Inhibition, Na+/I- symporter (NIS)

14797-73-0

VLTRZXGMWDSKGL-UHFFFAOYSA-M

Perchlorate

Perchlorate ion Perchlorate ion (ClO41-) Perchlorate ion(1-) Perchlorate(1-) Perchloric acid, ion(1-)

DTXSID6024252

14797-55-8

NHNBFGGVMKEFGY-UHFFFAOYSA-N

Nitrate

Nitrate (NO3-) Nitrate ion Nitrate ion (NO3-) Nitrate ion(1-) Nitrate(1-) Nitrates/nitrites Nitrato Nitric acid, ion(1-)

DTXSID5024217

302-04-5

ZMZDMBWJUHKJPS-UHFFFAOYSA-M

Thiocyanate

Thiocyanates Isothiocyanic acid, ion(1-) Rhodanide Thiocyanate (NCS1-) Thiocyanate anion Thiocyanate ion Thiocyanic acid, ion(1-) Thiocyanide

DTXSID8047763

51-52-5

KNAHARQHSZJURB-UHFFFAOYSA-N

6-Propyl-2-thiouracil

6-Propyl-2 thiouracil (PTU) 4(1H)-Pyrimidinone, 2,3-dihydro-6-propyl-2-thioxo- 2,3-Dihydro-6-propyl-2-thioxo-4(1H)-pyrimidinone 2-Mercapto-4-hydroxy-6-n-propylpyrimidine 2-Mercapto-4-hydroxy-6-propylpyrimidine 2-Mercapto-6-propylpyrimidin-4-ol 2-Thio-4-oxo-6-propyl-1,3-pyrimidine 2-Thio-6-propyl-1,3-pyrimidin-4-one 6-n-Propyl-2-thiouracil 6-n-Propylthiouracil 6-Propyl-2-thio-2,4(1H,3H)pyrimidinedione 6-Propylthiouracil NSC 6498 NSC 70461 Procasil Propacil propiltiouracilo Propycil Propyl-Thiorist Propylthiorit propylthiouracil Propylthiouracile Propyl-Thyracil Prothiucil Prothiurone Prothycil Prothyran Protiural Thiuragyl Thyreostat II URACIL, 4-PROPYL-2-THIO- Uracil, 6-propyl-2-thio-

DTXSID5021209

60-56-0

PMRYVIKBURPHAH-UHFFFAOYSA-N

Methimazole

2H-Imidazole-2-thione, 1,3-dihydro-1-methyl- 1,3-Dihydro-1-methyl-2H-imidazole-2-thione 1-Methyl-1,3-dihydroimidazole-2-thione 1-Methyl-1H-imidazole-2-thiol 1-Methyl-2-mercapto-1H-imidazole 1-Methyl-2-mercaptoimidazole 1-Methyl-4-imidazoline-2-thione 1-Methylimidazole-2(3H)-thione 1-Methylimidazole-2-thiol 1-Methylimidazole-2-thione 2-Mercapto-1-methyl-1H-imidazole 2-Mercapto-1-methylimidazole 2-Mercapto-N-methylimidazole 4-Imidazoline-2-thione, 1-methyl- Basolan Danantizol Favistan Frentirox Imidazole-2-thiol, 1-methyl- Mercaptazole Mercazole Mercazolyl Metazolo Methimazol Methylmercaptoimidazole Metothyrin Metothyrine Metotirin N-Methyl-2-mercaptoimidazole N-Methylimidazolethiol NSC 38608 Strumazol Tapazole Thacapzol Thiamazol thiamazole Thycapzol Thymidazol Thymidazole tiamazol

DTXSID4020820

149-30-4

YXIWHUQXZSMYRE-UHFFFAOYSA-N

2-Mercaptobenzothiazole

(2(3H)-Benzothiazolethione) 2(3H)-Benzothiazolethione 1,3-Benzothiazole-2-thiol 1,3-Benzothiazole-2-thione 2,3-Dihydrobenzothiazole-2-thione 2-Benzothiazolethiol 2-Benzothiazolinethione 2-BENZOTHIAZOLTHIOL 2-Benzothiazolyl mercaptan 2-Mercapthobenzothiazole Technical 2-Mercapto-1H-benzothiazole 2-Mercaptobenzthiazole 2-Sulfanylbenzothiazole Accel M Accelerator M Aero Promoter 412 Benz-1,3-thiazolidine-2-thione Benzo[d]thiazole-2-thiol Benzothiazol-2-thiol BENZOTHIAZOLE, 2-MERCAPTO- Benzothiazole-2-thiol Benzothiazole-2-thione Benzothiazolethiol benzotiazol-2-tiol Dermacid Ekagom G Kaptaks Mebetizol Mebetizole Mebithizol MERCAPTOBENZOTHIAZOLE Mercaptobenzthiazole Nocceler M Nocceler M-P Nonflex NB NSC 2041 Perkacit MBT Pneumax MBT Royal MBT Sanceler M Sanceler M-G Soxinol M Thiotax Vulkacit M Vulkacit Mercapto Vulkacit Mercapto MG/C Vulkacit Mercapto/C Vulkacit Mercapto/MG Vulkafil ZN 94TT01 Wobezit M

DTXSID1020807

PR:000015171

PR sodium/iodide cotransporter

CHEBI:16382

CHEBI iodide

CHEBI:60311

CHEBI thyroid hormone

CHEBI:30660

CHEBI thyroxine

GO:0008507

GO sodium:iodide symporter activity

GO:0015705

GO iodide transport

GO:0006590

GO thyroid hormone generation

MP:0005475

MP abnormal circulating thyroxine level

GO:0007552

GO metamorphosis

WIKI decreased

WIKI delayed

WIKI increased

Perchlorate

2016-11-29T18:42:26

Nitrate

2016-11-29T18:42:26

Thiocyanate

2016-11-29T18:42:26

Dysidenin

2016-11-29T18:42:26

Aryltrifluoroborates

2016-11-29T18:42:26

Econazole

2016-12-07T11:06:26

5-(N,N-hexamethylene) amiloride (HMA)

2016-12-07T11:06:59

Small molecules: ITB3, ITB4, ITB5, ITB9

2016-12-07T11:16:09

Propylthiouracil

2016-11-29T18:42:22

Methimazole

2016-11-29T18:42:19

Stressor:48 Propylthiouracil

2020-08-28T17:00:54

Mercaptobenzothiazole

2016-11-29T18:42:17

WCS_9606

common toxicological species human

10116

NCBI rat

10090

NCBI mouse

WikiUser_24

Wikiuser:Migration Pig

7955

NCBI zebra fish

224340

NCBI Xenopus (Silurana) epitropicalis

WCS_8355

common ecological species African clawed frog

WikiUser_17

mammals

WikiUser_6

ApacheUser fish

8292

NCBI Amphibia

451443

NCBI Xenopus (Silurana) n. sp. tetraploid-1

WCS_8355

common ecological species Xenopus laevis

WCS_7955

common ecological species zebrafish

WCS_90988

common ecological species fathead minnow

9823

NCBI Sus scrofa

WCS_9031

common ecological species chicken

443947

NCBI Xenopus laevis laevis

Inhibition, Na+/I- symporter (NIS)

Molecular

<h4>Evidence for Perturbation by Stressor</h4> <h5>Overview for Molecular Initiating Event</h5> Thyroid Disrupting Chemicals (TDCs) are defined as the xenobiotics that interfere with the thyroid axis with different outcomes for the organism. A very well-studied mechanism of action of the TDCs is the reduction of the circulating levels of THs by inhibiting hormone synthesis in the thyroid gland. For example, perchlorate is a very potent inhibitor of iodide uptake through the sodium/iodide symporter (Tonacchera et al., 2004). Perchlorate has been detected in human breast milk ranging from 1.4 to 92.2 mg μl–1 (10.5 μg l–1 mean) in 18 US states (Kirk et al. 2005), and 1.3 to 411 μg l–1 (9.1 μg l–1 median) in the Boston area, United States (Pearce et al. 2007). Perchlorate has also been detected in human colostrum of 46 women in the Boston area (from < 0.05 to 187.2 μmol l–1 (Leung et al. 2009)). The mechanism of perchlorate action is quite simple, as it is believed to be mediated only by the NIS inhibition (Dohan et al., 2007; Wolff, 1998). Additionally, thiocyanate and nitrate are two known inhibitors that have been found to reduce circulating TH levels (Blount et al., 2006; Steinhaus et al., 2007), but they are both less potent than perchlorate (Tonacchera et al., 2004). However, there are also contradictory results from other studies that showed no correlation between thyroid parameters and perchlorate levels in humans (Pearce et al., 2010; Amitai et al., 2007; Tellez et al., 2005). Co-occurrence of perchlorate, nitrate, and thiocyanate can alter thyroid function in pregnant women. Horton et al. (2015) have shown positive associations between the weighted sum of urinary concentrations of these three analytes and increased TSH, with perchlorate showing the largest weight in the index. Interestingly, De Groef et al. 2006 showed that nitrate and thiocyanate, acquired through drinking water or food, account for a much larger proportion of iodine uptake inhibition than perchlorate, suggesting that NIS inhibition and any potential downstream effect by perchlorate are highly dependent on the presence of other environmental NIS inhibitors and iodine intake itself (Leung et al., 2010). In particular, Tonacchera et al. (2004) showed that the relative potency of perchlorate to inhibit radioactive I− uptake by NIS is 15, 30 and 240 times that of thiocyanate, iodide, and nitrate respectively on a molar concentration basis. These data are in line with earlier studies in rats (Alexander and Wolff, 1996; Greer et al. 1966). Contradictory findings in these studies may therefore be a result of the confounding mixtures in the environment, masking the primary effect of perchlorate. Decreased iodine intake can decrease TH production, and therefore exposure to perchlorate might be particularly detrimental in iodine-deficient individuals (Leung et al. 2010). Moreover, biologically based dose-response modeling of the relationships among iodide status (e.g., dietary iodine levels), perchlorate dose, and TH production in pregnant women has shown that iodide intake has a profound effect on the likelihood that exposure to goitrogens will produce hypothyroxinemia (Lewandowski et al. 2015). During pregnancy TH requirements increase, particularly during the first trimester (Alexander et al. 2004; Leung et al. 2010), due to higher concentrations of thyroxine-binding globulin, placental T4 inner-ring deiodination leading to the inactive reverse T3 (rT3), and transfer of small amounts of T4 to the foetus (during the first trimester foetal thyroid function is absent). Moreover, glomerular filtration rate and clearance of proteins and other molecules are both increased during pregnancy, possibly causing increased renal iodide clearance and a decreased of circulating plasma iodine (Glinoer, 1997). Thus, even though the foetal thyroid can trap iodide by about 12 week of gestation (Fisher and Klein, 1981), high concentrations of maternal perchlorate may potentially decrease thyroidal iodine available to the foetus by inhibiting placental NIS (Leung et al. 2010). Consequences of TH deficiency depend on the developmental timing of the deficiency (Zoeller and Rovet, 2004). For instance, if the TH deficiency occurs during early pregnancy, offspring show visual attention, visual processing and gross motor skills deficits, while if it occurs later, offspring may show subnormal visual and visuospatial skills, along with slower response speeds and motor deficits. If TH insufficiency occurs after birth, language and memory skills are most predominantly affected (Zoeller and Rovet, 2004). Along this line, age and developmental stage are crucial in determining sensitivity to NIS inhibitors (e.g., perchlorate, thiocyanate, and nitrate). In this regard, McMullen et al. (2017) have shown that adolescent boys and girls, more than adults, represent vulnerable subpopulations to NIS symporter inhibitors. Altogether these studies indicate that age, gender, developmental stage, and dietary iodine levels can affect the impact of NIS inhibitors. Finally, ten more small simple-structured molecules were identified in a large screening study (Lecat-Guillet et al., 2008b) that could block iodide uptake by specifically disrupting NIS in a dose-dependent manner. These molecules were named Iodide Transport Blockers (ITBs). There are few organic molecules that lead to NIS inhibition but no direct interaction with NIS has been determined (Gerard et al., 1994; Kaminsky et al., 1991, Lindenthal et al., 2009). Up to date, only dysidenin, a toxin isolated from the marine sponge Dysidea herbacea, has been reported to specifically inhibit NIS (Van Sande et al., 2003). Finally, the aryltrifluoroborates were found to inhibit iodide uptake with an IC50 value of 0.4 μM on rat-derived thyroid cells (Lecat-Guillet et al., 2008a). The biological activity is rationalized by the presence of the BF3− ion as a minimal binding motif for substrate recognition at the iodide binding site. It has been also shown that many anions, such as ClO3-, SCN-, NO3-, ReO4-, TcO4- and in a lower extent Br- and BF4-, are also acting as NIS substrates and they enter the cell by the same transporter mechanism (Van Sande et al., 2003). It has been also shown that ClO4- is transferred by NIS with high affinity and is considered as one of its most potent inhibitors (Dohan et al., 2007). Most recently, the aryltrifluoroborates were also shown to inhibit NIS function (Lecat-Guillet et al., 2008a). A library of 17,020 compounds was tested by a radioactive screening method with high specificity using transfected mammalian cells (Lecat-Guillet et al., 2008b; 2007) for NIS inhibition evaluation. Further studies with the most powerful iodide transport blockers showed a high diversity in their structure and mode of action (Lindenthal et al., 2009). Apart from the human, functional NIS protein has been also identified in different species, including the rat (Dai et al., 1996), the mouse (Perron et al., 2001), the pig (Selmi-Ruby et al., 2003), zebrafish (Thienpont et al., 2011) and xenopus (amphibian) (Lindenthal et al., 2009). Mouse and rat NIS proteins contain 618 amino acid residues, while the human and pig variants contain 643. There are several NIS variants that produce three active proteins in the pig due to alternative splicing at mRNA sites that are not present on the other species (Selmi-Ruby et al., 2003). NIS orthologs are discussed in the review by Darrouzet's group (Darrouzet et al., 2014). Interestingly, functional differences have been identified between mouse or rat NIS (mNIS or rNIS, respectively) and human NIS (hNIS). The rat and themouse orthologs were shown to accumulate radioisotopes more efficiently than the human protein (Dayem et al., 2008; Heltemes et al., 2003). The molecular basis of these functional differences could be helpful for further characterization of NIS. Zhang and collaborators showed that rNIS is localized in a higher proportion at the plasma membrane than hNIS and the N-terminal region up to putative transmembrane helix TM7 appears to be involved in this difference (Zhang et al., 2005). These authors also reported differences in the kinetics of the Na+ binding, implicating the region spanning from TM4 to TM6 and Ser200 of hNIS. They, thus, proposed that this region could be involved in sodium binding (Zhang et al., 2005). In our laboratory, it was shown that the Vmax of the mouse protein is four times higher than the Vmax of the human protein when expressed in the same cell line (HEK-293) (Dayem et al., 2008; Darrouzet et al., 2014). The KmI value determined for hNIS (9.0 ± 0.8 μM) was significantly lower than the KmI for the mouse protein (26.4 ± 3.5 μM) whereas the KmNa values were not significantly different indicating that mNIS has a lower iodide affinity than hNIS. Similarly to the rat protein, mNIS is predominantly localized in the plasma membrane whereas the human ortholog is detected intracellularly in 40% of the cells in which it is expressed (Darrouzet et al., 2014). However, the difference in the Vmax values does not only seem to be related to the higher intracellular localization of hNIS. Using chimeric proteins between human and mouse NIS, we showed that the N-terminal region up to TM8 is most probably involved in iodide binding, and that the region from TM5 to the C terminus could play an important role in targeting the protein to the plasma membrane (Dayem et al., 2008). One of the long-term goals of these studies is the engineering of a chimeric NIS protein most suitable for gene therapy, i.e. preserving regions responsible for the high turnover rate and the efficient plasma membrane localization of the mouse proteinwhile replacing the immunogenic extracellular regions with those of the human ortholog. The porcine NIS gene gives rise to splice variants leading to three active NIS proteins with differences in their C-terminal extremities [4]. However, it is not known if these differences lead to distinct properties (Darrouzet et al., 2014). There is evidence that the MIE (NIS inhibition) is of relevance also for fish as an expression of the slc5a5 transcript (sodium/iodide co-transporter) has been described by various publications for the zebrafish embryo (see <a href="http://www.zfin.org/">www.zfin.org</a>). It has been demonstrated that NIS inhibitors in zebra fish lead also to a strong repression of thyroid hormone levels (Thienpont et al., 2011) and in xenopus (amphibian) to  inhibition of the iodide-induced current  (Lindenthal et al., 2009). Biological state: Sodium/Iodide symporter (NIS) is a key protein in the thyroid function and its role has been thoroughly investigated after the determination of its molecular identity a few decades ago (Dai et al., 1996). NIS is an intrinsic membrane glycoprotein and it belongs to the superfamily of sodium /solute symporters (SSS) and to the family of human transporters SLC5 (De La Vieja, 2000; Jung, 2002). Its molecular weight is 87 kDa and it contains 13 transmembrane domains that transport 2 sodium cations (Na+) for each iodide anion (I-) into the follicular thyroid cell (Dohan et al., 2003). The regulation of NIS protein function is usually cell- and tissue-specific (Hingorani et al., 2010) and it is done at the transcriptional and posttranslational levels, including epigenetic regulation (Darrouzet et al., 2014; Russo et al., 2011a). One of the major NIS regulators is the thyroid stimulating hormone (TSH), which has been shown to enhance NIS mRNA and protein expression, therefore it can contribute to restore and maintain iodide uptake activity (Saito et al., 1997; Kogai et al., 2000). At the posttranslational level TSH also contributes to NIS regulation but the specific mechanisms that underlie these effects are still under investigation (Riedel et al., 2001). Biological compartments: NIS protein is mainly found at the basolateral plasma membrane of the thyroid follicular cells (Dai et al., 1996), where it actively mediates the accumulation of iodide that is the main component of thyroid hormone synthesis and therefore is considered as a major regulator of thyroid homeostasis. NIS also mediates active I- transport in extrathyroidal tissues but it is commonly agreed that is regulated and processed differently in each tissue. Functional NIS protein has been found in salivary gland ductal cells (Jhiang et al., 1998; La Perle et al., 2013), in the mammary gland during lactation (Perron et al., 2001; Cho et al., 2000), lung epithelial cells (Fragoso et la., 2004), intestinal enterocytes (Nicola et al., 2009), stomach cells (Kotani et al., 1998), placenta (Bidart et al., 2000) and testicular cells (Russo et al. 2011b). Additionally, contradictory results have been obtained regarding the NIS expression in human kidney tissue (Lacroix et al., 2001; Spitzweg et al., 2001). In the case of the lactating breast, it is suggested that NIS serves the transfer of iodide in the cells and its subsequent accumulation in the milk, thereby supplying newborns with this component during this sensitive developmental period (Tazebay et al., 2000). Additionally, NIS mRNA has been detected in various other tissues, such as colon, ovaries, uterus, and spleen (Perron et al., 2001; Spitzweg et al., 1998; Vayre et al., 1999), but the functional NIS protein and the site of its localization has not been verified. General role in biology: The NIS is known in the field of thyroidology because of its ability to mediate the active transport of I- into the thyrocytes, which is the first and most crucial step for T3 and T4 biosynthesis (Dohan et al., 2000). NIS is located on the basolateral membrane of the thyrocytes and co-transports 2 sodium ions along with 1 iodide (2:1 stoichiometry). The electrochemical gradient of sodium serves as the driving force for iodide uptake and it is generated and maintained by the Na+/K+ ATPase pump, which is located in the same membrane of the thyrocytes. The iodide molecules, after their active transport in the cytoplasm, are passively translocated in the follicular lumen via the transporter protein pendrin and possibly other unknown efflux proteins that are located on the apical membrane (Bizhanova and Kopp, 2009). Subsequently, the thyroid hormones are synthesized in the follicular lumen by incorporating the accumulated iodide, a process which is significantly suppressed in case of NIS dysfunction or inhibition (reviewed in Spitzweg and Morris, 2010). NIS is the last thyroid-related component to be expressed during development at the 10th gestational week, which temporaly coincides with the onset of thyroid function and hormonogenesis (Szinnai et al., 2007). Albeit the localization of NIS is not fully completed at this stage, the iodide accumulation has already started. Mutations of NIS gene (SLCA5A) cause expression of non-functional NIS molecule leading to inability of the thyrocyte to accumulate iodide (Matsuda and Koshugi, 1997; Pohlenz et al., 1998), a condition called iodide transport defect (ITD). This is a rear autosomic recessive disease, which if not properly treated is clinically identified by congenital hypothyroidism, goiter, low I- uptake, low saliva/plasma I- ratio and mental impairment of varying degrees (Dohan et al., 2003). Up to date 13 mutations have been described in the NIS gene (Spitzweg and Morris, 2010) and each one of them produces mutants with different structure but in all cases non-functional. The extensive study after NIS molecular characterization and the numerous findings have convinced the scientists that is one of the most crucial components of the entire thyroid system. Additionally, after the realization that NIS could be also used as diagnostic and therapeutic tool for thyroid and non-thyroid cancers (Portulano et al., 2013) a new research activity concerning this specific mechanism has been initiated.

