net effects of increasing the amount of iodine available to the cells and of making and releasing a larger amount and a more effective type of thyroid hormone (T3).

Anti-thyroid drugs are external compounds influencing thyroid hormone synthesis. The major inhibitory drugs are the thionamides: propylthiouracil and methimazole. In the thyroid, they appear to act by competing with tyrosyl residues of Tg for oxidized iodine, at least in the rat (219). Iodotyrosyl coupling is also inhibited by these drugs and appears more sensitive to their effects than does tyrosyl iodination.

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3) Miot, Françoise, et al. “Thyroid hormone synthesis and secretion.” Thyroid disease manager (2010).

Central:Thyroid hormones (T4 and T3) are produced by the follicular cells of the thyroid gland and are regulated by TSH made by the thyrotropes of the anterior pituitary gland. The effects of T4 in vivo are mediated via T3 (T4 is converted to T3 in target tissues). T3 is 3- to 5- fold more active than T4.

Thyroxine (3,5,3′,5′-tetraiodothyronine) is produced by follicular cells of the thyroid gland. It is produced as the precursor thyroglobulin (this is not the same as TBG), which is cleaved by enzymes to produce active T4.

The steps in this process are as follows:

The Na+/I- symporter transports two sodium ions across the basement membrane of the follicular cells along with an iodine ion. This is a secondary active transporter that utilises the concentration gradient of Na+ to move I- against its concentration gradient.

I- is moved across the apical membrane into the colloid of the follicle.

Thyroperoxidase oxidises two I- to form I2. Iodide is non-reactive, and only the more reactive iodine is required for the next step.

The thyroperoxidase iodinates the tyrosyl residues of the thyroglobulin within the colloid. The thyroglobulin was synthesised in the ER of the follicular cell and secreted into the colloid.

Thyroid-stimulating hormone (TSH) released from the pituitary gland binds the TSH receptor ( a Gs protein-coupled receptor) on the basolateral membrane of the cell and stimulates the endocytosis of the colloid.

The endocytosed vesicles fuse with the lysosomes of the follicular cell. The lysosomal enzymes cleave the T4 from the iodinated thyroglobulin.

These vesicles are then exocytosed, releasing the thyroid hormones.

Thyroxine is produced by attaching iodine atoms to the ring structures of tyrosine molecules. Thyroxine (T4) contains four iodine atoms. Triiodothyronine (T3) is identical to T4, but it has one less iodine atom per molecule.

Iodide is actively absorbed from the bloodstream by a process called iodide trapping. In this process, sodium is cotransported with iodide from the basolateral side of the membrane into the cell and then concentrated in the thyroid follicles to about thirty times its concentration in the blood. Via a reaction with the enzyme thyroperoxidase, iodine is bound to tyrosine residues in the thyroglobulin molecules, forming monoiodotyrosine (MIT) and diiodotyrosine (DIT). Linking two moieties of DIT produces thyroxine. Combining one particle of MIT and one particle of DIT produces triiodothyronine.

DIT + MIT → r-T3 (biologically inactive)

MIT + DIT → triiodothyronine (usually referred to as T3)

DIT + DIT → thyroxine (referred to as T4)

 

Proteases digest iodinated thyroglobulin, releasing the hormones T4 and T3, the biologically active agents central to metabolic regulation.

 

Peripheral:

Thyroxine is believed to be a prohormone and a reservoir for the most active and main thyroid hormone T3. T4 is converted as required in the tissues by iodothyronine deiodinase. Deficiency of deiodinase can mimic an iodine deficiency. T3 is more active than T4 and is the final form of the hormone, though it is present in less quantity than T4.

4) Song, Yue, et al. “Roles of hydrogen peroxide in thyroid physiology and disease.” The Journal of Clinical Endocrinology & Metabolism 92.10 (2007): 3764-3773.

H2O2 in various cell types, and presumably in thyroid
cells, is a signal, a mitogen, a mutagen, a carcinogen, and a killer. The

To synthesize thyroid hormones, the thyrocyte takes up iodide from the blood and extracellular fluid and oxidizes it to bind it to selected tyrosines of thyroglobulin. Iodide is actively transported by the Na+/I− symporter in the cell at the basal membrane and leaked out along the electrical gradient (from the negative interior to the positive exterior) by an iodide channel at the apical membrane. Pendrin is a candidate for this role. Iodide in the follicular lumen is oxidized at the apical membrane by thyroperoxidase (TPO) using H2O2 as the other substrate. The latter originates from an H2O2-generating system whose main enzymes are the recently cloned thyroid DUOX1 and DUOX2 (45, 46). Oxidized iodide is linked covalently to tyrosines of thyroglobulin by TPO (47). The same system, by an oxidizing reaction, links covalently some iodotyrosines into iodothyronines within thyroglobulin. H2O2 is produced in large excess compared with the amounts of iodide incorporated into proteins. This may be necessary owing to the relatively high Michaelis Menten constant (Km) of TPO for H2O2 (48, 49). It is interesting that iodide leakage, i.e. presumably the iodide channel that releases iodide at the apical membrane, is acutely regulated by the same cascades and with the same timing as H2O2 generation (50).

In all species studied, iodide at high concentrations presumably through an iodinated lipid, iodohexadecanal, inhibits H2O2 generation (the Wolff-Chaikoff effect) and adenylate cyclase (62–64).

The dynamics of TPO localization in the rat thyrocyte has been beautifully demonstrated by the groups of Wollman and Ekholm (67, 68). We presume DUOX behaves similarly (Fig. 2). TPO in the thyrocyte at rest is concentrated in secretory granules just inside of the apical membrane. The iodination system is inactive there. After TSH stimulation, the granules fuse to the membrane between microvilli, allowing TPO to migrate to the microvillous membrane. It never locates on the pseudopods engulfing thyroglobulin. The main location of TPO and iodination and therefore presumably DUOX is thus in the microvillous membranes (68–70). This is supported by the demonstration by histochemistry-electron microscopy of NOX activity in the microvilli (71). Such a localization already appears, early in evolution, in the endostyle of larval amphioxus (72). In lung epithelium and in the gastrointestinal mucosa, DUOX also preferentially localizes at the brush border (51, 52).

At high concentration (above 0.1 mm), H2O2 induces apoptosis in thyroid cells, and at even higher levels (above 0.4 mm), necrosis (80–82), an effect that is potentiated by selenium deprivation and consequent GSH peroxidase depletion.

Also, iodide, at high concentration through an iodinated lipid derivative, most probably iodohexadecanal, inhibits H2O2 generation by open follicles (73, 93).

The thyroid cell contains all the biochemical systems that detoxify H2O2 in other cells, notably the selenoproteins: GSH peroxidases (cytoplasmic, plasma, and phospholipid) and thioredoxin reductases (104, 105). The concept of the TSH role in the activation of H2O2 generation and of the protective role of GSH peroxidase coupled to the hexose monophosphate pathway had already been developed in 1971 (84).

On the other hand, although the DUOX defect would suppress H2O2 generation, a TPO defect might increase H2O2 generation by decreasing its inactivation and allowing an increased generation in response to TSH.

We have repeatedly suggested that the important generation of H2O2 in thyroid cells might account for mutagenesis and the important generation of nodules in the thyroid (79, 118). This would also explain in part why more nodules are found in iodine-deficient areas.

Excess H2O2 in thyroid is neutralized in the thyrocytes by second-line mechanisms, the most efficient ones being GSH peroxidases, peroxiredoxin, and other Se-containing enzymes. Se deficiency should weaken such defenses. Indeed, low levels of serum Se have been associated with thyroid cancer

Myxedematous endemic cretinism, caused by thyroid destruction after birth, has been linked to low iodine supply in early life, leading to intense stimulation and presumably H2O2 generation, to passage from low O2 to high O2 at birth, to selenium deficiency, and thus to decreases in GSH peroxidase and thioredoxin reductase activity and to dietary thiocyanate. The experimental reproduction of this scenario in newborn rats confirms the validity of these conclusions (105, 123).

Interestingly, a similar scenario has been proposed for the physiopathology of thyroiditis (120, 124). Selenium dietary supplementation has therefore been proposed for prevention and treatment of thyroiditis and has indeed alleviated it (124, 125).

 

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5) Nakamura, Masao, et al. “Iodination and oxidation of thyroglobulin catalyzed by thyroid peroxidase.” Journal of Biological Chemistry 259.1 (1984): 359-364.

Thyroid peroxidase plays catalytic roles in two steps of the biosynthesis of thyroxine:

(a) the iodination of tyrosine residues of thyroglobulin and

(b) the coupling of diiodotyrosine to produce thyroxine

thyroid peroxidase oxidizes iodide to atomic iodine (I) or iodinium (I+).

The “organification of iodine,” the incorporation of iodine into thyroglobulin for the production of thyroid hormone, is nonspecific; that is, there is no TPO-bound intermediate, but iodination occurs via reactive iodine species released from TPO.[4]

The chemical reactions catalyzed by thyroid peroxidase occur on the outer apical membrane surface and are mediated by hydrogen peroxide.

6) Czarnocka, Barbara. “Thyroperoxidase, thyroglobulin, Na (+)/I (-) symporter, pendrin in thyroid autoimmunity.” Frontiers in Bioscience-Landmark 16.2 (2011): 783-802.

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Location of thyro peroxidase

7) Strum, Judy M., and Morris J. Karnovsky. “Cytochemical localization of endogenous peroxidase in thyroid follicular cells.” The Journal of cell biology 44.3 (1970): 655-666.

Peroxidase activity is found in thyroid follicular cells in the following sites: apical vesicles, associated with the external surfaces of the microvilli that project apically from the cell into the colloid.

In keeping with the radioautographic evidence of others and the postulated role of thyroid peroxidase in iodination, it is suggested that the microvillous apical cell border is the major site where iodination occurs.

 

Mutation in the ThYROperoxides Gene- cannot organify iodine

8) Wu, J. Y., et al. “Mutation analysis of thyroid peroxidase gene in Chinese patients with total iodide organification defect: identification of five novel mutations.” Journal of Endocrinology 172.3 (2002): 627-635.

Total iodide organification defect (TIOD), where the iodide in the thyroid gland cannot be oxidized and/or bound to the protein, is caused by a defect in the thyroid peroxidase (TPO) gene. Single strand conformation polymorphism analysis was used to screen for mutations in the TPO gene from five unrelated TIOD patients in Taiwan, and five novel mutations were detected.

 

9) NAGASAKA, AKIO, and HIROYOSHI HIDAKA. “Effect of antithyroid agents 6-propyl-2-thiouracil and l-methyl-2-mercaptoimidazole on human thyroid iodide peroxidase.” The Journal of Clinical Endocrinology & Metabolism 43.1 (1976): 152-158.

The mechanism of inhibition of human thyroid iodide peroxidase (TPO) by 6-propyl-2-thiouracil (PTU) and 1-methyl-2-mercaptoimidazole (MMI) used in the therapy of hyperthyroid patients was studied in vitro. MMI interacted directly with TPO and inhibited enzyme activity, rather than interacting with the product (oxidized iodide). The inhibition was irreversible with MMI, but reversible with PTU. These data may explain why MMI is a more potent inhibitor of iodination than PTU and may fit the clinical results observed when hyperthyroid patients are treated with these agents.

10) Roy, Gouriprasanna, and Govindasamy Mugesh. “Selenium analogues of antithyroid drugs–recent developments.” Chemistry & Biodiversity 5.3 (2008): 414-439.

Thyroxine (T4), the main secretory hormone of the thyroid gland, is produced on thyroglobulin by thyroid peroxidase (TPO)/H(2)O(2)/iodide system and deiodinated to its active form (T3) by a selenocysteine-containing enzyme, iodothyronine deiodinase (ID). The activation of thyroid-stimulating hormone (TSH) receptor by auto-antibodies leads to ‘hyperthyroidism’, a life-threatening disease which is treated by antithyroid drugs such as 6-propyl-2-thiouracil (PTU) and methimazole (MMI). The present review describes the biological activities of a number of S/Se derivatives bearing the methimazole pharmacophore. It is shown that the isosteric substitutions in the existing drugs lead to compounds that can effectively and reversibly inhibit the heme-containing lactoperoxidase (LPO). In contrast to methimazole, the selenium analogue, MSeI, does not interfere with the enzyme directly, but it inhibits LPO by reducing the H(2)O(2) that is required for the oxidation of the Fe-center in LPO.

 

11) Schmutzler, Cornelia, et al. “Selenoproteins of the thyroid gland: expression, localization and possible function of glutathione peroxidase 3.” (2007): 1053-1059.

The thyroid gland has an exceptionally high selenium content, even during selenium deficiency. At least 11 selenoproteins are expressed, which may be involved in the protection of the gland against the high amounts of H2O2 produced during thyroid hormone biosynthesis. As determined here by in situ hybridization and Northern blotting experiments, glutathione peroxidases (GPx) 1 and 4 and selenoprotein P were moderately expressed, occurring selectively in the follicular cells and in leukocytes of germinal follicles of thyroids affected by Hashimoto’s thyroiditis. Selenoprotein 15 was only marginally expressed and distributed over all cell types. GPx3 mRNA was exclusively localized to the thyrocytes, showed the highest expression levels and was down-regulated in 5 of 6 thyroid cancer samples as compared to matched normal controls. GPx3 could be extracted from thyroidal colloid by incubation with 0.5% sodium dodecyl sulfate indicating that this enzyme is (i) secreted into the follicular lumen and (ii) loosely attached to the colloidal thyroglobulin. These findings are consistent with a role of selenoproteins in the protection of the thyroid from possible damage by H2O2. Particularly, GPx3 might use excess H2O2 and catalyze the polymerization of thyroglobulin to the highly cross-linked storage form present in the colloid.

 

12) Schweizer, Ulrich, Jazmin Chiu, and Josef Köhrle. “Peroxides and peroxide-degrading enzymes in the thyroid.” Antioxidants & redox signaling 10.9 (2008): 1577-1592.