There are several methods that are used nowadays to detect the functionality of NIS but none of these methods is OECD validated (OECD Scoping document, 2017). The most well established methods are the following: 1. Measurement of radioiodide uptake (125I-) in NIS expressing cells. For this method the FRTL5 cell line is the most commonly used, as it endogenously express the NIS protein, but also NIS transfected cell lines have been successfully implemented in many cases (Lecat-Guillet et al., 2007; 2008b; Lindenthal et al., 2009). Once inhibitory activity is identified for a compound then further tests are performed in order to verify that the observed effect is specific due to NIS inhibition. This method has been also adapted in a high throughput format and has been already used for the screening of a chemical library of 17.020 compounds (Lecat-Guillet et al., 2008b). 2. More recently a non-radioactive method has been developed, which has been also adapted in a high throughput format (Waltz et al., 2010). It is a simple spectrophotometric assay for the determination of iodide uptake  using  rat thyroid-derived cells (FRTL5) based on the catalytic effect of iodide on the reduction of yellow cerium(IV) to colorless cerium(III) in the presence of arsenious acid (Sandell-Kolthoff reaction). The assay is fast, highly reproducible and equally sensitive with the radioiodine detection method. 3. A fluorescence-based method has been developed, which uses the variant  YFP-H148Q/I152L of the Yellow Fluorescent Protein (YFP) in order to detect the efflux of iodide into the rat FRTL5 cells. As a positive control perchlorate is used  as it is a well known  competitive inhibitor of iodide transport by NIS. Fluorescence of recombinant YFP-H148Q/I152L is  suppressed by perchlorate and iodide with similar affinities. Fluorescence changes in FRTL-5 cells are  Na+-dependent, consistent with the Na+-dependence of NIS activity.   It is supposed to be an innovative approach to detect the cellular uptake of perchlorate and characterize the kinetics of transport by NIS. This method needs further optimization, as YFP is not specific for iodide and thus binding of other ionic molecules could affect the results of the assay (Cianchetta et al., 2010; Rhoden et al., 2008; Di Bernarde et al., 2011). 4. In vivo 125I uptake assays is based on  immunofluorescence analyses of thyroid glands after the treatment of rat with excess I−,  injected with Ci Na125I  as previously described by Ferreira et al., 2005. Then the thyroid glands are  removed and weighed, and the amount of 125I in the thyroid gland is  measured in a γ-counter (PerkinElmer; model Wizard). The counts per minute in the thyroid gland are used to calculate the percentage of 125I in the thyroid gland, having in account that 100% corresponded to the counts per minute injected I− into the rat (Arriagada et al., 2015). 5. The U.S. EPA's Endocrine Disruptor Screening Program aims to use high-throughput assays and computational toxicology models to screen and prioritize chemicals that may disrupt the thyroid signaling pathway. Thyroid hormone biosynthesis requires active iodide uptake mediated by the sodium/iodide symporter (NIS). Monovalent anions, such as the environmental contaminant perchlorate, are competitive inhibitors of NIS, yet limited information exists for more structurally diverse chemicals. A novel cell line expressing human NIS, hNIS-HEK293TEPA, was used in a radioactive iodide uptake (RAIU) assay to identify inhibitors of NIS-mediated iodide uptake. The RAIU assay was optimized and performance evaluated with 12 reference chemicals comprising known NIS inhibitors and inactive compounds. An additional 39 chemicals including environmental contaminants were evaluated, with 28 inhibiting RAIU over 20% of that observed for solvent controls. Cell viability assays were performed to assess any confounding effects of cytotoxicity. RAIU and cytotoxic responses were used to calculate selectivity scores to group chemicals based on their potential to affect NIS. RAIU IC50 values were also determined for chemicals that displayed concentration-dependent inhibition of RAIU (≥50%) without cytotoxicity. Strong assay performance and highly reproducible results support the utilization of this approach to screen large chemical libraries for inhibitors of NIS-mediated iodide uptake (Hallinger et al., 2017). 6. This study (Wang et al., 2018) applied a previously validated high-throughput approach to screen for NIS inhibitors in the ToxCast phase I library, representing 293 important environmental chemicals. Here 310 blinded samples were screened in a tiered-approach using an initial single-concentration (100 μM) radioactive-iodide uptake (RAIU) assay, followed by 169 samples further evaluated in multi-concentration (0.001 μM−100 μM) testing in parallel RAIU and cell viability assays. A novel chemical ranking system that incorporates multi-concentration RAIU and cytotoxicity responses was also developed as a standardized method for chemical prioritization in current and future screenings. Representative chemical responses and thyroid effects of high-ranking chemicals are further discussed. This study significantly expands current knowledge of NIS inhibition potential in environmental chemicals and provides critical support to U.S. EPA’s Endocrine Disruptor Screening Program (EDSP) initiative to expand coverage of thyroid molecular targets, as well as the development of thyroid adverse outcome pathways (AOPs).

Apart from the human, functional NIS protein has been also identified in different species, including  the rat (Dai et al., 1996), the mouse (Perron et al., 2001), the pig (Selmi-Ruby et al., 2003), zebrafish (Thienpont et al., 2011) and in xenopus (amphibian)  (Lindenthal et al., 2009). Mouse and rat contain 618 amino acid residues, while the human and pig contain 643. There are several NIS variants that produce three active proteins in the pig due to alternative splicing at mRNA sites that are not present on the other species (Selmi-Ruby et al., 2003). NIS orthologs are discussed in the review by Darrouzet's group ( Darrouzet et al., 2014). Interestingly, functional differences have been identified between mouse or rat NIS (mNIS or rNIS, respectively) and human NIS (hNIS). The rat and themouse orthologs were shown to accumulate radioisotopes more efficiently than the human protein (Dayem et al., 2008; Heltemes et al., 2003). The molecular basis of these functional differences could be helpful for further characterization of NIS. Zhang and collaborators showed that rNIS is localized in a higher proportion at the plasma membrane than hNIS and the N-terminal region up to putative TM7 appears to be involved in this difference (Zhang et al., 2005). These authors also reported differences in the kinetics of the Na+ binding, implicating the region spanning from TM4 to TM6 and Ser200 of hNIS. They, thus, proposed that this region could be involved in sodium binding (Zhang et al., 2005). In our laboratory, it was shown that the Vmax of the mouse protein is four times higher than the Vmax of the human protein when expressed in the same cell line (HEK-293) (Dayem et al., 2008; Darrouzet et al., 2014). The KmI value determined for hNIS (9.0 ± 0.8 μM) was significantly lower than the KmI for the mouse protein (26.4 ± 3.5 μM) whereas the KmNa values were not significantly different. Similarly to the rat protein, mNIS is predominantly localized in the plasma membrane whereas the human ortholog is detected intracellularly in 40% of the cells in which it is expressed (Darrouzet et al., 2014). However, the difference in the Vmax values does not only seem to be related to the higher intracellular localization of hNIS. Using chimeric proteins between human and mouse NIS, we showed that the N-terminal region up to TM8 is most probably involved in iodide binding, and that the region from TM5 to the C terminus could play an important role in targeting the protein to the plasma membrane (Dayem et al., 2008). One of the long-term goals of these studies is the engineering of a chimeric NIS protein most suitable for gene therapy, i.e. preserving regions responsible for the high turnover rate and the efficient plasma membrane localization of the mouse proteinwhile replacing the immunogenic extracellular regions with those of the human ortholog. The porcine NIS gene gives rise to splice variants leading to three active NIS proteins with differences in their C-terminal extremities [4]. However, it is not known if these differences lead to distinct properties (Darrouzet et al., 2014). There is evidence that the MIE (NIS inhibition) is of relevance also for fish as an expression of the slc5a5 transcript (sodium/iodide co-transporter) has been described by various publications for the zebrafish embryo (see <a href="http://www.zfin.org">www.zfin.org</a>). It has been demonstrated that NIS inhibitors in zebrafish lead also to a strong repression of thyroid hormone levels (Thienpont et al., 2011) and in xenopus (amphibian) to  inhibition of the iodide-induced current  (Lindenthal et al., 2009). According to the evaluation of the empirical taxonomic domain of applicability (tDOA) of an adverse outcome pathway network for thyroid hormone system disruption (THSD) by Haigis et al., 2023, the level of confidence for a linkage between NIS inhibition and reduced thyroid hormone (TH) levels was considered high for mammals (Buckalew et al., 2020, Concilio et al., 2020, Dayem et al., 2008, Hallinger et al., 2017, Heltemes et al., 2003, Schmutzler et al., 2007, Selmi-Ruby et al., 2003, Wang et al., 2018) and moderate for fish and amphibians (Concilio et al., 2020, Hornung et al., 2010, Lindenthal et al., 2009, McMullen et al., 2017, Opitz et al., 2006; Opitz and Kloas, 2010, Thienpont et al., 2011). This was supported by structural protein conservation analysis by Lalone et al., 2018 and Haigis et al., 2023. Structural protein conservation of mammalian, fish, amphibian, reptilian and avian NIS was found compared to the human (Homo sapiens) protein target using the U.S. Environmental Protection Agency’s Sequence Alignment to Predict Across Species Susceptibility (SeqAPASS v6.0; seqapass.epa.gov/seqapass/) tool, while acknowledging the potential existence of interspecies differences in conservation. No empirical evidence linking NIS inhibition to THSD was found for reptiles and birds.

CL:0002258

CL thyroid follicular cell

High Mixed

High

Pregnancy

High

Birth to < 1 month

High

During brain development

High High High High High Moderate Not Specified High Moderate Moderate

Alexander EK, Marqusee E, Lawrence J, Jarolim P, Fischer GA, Larsen PR (2004). Timing and magnitude of increases in levothyroxine requirements during pregnancy in women with hypothyroidism. N Engl J Med. Jul 15; 351(3):241-9. Alexander WD and Wolff J (1996). Thyroidal iodide transport VIII, Relation between transport goitrogenic and antigoitrogenic properties of certain anions. Endocrinology 78 581–590. Amitai Y, Winston G, Sack J, Wasser J, Lewis M, Blount BC, Valentin-Blasini L, Fisher N, Israeli A, Leventhal A. (2007). Gestational exposure to high perchlorate concentrations in drinking water and neonatal thyroxine levels. Thyroid. 17:843-850. Arriagada A.A, Eduardo Albornoz, Ma. Cecilia Opazo, Alvaro Becerra, Gonzalo Vidal, Carlos Fardella, Luis Michea, Nancy Carrasco, Felipe Simon, Alvaro A. Elorza, Susan M. Bueno, Alexis M. Kalergis, and Claudia A.  (2015).Excess Iodide Induces an Acute Inhibition of the Sodium/Iodide Symporter in Thyroid Male Rat Cells by Increasing Reactive Oxygen Species. Endocrinology. 2015 Apr; 156(4): 1540–1551. Bidart JM, Lacroix L, Evain-Brion D, Caillou B, Lazar V, Frydman R, Bellet D, Filetti S, Schlumberger M. (2000). Expression of Na+/I− symporter and Pendred syndrome genes in trophoblast cells. J Clin Endocrinol Metab 85:4367–4372. Bizhanova A, Kopp P. (2009). The sodium-iodide symporter NIS and pendrin in iodide homeostasis of the thyroid. Endocrinol 150:1084-1090. Blount BC, Pirkle JL, Osterloh JD, Valentin-Blasini L, Caldwell KL. (2006). Urinary perchlorate and thyroid hormone levels in adolescent and adult men and women living in the United States. Env Health Persp. 114:1865-1871. Buckalew, A. R., Wang, J., Murr, A. S., Deisenroth, C., Stewart, W. M., Stoker, T. E., and Laws, S. C. (2020). Evaluation of potential sodium-iodide symporter (NIS) inhibitors using a secondary Fischer rat thyroid follicular cell (FRTL-5) radioactive iodide uptake (RAIU) assay. Arch. Toxicol. 94, 873–885. Cho JY, Leveille R, Kao R, Rousset B, Parlow AF, Burak WE Jr, Mazzaferri EL, Jhiang SM. (2000). Hormonal regulation of radioiodide uptake activity and Na+/I− symporter expression in mammary glands. J Clin Endocrinol Metab 85:2936–2943. Cianchetta S, di Bernardo J, Romeo G, Rhoden KJ. (2010). Perchlorate transport and ihnibition of the sodium iodide symporter measured with the yellow fluorescent protein variant YFP-H148Q/I152L. Toxicol App Pharmacol. 243:372-380. Concilio, S. C., Zhekova, H. R., Noskov, S. Y., and Russell, S. J. (2020). Inter-species variation in monovalent anion substrate selectivity and inhibitor sensitivity in the sodium iodide symporter (NIS). PLoS One 15, e0229085. Dai G, Levy O, Carrasco N. (1996). Cloning and characterization of the thyroid iodide transporter. Nature 379:458–460. Darrouzet E, Lindenthal S, Marcellin D, Pellequer J, Pourcher T. (2014). The sodium/iodide symporter: state of the art of its molecular characterization. Biochim Biophys Acta 1838:244-253. Dayem M, Basquin C, Navarro V, Carrier P, Marsault R, Chang P, Huc S, Darrouzet E, Lindenthal S, Pourcher T. (2008). Comparison of expressed human and mouse sodium/iodide symporters reveals differences in transport properties and subcellular localization. J Endocrinol. 197:95–109. De Groef B, Decallonne BR, Van der Geyten S, Darras VM, Bouillon R (2006). Perchlorate versus other environmental sodium/iodide symporter inhibitors: potential thyroid-related health effects. Eur J Endocrinol. Jul;155(1):17-25. De La Vieja A, Dohan O, Levy O, Carrasco N. (2000). Molecular analysis of the sodium/iodide symporter: impact on thyroid and extrathyroid pathophysiology. Physiol. Rev. 80: 1083–105. Di Bernardo J, Iosco C, Rhoden KJ. (2011). Intracellular anion fluorescence assay for sodium/iodide symporter substrates. Analyt Biochem. 415:32-38. Dohan O, De la Vieja A, Carrasco N. (2000). Molecular study of the sodium-iodide symporter (NIS): a new field in thyroidology. Trends Endocrinol Metab. 11:99–105. Dohan O, De la Vieja A, Paroder V, Riedel C, Artani M, Reed M, Ginter CS, Carrasco N. (2003). The sodium/iodide Symporter (NIS): characterization, regulation, and medical significance. Endocr Rev. 24:48–77. Dohan O, Portulano C, Basquin C, Reyna-Neyra A, Amzel LM, Carrasco N. (2007). The Na+/I symporter (NIS) mediates electroneutral active transport of the environmental pollutant perchlorate. Proc. Natl. Acad. Sci. U. S. A. 104:20250–20255. Ferreira ACF, Lima LIVP, Araujo RL, et al. Rapid regulation of thyroid sodium-iodide symporter activity by thyrotrophin and iodine. J Endocrinol. 2005;184:69–76. Fisher DA and Klein AH (1981). Thyroid development and disorders of thyroid function in the newborn. N Engl J Med. Mar 19; 304(12):702-12. Fragoso MA, Fernandez V, Forteza R, Randell SH, Salathe M, Conner GE. (2004).Transcellular thiocyanate transport by human airway epithelia. J Physiol 561:183–194. Gerard C, Rigot V, Penel C. (1994). Chloride channel blockers inhibit the Na+/I- symporter in thyroid follicles in culture. Biochem Biophys Res Communic. 204: 1265-1271. Glinoer D (1997). The regulation of thyroid function in pregnancy: pathways of endocrine adaptation from physiology to pathology. Endocr Rev. Jun; 18(3):404-33. Greer MA, Stott AK & Milne KA (1966). Effect of thiocyanate, perchlorate and other anions on thyroidal iodine metabolism. Endocrinology 79 237–247. Haigis A-C., Vergauwen L., LaLone C.A., Villeneuve D.L., O'Brien J.M., Knapen D. (2023). Cross-species applicability of an adverse outcome pathway network for thyroid hormone system disruption. Toxicol Sci. 195, 1-27. Hallinger DR, Murr AS, Buckalew AR, Simmons SO, Stoker TE, Laws SC.  (2017).  Development of a screening approach to detect thyroid disrupting chemicals that inhibit the human sodium iodide symporter (NIS). Toxicol In Vitro. Apr;40:66-78. Heltemes LM, Hagan CR, Mitrofanova EE, Panchal RG, Guo J, Link CJ. (2003). The rat sodium iodide symporter gene permits more effective radioisotope concentration than the human sodium iodide symporter gene in human and rodent cancer cells. Cancer Gene Ther. 10:14–22. Hingorani M, Spitzweg C, Vassaux G, Newbold K, Melcher A, Pandha H, Vile R, Harrington K. (2010). The biology of the sodium iodide symporter and its potential for targeted gene delivery. Curr Cancer Drug Targets 10:242–267. Hornung, M. W., Degitz, S. J., Korte, L. M., Olson, J. M., Kosian, P. A., Linnum, A. L., and Tietge, J. E. (2010). Inhibition of thyroid hormone release from cultured amphibian thyroid glands by methimazole, 6-propylthiouracil, and perchlorate. Toxicol. Sci. 118, 42–51. Horton MK, Blount BC, Valentin-Blasini L, Wapner R, Whyatt R, Gennings C, Factor-Litvak P (2015). CO-occurring exposure to perchlorate, nitrate and thiocyanate alters thyroid function in healthy pregnant women. Environ Res. Nov;143(Pt A):1-9. Jhiang SM, Cho JY, Ryu KY, DeYoung BR, Smanik PA, McGaughy VR, Fischer AH, Mazzaferri EL. (1998). An immunohistochemical study of Na+/I− symporter in human thyroid tissues and salivary gland tissues. Endocrinology 139:4416–4419. Jung H. (2002). The sodium/substrate symporter family: structural and functional features. FEBS Lett. 529:73–77. Kaminsky SM, Levy O, Garry MT, Carrasco N. (1991). Inhibition of the Na+/I- symporter by harmaline and 3-amino-1-methyl-5H-pyridol(4,3-b)indole acetate in thyroid cells and membrane vesicles. Eur J Biochem. 200:203-207. Kirk AB, Martinelango PK, Tian K, Dutta A, Smith EE, Dasgupta PK (2005). Perchlorate and iodide in dairy and breast milk. Environ Sci Technol. Apr 1; 39(7):2011-7. Kogai T, Curcio F, Hyman S, Cornford EM, Brent GA, Hershman JM. (2000). Induction of follicle formation in long-term cultured normal human thyroid cells treated with thyrotropin stimulates iodide uptake but not sodium/iodide symporter messenger RNA and protein expression. J Endocrinol 167:125–135. Kotani T, Ogata Y, Yamamoto I, Aratake Y, Kawano JI, Suganuma T, Ohtaki S. (1998). Characterization of gastric Na+/I− symporter of the rat. Clin Immunol Immunopathol 89:271–278. Lalone, C. A., Villeneuve, D. L., Doering, J. A., Blackwell, B. R., Transue, T. R., Simmons, C. W., Swintek, J., Degitz, S. J., Williams, A. J., and Ankley, G. T. (2018). Evidence for cross species extrapolation of mammalian-based high-throughput screening assay results. Environ. Sci. Technol. 52, 13960–13971. La Perle KM, Kim DC, Hall NC, Bobbey A, Shen DH, Nagy R, Wakely PE Jr, Leman A, Jarjoura D, Jhiang SM. (2013). Modulation of sodium/iodide symporter expression in the salivary gland. Thyroid 23:1029-1036. Lacroix L, Mian C, Caillou B, Talbot M, Filetti S, Schlumberger M, Bidart JM. (2001). Na+/I- symporter and pendred syndrome gene and protein expressions in human extra-thyroidal tissues. Eur J Endocrinol 144:297-302. Lecat-Guillet N, Ambroise Y. (2008a). Discovery of aryltrifluoroborates as potent sodium/iodide symporter (NIS) inhibitors. Chem Med Chem 3:1207–1209. Lecat-Guillet N, Merer G, Lopez R, Pourcher T, Rousseau B, Ambroise Y. (2008b). Small-molecule inhibitors of sodium iodide symporter function. Chembiochem 9:889–895. Lecat-Guillet N, Merer G, Lopez R, Pourcher T, Rousseau B, Ambroise Y. (2007). A 96-well automated radioiodide uptake assay for sodium/iodide symporter inhibitors. Assay Drug Dev Technol 5:535-540. Leung AM, Pearce EN, Hamilton T, He X, Pino S, Merewood A, Braverman LE (2009). Colostrum iodine and perchlorate concentrations in Boston-area women: a cross-sectional study. Clin Endocrinol (Oxf). Feb; 70(2):326-30. Leung AM, Pearce EN, Braverman LE (2010). Perchlorate, iodine and the thyroid. Best Pract Res Clin Endocrinol Metab. Feb;24(1):133-41. Lewandowski TA, Peterson MK2, Charnley G (2015). Iodine supplementation and drinking-water perchlorate mitigation. Food Chem Toxicol. Jun;80:261-70. Lindenthal S, Lecat-Guillet N, Ondo-Mendez A, Ambroise Y, Rousseau B, Pourcher T. (2009). Characterization of small-molecule inhibitors of the sodium iodide symporter. J Endocrinol 200:357–365. Matsuda A, Kosugi S. (1997). A homozygous missense mutation of the sodium/iodide symporter gene causing iodide transport defect. J Clin Endocrinol Metab 82:3966–3971. McMullen J, Ghassabian A, Kohn B, Trasande L (2017). Identifying Subpopulations Vulnerable to the Thyroid-Blocking Effects of Perchlorate and Thiocyanate. J Clin Endocrinol Metab. Jul 1;102(7):2637-2645. Nicola JP, Basquin C, Portulano C, Reyna-Neyra A, Paroder M, Carrasco N. (2009). The Na+/I− symporter mediates active iodide uptake in the intestine. Am J Physiol Cell Physiol 296:C654–C662. OECD Series on Testing and Assessment (2017). New Scoping Document on in vitro and ex vivo Assays for the Identification of Modulators of Thyroid Hormone Signalling (page 36 - 38). Opitz, R., Trubiroha, A., Lorenz, C., Lutz, I., Hartmann, S., Blank, T., Braunbeck, T., and Kloas, W. (2006). Expression of sodium-iodide symporter mRNA in the thyroid gland of Xenopus laevis tadpoles: Developmental expression, effects of antithyroidal compounds, and regulation by TSH. J. Endocrinol. 190, 157–170. Opitz, R., and Kloas, W. (2010). Developmental regulation of gene expression in the thyroid gland of Xenopus laevis tadpoles. Gen. Comp. Endocrinol. 168, 199–208. Pearce EN, Lazarus JH, Smyth PPA, He X, Dall'amico D, Parkes AB, Burns R, Smith DF, Maina A, Bestwick JP, Jooman M, Leung AM, Braverman LE. (2010). Perchlorate and thiocyanante exposure and thyroid function in first-trimester pregnant women. J Clin Endocrinol Metab. 95:3207-3215. Pearce EN, Leung AM, Blount BC, Bazrafshan HR, He X, Pino S, Valentin-Blasini L, Braverman LE (2007). Breast milk iodine and perchlorate concentrations in lactating Boston-area women. J Clin Endocrinol Metab. May; 92(5):1673-7. Perron B, Rodriguez AM, Leblanc G, Pourcher T. (2001). Cloning of the mouse sodium iodide symporter and its expression in the mammary gland and other tissues. J Endocrinol 170:185–196. Pohlenz J, Rosenthal IM, Weiss RE, Jhiang SM, Burant C, Refetoff S. (1998). Congenital hypothyroidism due to mutations in the sodium/iodide symporter. Identification of a nonsense mutation producing a downstream cryptic 3' splice site. J Clin Invest 101:1028-1035. Portulano C, Paroder-Belenitsky M, Carrasco N. (2014). The Na+/I- symporter (NIS): mechanism and medical impact. Endocr Rev. 35:106-49. Rhoden KJ, Cianchetta S, Duchi S, Romeo G. (2008). Fluorescence quantitation of thyrocyte iodide accumulation with the yellow fluorescent protein variant YFP-H148Q/I152L. 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Endocrine disruptors and the thyroid gland-a combined in vitro and in vivo analysis of potential new biomarkers. Environ. Health Perspect. 115, 77–83. Selmi-Ruby S, Watrin C, Trouttet-Masson S, Bernier-Valentin F, Flachon V, Munari-Silem Y, Rousset B. (2003). The porcine sodium/iodide symporter gene exhibits an uncommon expression pattern related to the use of alternative splice sites not present in the human or murine species. Endocrinology. 144:1074–1085. Spitzweg C, Dutton CM, Castro MR, Bergert ER, Goellner JR, Heufelder AE, Morris JC. (2001). Expression of the sodium iodide symporter in human kidney. Kidney Int 59:1013-1023. Spitzweg C, Joba W, Eisenmenger W, Heufelder AE. (1998). Analysis of human sodium iodide symporter gene expression in extrathyroidal tissues and cloning of its complementary deoxyribonucleic acids from salivary gland, mammary gland, and gastric mucosa. J Clin Endocrinol Metab. 83:1746–1751. Spitzweg C, Morris JC. (2010). Genetics and phenomics of hypothyroidism and goiter due to NIS mutations. Mol Cell Endocrinol 322: 56-63. Steinmaus C, Miller MD, Howd R. (2007). Impact of smoking and thiocyanate on perchlorate and thyroid hormone associations in the 2001-2002 National Health and Nutrition Examination Survey. Env Health Persp. 115:1333-1338. Szinnai G, Lacroix L, Carré A, Guimiot F, Talbot M, Martinovic J, Delezoide AL, Vekemans M, Michiels S, Caillou B, Schlumberger M, Bidart JM, Polak M. (2007). Sodium/iodide symporter (NIS) gene expression is the limiting step for the onset of thyroid function in the human fetus. J Clin Endocrinol Metab. 92:70–76. Tazebay UH, Wapnir IL, Levy O, Dohan O, Zuckier LS, Hua Zhao Q, Fu Deng H, Amenta PS, Fineberg S, Pestell RG, Carrasco N. (2000). The mammary gland iodide transporter is expressed during lactation and in breast cancer. Nat. Med. 6:871–878. Tellez RT, Chacon PM, Abarca CR, Blount BC, Van Landingham CB, Crump KS, Gibbs JP. (2005). 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Immunohistochemical analysis of Na-/I- symporter distribution in human extra-thyroidal tissues. Eur J Endocrinol. 141:382–386. Waltz F, Pillette L, Ambroise Y. (2010). A nonradioactive iodide uptake assay for sodium iodide symporter function. Analytic Biochem. 396:91-95. Wolff J. (1998). Perchlorate and the thyroid gland. Pharmacol Rev. 50:89-105. Wang J, Hallinger DR, Murr AS, Buckalew AR, Simmons SO, Laws SC, Stoker TE (2018). High-Throughput Screening and Quantitative Chemical Ranking for Sodium-Iodide Symporter Inhibitors in ToxCast Phase I Chemical Library. Environ Sci Technol. 2018 May 1;52(9):5417-5426. Zhang Z, Liu YY, Jhiang SM. (2005). Cell surface targeting accounts for the difference in iodide uptake activity between human Na+/I− symporter and rat Na+/I− symporter. J Clin Endocrinol Metab. 90:6131–6140. Zoeller RT, Rovet J (2004). Timing of thyroid hormone action in the developing brain: clinical observations and experimental findings. J Neuroendocrinol. Oct;16(10):809-18. AOPWiki 2016-11-29T18:41:24

2026-04-27T05:31:38

Decrease of Thyroidal iodide

Thyroidal Iodide, Decreased

Cellular

Biological state: Iodine (I2) is a non-metallic chemical element which is required for the normal cellular metabolism. It is one of the essential components of the TH, comprising 65% and 58% of T4's and T3's weight, respectively and therefore it is crucial for the normal thyroid function. It is a trace element and a healthy human body contains 15-20 mg of iodine, most of which is concentrated in the thyroid gland (Dunn, 1998). Iodide (I-) that enters the thyroid gland remains in the free state only briefly and subsequently it bounds to the tyrosine residues of thyroglobulin to form the precursors of the thyroid hormones mono-iodinated tyrosine (MIT) or di-iodinated tyrosine (DIT) (Berson and Yalow, 1955). The bounding rate of iodide is 50-100% of the intra-thyroidal iodide pool, meaning that only a very small proportion of this element is free in the thyroid and this comes mainly by the deiodination of MIT and DIT. The body is not able to produce or make iodine, thus the diet is the only source of this element. Iodine is found in nature in various forms, such as inorganic sodium and potassium salts (iodides and iodates), inorganic diatomic iodine and organic monoatomic iodine (Patrick, 2008). Thus, it is widely distributed in the environment but in many regions of the world the soil's iodine has been depleted due to different environmental phenomena. In these regions, the incidence of iodine deficiency is greatly increased (Ahad and Ganie, 2010). The daily iodine intake of adult humans varies greatly due to the different dietary habits between the different regions on earth (Dunn, 1993). In any case, the ingested iodine is absorbed through the intestine and transported into the plasma to reach the thyroid gland. However, thyroid is not the only organ of the body that concentrates iodide. It has been shown that other tissues have also the ability of iodide concentration, such as the salivary glands, the gastric mucosa, the mammary glands and the choroid plexus, all of which express NIS, the iodine transporter protein (Jhiang et al., 1998; Cho et al., 2000). Biological compartments: A sodium-iodide (Na/I) symporter pumps iodide (IO) actively into the cell, which previously has crossed the endothelium by largely unknown mechanisms. This iodide enters the follicular lumen from the cytoplasm by the transporter pendrin, in a purportedly passive manner. In the colloid, iodide (I−) is oxidized to iodine (I0) by an enzyme called thyroid peroxidase (TPO). IO is very reactive and iodinates the thyroglobulin at tyrosyl residues in its protein chain. In conjugation, adjacent tyrosyl residues are paired together. Thyroglobulin binds the megalin receptor for endocytosis back into the follicular cell. Proteolysis by various proteases liberates thyroxine (T4) and triiodothyronine molecules (T3), which enter the bloodstream where they are bound to thyroid hormone binding proteins, mainly thyroxin binding globulin (TBG) which accounts for about 75% of the bound hormone. The adult thyroid absorbs 60-80 μg of iodide per day to maintain the thyroid homeostasis (Degroot, 1966). Inadequate amount of iodide results to deficient production of thyroid hormones, which consequently leads to an increase of TSH secretion and goiter, as compensating effect (Delange, 2000). On the other hand, excess iodide could also inhibit TH synthesis (Wolff and Chaikoff, 1948). The proposed mechanism for this latter effect is the possible formation of 2-iodohexadecanal that inhibits the generation of H2O2 and the subsequent oxidation of iodide in the thyroid follicular cells. The lack of oxidized free radicals of iodide affects the reaction with the tyrosine residues of Thyroglobulin (Tg) (Panneels et al., 1994). During pregnancy, the organism of the mother is also supporting the needs of the foetus and therefore the iodide requirements are greatly increased (Glinoer, 1997). Additionally, small iodine concentrations have been found to have significant antioxidant effects that resembles to ascorbic acid (Smyth, 2003). General role in biology: The most important role of iodine is the formation of the thyroid hormones (T4 and T3). The thyroid actively concentrates the circulating iodide through the basolateral membrane of the thyrocytes by the sodium/iodide symporter protein (NIS). The concentrated thyroid-iodine is oxidized in the follicular cells of the gland and consequently binds to tyrosines to form mono- or di-iodotyrosines (MIT and DIT respectively), being incorporated into thyroglobulin. This newly formed iodothyroglobulin forms one of the most important constituents of the colloid material, present in the follicle of the thyroid unit. If two di-iodotyrosine molecules couple together, the result is the formation of thyroxin (T4). If a di-iodotyrosine and a mono-iodotyrosine are coupled together, the result is the formation of tri-iodothyronine (T3). From the perspective of the formation of thyroid hormone, the major coupling reaction is the di-iodotyrosine coupling to produce T4.

The radioactive iodine uptake test, or RAIU test, is a type of scan used in the diagnosis of thyroid gland dysfunction (<a href="http://www.thyca.org/pap-fol/rai/">http://www.thyca.org/pap-fol/rai/</a>; Kwee, et al., 2007). The patient swallows radioactive iodine in the form of capsule or fluid, and its absorption by the thyroid is studied after 4–6 hours and after 24 hours with the aid of a gamma scintillation counter. The percentage of RAIU 24 hours after the administration of radioiodide is the most useful, since this is the time when the thyroid gland has reached the plateau of isotope accumulation, and because it has been shown that at this time, the best separation between high, normal, and low uptake is obtained. The test does not measure hormone production and release but merely the avidity of the thyroid gland for iodide and its rate of clearance relative to the kidney.

Various species express functional NIS  encoded by the following genes: Human SLC5A5 (6528), Mouse Slc5a5 (114479), Rat Slc5a5 (114613), Zebrafish slc5a5 (561445), chicken SLC5A5 (431544), domestic cat SLC5A5 (101092587), dog SLC5A5 (484830), domestic guinea pig Slc5a5 (100714457), naked mole-rat Slc5a5 (101701995), cow SLC5A5 (505310), sheep SLC5A5 (101112315). The encoded protein is responsible for the uptake of iodine in tissues such as the thyroid and lactating breast tissue. The iodine taken up by the thyroid is incorporated into the metabolic regulators triiodothyronine (T3) and tetraiodothyronine (T4). Mutations in this gene are associated with thyroid dyshormonogenesis that significantly influences phenotypic expressions such as severity of hypothyroidism, goiter rates, and familial clustering demonstrating essentiality of NIS function to maintain TH status (Bakker et al., 2000; Spitzweg and Morris, 2010; Ramesh et al., 2016) . Animal studies have also proven that iodine normalizes elevated adrenal corticosteroid hormone secretion and has the ability to reverse the effects of hypothyroidism in the ovaries, testicles and thymus in thyroidectomized rats (Nolan et al., 2000).

UBERON:0002046

UBERON thyroid gland

CL:0002258

CL thyroid follicular cell

Moderate Mixed

Moderate

Birth to < 1 month

Moderate

Pregnancy

Moderate

During brain development

High High High High High Moderate Not Specified

Ahad F, Ganie SA. (2010). Iodine, iodine metabolism and iodine deficiency disorders revisited. Indian J Endocrinol Metab. 14: 13-17. Bakker B, Bikker H, Vulsma T, de Randamie JS, Wiedijk BM, De Vijlder JJ. 2000. Two decades of screening for congenital hypothyroidism in The Netherlands: TPO gene mutations in total iodide organification defects (an update). The Journal of clinical endocrinology and metabolism. Oct;85:3708-3712. Berson SA, Yalow RS. (1955). The iodide trapping and binding functions of the thyroid. J Clin Invest. 34: 186-204. Cho JY, Leveille R, Kao R, Rousset B, Parlow AF, Burak WE Jr, Mazzaferri EL, Jhiang SM.(2000). Hormonal regulation of radioiodide uptake activity and Na+/I- symporter expression in mammary glands. J Clin Endocrinol Metab. 85:2936-2943. Degroot LJ.(1966). Kinetic analysis of iodine metabolism. J Clin Endocrinol Metab. 26: 149-173. Delange F. (2000). Iodine deficiency. In: Braverman L, Utiger R, editors. Werner and Ingbar's the thyroid: a fundamental and clinical text. Philadelphia: JD Lippincott. pp 295-316. Dunn JT. (1993). Sources of dietary iodine in industrialized countries. In: Delange F, Dunn JT, Glinoer D, editors. Iodine deficiency in Europe. A continuing concern. New York: Plenum press. pp 17-21. Dunn JT. (1998). What's happening to our iodine? J Clin Endocrinol Metab. 83: 3398-3400. Glinoer D. (1997). The regulation of thyroid function in pregnancy: pathways of endocrine adaptation from physiology to pathology. Endocr Rev. 18: 404-433. <a href="http://www.thyca.org/pap-fol/rai/">http://www.thyca.org/pap-fol/rai/</a>: Thyroid Cancer Survivors' Association, Inc.,Radioactive Iodine (RAI) Jhiang SM, Cho JY, Ryu KY, DeYoung BR, Smanik PA, McGaughy VR, Fischer AH, Mazzaferri EL.(1998). An immunohistochemical study of Na+/I- symporter in human thyroid tissues and salivary gland tissues. Endocrinology. 139:4416-4419. Kwee, Sandi A.; Coel, Marc N.; Fitz-Patrick, David (2007). Eary, Janet F.; Brenner, Winfried, eds. "Iodine-131 Radiotherapy for Benign Thyroid Disease". Nuclear Medicine Therapy. CRC Press: 172. <a href="https://en.wikipedia.org/wiki/International_Standard_Book_Number" title="International Standard Book Number">ISBN</a> <a href="https://en.wikipedia.org/wiki/Special:BookSources/978-0-8247-2876-2" title="Special:BookSources/978-0-8247-2876-2">978-0-8247-2876-2</a>. Nolan LA, Windle RJ, Wood SA, Kershaw YM, Lunness HR, Lightman SL, Ingram CD, Levy A. (2000). Chronic iodine deprivation attenuates stress-induced and diurnal variation in corticosterone secretion in female Wistar rats. J Neuroendocrinol. 12:1149-1159. Panneels V, Van den Bergen H, Jacoby C, Braekman JC, Van Sande J, Dumont JE, Boeynaems JM. (1994). Inhibition of H2O2 production by iodoaldehydes in cultured dog thyroid cells. Mol Cell Endocrinol. 102:167-176. Patrick L. (2008).Iodine:Deficiency and therapeutic considerations. Altern MedRev. 13:166-127. Ramesh BG, Bhargav PR, Rajesh BG, Devi NV, Vijayaraghavan R, Varma BA.(2016). Genotype‑phenotype correlations of dyshormonogenetic goiter in children and adolescents from South India . I J Endocrinol and Metab. 20: 816-824. Smyth PA. (2003). Role of iodine in antioxidant defense in thyroid and breast disease. Biofactors. 19:121-130. Spitzweg C, Morris JC. 2010. Genetics and phenomics of hypothyroidism and goiter due to NIS mutations. Molecular and cellular endocrinology. Jun 30;322:56-63. Wolff J, Chaikoff IL. (1948). Plasma inorganic iodide as a homeostatic regulator of thyroid function. J Biol Chem. 174: 555-564. AOPWiki 2016-11-29T18:41:24