Iodination of thyroglobulin is the key step of thyroid hormone biosynthesis. It is catalyzed by thyroid peroxidase and occurs within the follicular space at the apical plasma membrane. Hydrogen peroxide produced by thyrocytes as an oxidant for iodide may compromise cellular and genomic integrity of the surrounding cells, unless these are sufficiently protected by peroxidases. Thus, peroxidases play two opposing roles in thyroid biology. Both aspects of peroxide biology in the thyroid are separated in space and time and respond to the different physiological states of the thyrocytes. Redox-protective peroxidases in the thyroid are peroxiredoxins, glutathione peroxidases, and catalase. Glutathione peroxidases are selenoenzymes, whereas selenium-independent peroxiredoxins are functionally linked to the selenoenzymes of the thioredoxin reductase family through their thioredoxin cofactors. Thus, selenium impacts directly and indirectly on protective enzymes in the thyroid, a link that has been supported by animal experiments and clinical observations. In view of this relationship, it is remarkable that rather little is known about selenoprotein expression and their potential functional roles in the thyroid. Moreover, selenium-dependent and -independent peroxidases have rarely been examined in the same studies. Therefore, we review the relevant literature and present expression data of both selenium-dependent and -independent peroxidases in the murine thyroid.

13) Mizukami, Y., F. Matsubara, and S. Matsukawa. “Cytochemical localization of peroxidase and hydrogen-peroxide-producing NAD (P) H-oxidase in thyroid follicular cells of propylthiouracil-treated rats.” Histochemistry 82.3 (1985): 263-268.

Abstract  The distribution of endogenous peroxidase and hydrogen-peroxide-producing NAD(P)H-oxidase, which are essential enzymes for the iodination of thyroglobulin, was cytochemically determined in the thyroid follicular cells of propylthiouracil (PTU)-treated rats. Peroxidase activity was determined using the diaminobenzidine technique. The presence of NAD(P)H-oxidase was determined using H2O2 generated by the enzyme; the reaction requires NAD(P)H as a substrate and cerous ions for the formation of an electron-dense precipitate. Peroxidase activity was found in the developed rough endoplasmic reticulum (rER) and Golgi apparatus, but it was also associated with the apical plasma membrane; NAD(P)H-oxidase activity was localized on the apical plasma membrane. The presence of both enzymes on the apical plasma membrane implies that the iodination of thyroglobulin occurs at the apical surface of the follicular cell in the TSH-stimulated state which follows PTU treatment.

 

14) Labato, Mary Anna. “Cytochemical localization of hydrogen peroxide generating sites in the rat thyroid gland.” Tissue and Cell 17.6 (1985): 889-900.

Abstract Sites of H2O2 generation in lightly prefixed, intact thyroid follicles were studied by two cytochemical reactions: peroxidase-dependent DAB oxidation and cerium precipitation. In both cases reaction product accumulated on the apical surface of the follicle cell at the membrane-colloid interface. The former reaction was inhibited by the peroxidase inhibitor, aminotriazole; both reactions were blocked by the presence of catalase. NADH in the medium slightly increased the amount of cerium precipitation. The ferricyanide technique for oxidoreductase activity was also applied; reaction product again was associated with the apical surface. These results strongly imply that the follicle cells have a NADH oxidizing system generating H2O2 at the apical plasma membrane.

15) De la Vieja, Antonio, et al. “Molecular analysis of the sodium/iodide symporter: impact on thyroid and extrathyroid pathophysiology.” Physiological reviews 80.3 (2000): 1083-1105.

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16) Ohye, Hidemi, and Masahiro Sugawara. “Dual oxidase, hydrogen peroxide and thyroid diseases.” Experimental biology and medicine 235.4 (2010): 424-433.

The thyroid gland is a unique endocrine organ that requires hydrogen peroxide (H(2)O(2)) for thyroid hormone formation. The molecule for H(2)O(2) production in the thyroid gland has been known as dual oxidase 2 (DUOX2). Recently, NADPH oxidase 4 (NOX4), a homolog of the NOX family, was added as a new intracellular source of reactive oxygen species (ROS) in the human thyroid gland. This review focuses on the recent progress of the DUOX system and its possible contribution to human thyroid diseases. Also, we discuss human thyroid diseases related to abnormal H(2)O(2) generation. The DUOX molecule contains peroxidase-like and NADPH oxidase-like domains. Human thyroid gland also contains DUOX1 that shares 83% similarity with the DUOX2 gene. However, thyroid DUOX1 protein appears to play a minor role in H(2)O(2) production. DUOX proteins require DUOX maturation or activation factors (DUOXA1 or 2) for proper translocation of DUOX from the endoplasmic reticulum to the apical plasma membrane, where H(2)O(2) production takes place. Thyroid cells contain antioxidants to protect cells from the H(2)O(2)-mediated oxidative damage. Loss of this balance may result in thyroid cell dysfunction and thyroid diseases. Mutation of either DUOX2 or DUOXA2 gene is a newly recognized cause of hypothyroidism due to insufficient H(2)O(2) production. Papillary thyroid carcinoma, the most common thyroid cancer, is closely linked to the increased ROS production by NOX4. Hashimoto’s thyroiditis, a common autoimmune thyroid disease in women, becomes conspicuous when iodide intake increases. This phenomenon may be explained by the abnormality of iodide-induced H(2)O(2) or other ROS in susceptible individuals. Discovery of DUOX proteins and NOX4 provides us with valuable tools for a better understanding of pathophysiology of prevalent thyroid diseases.

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In dog thyroid slices thyrotropin [TSH] and carbamylcholine greatly enhance protein iodination and H2O2 generation. 

17) Corvilain, Bernard, Jacqueline Van Sande, and Jacques E. Dumont. “Inhibition by iodide of iodide binding to proteins: the “Wolff-Chaikoff” effect is caused by inhibition of H2O2 generation.” Biochemical and biophysical research communications 154.3 (1988): 1287-1292.H2O2 generation is limiting the oxidation and binding to proteins of iodide.

In dog thyroid slices thyrotropin [TSH] and carbamylcholine greatly enhance protein iodination and H2O2 generation. The action of thyrotropin is mimicked by dibutyryl cyclic AMP and forskolin which suggests that it is mediated by cyclic AMP. The action of carbamylcholine was mimicked by ionomycin and by phorbol myristate ester. This suggests that the effect of carbamylcholine is mediated by the two intracellular signals generated by the Ca++ phosphatidylinositol cascade: Ca++ and diacylglycerol. The Wolff-Chaikoff effect is the inhibition by iodide of its own organification.

In dog thyroid slices, iodide greatly inhibited H2O2 generation stimulated by thyrotropin and by carbamylcholine. Iodide decreased the production of intracellular signals induced by TSH and carbamylcholine but it also inhibited the action of probes of these intracellular signals (dibutyryl cAMP, forskolin, ionomycin, phorbol-myristate ester) on the H2O2 generating system itself. These effects were suppressed by methimazole an inhibitor of iodide oxidation.

17B) Corvilain, Bernard, et al. “Stimulation by iodide of H2O2generation in thyroid slices from several species.” American Journal of Physiology-Endocrinology and Metabolism 278.4 (2000): E692-E699.

In comparison with conditions in which an inhibitory effect of iodide on H2O2 generation is observed, the stimulating effect was observed for lower concentrations and for a shorter incubation time with iodide. Such a dual control of H2O2 generation by iodide has the physiological interest of promoting an efficient oxidation of iodide when the substrate is provided to a deficient gland and of avoiding excessive oxidation of iodide and thus synthesis of thyroid hormones when it is in excess. The activation of H2O2 generation may also explain the well described toxic effect of acute administration of iodide on iodine-depleted thyroids.

17C)Poncin, Sylvie, et al. “Oxidative stress in the thyroid gland: from harmlessness to hazard depending on the iodine content.” Endocrinology 149.1 (2008): 424-433.