2019-04-04T09:00:11

Thyroid hormone synthesis, Decreased

TH synthesis, Decreased

Cellular

The thyroid hormones (TH), triiodothyronine (T3) and thyroxine (T4) are thyrosine-based hormones. Synthesis of THs is regulated by thyroid-stimulating hormone (TSH) binding to its receptor and thyroidal availability of iodine via the sodium iodide symporter (NIS). Other proteins contributing to TH production in the thyroid gland, including thyroperoxidase (TPO), dual oxidase enzymes (DUOX), and the transport protein pendrin are also necessary for iodothyronine production (Zoeller et al., 2007). The production of THs in the thyroid gland and resulting serum concentrations are controlled by a negatively regulated feedback mechanism. Decreased T4 and T3 serum concentrations activates the hypothalamus-pituitary-thyroid (HPT) axis which upregulates thyroid-stimulating hormone (TSH) that acts to increase production of additional THs (Zoeller and Tan, 2007). This regulatory system includes: 1) the hypothalamic secretion of the thyrotropin-releasing hormone (TRH); 2) the thyroid-stimulating hormone (TSH) secretion from the anterior pituitary; 3) hormonal transport by the plasma binding proteins; 4) cellular uptake mechanisms at the tissue level; 5) intracellular control of TH concentrations by deiodinating mechanisms; 6) transcriptional function of the nuclear TH receptor; and 7) in the fetus, the transplacental passage of T4 and T3 (Zoeller et al., 2007). TRH and the TSH primarily regulate the production of T4, often considered a “pro-hormone,” and to a lesser extent of T3, the transcriptionally active TH. Most of the hormone released from the thyroid gland into circulation is in the form of T4, while peripheral deiodination of T4 is responsible for the majority of circulating T3. Outer ring deiodination of T4 to T3 is catalyzed by the deiodinases 1 and 2 (DIO1 and DIO2), with DIO1 expressed mainly in liver and kidney, and DIO2 expressed in several tissues including the brain (Bianco et al., 2006). Conversion of T4 to T3 takes place mainly in the liver and kidney, but also in other target organs such as in the brain, the anterior pituitary, brown adipose tissue, thyroid and skeletal muscle (Gereben et al., 2008; Larsen, 2009).  In mammals, most evidence for the ontogeny of TH synthesis comes from measurements of serum hormone concentrations. And, importantly, the impact of xenobiotics on fetal hormones must include the influence of the maternal compartment since a majority of fetal THs are derived from maternal blood early in fetal life, with a transition during mid-late gestation to fetal production of THs that is still supplemented by maternal THs. In humans, THs can be found in the fetus as early as gestational weeks 10-12, and concentations rise continuously until birth. At term, fetal T4 is similar to maternal levels, but T3 remains 2-3 fold lower than maternal levels. In rats, THs can be detected in the fetus as early as the second gestational week, but fetal synthesis does not start until gestational day 17 with birth at gestational day 22-23. Maternal THs continue to supplement fetal production until parturition. (see Howdeshell, 2002; Santisteban and Bernal, 2005 for review). Due to the maternal factor, the life stage specific impact of TPO inhibition after exposure to environmental chemicals is complex (Ramhoj et al., 2022). Decreased TH synthesis in the thyroid gland may result from several possible molecular-initiating events (MIEs) including: 1) Disruption of key catalytic enzymes or cofactors needed for TH synthesis, including TPO, NIS, or dietary iodine insufficiency. Theoretically, decreased synthesis of Tg could also affect TH production (Kessler et al., 2008; Yi et al., 1997). Mutations in genes that encode requisite proteins in the thyroid may also lead to impaired TH synthesis, including mutations in pendrin associated with Pendred Syndrome (Dossena et al., 2011), mutations in TPO and Tg (Huang and Jap 2015), and mutations in NIS (Spitzweg and Morris, 2010). 2) Decreased TH synthesis in cases of clinical hypothyroidism may be due to Hashimoto's thyroiditis or other forms of thyroiditis, or physical destruction of the thyroid gland as in radioablation or surgical treatment of thyroid lymphoma. 3) It is possible that TH synthesis may also be reduced subsequent to disruption of the negative feedback mechanism governing TH homeostasis, e.g. pituitary gland dysfunction may result in a decreased TSH signal with concomitant T3 and T4 decreases. 4) More rarely, hypothalamic dysfunction can result in decreased TH synthesis.  Increased fetal TH levels are also possible. Maternal Graves disease, which results in fetal thyrotoxicosis (hyperthyroidism and increased serum T4 levels), has been successfully treated by maternal administration of TPO inhibitors (c.f., Sato et al., 2014).   It should be noted that different species and different life stages store different amounts of TH precursors and iodine within the thyroid gland. Thus, decreased TH synthesis via transient iodine insufficiency or inhibition of TPO may not affect TH release from the thyroid gland until depletion of stored iodinated Tg. Adult humans may store sufficient Tg-DIT residues to serve for several months to a year of TH demand (Greer et al., 2002; Zoeller, 2004). Neonates and infants have a much more limited supply of less than a week. While the TH system is highly conserved across vertebrates, there are some taxon-specific considerations. Zebrafish and fathead minnows are oviparous fish species in which maternal THs are transferred to the eggs and regulate early embryonic developmental processes during external (versus intra-uterine in mammals) development (Power et al., 2001; Campinho et al., 2014; Ruuskanen and Hsu, 2018) until embryonic TH synthesis is initiated. Maternal transfer of THs to the eggs has been demonstrated in zebrafish (Walpita et al., 2007; Chang et al., 2012) and fathead minnows (Crane et al., 2004; Nelson et al., 2016). Decreases in TH synthesis can only occur after initiation of embryonic TH synthesis. The components of the TH system responsible for TH synthesis are highly conserved across vertebrates and therefore interference with the same molecular targets compared to mammals can lead to decreased TH synthesis (TPO, NIS, etc.) in fish. Endogenous transcription profiles of thyroid-related genes in zebrafish and fathead minnow showed that mRNA coding for these genes is also maternally transferred and increasing expression of most transcripts during hatching and embryo-larval transition indicates a fully functional HPT axis in larvae (Vergauwen et al., 2018). Although the HPT axis is highly conserved, there are some differences between fish and mammals (Blanton and Specker, 2007; Deal and Volkoff, 2020). For example, in fish, corticotropin releasing hormone (CRH) often plays a more important role in regulating thyrotropin (TSH) secretion by the pituitary and thus TH synthesis compared to TSH-releasing hormone (TRH). Also, in most fish species thyroid follicles are more diffusely located in the pharyngeal region rather than encapsulated in a gland.

Decreased TH synthesis is often implied by measurement of TPO and NIS inhibition measured clinically and in laboratory models as these enzymes are essential for TH synthesis. Rarely is decreased TH synthesis measured directly, but rather the impact of chemicals on the quantity of T4 produced in the thyroid gland, or the amount of T4 present in serum is used as a marker of decreased T4 release from the thyroid gland (e.g., Romaldini et al., 1988). Methods used to assess TH synthesis include, incorporation of radiolabeled tracer compounds, radioimmunoassay, ELISA, and analytical detection.    Recently, amphibian thyroid explant cultures have been used to demonstrate direct effects of chemicals on TH synthesis, as this model contains all necessary synthesis enzymes including TPO and NIS (Hornung et al., 2010). For this work THs was measured by HPLC/ICP-mass spectometry. Decreased TH synthesis and release, using T4 release as the endpoint, has been shown for thiouracil antihyperthyroidism drugs including MMI, PTU, and the NIS inhibitor perchlorate (Hornung et al., 2010). Techniques for in vivo analysis of TH system disruption among other drug-related effects in fish were reviewed by Raldua and Piña (2014). TIQDT (Thyroxine-immunofluorescence quantitative disruption test) is a method that provides an immunofluorescent based estimate of thyroxine in the gland of zebrafish (Raldua and Babin, 2009; Thienpont et al., 2011; Jomaa et al., 2014; Rehberger et al., 2018).  Thienpont used this method with ~25 xenobiotics (e.g., amitrole, perchlorate, methimazole, PTU, DDT, PCBs). The method detected changes for all chemicals known to directly impact TH synthesis in the thyroid gland (e.g., NIS and TPO inhibitors), but not those that upregulate hepatic catabolism of T4. Rehberger et al. (2018) updated the method to enable simultaneous semi-quantitative visualization of intrafollicular T3 and T4 levels. Most often, whole body TH level measurements in fish early life stages are used as indirect evidence of decreased TH synthesis (Nelson et al., 2016; Stinckens et al., 2016; Stinckens et al., 2020). Analytical determination of TH levels by LC-MS is becoming increasingly available (Hornung et al., 2015). More recently, transgenic zebrafish with fluorescent thyroid follicles are being used to visualize the compensatory proliferation of the thyroid follicles following inhibition of TH synthesis among others (Opitz et al., 2012).

Taxonomic: This KE is plausibly applicable across vertebrates. Decreased TH synthesis resulting from TPO or NIS inhibition is conserved across vertebrate taxa, with in vivo evidence from humans, rats, amphibians, some fish species, and birds, and in vitro evidence from rat and porcine microsomes. Indeed, TPO and NIS mutations result in congenital hypothyroidism in humans (Bakker et al., 2000; Spitzweg and Morris, 2010), demonstrating the essentiality of TPO and NIS function toward maintaining euthyroid status. Though decreased serum T4 is used as a surrogate measure to indicate chemical-mediated decreases in TH synthesis, clinical and veterinary management of hyperthyroidism and Graves’ disease using propylthiouracil and methimazole, known to decrease TH synthesis, indicates strong evidence for chemical inhibition of TPO (Zoeller and Crofton, 2005). Life stage: Applicability to certain life stages may depend on the species and their dependence on maternally transferred THs during the earliest phases of development. The earliest life stages of teleost fish (e.g., fathead minnow, zebrafish) rely on maternally transferred THs to regulate certain developmental processes until embryonic TH synthesis is active (Power et al., 2001). In externally developing fish species, decreases in TH synthesis can only occur after initiation of embryonic TH synthesis. In zebrafish, Opitz et al. (2011) showed the formation of a first thyroid follicle at 55 hours post fertilization (hpf), Chang et al. (2012) showed a first significant TH increase at 120 hpf and Walter et al. (2019) showed clear TH production already at 72 hpf but did not analyse time points between 24 and 72 hpf. TPO inhibition in a homozygous knockout line abolished the T4 production in thyroid follicles of mutant zebrafish with phenotypic abnormalities occurring from 20 dpf onwards but not before 10 dpf (Fang et al., 2022). Therefore, it is still uncertain when exactly embryonic TH synthesis is activated and thus when exactly this process becomes sensitive to disruption. In fathead minnows, a significant increase of whole body TH levels was already observed between 1 and 2 dpf, which corresponds to the appearance of the thyroid anlage at 35 hpf prior to the first observation of thyroid follicles at 58 hpf (Wabuke-Bunoti and Firling, 1983). It currently remains unclear when exactly embryonic TH production is initiated in zebrafish. Sex: The KE is plausibly applicable to both sexes. THs are essential in both sexes and the components of the HPT-axis are identical in both sexes. There can however be sex-dependent differences in the sensitivity to the disruption of TH levels and the magnitude of the response. In humans, females appear more susceptible to hypothyroidism compared to males when exposed to certain halogenated chemicals (Hernandez‐Mariano et al., 2017; Webster et al., 2014). In adult zebrafish, Liu et al. (2019) showed sex-dependent changes in TH levels and mRNA expression of regulatory genes including corticotropin releasing hormone (crh), thyroid stimulating hormone (tsh) and deiodinase 2 after exposure to organophosphate flame retardants. The underlying mechanism of any sex-related differences remains unclear.

UBERON:0002046

UBERON thyroid gland

CL:0002258

CL thyroid follicular cell

High Male High Female

High

All life stages

High High Moderate High Moderate High

Bakker B, Bikker H, Vulsma T, de Randamie JS, Wiedijk BM, De Vijlder JJ. 2000. Two decades of screening for congenital hypothyroidism in The Netherlands: TPO gene mutations in total iodide organification defects (an update). The Journal of clinical endocrinology and metabolism.  85:3708-3712. Bianco AC, Kim BW. (2006). Deiodinases: implications of the local control of thyroid hormone action. J Clin Invest. 116: 2571–2579. Blanton ML, Specker JL. 2007. The hypothalamic-pituitary-thyroid (hpt) axis in fish and its role in fish development and reproduction. Crit Rev Toxicol. 37(1-2):97-115. Campinho MA, Saraiva J, Florindo C, Power DM. 2014. Maternal thyroid hormones are essential for neural development in zebrafish. Molecular Endocrinology. 28(7):1136-1149. Chang J, Wang M, Gui W, Zhao Y, Yu L, Zhu G. 2012. Changes in thyroid hormone levels during zebrafish development. Zoological Science. 29(3):181-184. Crane HM, Pickford DB, Hutchinson TH, Brown JA. 2004. Developmental changes of thyroid hormones in the fathead minnow, pimephales promelas. General and Comparative Endocrinology. 139(1):55-60. Deal CK, Volkoff H. 2020. The role of the thyroid axis in fish. Frontiers in Endocrinology. 11. Dossena S, Nofziger C, Brownstein Z, Kanaan M, Avraham KB, Paulmichl M. (2011). Functional characterization of pendrin mutations found in the Israeli and Palestinian populations. Cell Physiol Biochem. 28: 477-484.Gereben B, Zavacki AM, Ribich S, Kim BW, Huang SA, Simonides WS, Zeöld A, Bianco AC. (2008). Cellular and molecular basis of deiodinase-regulated thyroid hormone signalling. Endocr Rev. 29:898–938. Fang, Y., Wan, J. P., Zhang, R. J., Sun, F., Yang, L., Zhao, S. X., Dong, M., & Song, H. D. (2022). Tpo knockout in zebrafish partially recapitulates clinical manifestations of congenital hypothyroidism and reveals the involvement of TH in proper development of glucose homeostasis. General and Comparative Endocrinology, 323–324. https://doi.org/10.1016/j.ygcen.2022.114033 Gereben B, Zeöld A, Dentice M, Salvatore D, Bianco AC.  Activation and inactivation of thyroid hormone by deiodinases: local action with general consequences.  Cell Mol Life Sci. 2008 Feb;65(4):570-90 Greer MA, Goodman G, Pleus RC, Greer SE. Health effects assessment for environmental perchlorate contamination: the dose response for inhibition of thyroidal radioiodine uptake in humans. Environ Health Perspect. 2002. 110:927-937. Hernandez-Mariano JA, Torres-Sanchez L, Bassol-Mayagoitia S, Escamilla-Nunez M, Cebrian ME, Villeda-Gutierrez EA, Lopez-Rodriguez G, Felix-Arellano EE, Blanco-Munoz J. 2017. Effect of exposure to p,p '-dde during the first half of pregnancy in the maternal thyroid profile of female residents in a mexican floriculture area. Environmental Research. 156:597-604. Hornung MW, Degitz SJ, Korte LM, Olson JM, Kosian PA, Linnum AL, Tietge JE. 2010. Inhibition of thyroid hormone release from cultured amphibian thyroid glands by methimazole, 6-propylthiouracil, and perchlorate. Toxicol Sci 118:42-51. Hornung MW, Kosian PA, Haselman JT, Korte JJ, Challis K, Macherla C, Nevalainen E, Degitz SJ. 2015. In vitro, ex vivo, and in vivo determination of thyroid hormone modulating activity of benzothiazoles. Toxicological Sciences. 146(2):254-264. Howdeshell KL. 2002. A model of the development of the brain as a construct of the thyroid system. Environ Health Perspect. 110 Suppl 3:337-48. Huang CJ and Jap TS. 2015. A systematic review of genetic studies of thyroid disorders in Taiwan. J Chin Med Assoc. 78: 145-153. Jomaa B, Hermsen SAB, Kessels MY, van den Berg JHJ, Peijnenburg AACM, Aarts JMMJG, Piersma AH, Rietjens IMCM. 2014. Developmental toxicity of thyroid-active compounds in a zebrafish embryotoxicity test. Altex-Alternatives to Animal Experimentation. 31(3):303-317. Kessler J, Obinger C, Eales G. Factors influencing the study of peroxidase-generated iodine species and implications for thyroglobulin synthesis. Thyroid. 2008 Jul;18(7):769-74. doi: 10.1089/thy.2007.0310 Larsen PR. (2009). Type 2 iodothyronine deiodinase in human skeletal muscle: new insights into its physiological role and regulation. J Clin Endocrinol Metab. 94:1893-1895. Liu XS, Cai Y, Wang Y, Xu SH, Ji K, Choi K. 2019. Effects of tris(1,3-dichloro-2-propyl) phosphate (tdcpp) and triphenyl phosphate (tpp) on sex-dependent alterations of thyroid hormones in adult zebrafish. Ecotoxicology and Environmental Safety. 170:25-32. Nelson K, Schroeder A, Ankley G, Blackwell B, Blanksma C, Degitz S, Flynn K, Jensen K, Johnson R, Kahl M et al. 2016. Impaired anterior swim bladder inflation following exposure to the thyroid peroxidase inhibitor 2-mercaptobenzothiazole part i: Fathead minnow. Aquatic Toxicology. 173:192-203. Opitz R, Maquet E, Huisken J, Antonica F, Trubiroha A, Pottier G, Janssens V, Costagliola S. 2012. Transgenic zebrafish illuminate the dynamics of thyroid morphogenesis and its relationship to cardiovascular development. Developmental Biology. 372(2):203-216. Opitz R, Maquet E, Zoenen M, Dadhich R, Costagliola S. 2011. Tsh receptor function is required for normal thyroid differentiation in zebrafish. Molecular Endocrinology. 25(9):1579-1599. Power DM, Llewellyn L, Faustino M, Nowell MA, Bjornsson BT, Einarsdottir IE, Canario AV, Sweeney GE. 2001. Thyroid hormones in growth and development of fish. Comp Biochem Physiol C Toxicol Pharmacol. 130(4):447-459. Raldua D, Babin PJ. 2009. Simple, rapid zebrafish larva bioassay for assessing the potential of chemical pollutants and drugs to disrupt thyroid gland function. Environmental Science & Technology. 43(17):6844-6850. Raldua D, Pina B. 2014. In vivo zebrafish assays for analyzing drug toxicity. Expert Opinion on Drug Metabolism & Toxicology. 10(5):685-697. Ramhoj, L., Svingen, T., Fradrich, C., Rijntjes, E., Wirth, E.K., Pedersen, K., Kohrle, J., Axelstad, M., 2022. Perinatal exposure to the thyroperoxidase inhibitors methimazole and amitrole perturbs thyroid hormone system signaling and alters motor activity in rat offspring. Toxicology Letters 354, 44-55. Rehberger K, Baumann L, Hecker M, Braunbeck T. 2018. Intrafollicular thyroid hormone staining in whole-mount zebrafish (danio rerio) embryos for the detection of thyroid hormone synthesis disruption. Fish Physiology and Biochemistry. 44(3):997-1010. Romaldini JH, Farah CS, Werner RS, Dall'Antonia Júnior RP, Camargo RS. 1988.  "In vitro" study on release of cyclic AMP and thyroid hormone in autonomously functioning thyroid nodules.  Horm Metab Res.20:510-2. Ruuskanen S, Hsu BY. 2018. Maternal thyroid hormones: An unexplored mechanism underlying maternal effects in an ecological framework. Physiological and Biochemical Zoology. 91(3):904-916. Santisteban P, Bernal J. Thyroid development and effect on the nervous system. Rev Endocr Metab Disord. 2005 Aug;6(3):217-28. Spitzweg C, Morris JC. 2010. Genetics and phenomics of hypothyroidism and goiter due to NIS mutations. Molecular and cellular endocrinology. 322:56-63. Stinckens E, Vergauwen L, Blackwell BR, Anldey GT, Villeneuve DL, Knapen D. 2020. Effect of thyroperoxidase and deiodinase inhibition on anterior swim bladder inflation in the zebrafish. Environmental Science & Technology. 54(10):6213-6223. Stinckens E, Vergauwen L, Schroeder A, Maho W, Blackwell B, Witters H, Blust R, Ankley G, Covaci A, Villeneuve D et al. 2016. Impaired anterior swim bladder inflation following exposure to the thyroid peroxidase inhibitor 2-mercaptobenzothiazole part ii: Zebrafish. Aquatic Toxicology. 173:204-217. Thienpont B, Tingaud-Sequeira A, Prats E, Barata C, Babin PJ, Raldúa D.  Zebrafish eleutheroembryos provide a suitable vertebrate model for screening chemicals that impair thyroid hormone synthesis.  Environ Sci Technol. 2011. 45(17):7525-32. Vergauwen L, Cavallin JE, Ankley GT, Bars C, Gabriels IJ, Michiels EDG, Fitzpatrick KR, Periz-Stanacev J, Randolph EC, Robinson SL et al. 2018. Gene transcription ontogeny of hypothalamic-pituitary-thyroid axis development in early-life stage fathead minnow and zebrafish. General and Comparative Endocrinology. 266:87-100. Wabukebunoti MAN, Firling CE. 1983. The prehatching development of the thyroid-gland of the fathead minnow, pimephales-promelas (rafinesque). General and Comparative Endocrinology. 49(2):320-331. Walpita CN, Van der Geyten S, Rurangwa E, Darras VM. 2007. The effect of 3,5,3'-triiodothyronine supplementation on zebrafish (danio rerio) embryonic development and expression of iodothyronine deiodinases and thyroid hormone receptors. Gen Comp Endocrinol. 152(2-3):206-214. Walter KM, Miller GW, Chen XP, Yaghoobi B, Puschner B, Lein PJ. 2019. Effects of thyroid hormone disruption on the ontogenetic expression of thyroid hormone signaling genes in developing zebrafish (danio rerio). General and Comparative Endocrinology. 272:20-32. Webster GM, Venners SA, Mattman A, Martin JW. 2014. Associations between perfluoroalkyl acids (pfass) and maternal thyroid hormones in early pregnancy: A population-based cohort study. Environmental Research. 133:338-347. Yi X, Yamamoto K, Shu L, Katoh R, Kawaoi A. Effects of Propyithiouracil (PTU) Administration on the Synthesis and Secretion of Thyroglobulin in the Rat Thyroid Gland: A Quantitative Immuno-electron Microscopic Study Using Immunogold Technique. Endocr Pathol. 1997 Winter;8(4):315-325. Zoeller RT, Crofton KM. 2005.  Mode of action: developmental thyroid hormone insufficiency--neurological abnormalities resulting from exposure to propylthiouracil. Crit Rev Toxicol. 35:771-81 Zoeller RT, Tan SW, Tyl RW. 2007. General background on the hypothalamic-pituitary-thyroid (HPT) axis. Critical reviews in toxicology.  37:11-53. Zoeller RT.  Interspecies differences in susceptibility to perturbation of thyroid hormone homeostasis requires a definition of "sensitivity" that is informative for risk analysis. Regul Toxicol Pharmacol. 2004 Dec;40(3):380.   AOPWiki 2016-11-29T18:41:23