17D Sun, Rong, et al. “Protection of Vitamin C on Oxidative Damage Caused by Long-Term Excess Iodine Exposure in Wistar Rats.” Nutrients 14.24 (2022): 5245.

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18) Markou, K., et al. “Iodine-induced hypothyroidism.” Thyroid 11.5 (2001): 501-510. Markou K Iodine induced hypothyroidism Thyroid 2001

19) Benvenga, Salvatore, et al. “Thyroid gland: anatomy and physiology.” Encyclopedia of Endocrine Diseases 4 (2018): 382-390.

20) Harper, Mary-Ellen, and Erin L. Seifert. “Thyroid hormone effects on mitochondrial energetics.” Thyroid 18.2 (2008): 145-156.

thyroid hormones stimulate mitochondriogenesis and thereby augment cellular oxidative capacity. Thyroid hormones induce substantial modifications in mitochondrial inner membrane protein and lipid compositions. Results are consistent with the idea that thyroid hormones activate the uncoupling of oxidative phosphorylation through various mechanisms involving inner membrane proteins and lipids. Increased uncoupling appears to be responsible for some of the hypermetabolic effects of thyroid hormones. ATP synthesis and turnover reactions are also affected. There appear to be complex relationships between mitochondrial proton leak mechanisms, reactive oxygen species production, and thyroid status

21) Cioffi, Federica, et al. “Bioenergetic Aspects of Mitochondrial Actions of Thyroid Hormones.” Cells 11.6 (2022): 997.

almost all components of the respiratory chain are directly or indirectly affected by iodothyronines (see, Table 1). In some cases, the actions result in an activation of specific biochemical pathways while in others the effects would result into an increase in mRNA or protein levels of specific components of respiratory chain.

22) Szanto, Ildiko, Marc Pusztaszeri, and Maria Mavromati. “H2O2 metabolism in normal thyroid cells and in thyroid tumorigenesis: focus on NADPH oxidases.” Antioxidants 8.5 (2019): 126.

23) Ghaddhab, Chiraz, et al. “Factors contributing to the resistance of the thyrocyte to hydrogen peroxide.” Molecular and cellular endocrinology 481 (2019): 62-70.

24) Fortunato, Rodrigo Soares, et al. “Functional consequences of dual oxidase-thyroperoxidase interaction at the plasma membrane.” The Journal of Clinical Endocrinology & Metabolism 95.12 (2010): 5403-5411.

25) Lanzolla, Giulia, Michele Marinò, and Claudio Marcocci. “Selenium in the treatment of Graves’ hyperthyroidism and eye disease.” Frontiers in Endocrinology 11 (2021): 608428.

26) Ruggeri, Rosaria M., et al. “Selenium exerts protective effects against oxidative stress and cell damage in human thyrocytes and fibroblasts.” Endocrine 68.1 (2020): 151-162.

27) Schomburg, Lutz, and Josef Köhrle. “On the importance of selenium and iodine metabolism for thyroid hormone biosynthesis and human health.” Molecular nutrition & food research 52.11 (2008): 1235-1246.

28) Brix, Klaudia, et al. “Thyroglobulin storage, processing and degradation for thyroid hormone liberation.” The Thyroid and Its Diseases. Springer, Cham, 2019. 25-48.

29) Fallahi, Poupak, et al. “Myo-inositol in autoimmune thyroiditis, and hypothyroidism.” Reviews in Endocrine and Metabolic Disorders 19.4 (2018): 349-354.

30) Ferrari, Silvia Martina, et al. “The protective effect of myo-inositol on human thyrocytes.” Reviews in Endocrine and Metabolic Disorders 19.4 (2018): 355-362.

31) Benvenga, Salvatore, et al. “The role of inositol in thyroid physiology and in subclinical hypothyroidism management.” Frontiers in Endocrinology (2021): 458.

32) Portulano, Carla, Monika Paroder-Belenitsky, and Nancy Carrasco. “The Na+/I− symporter (NIS): mechanism and medical impact.” Endocrine reviews 35.1 (2014): 106-149.

The Na+/I− symporter (NIS) is the plasma membrane glycoprotein that mediates active I− transport in the thyroid and other tissues, such as salivary glands, stomach, lactating breast, and small intestine.

Since the cloning of NIS in 1996, NIS research has become a major field of inquiry

The significance of the thyroid gland for human health is difficult to overstate, given the wide-ranging effects of the thyroid hormones (THs) on prenatal and early development as well as on intermediary metabolism at all stages of life. Thyroid function, in turn, depends on an adequate supply of iodide (I−), an essential constituent of the THs. I− reaches the thyroid via a highly specialized I−-concentrating system, the Na+/I− symporter (NIS), the key plasma membrane transport protein located on the basolateral surface of thyrocytes.

Inadequate dietary I− intake results in hypothyroidism and goiter at all ages. Mild ID also has a significant impact on reproductive health, increasing miscarriages and stillbirths, and is the most common cause of reproductive failure worldwide (9). In addition, it is associated with increased perinatal mortality in the population (9, 10).

Insufficient I− intake in critical windows of development during fetal life and childhood results in the most dramatic manifestation of ID disorders (IDDs), ie, neurological cretinism, characterized by severe and permanent growth and cognitive impairment. The TH T4, which includes 4 iodine atoms in its structure (Figure 1 and Section III.B.), has a profound effect on brain development; it is required for myelination, somatogenesis, neuronal differentiation, and formation of neural processes, and its function is critical for proper development of the cerebral cortex, cochlea, and basal ganglia during the first trimester of gestation and for brain growth and differentiation in the third trimester

More subtly, suboptimal I− intake is a cause of reduced mental ability, lower school and work performance, and loss of an estimated 13.5 intelligence quotient (IQ) points at a population level (11) and intellectual potential (for a brief overview, see Refs. 11–13). The urinary I− (UI) concentration (normal value >100 μg/L) is a significant indicator of thyroid function and the most useful measure of I− status of a given population. Even in I−-sufficient countries, suboptimal I− intake still affects a significant number of people. In the United States, ∼8% of the population has a UI <50 μg/L. This percentage increases in women of reproductive age to 14.8%, and ∼11% of pregnant women have also been reported to have a UI <50 μg/L, both numbers higher than in previous years (14, 15). Strikingly, ID is still the most common cause of preventable brain damage after starvation.

NIS (5) (Figure 2), which is located on the basolateral membrane of thyroid epithelial cells

As indicated above, THs thyroid hormones are required for the development and maturation of the central nervous system (CNS), lungs, and skeleton (21, 24–26). They are master regulators of cell metabolism (lipid, protein, and carbohydrate anabolism and catabolism). Thus, their deficiency or excess perturbs homeostasis and leads to disease.

The functional units of the mature thyroid gland are the thyroid follicles, which are enclosed by a basement membrane. Histologically, the spherical follicles are lined by a monolayer of polarized epithelial cells, with the basolateral surface facing the bloodstream and the apical surface the follicular lumen (Figure 1). The lumen is filled with colloid, which is mainly composed of thyroglobulin (Tg), a protein present at a very high concentration (∼100–750 mg/mL) (27) that serves as the backbone for the THs. I− is covalently incorporated into Tg, a process called I− organification; iodinated Tg is the precursor and storage form of the THs, which are released into the bloodstream upon stimulation by TSH (see Section III.B. for more details). The thyroid is unique among endocrine glands in that its hormonal products are stored extracellularly in the colloid.

One candidate to be an apical I− transporter is pendrin, In conclusion, although various candidate proteins that may mediate I− translocation across the apical membrane of thyrocytes have been proposed, at the present time, the identity of the I− apical translocation pathway remains poorly understood.

The oxidative power for I− organification is provided by H2O2 generated by dual oxidase 2 (Duox2), a multispanning membrane protein belonging to the family of the nicotinamide adenine dinucleotide phosphate-dependent flavin adenine dinucleotide oxidases. The name dual oxidase stems from the presence of a lumen-facing domain that exhibits >45% amino acid similarity with TPO, with putative and controversial peroxidase activity (44–46). Interestingly, Duox2 shares 47% amino acid similarity to gp91phox (Nox2), which is responsible for the respiratory burst and the production of reactive oxygen species (ROS) in neutrophils (45). In contrast to most of the proteins involved in TH biosynthesis, Duox2 expression and activity do not appear to be regulated by TSH.

When TSH serum levels increase, all steps of TH biosynthesis and release are stimulated, including Tg internalization.

Extrathyroidal accumulation of I− has been reported in the salivary glands, gastric mucosa, small intestine, lactating mammary gland, choroid plexus, and the ciliary body of the eye (127, 186). However, in contrast to the thyroid, I− is not organified and I− uptake is not regulated by TSH in these tissues

 

In addition to NIS, TSH also upregulates expression of TPO and Tg as well as iodinated Tg endocytosis. Levy et al (113) have shown that thyroid NIS protein expression is upregulated by TSH in vivo.

 

In addition to TSH, I− itself is a major regulator of I− accumulation in the thyroid. The ability to block thyroid function with high I− was described by Plummer (219) as early as 1923. In 1948, Wolff and Chaikoff (220) reported that high plasma I− levels blocked I− organification in rat thyroid in vivo. The ability of high doses of I− to decrease thyroid function became known as the Wolff-Chaikoff effect (Figure 11A). However, this inhibitory effect of I− lasts 2 days in the presence of high plasma I, after which TH biosynthesis is restored (221). This adaptation to persistent high I was termed the escape from the Wolff-Chaikoff effect (Figure 11B).

What is the mechanism for the escape/decrease in I− transport? In 1986, it was reported that I− preincubation suppressed I− uptake in FRTL-5 cells in a time- and dose-dependent manner, an effect preventable with methimazole (MMI), which inhibits I− organification (223). Inhibition by MMI suggested that the Wolff-Chaikoff effect was mediated by an intracellular iodinated compound. Consistent with the observed reduction in I− transport, later studies demonstrated an I−-induced reduction in NIS and TPO mRNA in dog thyroid (224) and NIS mRNA and I− transport in FRTL-5 cells (225). A decrease in NIS mRNA and protein as well was demonstrated 24 hours after incubation with high I−, but no correlation was made with NIS activity (226).

High concentrations of I−, presumably in the form of iodolipids, inhibit the generation of H2O2 (229–231). Interestingly, iodolipids have been found in rat, horse, dog, and human thyroid. These iodolipids inhibit Duox and adenylyl cyclase, suggesting that they are involved in regulatory actions of I− in the thyroid (232, 233).

33) Serrano-Nascimento, Caroline, et al. “The acute inhibitory effect of iodide excess on sodium/iodide symporter expression and activity involves the PI3K/Akt signaling pathway.” Endocrinology 155.3 (2014): 1145-1156.

34) Eleftheriadou, Anna-Maria, et al. “Re-visiting autoimmunity to sodium-iodide symporter and pendrin in thyroid disease.” European Journal of Endocrinology 183.6 (2020): 571-580..

35) Ihnatowicz, Paulina, Paweł Wątor, and Małgorzata Drywień. “Supplementation in Autoimmune Thyroid Hashimoto’s Disease. Vitamin D and Selenium.” J. Food Nutr. Res 7 (2019): 584-591.

36) Tsatsoulis, Agathocles. “The role of iodine vs selenium on the rising trend of autoimmune thyroiditis in iodine sufficient countries-an opinion article.” Open Acc J Thy Res 2.1 (2018): 12-14.

37) Rostami, Rahim, et al. “Serum selenium status and its interrelationship with serum biomarkers of thyroid function and antioxidant defense in Hashimoto’s thyroiditis.” Antioxidants 9.11 (2020): 1070.\\

38) Tian, Xun, et al. “Selenium supplementation may decrease thyroid peroxidase antibody titer via reducing oxidative stress in euthyroid patients with autoimmune thyroiditis.” International Journal of Endocrinology 2020 (2020).

39) Vasiliu, Ioana, et al. “Experimental induced autoimmune thyroiditis in wistar rats: possible protective role of selenium.” Endocrine Abstracts. Vol. 70. Bioscientifica, 2020.

40) Dashdamirova, Gulnara, et al. “Pathogenic Mechanisms of Autoimmune Thyroid Disease.” Int J Med Sci Health Res 6 (2022): 26-33

41) Moncayo, Roy, Helga Moncayo, and Juliana Reisenzahn. “Global view on the pathogenesis of benign thyroid disease based on historical, experimental, biochemical and genetic data, identifying the role of magnesium, selenium, coenzyme Q10 and iron in the context of the unfolded protein response and protein quality control of thyroglobulin.” Journal of Translational Genetics and Genomics 4.4 (2020): 356-383.

42) Xu, Bin, et al. “A pilot study on the beneficial effects of additional selenium supplementation to methimazole for treating patients with Graves’ disease.” Turkish journal of medical sciences 49.3 (2019): 715-722.

43) Zhang, Xiao-Hong, Gao-Pin Yuan, and Ting-Li Chen. “Clinical effect of methimazole combined with selenium in the treatment of toxic diffuse goiter in children.” World Journal of Clinical Cases 10.4 (2022): 1190.

44) Gallo, Daniela, et al. “Add-on effect of selenium and vitamin D combined supplementation in early control of Graves’ disease hyperthyroidism during methimazole treatment.” Frontiers in Endocrinology 13 (2022).

45) Barbaro, Daniele, Beatrice Orrù, and Vittorio Unfer. “Iodine and myo-inositol: a novel promising combination for iodine deficiency.” Frontiers in Endocrinology 10 (2019): 457.

Myo-inositol, a carbocyclic polyol, regulates the generation of hydrogen peroxide (H2O2) in thyrocytes, crucial for iodine organification and thyroid hormone biosynthesis.

Either in vitro and in vivo iodide main roles are:

(1) decreasing the response of TSH;

(2) inhibiting its own oxidation (the Wolff-Chaikoff effect);

(3) reducing its trapping after a delay;

(4) at high levels, inhibiting thyroid hormone secretion (28).