2022-11-04T09:25:39

Thyroxine (T4) in serum, Decreased

T4 in serum, Decreased

Tissue

All iodothyronines are derived from the modification of tyrosine molecules (Taurog, 2000). There are two biologically active thyroid hormones (THs) in serum, triiodothyronine (T3) and T4, and a few less active iodothyronines, reverse T3 (rT3),  and 3,3'-Diiodothyronine (3,5-T2). T4 is the predominant TH in circulation, comprising approximately 80% of the TH excreted from the thyroid gland in mammals and is the pool from which the majority of T3 in serum is generated (Zoeller et al., 2007). As such, serum T4 changes usually precede changes in other serum THs. Decreased thyroxine (T4) in serum results from one or more MIEs upstream and is considered a key biomarker of altered TH homeostasis (DeVito et al., 1999). Serum T4 is used as a biomarker of TH status because the circulatory system serves as the major transport and delivery system for TH delivery to tissues. The majority of THs in the blood are bound to transport proteins (Bartalena and Robbins, 1993). In serum, it is the unbound, or ‘free’ form of the hormone that is thought to be available for transport into tissues. Free hormones are approximately 0.03 and 0.3 percent for T4 and T3, respectively. There are major species differences in the predominant binding proteins and their affinities for THs (see below). However, there is broad agreement that changes in serum concentrations of THs is diagnostic of thyroid disease or chemical-induced disruption of thyroid homeostasis across vertebrates (DeVito et al., 1999; Miller et al., 2009; Zoeller et al., 2007; Carr and Patiño, 2011). Normal serum T4 reference ranges can be species and lifestage specific. In rodents, serum THs are low in the fetal circulation, increasing as the fetal thyroid gland becomes functional on gestational day 17, just a few days prior to birth. After birth serum hormones increase steadily, peaking at two weeks, and falling slightly to adult levels by postnatal day 21 (Walker et al., 1980; Harris et al., 1978; Goldey et al., 1995; Lau et al., 2003). Similarly, in humans, adult reference ranges for THs do not reflect the normal ranges for children at different developmental stages, with TH concentrations highest in infants, still increased in childhood, prior to a decline to adult levels coincident with pubertal development (Corcoran et al. 1977; Kapelari et al., 2008). In some frog species, there is an analogous peak in THs in tadpoles that starts around embryonic NF stage 56, peaks at stage 62 and the declines to lower levels by stage 56 (Sternberg et al., 2011; Leloup and Buscaglia, 1977).  Additionally, ample evidence is available from studies investigating responses to inhibitors of TH synthesis in fish. For example, Stinckens et al. (2020) showed reduced whole body T4 concentrations in zebrafish larvae exposed to 50 or 100 mg/L methimazole, a potent TPO inhibitor, from immediately after fertilization until 21 or 32 days of age. Exposure to 37 or 111 mg/L propylthiouracil also reduced T4 levels after exposure up to 14, 21 and 32 days in the same study. Walter et al. (2019) showed that propylthiouracil had no effect on T4 levels in 24h old zebrafish, but decreased T4 levels of 72h old zebrafish. This difference is probably due to the onset of embryonic TH production between the age of 24 and 72 hours (Opitz et al., 2011). Stinckens et al. (2016) showed that exposure to 2-mercaptobenzothiazole (MBT), an environmentally relevant TPO inhibitor, decreased whole body T4 levels in continuously exposed 5 and 32 day old zebrafish larvae. A high concentration of MBT also decreased whole body T4 levels in 6 day old fathead minnows, but recovery was observed at the age of 21 days although the fish were kept in the exposure medium (Nelson et al., 2016). Crane et al. (2006) showed decreased T4 levels in 28 day old fathead minnows continuously exposed to 32 or 100 µg/L methimazole.

Serum T3 and T4 can be measured as free (unbound) or total (bound + unbound). Free hormone concentrations are clinically considered more direct indicators of T4 and T3 activities in the body, but in animal studies, total T3 and T4 are typically measured. Historically, the most widely used method in toxicology is the radioimmunoassay (RIA). The method is routinely used in rodent endocrine and toxicity studies. The ELISA method is commonly used as a human clinical test method. Analytical determination of iodothyronines (T3, T4, rT3, T2) and their conjugates, through methods employing HPLC, liquid chromatography, immuno luminescence, and mass spectrometry are less common, but are becoming increasingly available (Hornung et al., 2015; DeVito et al., 1999; Baret and Fert, 1989; Spencer, 2013; Samanidou V.F et al., 2000; Rathmann D. et al., 2015 ). In fish early life stages most evidence for the ontogeny of thyroid hormone synthesis comes from measurements of whole body thyroid hormone levels using LC-MS techniques (Hornung et al., 2015) which are increasingly used to accurately quantify whole body thyroid hormone levels as a proxy for serum thyroid hormone levels (Nelson et al., 2016; Stinckens et al., 2016; Stinckens et al., 2020). It is important to note that thyroid hormones concentrations can be influenced by a number of intrinsic and extrinsic factors (e.g., circadian rhythms, stress, food intake, housing, noise) (see for example, Döhler et al., 1979). Any of these measurements should be evaluated for the relationship to the actual endpoint of interest, repeatability, reproducibility, and lower limits of quantification using a fit-for-purpose approach. This is of particular significance when assessing the very low levels of TH present in fetal serum. Detection limits of the assay must be compatible with the levels in the biological sample. All three of the methods summarized above would be fit-for-purpose, depending on the number of samples to be evaluated and the associated costs of each method. Both RIA and ELISA measure THs by an indirect methodology, whereas analytical determination is the most direct measurement available. All these methods, particularly RIA, are repeatable and reproducible.

Taxonomic: This KE is plausibly applicable across vertebrates and the overall evidence supporting taxonomic applicability is strong. THs are evolutionarily conserved molecules present in all vertebrate species (Hulbert, 2000; Yen, 2001). Moreover, their crucial role in zebrafish development, embryo-to-larval transition and larval-to-juvenile transition (Thienpont et al., 2011; Liu and Chan, 2002), and amphibian and lamprey metamorphoses is well established (Manzon and Youson, 1997; Yaoita and Brown, 1990; Furlow and Neff, 2006). Their role as environmental messenger via exogenous routes in echinoderms confirms the hypothesis that these molecules are widely distributed among the living organisms (Heyland and Hodin, 2004). However, the role of THs in the different species depends on the expression and function of specific proteins (e.g receptors or enzymes) under TH control and may vary across species and tissues. As such, extrapolation regarding TH action across species and developmental stages should be done with caution. With few exceptions, vertebrate species have circulating T4 (and T3) that are bound to transport proteins in blood. Clear species differences exist in serum transport proteins (Dohler et al., 1979; Yamauchi and Isihara, 2009). There are three major transport proteins in mammals; thyroid binding globulin (TBG), transthyretin (TTR), and albumin. In adult humans, the percent bound to these proteins is about 75, 15 and 10 percent, respectively (Schussler 2000).  In contrast, in adult rats the majority of THs are bound to TTR. Thyroid- binding proteins are developmentally regulated in rats. TBG is expressed in rats until approximately postnatal day (PND) 60, with peak expression occurring during weaning (Savu et al., 1989). However, low levels of TBG persist into adult ages in rats and can be experimentally induced by hypothyroidism, malnutrition, or caloric restriction (Rouaze-Romet et al., 1992). While these species differences impact TH half-life (Capen, 1997) and possibly regulatory feedback mechanisms, there is little information on quantitative dose-response relationships of binding proteins and serum hormones during development across different species. Serum THs are still regarded as the most robust measurable key event causally linked to downstream adverse outcomes. Life stage: The earliest life stages of teleost fish rely on maternally transferred THs to regulate certain developmental processes until embryonic TH synthesis is active (Power et al., 2001). As a result, T4 levels are not expected to decrease in response to exposure to inhibitors of TH synthesis during these earliest stages of development. In zebrafish, Opitz et al. (2011) showed the formation of a first thyroid follicle at 55 hours post fertilization (hpf), Chang et al. (2012) showed a first significant TH increase at 120 hpf and Walter et al. (2019) showed clear TH production already at 72 hpf but did not analyse time points between 24 and 72 hpf. In fathead minnows, a significant increase of whole body TH levels was already observed between 1 and 2 dpf, which corresponds to the appearance of the thyroid anlage at 35 hpf prior to the first observation of thyroid follicles at 58 hpf (Wabuke-Bunoti and Firling, 1983). It is still uncertain when exactly embryonic TH synthesis is activated and how this determines sensitivity to TH system disruptors. Sex: The KE is plausibly applicable to both sexes. THs are essential in both sexes and the components of the HPT-axis are identical in both sexes. There can however be sex-dependent differences in the sensitivity to the disruption of TH levels and the magnitude of the response. In humans, females appear more susceptible to hypothyroidism compared to males when exposed to certain halogenated chemicals (Hernandez‐Mariano et al., 2017; Webster et al., 2014). In adult zebrafish, Liu et al. (2019) showed sex-dependent changes in TH levels and mRNA expression of regulatory genes including corticotropin releasing hormone (crh), thyroid stimulating hormone (tsh) and deiodinase 2 after exposure to organophosphate flame retardants. The underlying mechanism of any sex-related differences remains unclear.

UBERON:0001977

UBERON serum

High Female High Male

High

All life stages

High High High Moderate Moderate High High High

Axelrad DA, Baetcke K, Dockins C, Griffiths CW, Hill RN, Murphy PA, Owens N, Simon NB, Teuschler LK. Risk assessment for benefits analysis: framework for analysis of a thyroid-disrupting chemical. J Toxicol Environ Health A. 2005 68(11-12):837-55. Baret A. and Fert V.  T4 and ultrasensitive TSH immunoassays using luminescent enhanced xanthine oxidase assay. J Biolumin Chemilumin. 1989. 4(1):149-153 Bartalena L, Robbins J. Thyroid hormone transport proteins. Clin Lab Med. 1993 Sep;13(3):583-98. Bassett JH, Harvey CB, Williams GR. (2003). Mechanisms of thyroid hormone receptor-specific nuclear and extra nuclear actions. Mol Cell Endocrinol. 213:1-11. Capen CC. Mechanistic data and risk assessment of selected toxic end points of the thyroid gland. Toxicol Pathol. 1997 25(1):39-48. Carr JA, Patino R. 2011. The hypothalamus-pituitary-thyroid axis in teleosts and amphibians: Endocrine disruption and its consequences to natural populations. General and Comparative Endocrinology. 170(2):299-312. Chang J, Wang M, Gui W, Zhao Y, Yu L, Zhu G. 2012. Changes in thyroid hormone levels during zebrafish development. Zoological Science. 29(3):181-184. Cope RB, Kacew S, Dourson M. A reproductive, developmental and neurobehavioral study following oral exposure of tetrabromobisphenol A on Sprague-Dawley rats. Toxicology. 2015 329:49-59. Corcoran JM, Eastman CJ, Carter JN, Lazarus L. (1977). Circulating thyroid hormone levels in children. Arch Dis Child. 52: 716-720. Crane HM, Pickford DB, Hutchinson TH, Brown JA. 2006. The effects of methimazole on development of the fathead minnow, pimephales promelas, from embryo to adult. Toxicological Sciences. 93(2):278-285. Crofton KM. Developmental disruption of thyroid hormone: correlations with hearing dysfunction in rats. Risk Anal. 2004 Dec;24(6):1665-71. DeVito M, Biegel L, Brouwer A, Brown S, Brucker-Davis F, Cheek AO, Christensen R, Colborn T, Cooke P, Crissman J, Crofton K, Doerge D, Gray E, Hauser P, Hurley P, Kohn M, Lazar J, McMaster S, McClain M, McConnell E, Meier C, Miller R, Tietge J, Tyl R. (1999). Screening methods for thyroid hormone disruptors. Environ Health Perspect. 107:407-415. Döhler KD, Wong CC, von zur Mühlen A (1979).   The rat as model for the study of drug effects on thyroid function: consideration of methodological problems.  Pharmacol Ther B. 5:305-18. Eales JG. (1997). Iodine metabolism and thyroid related functions in organisms lacking thyroid follicles: Are thyroid hormones also vitaminsProc Soc Exp Biol Med. 214:302-317. Furlow JD, Neff ES. (2006). A developmental switch induced by thyroid hormone: Xenopus laevis metamorphosis. Trends Endocrinol Metab. 17:40–47. Goldey ES, Crofton KM. Thyroxine replacement attenuates hypothyroxinemia, hearing loss, and motor deficits following developmental exposure to Aroclor 1254 in rats. Toxicol Sci. 1998 45(1):94-10 Goldey ES, Kehn LS, Lau C, Rehnberg GL, Crofton KM.  Developmental exposure to polychlorinated biphenyls (Aroclor 1254) reduces circulating thyroid hormone concentrations and causes hearing deficits in rats. Tox Appl Pharmacol. 1995 135(1):77-88. Harris AR, Fang SL, Prosky J, Braverman LE, Vagenakis AG.  Decreased outer ring monodeiodination of thyroxine and reverse triiodothyronine in the fetal and neonatal rat.  Endocrinology. 1978 Dec;103(6):2216-22 Hernandez-Mariano JA, Torres-Sanchez L, Bassol-Mayagoitia S, Escamilla-Nunez M, Cebrian ME, Villeda-Gutierrez EA, Lopez-Rodriguez G, Felix-Arellano EE, Blanco-Munoz J. 2017. Effect of exposure to p,p '-dde during the first half of pregnancy in the maternal thyroid profile of female residents in a mexican floriculture area. Environmental Research. 156:597-604. Heyland A, Hodin J. (2004). Heterochronic developmental shift caused by thyroid hormone in larval sand dollars and its implications for phenotypic plasticity and the evolution of non-feeding development. Evolution. 58: 524-538. Heyland A, Moroz LL. (2005). Cross-kingdom hormonal signaling: an insight from thyroid hormone functions in marine larvae. J Exp Biol. 208:4355-4361. Hill RN, Crisp TM, Hurley PM, Rosenthal SL, Singh DV. Risk assessment of thyroid follicular cell tumors.  Environ Health Perspect. 1998 Aug;106(8):447-57. Hornung MW, Kosian P, Haselman J, Korte J, Challis K, Macherla C, Nevalainen E, Degitz S (2015) In vitro, ex vivo and in vivo determination of thyroid hormone modulating activity of benzothiazoles. Toxicol Sci 146:254-264. Hulbert AJ. Thyroid hormones and their effects: a new perspective. Biol Rev Camb Philos Soc. 2000 Nov;75(4):519-631. Review. Kapelari K, Kirchlechner C, Högler W, Schweitzer K, Virgolini I, Moncayo R. 2008. Pediatric reference intervals for thyroid hormone levels from birth to adulthood: a retrospective study. BMC Endocr Disord. 8: 15. Lau C, Thibodeaux JR, Hanson RG, Rogers JM, Grey BE, Stanton ME, Butenhoff JL, Stevenson LA.  Exposure to perfluorooctane sulfonate during pregnancy in rat and mouse. II: postnatal evaluation.  Toxicol Sci. 2003 Aug;74(2):382-92. Leloup, J., and M. Buscaglia. La triiodothyronine: hormone de la métamorphose des amphibiens. CR Acad Sci 284 (1977): 2261-2263. Liu J, Liu Y, Barter RA, Klaassen CD.: Alteration of thyroid homeostasis by UDP-glucuronosyltransferase inducers in rats: a dose-response study. J Pharmacol Exp Ther 273, 977-85, 1994 Liu XS, Cai Y, Wang Y, Xu SH, Ji K, Choi K. 2019. Effects of tris(1,3-dichloro-2-propyl) phosphate (tdcpp) and triphenyl phosphate (tpp) on sex-dependent alterations of thyroid hormones in adult zebrafish. Ecotoxicology and Environmental Safety. 170:25-32. Liu YW, Chan WK. 2002. Thyroid hormones are important for embryonic to larval transitory phase in zebrafish. Differentiation. 70(1):36-45. Manzon RG, Youson JH. (1997). The effects of exogenous thyroxine (T4) or triiodothyronine (T3), in the presence and absence of potassium perchlorate, on the incidence of metamorphosis and on serum T4 and T3 concentrations in larval sea lampreys (Petromyzon marinus L.). Gen Comp Endocrinol. 106:211-220.  McClain RM. Mechanistic considerations for the relevance of animal data on thyroid neoplasia to human risk assessment. Mutat Res. 1995 Dec;333(1-2):131-42 Miller MD, Crofton KM, Rice DC, Zoeller RT.  Thyroid-disrupting chemicals: interpreting upstream biomarkers of adverse outcomes. Environ Health Perspect. 2009 117(7):1033-41 Morse DC, Wehler EK, Wesseling W, Koeman JH, Brouwer A. Alterations in rat brain thyroid hormone status following pre- and postnatal exposure to polychlorinated biphenyls (Aroclor 1254). Toxicol Appl Pharmacol. 1996 Feb;136(2):269-79. Nelson K, Schroeder A, Ankley G, Blackwell B, Blanksma C, Degitz S, Flynn K, Jensen K, Johnson R, Kahl M et al. 2016. Impaired anterior swim bladder inflation following exposure to the thyroid peroxidase inhibitor 2-mercaptobenzothiazole part i: Fathead minnow. Aquatic Toxicology. 173:192-203. NTP National Toxicology Program.: NTP toxicology and carcinogenesis studies of 3,3'-dimethylbenzidine dihydrochloride (CAS no. 612-82-8) in F344/N rats (drinking water studies). Natl Toxicol Program Tech Rep Ser 390, 1-238, 1991. O'Connor, J. C., J. C. Cook, et al. (1998). "An ongoing validation of a Tier I screening battery for detecting endocrine-active compounds (EACs)." Toxicol Sci 46(1): 45-60. O'Connor, J. C., L. G. Davis, et al. (2000). "Detection of dopaminergic modulators in a tier I screening battery for identifying endocrine-active compounds (EACs)." Reprod Toxicol 14(3): 193-205. Opitz R, Maquet E, Zoenen M, Dadhich R, Costagliola S. 2011. Tsh receptor function is required for normal thyroid differentiation in zebrafish. Molecular Endocrinology. 25(9):1579-1599. Power DM, Llewellyn L, Faustino M, Nowell MA, Bjornsson BT, Einarsdottir IE, Canario AV, Sweeney GE. 2001. Thyroid hormones in growth and development of fish. Comp Biochem Physiol C Toxicol Pharmacol. 130(4):447-459. Rathmann D, Rijntjes E, Lietzow J, Köhrle J. (2015) Quantitative Analysis of Thyroid Hormone Metabolites in Cell Culture Samples Using LC-MS/MS. Eur Thyroid J. Sep;4(Suppl 1):51-8. Rouaze-Romet M, Savu L, Vranckx R, Bleiberg-Daniel F, Le Moullac B, Gouache P, Nunez EA. 1992. Reexpression of thyroxine-binding globulin in postweaning rats during protein or energy malnutrition. Acta Endocrinol (Copenh).127:441-448. Samanidou VF, Kourti PV. (2009) Rapid HPLC method for the simultaneous monitoring of duloxetine, venlaflaxine, fluoxetine and paroxetine in biofluids. Bioanalysis. 2009 Aug;1(5):905-17. Savu L, Vranckx R, Maya M, Gripois D, Blouquit MF, Nunez EA. 1989. Thyroxine-binding globulin and thyroxinebinding prealbumin in hypothyroid and hyperthyroid developing rats. BiochimBiophys Acta. 992:379-384. Schneider S, Kaufmann W, Strauss V, van Ravenzwaay B.    Vinclozolin: a feasibility and sensitivity study of the ILSI-HESI F1-extended one-generation rat reproduction protocol. Regul Toxicol Pharmacol. 2011 Feb;59(1):91-100. Schussler, G.C. (2000). The thyroxine-binding proteins. Thyroid 10:141–149. Spencer, CA. (2013). Assay of thyroid hormone and related substances. In De Groot, LJ et al. (Eds). Endotext. South Dartmouth, MA Sternberg RM, Thoemke KR, Korte JJ, Moen SM, Olson JM, Korte L, Tietge JE, Degitz SJ Jr. Control of pituitary thyroid-stimulating hormone synthesis and secretion by thyroid hormones during Xenopus metamorphosis. Gen Comp Endocrinol. 2011. 173(3):428-37 Stinckens E, Vergauwen L, Blackwell BR, Anldey GT, Villeneuve DL, Knapen D. 2020. Effect of thyroperoxidase and deiodinase inhibition on anterior swim bladder inflation in the zebrafish. Environmental Science & Technology. 54(10):6213-6223. Stinckens E, Vergauwen L, Schroeder A, Maho W, Blackwell B, Witters H, Blust R, Ankley G, Covaci A, Villeneuve D et al. 2016. Impaired anterior swim bladder inflation following exposure to the thyroid peroxidase inhibitor 2-mercaptobenzothiazole part ii: Zebrafish. Aquatic Toxicology. 173:204-217. Taurog A. 2005. Hormone synthesis. In: Werner and Ingbar’s The Thyroid: A Fundamental and Clinical Text (Braverman LE, Utiger RD, eds). Philadelphia:Lippincott, Williams and Wilkins, 47–81Walker P, Dubois JD, Dussault JH.  Free thyroid hormone concentrations during postnatal development in the rat.  Pediatr Res. 1980 Mar;14(3):247-9. Thienpont B, Tingaud-Sequeira A, Prats E, Barata C, Babin PJ, Raldúa D. Zebrafish eleutheroembryos provide a suitable vertebrate model for screening chemicals that impair thyroid hormone synthesis. Environ Sci Technol. 2011 Sep 1;45(17):7525-32. Wabukebunoti MAN, Firling CE. 1983. The prehatching development of the thyroid-gland of the fathead minnow, pimephales-promelas (rafinesque). General and Comparative Endocrinology. 49(2):320-331. Walter KM, Miller GW, Chen XP, Yaghoobi B, Puschner B, Lein PJ. 2019. Effects of thyroid hormone disruption on the ontogenetic expression of thyroid hormone signaling genes in developing zebrafish (danio rerio). General and Comparative Endocrinology. 272:20-32. Webster GM, Venners SA, Mattman A, Martin JW. 2014. Associations between perfluoroalkyl acids (pfass) and maternal thyroid hormones in early pregnancy: A population-based cohort study. Environmental Research. 133:338-347. Yamauchi K1, Ishihara A. Evolutionary changes to transthyretin: developmentally regulated and tissue-specific gene expression. FEBS J. 2009. 276(19):5357-66. Yaoita Y, Brown DD. (1990). A correlation of thyroid hormone receptor gene expression with amphibian metamorphosis. Genes Dev. 4:1917-1924. Yen PM. (2001). Physiological and molecular basis of thyroid hormone action. Physiol Rev. 81:1097-1142. Zoeller RT, Tan SW, Tyl RW. General background on the hypothalamic-pituitary-thyroid (HPT) axis. Crit Rev Toxicol. 2007 Jan-Feb;37(1-2):11-53 Zoeller, R. T., R. Bansal, et al. (2005). "Bisphenol-A, an environmental contaminant that acts as a thyroid hormone receptor antagonist in vitro, increases serum thyroxine, and alters RC3/neurogranin expression in the developing rat brain." Endocrinology 146(2): 607-612. AOPWiki 2016-11-29T18:41:23