The Wolff–Chaikoff effect is an autoregulatory phenomenon that prevents the thyroid gland from synthesizing and releasing large quantities of thyroid hormones (19, 29) through the inhibition of organification when inorganic iodide levels in thyrocytes are too high (30, 31). The mechanism responsible for such effect is still unknown, but it may be ascribable to the inhibitory effect of iodide on TPO or other enzymes (19). Indeed, iodide is able to block the cyclic adenosine monophosphate cascade and the Ca2+-phosphatidylinositol 4, 5-bisphosphate (PIP2) cascade in thyrocytes and to induce H2O2 generation (28). H2O2 is crucial for the TPO-catalyzed thyroid hormone formation (32). In 1993, Chen at al. demonstrated in vitro that H2O2 specifically regulates iodide transport and organification in a dose-dependent manner (33). High H2O2 concentrations inhibit these functions and can be detrimental to the thyroid, although protection mechanisms prevents damages to the thyrocytes under physiological conditions (34). Intrathyroidal H2O2 generation, first reported about 50 years ago (35), and iodination are both stimulated by TSH. H2O2 can regulate iodination either directly, as a substrate of TPO, or indirectly by regulating the activity of TPO when iodide and Tg concentrations are kept constant.

As precursor also of the second messenger phosphoinositide, myo-inositol is involved in cell signaling and regulates the activities of hormones such as TSH, follicle-stimulating hormone (FSH) and insulin (37–39).

It recently proved to be a very efficacious and safe treatment for subclinical hypothyroid patients with Autoimmune Thyroiditis (40–46). In 2013, Nordio and Pajalich demonstrated that treatment with myo-inositol plus selenium for 6 months in patients with subclinical hypothyroidism reduced significantly TSH concentrations by 31% (4.4 ± 0.9 mIU/mL vs. 3.1 ± 0.6 mIU/mL), compared to the control group treated only with selenium.

 

Interestingly, myo-inositol helps thyroid-hormone-producing cells to become more efficient and faster at building T4 (32, 41). This might be ascribable to a higher availability of iodine, whose organification is boosted by myo-inositol action. Indeed, myo-inositol is involved in one of the first steps of thyroid hormone production and modulates the H2O2-mediated iodination through the phospholipase C-dependent inositol phosphate Ca2+/diacylglycerol pathway, resulting in increased H2O2 generation (Figure 1).

Differently, the cAMP cascade, induced by the TSH activity (through the TSH receptor activation), is more involved in cell growth and differentiation, and in thyroid hormones secretion.

As myo-inositol plays a crucial role in the regulation of iodine organification, supplementation may promote faster recovery from ID. Indeed, H2O2 generated under the stimulus of myo-inositol is available for iodine incorporation inside the thyroid (33, 48). Such activity makes myo-inositol very appealing as a novel molecule to increase iodine availability.

To date, endocrinologists and gynecologists recommend taking myo-inositol for several benefits in different pathologies, from thyroid diseases to polycystic ovary syndrome and gestational diabetes. Due to its action in regulating iodine organification and thyroid hormone biosynthesis, myo-inositol supplementation along with iodine may improve thyroid functionality and possibly lead to a faster recovery from ID.   … When iodine intake is insufficient, the T3:T4 ratio increases along with the decrease of T4 production,

46) Rus-Hrincu, Florentina, Alexandra Totan, and A. M. A. Stanescu. “DUOX2, a new player on the scene of thyroid hormones.” Practica Medicala 14.3 (2019): 67.

48) Eng, Peter HK, et al. “Escape from the acute Wolff-Chaikoff effect is associated with a decrease in thyroid sodium/iodide symporter messenger ribonucleic acid and protein.” Endocrinology 140.8 (1999): 3404-3410.

49) Bogusławska, Joanna, et al. “Cellular and molecular basis of thyroid autoimmunity.” European Thyroid Journal 11.1 (2022).

TPO autoantibodies (TPOAb) occur in almost all HT patients and approximately 75% of individuals with GD, whereas their positivity in euthyroid patients may be the risk factor for future thyroid disorders (15).

The immune response to iodide transporters, sodium iodide symporter (NIS) and pendrin, has been widely discussed (24, 25

In contrast to TPO and Tg, TSHR is widely expressed in extrathyroidal tissues and cells, including lymphocytes, adipose tissue, and fibroblasts. In consequence, the presence of TSHR antibodies contributes to the extrathyroidal GD manifestations, such as Graves’ ophthalmopathy (GO), Graves’ dermopathy, or GD-associated thymus hyperplasia (1, 6). TSHR autoantigen is presented by macrophages and B cells recruited to the orbit, which leads to T cell activation (5). In turn, activated T cells initiate immunological attack to the orbital fibroblasts expressing TSHR

50) Roy, Gouriprasanna, and G. Mugesh. “Thyroid hormone synthesis and anti-thyroid drugs: A bioinorganic chemistry approach.” Journal of Chemical Sciences 118.6 (2006): 619-625.

The most widely used anti-thyroid drug methimazole (MMI) inhibits the production of thyroid hormones by irreversibly inactivating the enzyme TPO.

51) Lane, Laura C., Claire Louise Wood, and Tim Cheetham. “Graves’ disease: moving forwards.” Archives of Disease in Childhood (2022).

Thionamides block thyroid hormone synthesis by inhibiting the TPO-mediated iodination of tyrosine      residues in thyroglobulin, a vital step in the synthesis of thyroxine

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52) Sugawara, M., Y. Sugawara, and K. Wen. “Methimazole and propylthiouracil increase cellular thyroid peroxidase activity and thyroid peroxidase mRNA in cultured porcine thyroid follicles.” Thyroid: Official Journal of the American Thyroid Association 9.5 (1999): 513-518.

In conclusion, MMI and PTU at therapeutic concentrations can increase TPO mRNA and cellular TPO activity, although the 2 drugs inhibit the TPO-H2O2-mediated catalytic reaction.

Explains the 10% rebound phenomenon when switching from MMI to Iodine in Pregnancy Graves

53) Landex, N. L., J. Thomsen, and L. Kayser. “Methimazole increases H2O2 toxicity in human thyroid epithelial cells.” Acta histochemica 108.6 (2006): 431-439.

54) Roy, Gouriprasanna, and G. Mugesh. “Thyroid hormone synthesis and anti-thyroid drugs: A bioinorganic chemistry approach.” Journal of Chemical Sciences 118.6 (2006): 619-625.

55) Manna, Debasish, Gouriprasanna Roy, and Govindasamy Mugesh. “Antithyroid drugs and their analogues: synthesis, structure, and mechanism of action.” Accounts of chemical research 46.11 (2013): 2706-2715.

The prohormone thyroxine (T4) is synthesized on thyroglobulin by thyroid peroxidase (TPO), a heme enzyme that uses iodide and hydrogen peroxide to perform iodination and phenolic coupling reactions. The monodeiodination of T4 to 3,3′,5-triiodothyronine (T3) by selenium-containing deiodinases (ID-1, ID-2) is a key step in the activation of thyroid hormones. The type 3 deiodinase (ID-3) catalyzes the deactivation of thyroid hormone in a process that removes iodine selectively from the tyrosyl ring of T4 to produce 3,3′,5′-triiodothyronine (rT3).

56) Ortiz de Montellano, Paul R. “Hemes in biology.” Wiley encyclopedia of chemical biology (2007): 1-10.

Heme is a ubiquitous prosthetic group in proteins with oxidation- reduction functions. The proteins that incorporate heme as an essential component include respiratory proteins, such as cytochrome c and cytochrome c oxidase. The oxygen carriers myoglobin and hemoglobin are members of a second class of hemoproteins that have some relationship to gas sensor hemoproteins, as represented for NO by guanylate cyclase; for O2 by Hap1, FixL, and HemAT; and for CO by CooA. The largest class of hemoproteins is made up of catalytic enzymes and includes the cytochromes P450, peroxidases, peroxygenases, catalases, NO synthases, prostaglandin synthases, and thromboxane/prostacyclin synthases.