2022-10-10T08:52:30

Altered, Amphibian metamorphosis

Organ

Vertebrate metamorphosis is a biological transformation process that transitions an organism from one life stage to another; it is defined by growth of new tissues, programmed death of other tissues and physiological transformation of yet other tissues (Laudet, 2011; Brown and Cai, 2007). In the case of most amphibians, metamorphosis mediates the transition from aquatic to terrestrial life, while in bony and jawless fish, metamorphosis mediates transitions between life stages that offer various advantages for survival and reproduction. In vertebrates, metamorphosis is orchestrated by the hypothalamus-pituitary-thyroid (HPT) axis involving complex timing of gene expression/repression within various tissues, whereas in some cases across taxonomic classes, metamorphosis has been shown to be controlled very differently by the HPT axis. Thyroid hormone-mediated amphibian metamorphosis can be characterized by three phases during larval development: (1) pre-metamorphosis, (2) pro-metamorphosis and (3) metamorphic climax. All three of these phases coincide with activity states of the HPT axis. Pre-metamorphosis is characterized by a fully aquatic organism with low-level function of the thyroid gland and very low circulating levels of thyroid hormone. Pro-metamorphosis is characterized by the onset of full thyroid axis function and the initiation of rising levels of thyroid hormone in the plasma, with consequential changes in anatomy and physiology defining the transition from aquatic to terrestrial life. Metamorphic climax occurs when circulating thyroid hormone levels peak, which subsequently decrease to levels maintained homeostatically as adults. This climax period also represents the time at which all anatomical and physiological changes induced by thyroid hormone have either been initiated or are already completed. Detailed descriptions of these processes are reviewed by Brown and Cai (2007). Altered metamorphosis occurs when these thyroid hormone-mediated processes are perturbed, primarily during pro-metamorphosis and metamorphic climax. These perturbations can lead to either, delayed/arrested development, accelerated development or asynchronous development depending on the xenobiotic mode of action or MIE. Genetic defects or xenobiotic exposure that reduce thyroid hormone synthesis can delay metamorphosis, and in extreme cases, can completely arrest development. The most profound impacts on TH-mediated metamorphosis have be demonstrated through inhibition of key proteins in the TH synthesis pathway including the sodium-iodide symporter (Tietge et al., 2005, 2010; Hornung et al., 2010) and thyroperoxidase (Degitz et al., 2005; Tietge et al., 2010, 2013; Hornung et al., 2010, 2015). Alternatively, agonism of the thyroid axis through inhibition of negative feedback at the level of the hypothalamus-pituitary, or premature activation of thyroid receptor-mediated transcription can accelerate metamorphosis (Degitz et al., 2005), which can lead to asynchronous development due to errors in gene expression timing across the various metamorphic tissues. Asynchronous development can also occur due to inhibition of deiodinase (DIO) enzymes in peripheral tissues. DIO enzymes are responsible for activation and catabolism of TH; when dio gene expression profiles are altered, or the enzymes themselves undergo chemical inhibition, the imbalance of prohormone (T4), active hormone (T3) and inactive hormone (rT3, T2) can cause aberrant tissue development.

Rates of metamorphosis in model amphibian species, Xenopus laevis, are measured multiple ways, both of which rely on a developmental staging atlas developed by Nieuwkoop and Faber (NF)(1994). The method utilized within the 21 d Amphibian Metamorphosis Assay regulatory test guideline (OECD, 2009; US EPA 2009) relate the distribution of developmental stage of control larvae to the distributions of developmental stages of treated/exposed larvae. These data are typically analyzed for differences from control using non-parametric statistical approaches such as the Kruskal-Wallis test followed by Dunn's test for pairwise comparisons. The method utilized within the Larval Amphibian Growth and Development Assay regulatory test guideline (OECD, 2015; US EPA 2015) relate the number of days to reach metamorphic climax (NF stage 62) in control larvae to the number of days to NF stage 62 in treated/exposed larvae. These data are typically analyzed for differences from control using a Cox mixed-effects proportional hazard model. Asynchronous development is identified as disruption of the relative timing of morphogenic milestones and/or somatic development within a single larvae undergoing metamorphosis. The inability to identify an organism's developmental stage based on accepted criteria, such as outlined in Nieuwkoop and Faber (1994) for Xenopus sp. or Gosner (1960) for anurans, constitutes evidence of asynchronous development and would be counted as an incidence. Evaluations of severity are possible but the accuracy and resolution of the results would depend on the experience of the observer. One possible statistical approach for analyzing these data collected from a regulatory test guideline (OECD, 2009, 2015) would be a Rao-Scott-Cochran-Armitage by slices test (Green et al., 2014), as is often used for analysis of histopathology incidence and severity data.

Anurans Xenopus laevis

High Unspecific

High

Development

High

Brown, D.D. and Cai, L., 2007. Amphibian metamorphosis. Developmental biology, 306(1), pp.20-33. Degitz, S.J., Holcombe, G.W., Flynn, K.M., Kosian, P.A., Korte, J.J. and Tietge, J.E., 2005. Progress towards development of an amphibian-based thyroid screening assay using Xenopus laevis. Organismal and thyroidal responses to the model compounds 6-propylthiouracil, methimazole, and thyroxine. Toxicological sciences, 87(2), pp.353-364. Gosner, K.L., 1960. A simplified table for staging anuran embryos and larvae with notes on identification. Herpetologica, 16(3), pp.183-190. Green, J.W., Springer, T.A., Saulnier, A.N. and Swintek, J., 2014. Statistical analysis of histopathological endpoints. Environmental toxicology and chemistry, 33(5), pp.1108-1116. Hornung, M.W., Degitz, S.J., Korte, L.M., Olson, J.M., Kosian, P.A., Linnum, A.L. and Tietge, J.E., 2010. Inhibition of thyroid hormone release from cultured amphibian thyroid glands by methimazole, 6-propylthiouracil, and perchlorate. Toxicological Sciences, 118(1), pp.42-51. Laudet, V., 2011. The origins and evolution of vertebrate metamorphosis. Current Biology, 21(18), pp.R726-R737. Nieuwkoop, P.D. and Faber, J., 1994. Normal Table of Xenopus laevis (Daudin) Garland Publishing. New York, 252. OECD. (2009). Test No. 231: Amphibian Metamorphosis Assay, OECD Guidelines for the Testing of Chemicals, Section 2. OECD Publishing, Paris. OECD. (2015). Test No. 241: The Larval Amphibian Growth and Development Assay (LAGDA), OECD Guidelines for the Testing of Chemicals, Section 2. OECD Publishing, Paris. Tietge, J.E., Butterworth, B.C., Haselman, J.T., Holcombe, G.W., Hornung, M.W., Korte, J.J., Kosian, P.A., Wolfe, M. and Degitz, S.J., 2010. Early temporal effects of three thyroid hormone synthesis inhibitors in Xenopus laevis. Aquatic Toxicology, 98(1), pp.44-50. Tietge, J.E., Holcombe, G.W., Flynn, K.M., Kosian, P.A., Korte, J.J., Anderson, L.E., Wolf, D.C. and Degitz, S.J., 2005. Metamorphic inhibition of Xenopus laevis by sodium perchlorate: effects on development and thyroid histology. Environmental Toxicology and Chemistry, 24(4), pp.926-933. Tietge, J.E., Degitz, S.J., Haselman, J.T., Butterworth, B.C., Korte, J.J., Kosian, P.A., Lindberg-Livingston, A.J., Burgess, E.M., Blackshear, P.E. and Hornung, M.W., 2013. Inhibition of the thyroid hormone pathway in Xenopus laevis by 2-mercaptobenzothiazole. Aquatic toxicology, 126, pp.128-136. U.S. EPA. (2009). OCSPP 890.1100: Amphibian Metamorphosis Assay (AMA), Endocrine Disruptor Screening Program Test Guidelines, 890 Series. Available at: www.regulations.gov, ID: EPA-HQ-OPPT-2009-0576-0002. Accessed March 20, 2020. U.S. EPA. (2015). OCSPP 890.2300: Larval Amphibian Growth and Development Assay (LAGDA), Endocrine Disruptor Screening Program Test Guidelines, 890 Series. Available at: www.regulations.gov, ID: EPA-HQ-OPPT-2014-0766-0020. Accessed March 20, 2020. AOPWiki 2016-11-29T18:41:29

2020-09-02T11:19:05

<upstream-id>b6682d36-e038-4045-9f96-0ab900427d06</upstream-id> <downstream-id>f94341d3-5471-4b66-9aae-b91ff0c46de5</downstream-id> NIS is a membrane protein implicated in iodide uptake into the follicular cells of the thyroid. Other large anions can be also bound by NIS and inhibit accumulation of iodide into the thyroid by competing binding with iodide (Wolff, 1964).

NIS is a membrane bound glycoprotein and its main physiological function is to transport one iodide ion along with two sodium ions across the basolateral membrane of thyroid follicular cells. It uses the sodium gradient generated by the Na+/K+ ATPase for the active transport of iodide into the thyrocytes (Eskandari et al., 1997). Extensive studies on NIS protein have identified 14 different mutations and each one of them is related to Iodine Transport Deficiencies (ITD) (reviewed in Spitzweg and Morris, 2010). Most of these mutations have been characterized and it is well known that they even lead to the synthesis of truncated protein (Pohlenz et al., 1997; Pohlenz et al., 1998), partial deletions (Kosugi et al., 2002; Tonacchera et al., 2003; Montanelli et al., 2009) or substitutions of amino acids (Matsuda and Kosugi, 1997; Kosugi et al., 1999; Szinnai et al., 2006) that eventually result in total or partial NIS dysfunction. While most of the NIS mutants have been further investigated and the functional relationship between the NIS dysfunction and ITD is well established (reviewed in Darrouzet et al., 2014; Portulano et al., 2014), the exact structural relationship between mutated NIS and ITD still needs to be elucidated and the molecular modelling of the protein would greatly benefit these studies. At the same time, causative link between  iodide deficiency, thyroid hormones, and neurodevelopment deffects is well documented (Gilbert et al., 2009). Recent revision of the affinity constant for perchlorate binding to the NIS symporter based on in vitro and human in vivo data, performed by refitting published in vitro data, in which perchlorate-induced inhibition of iodide uptake via the NIS was measured, yielding a Michaelis-Menten kinetic constant (Km) of 1.5 μm, showed that a 60% lower value for the Km, equal to 0.59 μm. Substituting this value into the PBPK model for an average adult human significantly improved model agreement with the human RAIU data for exposures <100 μg kg-1 day-1 (Schlosser PM, 2016). The effects of maternal hypothyroidism could also contribute to this KER.  During pregnancy TH requirements increase, particularly during the first trimester (Alexander et al. 2004; Leung et al. 2010), due to higher concentrations of thyroxine-binding globulin, placental T4 inner-ring deiodination leading to the inactive reverse T3 (rT3), and transfer of small amounts of T4 to the foetus (during the first trimester foetal thyroid function is absent). Moreover, glomerular filtration rate and clearance of proteins and other molecules are both increased during pregnancy, possibly causing increased renal iodide clearance and a decreased of circulating plasma iodine (Glinoer, 1997). Thus, even though the foetal thyroid can trap iodide by about 12 week of gestation (Fisher and Klein, 1981), high concentrations of maternal perchlorate may potentially decrease thyroidal iodine available to the foetus by inhibiting placental NIS (Leung et al. 2010). Consequences of TH deficiency depend on the developmental timing of the deficiency (Zoeller and Rovet, 2004). For instance, if the TH deficiency occurs during early pregnancy, offspring show visual attention, visual processing and gross motor skills deficits, while if it occurs later, offspring may show subnormal visual and visuospatial skills, along with slower response speeds and motor deficits. If TH insufficiency occurs after birth, language and memory skills are most predominantly affected (Zoeller and Rovet, 2004). There are limited data regarding low-level environmental perchlorate exposure and maternal thyroid function during pregnancy. A Chilean study found no increases in TSH or decreases in free thyroxine or urinary iodine concentrations in pregnant women living in three areas (all of which had more than adequate mean urinary iodine levels) with long-term environmental perchlorate exposure (Téllez Téllez et al. 2005). A follow-up analysis of this cohort also confirmed the lack of association between individual urinary iodide or perchlorate concentrations and thyroid function in the pregnant women (Gibbs and Van Landingham, 2008). Studies of large cohorts of first-trimester pregnant women from the U.S., Europe and Argentina found that environmental perchlorate exposure did not affect maternal thyroid function (Pearce et al. 2009).