TPO is a heme protein containing  a porphyrin ring

57) de Castro, Guilherme Vieira, et al. “Synthesis of Analogues of Thyroid Hormones: Nuclear Receptor Modulators.” Orbital: The Electronic Journal of Chemistry 7.3 (2015): 282-291.

58) Pavelka, S. “Metabolism of bromide and its interference with the metabolism of iodine.” Physiological Research 53 (2004): S81-90.

59) Kalderon, Albert E., and Hendrik A. Bogaars. “Immune complex deposits in Graves’ disease and Hashimoto’s thyroiditis.” The

60) Matsuta, Morimasa. “Immunohistochemical and electron microscopic studies on Hashimoto’s thyroiditis.” Pathology International 32.1 (1982): 41-56.

61) Fujiwara, Hiroshi, et al. “Immune complex deposits in thyroid glands of patients with Graves’ disease: II. Anti-thyroglobulin and anti-microsome activities of γ-globulin eluted from thyroid homogenates.” Clinical Immunology and Immunopathology 19.1 (1981): 109-117.

62) Solis-S, Juan C., et al. “Inhibition of intrathyroidal dehalogenation by iodide.” Journal of endocrinology 208.1 (2011): 89.

63) Dugrillon, A. “Iodolactones and iodoaldehydes—mediators of iodine in thyroid autoregulation.” Experimental and clinical endocrinology & diabetes 104.S 04 (1996): 41-45.

64) Laurent, Eric, et al. “Unlike thyrotropin, thyroid-stimulating antibodies do not activate phospholipase C in human thyroid slices.” The Journal of clinical investigation 87.5 (1991): 1634-1642.

65) Freitas Ferreira, Andrea C., et al. “Thyroid Ca2+/NADPH‐dependent H2O2 generation is partially inhibited by propylthiouracil and methimazole.” European Journal of Biochemistry 270.11 (2003): 2363-2368.

66) Molnár, Ildikó, et al. “Effect of antithyroid drugs on the occurrence of antibodies against type 2 deiodinase (DIO2), which are involved in hyperthyroid Graves’ disease influencing the therapeutic efficacy.” Clinical and Experimental Medicine 19.2 (2019): 245-254.

67) Molnár, Ildikó. “Relationship between autoantibodies against type 2 deiodinase and antithyroid autoantibodies in Graves’ disease using antithyroid drug therapy.” Medical Research Archives 10.12 (2022).

68) Morand, Stanislas, et al. “Effect of iodide on nicotinamide adenine dinucleotide phosphate oxidase activity and Duox2 protein expression in isolated porcine thyroid follicles.” Endocrinology 144.4 (2003): 1241-1248.

69) Cardoso, Luciene C., et al. “Ca2+/nicotinamide adenine dinucleotide phosphate-dependent H2O2 generation is inhibited by iodide in human thyroids.” The Journal of Clinical Endocrinology & Metabolism 86.9 (2001): 4339-4343.

70) Abraham, Guy E. “The safe and effective implementation of orthoiodosupplementation in medical practice.” The Original Internist 11.1 (2004): 17-36.

71) Suzuki, Hiroshi, Hiroshi Sano, and Hisashi Fukuzaki. “Decreased cytosolic free calcium concentration in lymphocytes of magnesium-supplemented DOCA-salt hypertensive rats.” Clinical and Experimental Hypertension. Part A: Theory and Practice 11.3 (1989): 487-500.

72) Yang, Ying, et al. “Magnesium deficiency enhances hydrogen peroxide production and oxidative damage in chick embryo hepatocyte in vitro.” Biometals 19.1 (2006): 71-81.

73) Betterle, Corrado, and Renato Zanchetta. “The immunofluorescence techniques in the diagnosis of endocrine autoimmune diseases.” Autoimmunity Highlights 3.2 (2012): 67-78.

74) Wolff, Jirj, et al. “The temporary nature of the inhibitory action of excess iodide on organic iodine synthesis in the normal thyroid.” Endocrinology 45.5 (1949): 504-513.

75) Leoni, Suzana G., et al. “Regulation of Thyroid Oxidative State by Thioredoxin Reductase Has a Crucial Role in Thyroid Responses to Iodide Excess.” MOLECULAR ENDOCRINOLOGY 25.11 (2011): 1924.

76) Murphy, Robyn, Caroline Turek, and Leigh Arseneau. “The role of iodine deficiency and subsequent repletion in autoimmune thyroid disease and thyroid cancer.” Journal of Restorative Medicine 5.1 (2016): 32.

The sodium-iodide symporter (NIS) transports iodide (I) into the thyrocyte to be organified to iodine (I2) and bound to thyroglobulin (TG) by thyroid peroxidase (TPO) and hydrogen peroxide (H2O2) (Figure 1). The iodine–thyroglobulin complexes then combine to form thyroid hormones. The nicotinamide adenine dinucleotide phosphate (NADPH) oxidase system is upregulated by intracellular calcium to generate H2O2, a reactive oxygen species (ROS). To prevent excess H2O2, iodolactones negatively inhibit NADPH oxidase, and glutathione peroxidase degrades H2O21214 (Figure 1). In iodine deficiency, the loss of negative feedback on H2O2 production has been implicated in thyroid dysfunction and as a possible mechanism in the generation of autoantibodies.

The import of iodide (I) by the sodium-iodide symporter (NIS) into the thyrocyte and the subsequent organification of I to form iodine (I2).15 I2 is then is bound to thyroglobulin (TG) to form thyroid hormones, thyroxine (T4), and triiodothyronine (T3). The organification process is catalyzed by thyroid peroxidase (TPO) and hydrogen peroxide (H2O2), produced by the nicotinamide adenine dinucleotide phosphate (NADPH) oxidase system. Iodinated lipids provide negative feedback on NADPH oxidase to reduce H2O2 and oxidative damage to the thyrocyte.

Studies suggest that when iodine intake is above 1.5 mg daily, TPO synthesizes iodolactones.16 TPO facilitates the iodination of arachidonic acid and other polyunsaturated fatty acids to produce iodolactones.17 Research shows that measurable concentrations of δ-iodolactone are found in the thyroid tissue, prevent excess iodide uptake and the generation of H2O2 by NADPH oxidase, and regulate thyroid function.18

However, in iodine deficiency, iodolactones are obsolete, and the thyroid gland is susceptible to oxidative damage and loss of cell-cycle control.

In chronic iodine deficiency, compensatory mechanisms fail to maintain iodine concentration, and the thyroid is susceptible to damage. As iodine concentrations decrease, the pituitary gland secretes thyroid-stimulating hormone (TSH) to induce NIS expression.18 TSH remains high, and the NIS fails to shut down, predisposing the gland to excess iodine during supplementation, a phenomenon found in individuals with preexisting autoimmune thyroiditis (AIT).4 In addition, thyroid activity remains high, leading to hyperplasia and goiter.12 Researchers in cross-sectional studies reported that patients with nodular goiters often have an increase in thyroid antibodies, which is more common in iodine deficiency.4,21 These observations suggest that immune stimulation to thyroid proteins occurs during iodine deficiency and may be a causative factor in the development of thyroid autoimmunity.

In iodine deficiency, the thyroid is susceptible to oxidative damage. As TSH stimulates NIS and TPO activity, low levels of iodinated lipids with high cytosolic free calcium allow for excess H2O2 to be produced.16 Iodine alone fails to initiate immune activation; rather, experimental studies show that iodine in the presence of inflammatory cytokines augments immune function. In an in vitro study, researchers observed increased ICAM-1 mRNA expression in the presence of potassium iodide only when low-dose IFN-γ was added.23,27 In the presence of excess H2O2, oxidative damage to TPO and TG cells leads to antigen presentation and lymphocytic infiltration, which facilitates the production of anti-TPO and anti-TG antibodies.16,28 This mechanism may account for the prevalence of AIT upon iodine supplementation through USI programs.

The lowest concentrations of iodine are found in individuals with thyroid autoimmunity. In 13 patients with overt symptoms of hypothyroidism and AIT, an average of 2.3 mg of iodine was found in the thyroid, as compared with 10 mg in healthy subjects.16 On the basis of data derived from the National Health and Nutrition Examination Survey, although iodine intake in the United States continues to decrease, the prevalence of AIT continues to increase.21,30,31 This correlation suggests that iodine deficiency may be a causative factor in patients with AIT.

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Na+/I− symporter  zzzzzzzzzzzzzzzzzzzzzzzz

Zuckier, Lionel S., et al. “The endogenous mammary gland Na+/I− symporter may mediate effective radioiodide therapy in breast cancer.” Journal of Nuclear Medicine 42.6 (2001): 987-987.

Ślebodziński, A. B. “Ovarian iodide uptake and triiodothyronine generation in follicular fluid: The enigma of the thyroid ovary interaction.” Domestic animal endocrinology 29.1 (2005): 97-103.

Monteleone, Patrizia, Pinuccia Faviana, and Paolo Giovanni Artini. “Thyroid peroxidase identified in human granulosa cells: another piece to the thyroid-ovary puzzle?.” Gynecological Endocrinology 33.7 (2017): 574-576.

Mutinati, M., et al. “Localization of thyrotropin receptor and thyroglobulin in the bovine corpus luteum.” Animal reproduction science 118.1 (2010): 1-6.

Ravera, Silvia, et al. “The sodium/iodide symporter (NIS): molecular physiology and preclinical and clinical applications.” Annual review of physiology 79 (2017): 261.

Functional NIS is found in several extrathyroidal tissues, such as the salivary glands, stomach, and lactating breast, as well as in primary and metastatic breast cancers (7, 8). The latter findings have raised the possibility that NIS-mediated 131I− treatment may be effective in breast cancer.

One of the most remarkable properties of NIS is that it transports different substrates with different stoichiometries. NIS transports I−, thiocynate (SCN−) and chlorate (ClO3−) with a 2 Na+:1 anion electrogenic stoichiometry. In contrast, NIS transports perrhenate (ReO4−) and perchlorate (ClO4−) with a 1 Na+:1 anion electroneutral stochiometry (9). The anion ClO4−, an environmental pollutant previously known only as a competitive inhibitor of NIS, is actively transported by NIS as a substrate (10). Furthermore, NIS is increasingly used as a highly effective reporter gene for imaging techniques (11, 12).

TSH is the primary regulator of NIS in the thyroid at both the transcriptional and post-transcriptional levels (25, 31–35) (see 36 for a recent review). Another regulator of NIS function is I− itself. Wolff & Chaikoff (37) showed that when I− reaches a critical high concentration in the plasma, TH biosynthesis decreases (a.k.a. the Wolff-Chaikoff effect). However, there is an “escape” from this acute Wolff-Chaikoff effect that restores normal TH biosynthesis even in the continued presence of high plasma I− concentrations (38) (see 14 for a recent review). At the molecular level, excess I− may have a deleterious effect on the thyroid by modifying NIS mRNA stability and increasing the production of reactive oxygen species (39, 40).

NIS function is traditionally associated with the thyroid. However, active I− transport has also been also demonstrated in other organs (Figures 2 and ​and3),3), including the lacrimal sac and nasolacrimal duct, salivary glands, choroid plexus, stomach, intestine, and lactating breast (14). Reverse transcription polymerase chain reaction (RT-PCR) and immunodetection have further uncovered NIS expression in the kidney, placenta, and ovary (49–53)

NIS is expressed in the normal breast only late in pregnancy and during lactation

NIS is also expressed in mammary gland tumors in animal models of breast cancer and, more importantly, in human breast cancer samples. As many as 87% of the breast tumor samples analyzed by Tazebay et al. (7) tested positive for NIS expression, in contrast with 0% of samples of normal nonlactating breast tissue

 

Kogai, Takahiko, et al. “Retinoic acid induces sodium/iodide symporter gene expression and radioiodide uptake in the MCF-7 breast cancer cell line.” Proceedings of the National Academy of Sciences 97.15 (2000): 8519-8524.

Tazebay, Uygar H., et al. “The mammary gland iodide transporter is expressed during lactation and in breast cancer.” Nature medicine 6.8 (2000): 871-878.

Zuckier, Lionel S., et al. “The endogenous mammary gland Na+/I− symporter may mediate effective radioiodide therapy in breast cancer.” Journal of Nuclear Medicine 42.6 (2001): 987-987.

Yao, Chen, et al. “Effect of sodium/iodide symporter (NIS)-mediated radioiodine therapy on estrogen receptor-negative breast cancer.” Oncology Reports 34.1 (2015): 59-66.

Elliyanti, Aisyah, et al. “Analysis natrium iodide symporter expression in breast cancer subtypes for radioiodine therapy response.” Nuclear Medicine and Molecular Imaging 54.1 (2020): 35-42.

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Toxic Effects of Iodine Excess
Leoni, Suzana G., et al. “Regulation of Thyroid Oxidative State by Thioredoxin Reductase Has a Crucial Role in Thyroid Responses to Iodide Excess.” Molecular Endocrinology 25.11 (2011): 1924.

Iodide excess induces reactive oxygen species production and cell toxicity.  Here we analyze the effects of high I doses in rat thyroids and in PCCl3 cells in the period comprising I autoregulation (i.e. 0–72 h after I administration), focusing on NIS expression, redox state, and the expression and activity of selenoproteins. Our results show that NIS mRNA inhibition by I does not occur at the transcriptional level, because neither NIS promoter activity nor Pax8 expression or its binding to DNA was modulated. Because I uptake was inhibited much earlier than NIS protein, and no effect was observed on its subcellular localization, we suggest that I is inhibiting NIS in the plasma membrane. The increased reactive oxygen species production leads to an increase in thioredoxin reductase mRNA levels and enzyme activity, which reduces the oxidative stress. Inhibition of thioredoxin reductase at either gene expression or activity levels prevented NIS recovery, thus illustrating a new role played by this selenoprotein in the regulation of cell homeostasis and consequently in I autoregulation.

Serrano-Nascimento, Caroline, Jamile Calil-Silveira, and Maria Tereza Nunes. “Posttranscriptional regulation of sodium-iodide symporter mRNA expression in the rat thyroid gland by acute iodide administration.” American Journal of Physiology-Cell Physiology 298.4 (2010): C893-C899.

Our data show for the first time that iodide regulates NIS mRNA expression at posttranscriptional level, providing a new mechanism by which iodide exerts its autoregulatory effect on thyroid.

Last updated on by Jeffrey Dach MD

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