Many studies have shown inhibition of radioactive iodide uptake by using different cell models and assays. However, there have been identified only few specific NIS inhibitors up to date, while all the others are thought to act through different inhibitory mechanisms. Monovalent anions, others than iodide, are also transported by NIS but Nitrate (NO3-), thiocyanate (SCN-), perchlorate (ClO-4), dysidenin and aryltrifluoroborates are of particular dietary and environmental importance (Jones et al., 1996; Tonacchera et al., 2004; De Groef et al., 2006). Recent revision of the affinity constant for perchlorate binding to the NIS symporter based on in vitro and human in vivo data, performed by refitting published in vitro data, in which perchlorate-induced inhibition of iodide uptake via the NIS was measured, yielding a Michaelis-Menten kinetic constant (Km) of 1.5 μm, showed that a 60% lower value for the Km, equal to 0.59 μm. Substituting this value into the PBPK model for an average adult human significantly improved model agreement with the human RAIU data for exposures <100 μg kg-1 day-1 (Schlosser PM, 2016). There are many studies showing the effect of inhibition of NIS on thyroidal iodide uptake: - Cianchetta et al., 2010 For this study the rat FRTL5 thyroid cell line endogenously expressing NIS, and the monkey kidney fibroblast-like cells (COS-7) transfected with hNIS were used. NIS functionality was assessed with the use of the Yellow Fluorescent Protein (YFP) variant YFP-H148Q/I152L, a genetically encodable biosensor of intracellular perchlorate concentration monitored by real-time fluorescence microscopy. Decrease of YFP-H148Q/I152L fluorescence in FRTL-5 cells occurs as a result of NIS-mediated uptake and binding to the intracellular fluorochrome (Rhoden et al., 2007). The biosensor was used to compare the kinetics of iodide and perchlorate transport by NIS, and to assess the ability of perchlorate to inhibit iodide transport. Additionally, perchlorate was shown to inhibit NIS function (competitive inhibition) by preventing iodide-induced changes in fluorescence of FRTL5 cells. Perchlorate caused a concentration-dependent inhibition of iodide uptake in the initial influx rate (IC50=1.6μM) and in the intracellular concentration of iodide (IC50=1.1μM). Also, both perchlorate and iodide (1–1000 μM) induced concentration-dependent decreases in YFP-H148Q/I152L fluorescence in COS-7 cells expressing hNIS, but had no effect (< 2%) in COS-7 cells lacking hNIS. Additionally, perchlorate induced a significantly smaller decrease in fluorescence (10.6% at 1 mM) than iodide (31.8% at 1 mM iodide). Thus, it was confirmed that the reduction of fluorescence was due to NIS-mediated anion transport into the cells, excluding non-specific effects. - Tonacchera et al., 2004 Chinese hamster ovary (CHO) cell line had been stably transfected with human NIS and the measurement of iodide uptake was performed with the use of radioactive iodide uptake (RAIU) method. It was shown that the inhibition of iodide uptake was dose-dependent when using the known NIS inhibitors (ClO-4, NO3-, SCN-). Additionally, unlabeled I- (non 125I) was used to investigate the inhibition level of radioiodide uptake and to compare it with the potency of the other monoions, which are known NIS inhibitors. The IC50 values for the studied monoions were the following: ClO4-: IC50 was 1.22 μΜ; SCN-: IC50 was 18.7 μΜ; NO3-: IC50 was 293 μΜ; I-: IC50 was 36.6 μΜ. Finally, the present study investigated the joint effects of simultaneous exposure to multiple RAIU inhibitors, by generating multiple dose-response curves in the presence of fixed concentrations of inhibitors. The results of those experiments indicated a competition between the four anions with similar size for access to the binding sites of the NIS. The prediction model developed in this study, actually suggests that thyroidal iodide uptake is approximately proportional to iodide nutrition for any fixed inhibitor concentration, answering to the question whether dietary iodide can modulate the inhibitory effects of the known environmental goitrogens. - Waltz et al., 2010 Measurement of iodide uptake was performed with a non-radioactive method. By using the rat thyroid low –serum 5 (FRTL5) cells, which endogenously express NIS, a spectrophotometric assay was developed and the iodide accumulation was determined based on the catalytic reduction of yellow cerium to colorless cerium in the presence of arsenious acid (Sandell-Kolthoff reaction). A dose-dependent inhibition of iodide uptake was shown. The IC50 values for the studied compounds were the following: Sodium perchlorate (NaClO4): IC50 was 0.1 μΜ Sodium thiocyanate (NaSCN): IC50 was 12 μΜ Sodium nitrate (NaNO3): IC50 was 800 μΜ Sodium Tetrafluoroborate (NaBF4): IC50 was 1.2 μΜ - Lecat-Guillet et al., 2007; 2008a A fully automated radioiodide uptake assay was developed and some known NIS inhibitors were tested. A dose-dependent inhibition of iodide uptake was shown. The IC50 values for the studied compounds were the following: Sodium perchlorate (NaClO4): IC50 was 1 μΜ; Sodium thiocyanate (NaSCN): IC50 was 14 μΜ; Sodium nitrate (NaNO3): IC50 was 250 μΜ; Sodium Tetrafluoroborate (NaBF4): IC50 was 0.75 μΜ.  Additionally, a library of 17020 compounds was screened for the identification of new human NIS inhibitors. The identification was based on the magnitude of changes in iodide uptake using Human Embryonic Kidney 293 (HEK293) cells, stably transfected with the hNIS. The same experiments and with similar results were also performed in rat thyroid derived cells (FRTL5), which endogenously express NIS. Compounds that inhibited iodide uptake in a time-dependent manner were considered to act through direct NIS inhibition. In contrast, those compounds that had a delayed effect on iodide uptake were thought to act through a sodium gradient disruption system resulting in indirect inhibition of iodide transport. Perchlorate was used as a positive control in these experiments and, as expected, it blocked iodide uptake immediately and totally throughout the experiment. Dysidenin was also used as a control and the IC50 value identified was 2 μΜ. All the compounds that were used for these experiments were small drug-like molecules that have not been detected in the environment and they were named as ITBs (Iodide Transport Blockers). - Lecat-Guillet et al., 2008b With the same fully automated radioiodide uptake assay, as described above, new NIS inhibitors were also identified. The organotrifluoroborate (BF3−) was found to inhibit iodide uptake with an IC50 value of 0.4 μM using rat-derived thyroid cells (FRTL5). The biological activity is rationalized by the presence of the ion BF3− as a minimal binding motif for substrate recognition at the iodide binding site. - Lindenthal et al., 2009 With the use of a patch-clamp technique an analysis of the NIS inhibitors identified by Lecat-Guillet et al., 2008 (named ITB-1 to ITB-10 for "Iodide Transport Blockers") was evaluated in Xenopus oocytes expressing NIS to further assess the inhibitory effect of those molecules specifically on NIS activity. Four of those molecules (ITB-3, ITB-9, ITB-5 and ITB-4) were identified as the most potent, non-competitive NIS inhibitors. The effects of dysidenin were also analyzed with the same technique, as it had been reported to be a specific inhibitor of NIS (Vroye et al., 1998). It was found that dysidenin (50 μM) induced a rapid and reversible inhibition of the iodide (about 40%) of induced current in mNIS-expressing oocytes, but did not evoke any currents in the absence of iodide, suggesting that this effect was due to the inhibition of NIS activity. - Greer et al., 2002 In human studies, potassium perchlorate was used to predict inhibition of thyroidal iodide uptake by applying the RAIU method. Greer et al., tested body weight adjusted doses of potassium perchlorate and an assessment of RAIU uptake was performed on day 2 and day 14 of treatment and 24 h following treatment termination (on day 15). The NOEL value for inhibition of thyroidal uptake was 0.007 mg/kg-day, while the true NEL value was estimated to be 0.0052 and 0.0064 mg/kg-day. According to the dose-response inhibition of iodide uptake the maximum percentage of iodide inhibition at the doses of 0.0052 and 0.0064 mg/kg-day is 8.3-9.5%, which is physiologically insignificant for a person with dietary sufficient iodine intake. - Wen et al., 2016 By using human MCF-7 cells, a breast adenocarcinoma cell line, which express inducible NIS in the presence of all-trans retinoic acid (ATRA) it has been shown that inhibition of sterol regulatory element-binding proteins (SREBP) maturation by treatment with 25-hydroxycholesterol (5 µM) for 48 hr reduced ATRA (1 µM)-induced mRNA concentration of NIS and decreased iodide uptake by approximately 20%. This study showed for the first time that the NIS gene and iodide uptake are regulated by SREBP in cultured human mammary epithelial cells. - Arriagada et al. 2015 This study showed that 2 hr or 5 hr exposure to excess I- (100 μM) respectively in FRTL-5 cells and in ex-vivo rat thyroid gland (removed after single in vivo i.p. injection of 100 μg of I− in 500 μL of distilled water, and analysis of 125I thyroid uptake), induced inhibition of I- uptake through the NIS (~ 30% uptake inhibition after 5 hr in vivo), a process known as the Wolff-Chaikoff effect, which was not associated with a decrease of NIS expression or a change in NIS localization. Incubation of FRTL-5 cells with excess I- for 2 hr increased hydrogen peroxide generation. Also incubation with hydrogen peroxide (100 μM) decreased NIS-mediated I- transport, effect that was reverted by ROS scavengers.

The thyroid system is quite complex and therefore some inconsistent results have been produced by recent studies. For example, it has been observed in healthy volunteers that a 6-month exposure to perchlorate at doses up to 3 mg/d (low doses) had no effect on thyroid function, including inhibition of thyroid iodide uptake as well as serum levels of thyroid hormones, TSH, and Tg (Braverman et al., 2006). However, this study was limited by the small sample size and is obviously underpowered. The review by Charnley (2008) examines a number of studies where association between perchlorate environmental (low) exposure and thyroid effects were analysed and many inconsistent conclusions have been drawn. For instance, no correlations were found between TH serum levels and urinary iodine concentrations among women exposed to perchlorate participating in the 2000-2001 National Health and Nutrition Examination Survey (NHANES). Available evidence does not support a causal relationship between changes in TH levels and current environmental levels of perchlorate exposure, but does support the conclusion that the US Environmental Protection Agency's reference dose (RfD) for perchlorate is conservatively health-protective. However, potential perchlorate risks are unlikely to be distinguishable from the ubiquitous background of naturally occurring substances present at much higher exposures that can affect the thyroid via the same biological mode of action as perchlorate, such as nitrate and thiocyanate. Therefore, risk management approaches that account for both aggregate and cumulative exposures and that consider the larger public health context in which exposures are occurring are desirable. Additionally, a more comprehensive study by Pearce et al. (2010) conducted during 2002-2006 on 22,000 women at less than 16-week gestation showed that while low-level perchlorate exposure was ubiquitous in these women (with a median urinary perchlorate concentration of 5 µg/liter in the Turin cohort and 2 µg/liter in the Cardiff cohort), no associations between urine perchlorate concentrations and serum TSH or free T4 in the individual euthyroid or hypothyroid/hypothyroxinemic cohorts were found. The data assessing the effect of maternal perchlorate exposure in neonates and children and thyroid function remain unclear (Leung et al., 2010). Decreased iodine intake can decrease TH production, and therefore exposure to perchlorate might be particularly detrimental in iodine-deficient individuals (Leung et al. 2010). Moreover, biologically based dose-response modeling of the relationships among iodide status (e.g., dietary iodine levels), perchlorate dose, and TH production in pregnant women has shown that iodide intake has a profound effect on the likelihood that exposure to goitrogens will produce hypothyroxinemia (Lewandowski et al. 2015). Consequences of TH deficiency depend on the developmental timing of the deficiency (Zoeller and Rovet, 2004). For instance, if the TH deficiency occurs during early pregnancy, offspring show problems in visual attention, visual processing and gross motor skills, while if it occurs later, offspring may show subnormal visual and visuospatial skills, slower response speeds and motor deficits. If TH insufficiency occurs after birth, language and memory skills are most predominantly affected (Zoeller and Rovet, 2004). Altogether these studies indicate that factors, such as age, gender, developmental stage, and iodide status can affect the impact of perchlorate and other NIS inhibitors. All these variables should be taken into account to explain possible inconsistencies in study findings.

For this relationship there is not enough data linking quantitatively the inhibition of NIS with the amount of thyroidal uptake. The NIS inhibition is possible to be directly measured by using the fact that the simultaneous transport of 2 Na+ and 1 I- generates a current, which could be easily measured with electrophysiological methods (Eskandari et al., 1997) or with patch clamp techniques (Van Sande et al., 2003). However, the exact stoichiometry of the molecules that are transferred is not yet known, meaning that in some cases it cannot be detected. For example, perchlorate does not cause depolarization of the cellular membrane, as it is thought to be transferred in 1 to 1 stoichiometry with the Na+ (Van Sande et al., 2003). However, I- uptake can also be measured in vivo, as shown in rats i.p. injected with 100 μg of I− in 500 μL of distilled water (known to cause an inhibition of NIS- mediated I- transport), followed by analysis of radioactive 125I thyroid uptake (Arriagada et al. 2015). Further studies are needed to support quantitative evaluation of this KER.

High Mixed

High

During brain development

High High High Moderate

Empirical evidence comes from in vitro works using rat follicular cells (Cianchetta et al., 2010; Waltz et al., 2010; Lecat-Guillet et al., 2007; 2008; Lecat-Guillet et al., 2008b), human in vitro cell models (Wen et al., 2016) and in vivo data (Arriagada et al. 2015), as well as Xenopus oocytes (Lindenthal et al., 2009) and Zebrafish (Thienpont et al., 2011).

Alexander EK, Marqusee E, Lawrence J, Jarolim P, Fischer GA, Larsen PR (2004). Timing and magnitude of increases in levothyroxine requirements during pregnancy in women with hypothyroidism. N Engl J Med. 2004 Jul 15;351(3):241-9. Arriagada AA, Albornoz E, Opazo MC, Becerra A, Vidal G, Fardella C, Michea L, Carrasco N, Simon F, Elorza AA, Bueno SM, Kalergis AM, Riedel CA. (2015). Excess iodide induces an acute inhibition of the sodium/iodide symporter in thyroid male rat cells by increasing reactive oxygen species. Endocrinology. Apr;156(4):1540-51. Braverman LE, Pearce EN, He X, Pino S, Seeley M, Beck B, Magnani B, Blount BC, Firek A. (2006). Effects of six months of daily low-dose perchlorate exposure on thyroid function in healthy volunteers. J Clin Endocrinol Metab. 91:2721-2724. Charnley G. (2008) Perchlorate: overview of risks and regulation. Food Chem Toxicol. 46(7):2307-15 (Review). Cianchetta S, di Bernardo J, Romeo G, Rhoden KJ (2010). Perchlorate transport and inhibition of the sodium iodide symporter measured with the yellow fluorescent protein variant YFP-H148Q/I152L. Toxicol Appl Pharmacol. 243:372-380. Darrouzet E, Lindenthal S, Marcellin D, Pellequer JL, Pourcher T. (2014). The sodium/iodide symporter: state of the art of its molecular characterization. Biocim Biophys Acta. 1838:244-253. De Groef B, Decallonne BR, Van der Geyten S, Darras VM, Bouillon R. (2006). Perchlorate versus other environmental sodium/iodide symporter inhibitors: potential thyroid-related health effects. Europ J Endocr. 155:17-25. Eskandari S, Loo DD, Dai G, Levy O, Wright M, Carrasco N. (1997). Thyroid Na+/I- symporter: mechanism, stoichiometry, and specificity. J Biol Chem 272: 27230-27238. Fisher DA, Klein AH (1981). Thyroid development and disorders of thyroid function in the newborn. N Engl J Med. 1981 Mar 19;304(12):702-12. Gibbs JP, Van Landingham C (2008). Urinary perchlorate excretion does not predict thyroid function among pregnant women. Thyroid.  Jul; 18(7):807-8. Gilbert ME, Hedge J, Grant K, Lyke D, Gitata I, Anderson W, et al. (2009) Marginal iodide deficiency, thyroid hormones, and neurodevelopment: developing a model. Toxicol Sci., 108 (S-1):32. Glinoer D (1997). The regulation of thyroid function in pregnancy: pathways of endocrine adaptation from physiology to pathology. Endocr Rev. Jun;18(3):404-33. Greer MA, Goodman G, Pleus RC, Greer SE. (2002). Health effects assessment for environmental perchlorate contamination: the dose response for inhibition of thyroidal radioiodine uptake in humans. Environm Health Persp. 110: 927-937. Jones PA, Pendlington RU, Earl LK, Sharma RK, Barrat MD. (1996). In vitro investigations of the direct effects of complex anions on thyroidal iodide uptake: identification of novel inhibitors. Toxicol. In Vitro. 10: 149-160. Kosugi S, Bhayana S, Dean HJ. (1999). A novel mutation in the sodium/iodide symporter gene in the largest family with iodide transport defect. J Clin Endocrinol Metab. 84: 3248-3253. Kosugi S, Okamoto H, Tamada A, Sanchez-Franco F. (2002). A novel peculiar mutation in the sodium/iodide symporter gene in Spanish siblings with iodide transport defect. J Clin Endocrinol Metab. 87: 3830–3836. Lecat-Guillet N, Merer G, Lopez R, Pourcher T, Rousseau B, Ambroise Y. (2008a). Small-molecule inhibitors of sodium iodide symporter function. Chembiochem 9:889–895. Lecat-Guillet N, Ambroise Y. (2008b). Discovery of aryltrifluoroborates as potent sodium/iodide symporter (NIS) inhibitors. Chem Med Chem 3:1207–1209. Lecat-Guillet N, Merer G, Lopez R, Pourcher T, Rousseau B, Ambroise Y. (2007). A 96-well automated radioiodide uptake assay for sodium/iodide symporter inhibitors. Assay Drug Dev Technol 5:535-540. Leung AM, Pearce EN, Braverman LE (2010). Perchlorate, iodine and the thyroid. Best Pract Res Clin Endocrinol Metab. Feb;24(1):133-41. Lewandowski TA, Peterson MK2, Charnley G (2015). Iodine supplementation and drinking-water perchlorate mitigation. Food Chem Toxicol. Jun;80:261-70. Lindenthal S, Lecat-Guillet N, Ondo-Mendez A, Ambroise Y, Rousseau B, Pourcher T. (2009). Characterization of small-molecule inhibitors of the sodium iodide symporter. J Endocrinol 200:357–365. Matsuda A, Kosugi S. (1997). A homozygous missense mutation of the sodium/iodide symporter gene causing iodide transport defect. J Clin Endocrinol Metab. 82: 3966-3971. Montanelli L, Agretti P, Marco G, Bagattini B, Ceccarelli C, Brozzi F, Lettiero T, Cerbone M, Vitti P, Salerno M, Pinchera A, Tonacchera M. (2009). Congenital hypothyroidism and late-onset goiter: identification and characterization of a novel mutation in the sodium/iodide symporter of the proband and family members. Thyroid 19: 1419-1425. Pearce EN, Lazarus JH, Smythe PP, et al. (2009). Thyroid Function is Not Affected by Environmental Perchlorate Exposure in First Trimester Pregnant Women. Endocrine Society 91st Annual Meeting; USA. Pearce EN, Lazarus JH, Smyth PP, et al. (2010). Perchlorate and thiocyanate exposure and thyroid function in first-trimester pregnant women. J Clin Endocrinol Metab. 95:3207–3215. Pohlenz J, Rosenthal IM, Weiss RE, Jhiang SM, Burant C, Refetoff S. (1998). Congenital hypothyroidism due to mutations in the sodium/iodide symporter. Identification of a nonsense mutation producing a downstream cryptic 3′ splice site. J Clin Invest. 101:1028-1035. Pohlenz J, Medeiros-Neto G, Gross JL, Silveiro SP, Knobel M, Refetoff S. (1997). Hypothyroidism in a Brazilian kindred due to iodide trapping defect caused by a homozygous mutation in the sodium/iodide symporter gene. Biochem Biophys Res Commun. 240: 488-491. Portulano C, Paroder-Belenitsky M, Carrasco N. (2014). The Na+/I- symporter (NIS): Mechanism and medical impact. Endocr Rev. 35: 106-149. Rhoden KJ, Cianchetta S, Stivani V, Portulano C, Galietta LJV, Romeo G. (2007). Cell-based imaging of sodium iodide symporter activity with the yellow fluorescent protein variant YFP-H148Q/I152L. Am. J. Physiol., 292, pp. C814–C823. Schlosser PM. (2016). Revision of the affinity constant for perchlorate binding to the sodium-iodide symporter based on in vitro and human in vivo data. J Appl Toxicol. Dec;36(12):1531-1535. Spitzweg C, Morris JC. (2010). Genetics and phenomics of hypothyroidism and goiter due to NIS mutations. Mol Cell Endocrinol. 322: 56-63. Szinnai G, Kosugi S, Derrien C, Lucidarme N, David V, Czernichow P, Polak M. (2006). Extending the clinical heterogeneity of iodide transport defect (ITD): a novel mutation R124H of the sodium/iodide symporter gene and review of genotype-phenotype correlations in ITD. J Clin Endocrinol Metab. 91: 1199–1204. Téllez Téllez R, Michaud Chacón P, Reyes Abarca C, Blount BC, Van Landingham CB, Crump KS, Gibbs JP (2005). Long-term environmental exposure to perchlorate through drinking water and thyroid function during pregnancy and the neonatal period. Thyroid. Sep; 15(9):963-75. Thienpont B, Tingaud-Sequeira A, Prats E, Barat, C., Babin P.J, Raldua D, (2011). Zebrafish eleutheroembryos provide a suitable vertebrate model for screening chemicals that impair thyroid hormone synthesis. Environ Sci Technol 45, 7525-7532. Tonacchera M, Agretti P, de Marco G, Elisei R, Perri A, Ambrogini E, De Servi M, Ceccarelli C, Viacava P, Refetoff S, Panunzi C, Bitti ML, Vitti P, Chiovato L, Pinchera A. (2003). Congenital hypothyroidism due to a new deletion in the sodium/iodide symporter protein. Clin Endocrinol. 59: 500–506. Tonacchera M, Pinchera A, Dimida A, Ferrarini E, Agretti P, Vitti P, Santini F, Crump K, Gibbs J. (2004). Relative potencies and additivity of perchlorate, thiocyanate, nitrate, and iodide on the inhibition of radioactive iodide uptake by the human sodium iodide symporter. Thyroid. 14: 1012-1019. Van Sande J, Massart C, Beauwens R, Schoutens A, Costagliola S, Dumont JE, Wolff J. (2003). Anion selectivity by the sodium iodide symporter. Endocrinology. 144: 247-252. Vroye L, Beauwens R, Van Sande J, Daloze D, Braekman JC, Golstein PE. (1998). The Na+/I- co-transporter of th e thyroid: characterization of new inhibitors. Pflugers Archiv. 435:259-266. Waltz F, Pillette L, Ambroise Y. (2010). A nonradioactive iodide uptake assay for sodium iodide symporter function. Anal Biochem. 396:91-95. Wen G, Pachner LI, Gessner DK, Eder K, Ringseis R. (2016). Sterol regulatory element-binding proteins are regulators of the sodium/iodide symporter in mammary epithelial cells. J Dairy Sci. Nov;99(11):9211-9226. Wolff J. (1964). Transport of iodide and other anions in the thyroid gland. Physiol Rev 44: 45-90. Zoeller RT, Rovet J (2004). Timing of thyroid hormone action in the developing brain: clinical observations and experimental findings. J Neuroendocrinol. Oct;16(10):809-18. AOPWiki 2016-11-29T18:41:34

2018-05-29T07:24:34

<upstream-id>b6682d36-e038-4045-9f96-0ab900427d06</upstream-id> <downstream-id>fa76c581-3528-4c8a-b62c-86b29dac93fb</downstream-id>

Not Specified High Moderate Moderate

Taxonomic: According to the evaluation of the empirical taxonomic domain of applicability (tDOA) of an adverse outcome pathway network for thyroid hormone system disruption (THSD) by Haigis et al., 2023, the level of confidence for a linkage between NIS inhibition and reduced thyroid hormone (TH) levels was considered high for mammals (Buckalew et al., 2020, Concilio et al., 2020, Dayem et al., 2008, Hallinger et al., 2017, Heltemes et al., 2003, Schmutzler et al., 2007, Selmi-Ruby et al., 2003, Wang et al., 2018) and moderate for fish and amphibians (Concilio et al., 2020, Hornung et al., 2010, Lindenthal et al., 2009, McMullen et al., 2017, Opitz et al., 2006; Opitz and Kloas, 2010, Thienpont et al., 2011). This was supported by structural protein conservation analysis by Lalone et al., 2018 and Haigis et al., 2023. Structural protein conservation of mammalian, fish, amphibian, reptilian and avian NIS was found compared to the human (Homo sapiens) protein target using the U.S. Environmental Protection Agency’s Sequence Alignment to Predict Across Species Susceptibility (SeqAPASS v6.0; seqapass.epa.gov/seqapass/) tool, while acknowledging the potential existence of interspecies differences in conservation. No empirical evidence linking NIS inhibition to THSD was found for reptiles and birds.

Buckalew, A. R., Wang, J., Murr, A. S., Deisenroth, C., Stewart, W. M., Stoker, T. E., and Laws, S. C. (2020). Evaluation of potential sodium-iodide symporter (NIS) inhibitors using a secondary Fischer rat thyroid follicular cell (FRTL-5) radioactive iodide uptake (RAIU) assay. Arch. Toxicol. 94, 873–885. Concilio, S. C., Zhekova, H. R., Noskov, S. Y., and Russell, S. J. (2020). Inter-species variation in monovalent anion substrate selectivity and inhibitor sensitivity in the sodium iodide symporter (NIS). PLoS One 15, e0229085. Dayem, M., Basquin, C., Navarro, V., Carrier, P., Marsault, R., Chang, P., Huc, S., Darrouzet, E., Lindenthal, S., and Pourcher, T. (2008). Comparison of expressed human and mouse sodium/iodide symporters reveals differences in transport properties and subcellular localization. J. Endocrinol. 197, 95–109. Haigis A-C., Vergauwen L., LaLone C.A., Villeneuve D.L., O'Brien J.M., Knapen D. (2023). Cross-species applicability of an adverse outcome pathway network for thyroid hormone system disruption. Toxicol Sci. 195, 1-27. Hallinger, D. R., Murr, A. S., Buckalew, A. R., Simmons, S. O., Stoker, T. E., and Laws, S. C. (2017). Development of a screening approach to detect thyroid disrupting chemicals that inhibit the human sodium iodide symporter (NIS). Toxicol. In Vitro 40, 66–78. Heltemes, L. M., Hagan, C. R., Mitrofanova, E. E., Panchal, R. G., Guo, J., and Link, C. J. (2003). The rat sodium iodide symporter gene permits more effective radioisotope concentration than the human sodium iodide symporter gene in human and rodent cancer cells. Cancer Gene Ther. 10, 14–22. Hornung, M. W., Degitz, S. J., Korte, L. M., Olson, J. M., Kosian, P. A., Linnum, A. L., and Tietge, J. E. (2010). Inhibition of thyroid hormone release from cultured amphibian thyroid glands by methimazole, 6-propylthiouracil, and perchlorate. Toxicol. Sci. 118, 42–51. Lalone, C. A., Villeneuve, D. L., Doering, J. A., Blackwell, B. R., Transue, T. R., Simmons, C. W., Swintek, J., Degitz, S. J., Williams, A. J., and Ankley, G. T. (2018). Evidence for cross species extrapolation of mammalian-based high-throughput screening assay results. Environ. Sci. Technol. 52, 13960–13971. Lindenthal, S., Lecat-Guillet, N., Ondo-Mendez, A., Ambroise, Y., Rousseau, B., and Pourcher, T. (2009). Characterization of small-molecule inhibitors of the sodium iodide symporter. J. Endocrinol. 200, 357–365. McMullen, J., Ghassabian, A., Kohn, B., and Trasande, L. (2017). Identifying subpopulations vulnerable to the thyroid-blocking effects of perchlorate and thiocyanate. J. Clin. Endocrinol. Metab. 102, 2637–2645. Opitz, R., Trubiroha, A., Lorenz, C., Lutz, I., Hartmann, S., Blank, T., Braunbeck, T., and Kloas, W. (2006). Expression of sodium-iodide symporter mRNA in the thyroid gland of Xenopus laevis tadpoles: Developmental expression, effects of antithyroidal compounds, and regulation by TSH. J. Endocrinol. 190, 157–170. Opitz, R., and Kloas, W. (2010). Developmental regulation of gene expression in the thyroid gland of Xenopus laevis tadpoles. Gen. Comp. Endocrinol. 168, 199–208. Schmutzler, C., Gotthardt, I., Hofmann, P. J., Radovic, B., Kovacs, G., Stemmler, L., Nobis, I., Bacinski, A., Mentrup, B., Ambrugger, P., et al. (2007). Endocrine disruptors and the thyroid gland-a combined in vitro and in vivo analysis of potential new biomarkers. Environ. Health Perspect. 115, 77–83. Selmi-Ruby, S., Watrin, C., Trouttet-Masson, S., Bernier-Valentin, F., Flachon, V., Munari-Silem, Y., and Rousset, B. (2003). The porcine sodium/iodide symporter gene exhibits an uncommon expression pattern related to the use of alternative splice sites not present in the human or murine species. Endocrinology 144, 1074–1085. Thienpont, B., Tingaud-Sequeira, A., Prats, E., Barata, C., Babin, P. J., and Raldua, D. (2011). Zebrafish eleutheroembryos provide a suitable vertebrate model for screening chemicals that impair thyroid hormone synthesis. Environ. Sci. Technol. 45, 7525–7532. Wang, J., Hallinger, D. R., Murr, A. S., Buckalew, A. R., Simmons, S. O., Laws, S. C., and Stoker, T. E. (2018). High-throughput screening and quantitative chemical ranking for sodium-iodide symporter inhibitors in ToxCast phase I chemical library. Environ. Sci. Technol. 52, 5417–5426. AOPWiki 2020-12-09T12:16:18

2025-11-19T05:42:21

<upstream-id>f94341d3-5471-4b66-9aae-b91ff0c46de5</upstream-id> <downstream-id>9bedfcac-14d1-49a0-b56e-8eb86d7db03f</downstream-id> Thyroid hormones (THs), thyroxine (T4) and triiodothyronine (T3) are synthesized in the thyroid gland in the presence of functional NIS and thyroid peroxidase (TPO) as iodinated thyroglobulin (Tg), and stored in the colloid of thyroid follicles. NIS is a membrane bound glycoprotein whose main physiological function is to transport one iodide ion along with two sodium ions across the basolateral membrane of thyroid follicular cells. Extensive studies on NIS protein have identified 14 different mutations and each one of them is related to Iodine Transport Deficiencies (ITD) (Spitzweg and Morris, 2010). Once inside the follicular cells, the iodide diffuses to the apical membrane, where it is metabolically oxidized through the action of TPO to iodinium (I+), which in turn iodinates tyrosine residues of the Tg proteins in the follicle colloid. Therefore, NIS is essential for the synthesis of thyroid hormones (T3 and T4). TPO is a heme-containing apical membrane protein within the follicular lumen of thyrocytes that acts as the enzymatic catalyst for TH synthesis (Taurog, 2005). Propylthiouracil (PTU) and methimazole (MMI), are thioureylene drugs that are known to inhibit the ability of TPO to: a) activate iodine and transfer it to thyroglobulin (Tg) (Davidson et al., 1978) and, b) couple thyroglobulin (Tg)-bound iodotyrosyls to produce Tg-bound T3 and T4 (Taurog, 2005). PTU and MMI have been found to decrease also the expression of NIS mRNA and consequently iodide accumulation, as shown in FRTL-5 cells (Spitzweg et al. 1999). Other compounds, such as triclosan, triclocarban, 2,2',4,4'-tetrabromodiphenyl ether (BDE-47), and bisphenol A (BPA) have been reported to decrease thyroid hormone (TH) levels by inducing an inhibition of NIS-mediated iodide uptake and altering the expression of genes involved in TH synthesis in rat thyroid follicular FRTL-5 cells, and on the activity of thyroid peroxidase (TPO), using rat thyroid microsomes (Wu Y et al. 2016). Perchlorate, thiocyanate, nitrate, and iodide, which are competitive inhibitors of iodide uptake, have been shown to inhibit radioactive iodide uptake by NIS (Tonacchera et al. 2004), consequentially resulting in inhibition of TH synthesis. In particular, perchlorate blocks iodide uptake into the thyroid through NIS inhibition and decreases the production of TH (Steinmaus, 2016a). More recent evidence also suggests that young children, pregnant women, foetuses, and people co-exposed to similarly acting agents may be especially susceptible to perchlorate-induced toxicity (Steinmaus et al., 2016b). Concern about environmental perchlorate exposure is focused on its inhibition of iodide uptake into the thyroid (MIE). Decreased iodine intake may decrease thyroid hormone production. Perchlorate exposure, therefore, might be particularly detrimental in iodine-deficient individuals. Median urinary iodine levels are used instead and reflect dietary iodine sufficiency across populations (International Council for the Control of Iodine Deficiency Disorders (ICCIDD); available from: <a href="http://www.iccidd.org">www.iccidd.org</a>). According to ICCIDD report Iodine deficiency continues to be an important global public health issue, with an estimated 2.2 million people (38% of the world's population) living in iodine-deficient areas. In 1990, the United Nations World Summit for Children set forth the goal of eliminating iodine deficiency worldwide (UNICEF World Summit for Children. Available from: <a href="http://www.unicef.org/wsc/declare.htm">http://www.unicef.org/wsc/declare.htm</a>; 1990).  Considerable progress has been achieved by programmes of universal salt iodisation (USI) in various countries, in line with the recommendations of the World Health Organization (WHO) (WHO, UNICEF, ICCIDD. A guide for programme managers. World Health Organization; Geneva: 2007. Assessment of the iodine deficiency disorders and monitoring their elimination.WHO/NHD/01.1). However, many countries remain iodine deficient (de Benoist et al., 2013; Lazarus and Delange, 2004). In the U.S., data from large population studies have shown that median urinary iodine levels decreased by approximately 50% between the early 1970s and the early 1990s, although the population overall remained iodine sufficient (Hollowell et al., 1998). Subsequent studies have shown that this decrease has stabilised (Caldwell et al., 2005). The WHO still considers iodine deficiency, which leads to hypothyroidism, the single most important preventable cause of brain damage worldwide (WHO/UNICEF/ICCIDD, 2007). The most vulnerable groups are pregnant and lactating women and their developing fetuses and neonates, given the crucial importance of iodine to ensure adequate levels of thyroid hormones for brain maturation. Iodine deficiency in pregnancy is a prevailing problem not only in developing countries, but also in western industrialized nations and other countries classified as free of iodine deficiency, and solution may be found in dietary changes (Moog et al., 2017).

The association between these two KEs is strong, and supported by in vitro, in vivo and epidemiological studies. Blocking iodide uptake into the thyroid follicular cells as a consequence of NIS inhibition or functional impairment, leads to reduced TH synthesis. Compounds that have been shown to inhibit NIS function (e.g., perchlorate, thiocyanate, nitrate, and iodide), has also been proven to decrease TH synthesis by inducing a downregulation of TPO gene expression and/or increase of TSH level, which are both indicative of a reduce TH biosynthesis. TSH receptor controls transcription and posttranslational modification of NIS (Dai et al., 1996). Stimulation of TSH receptor increases T3 and T4 production and secretion (Szkudlinski et al., 2002). NIS gene expression is suppressed by growth factors such as IGF-1 and TGF-β (the latter is induced by the BRAF-V600E oncogene), which prevent NIS to localize in the basolateral membrane (Riesco-Eizaguirre et al., 2009). The BRAF-V600E oncogene is also associated with downregulation TSH receptor (Kleiman et al. 2013). Altogether these studies support the association between NIS inhibition-induced decreased iodide uptake (KE up) and reduced TH synthesis (KE down).

Several in vitro and epidemiological studies have shown that iodide uptake blockade occurring as a consequence of NIS (and TPO) inhibition leads to reduced TH synthesis: - Spitzweg et al., 1999: In this in vitro study, a 48 hr treatment of FRTL-5 cells with MMI (100 µM), PTU (100 µM), and potassium iodide (40 µM) induced ~ 50% decrease of NIS mRNA steady-state levels. Incubation with MMI and PTU resulted in a 20% and 25% decrease of iodide accumulation, respectively, whereas potassium iodide suppressed iodide accumulation by approximately 50%. - Wu Y et al., 2016: This in vitro study showed that triclosan, triclocarban, 2,2',4,4'-tetrabromodiphenyl ether (BDE-47), and bisphenol A (BPA) induced a concentration-dependent inhibition of NIS-mediated iodide uptake. Moreover,  triclosan or triclocarban did not affect the expression of genes involved in TH synthesis (Slc5a5, TPO, and Tgo) or thyroid transcription factors (Pax8, Foxe1, and Nkx2-1), BDE-47 decreased the level of TPO, while BPA altered the expression of all six genes, as shown in rat thyroid follicular FRTL-5 cells. At the same time, triclosan and triclocarban also inhibited the activity of TPO at 166 and >300 μM, respectively.  - Steinmaus et al., 2016b: In 1,880 pregnant women from San Diego County, California, during 2000–2003, it has been found that the presence of high level of perchlorate, thiocyanate, nitrate, and iodide in water supply induced a decrease of total thyroxine (T4) [regression coefficient (β) = –0.70; 95% CI: –1.06, –0.34], a decrease of free thyroxine (fT4) (β = –0.053; 95% CI: –0.092, –0.013), and an increase of thyroid-stimulating hormone (TSH), all indicators of reduced TH synthesis. - Horton et al., 2015: in this study TSH levels measured in blood samples of 284 pregnant women at 12 (± 2.8) weeks gestation were found to positively correlate with the levels of urinary concentrations of perchlorate, nitrate and thiocyanate (NIS inhibitors), but perchlorate had the largest weight in the index, indicating the largest contribution to the weighted quantile sum regression. This indicates a perchlorate-dependent alteration of maternal thyroid function, through NIS inhibition. - Brechner et al., 2000: Median newborn TSH levels in a city where drinking water supply was perchlorate-contaminated (from the Colorado River below Lake Mead) were significantly higher than those in a city totally supplied with non-perchlorate-contaminated drinking water, even after adjusting for factors known or suspected to elevate newborn TSH levels. - Charatcharoenwitthaya et al. 2014: this cross-sectional epidemiological study conducted in 200 pregnant Thai women with a gestational age of 14 weeks or less, showed that low-level exposure to perchlorate (i.e., 1.9 μg/L of urinary perchlorate) was positively associated with TSH and negatively associated with free T4 using multivariate analyses in first-trimester pregnant women. Low thiocyanate urinary levels (510.5 μg/L) were also positively associated with TSH in a subgroup of pregnant women with low iodine excretion (less than 100 μg/L). Several other studies have proven that NIS inhibitors lead to a decrease of thyroidal iodide uptake (Jones et al., 1996; Tonacchera et al., 2004; De Groef et al., 2006; Waltz et al., 2010), leading to a reduction of TH synthesis.

Some studies have highlighted contradictory results in relation to response to chemicals. For instance, PTU and MMI have been shown to inhibit the activity of TPO in rats (Davidson et al., 1978), while inducing an increase of cellular TPO activity and TPO mRNA in cultured porcine thyroid follicles (Sugawara et al., 1999). PTU was also found to increase NIS gene expression, and the accumulation of 125I, as shown in in rat thyroid FRTL-5 cells, while MMI had no effect (Sue et al., 2012). Moreover, despite the well described effects of perchlorate, thiocyanate, nitrate, and iodide on iodide uptake into the thyroid, occupational and clinical dosing studies have not identified clear adverse effects, particularly in the case of perchlorate (Tarone et al. 2010). For instance, a longitudinal epidemiologic Chilean study found that there were no increases of thyroglobulin (Tg) or thyrotropin (TSH) levels, and no decreases of free T4 levels among either women during early pregnancy, late pregnancy, or the neonates at birth related to perchlorate in drinking water, suggesting that perchlorate in drinking water at 114 microg/L did not cause changes in neonatal thyroid function or fetal growth retardation (Téllez Téllez et al., 2005). Similarly, no associations between urine perchlorate concentrations and serum TSH or free T4 were found in individual euthyroid or hypothyroid/hypothyroxinemic cohorts of 261 hypothyroid/hypothyroxinemic and 526 euthyroid women from Turin and 374 hypothyroid/hypothyroxinemic and 480 euthyroid women from Cardiff (Pearce et al., 2010), suggesting that log perchlorate may not be a predictor of serum free T4 or TSH. However, it should be considered that these studies may be limited by short study durations, and the inclusion of mostly healthy adults (Steinmaus, 2016b). Charnley's (2008) review examines several studies pointing out a number of inconsistent conclusions regarding link between TH serum levels, urinary iodine concentrations, and environmental perchlorate exposure (Charnley et al. 2008). For instance, no correlations were found between TH serum levels and urinary iodine concentrations among women exposed to perchlorate participating in the 2000-2001 National Health and Nutrition Examination Survey (NHANES). Available evidence does not support a causal relationship between changes in TH levels and current environmental levels of perchlorate exposure, but does support the conclusion that the US EPA's reference dose (RfD) for perchlorate is conservatively health-protective. However, potential perchlorate risks are unlikely to be distinguishable from the ubiquitous background of naturally occurring substances present at much higher exposures that can affect the thyroid via the same biological mode of action as perchlorate, such as nitrate and thiocyanate. Therefore, risk management approaches that account for both aggregate and cumulative exposures and that consider the larger public health context in which exposures are occurring are desirable. In a cross-sectional analysis, McMullen et al. (2017) evaluated the exposure to perchlorate, thiocyanate, and nitrate in 3151 participants aged 12 to 80, to assess whether sensitivity  to perchlorate, thiocyanate, and nitrate (NIS inhibitors) could be a factor of age and sex. These results indicate that adolescent boys and girls represent the most vulnerable subpopulations to NIS symporter inhibitors. Therefore, discrepancies in results described in epidemiological studies may be due to difference in age of study participants.  Apart from age, relative source contribution of perchlorate exposure plays an important role in determining a significant reduction of serum TH levels. For instance, Lumen and George (2017) showed that there was no significant difference in geometric mean estimates of free T4 when perchlorate exposure from food only was compared to no perchlorate exposure in pregnant women. The reduction in maternal free T4 levels reached statistical significance when an added contribution from drinking water was assumed in addition to the 90th percentile of food intake for pregnant women. In particular, a daily intake of 0.45- 0.50μg/kg/day of perchlorate was necessary to produce results that were significantly different than those obtained from no perchlorate exposure. The authors comment that 'these modelling results can explain why findings from observational studies present inconsistent outcomes regarding the relationship between perchlorate exposure and thyroid hormone levels'."

In vitro and in vivo studies have specifically reported data supporting quantitative understanding of this KER. - Gilbert et al., 2011: This in vivo study examined the relationship between graded levels of iodine (ID) in rats and serum thyroid hormones levels, thyroid iodine content, and urinary iodide excretion. The study provided parametric and dose-response information for development of a quantitative model of the thyroid axis. Female Long Evans rats were fed casein-based diets containing varying iodine (I) concentrations for 8 weeks. Diets were created by adding 975, 200, 125, 25, or 0 μg/kg I to the base diet (~25 μg I/kg chow) to produce 5 nominal I levels, ranging from excess (basal+added I, Treatment 1: 1000 μg I/kg chow) to deficient (Treatment 5: 25 μg I/kg chow). Food intake and body weight were monitored throughout and on 2 consecutive days each week over the 8-week exposure period, animals were placed in metabolism cages to capture urine. Food, water intake, and body weight gain did not differ among treatment groups. Serum T4 was dose-dependently reduced relative to Treatment 1 with significant declines (19 and 48%) at the two lowest I groups, and no significant changes in serum T3 or TSH were detected. Increases in thyroid weight and decreases in thyroidal and urinary iodide content were observed as a function of decreasing ID in the diet. Data were compared with predictions from a published biologically based dose-response (BBDR) model for ID. These results challenged existing models and provide essential information for development of quantitative BBDR models for ID during pregnancy and lactation. - Spitzweg et al., 1999:  this in vitro study showed that inhibition of TH synthesis (induced by TPO specific inhibitors) decreases the expression of NIS. A 48 hr treatment of FRTL-5 cells with the TPO specific inhibitors MMI (100 µM), PTU (100 µM), and potassium iodide (40 µM), induced a ~ 50% decrease of NIS RNA steady-state levels. Incubation with MMI and PTU resulted in a 20% and 25% decrease of iodide accumulation, respectively, whereas potassium iodide suppressed iodide accumulation by approximately 50%. - Wu F et al., 2012: An in vivo study found that high dose of NIS inhibitor perchlorate (520 mg/kg b.wt.) in Sprague-Dawley rats (28-day old) caused a decrease of Tg (~ 50% lower than control), and TPO (~ 45% lower than control) gene expression, indicative of reduced TH biosynthesis, together with a decrease of free T3 (~ 50% lower than control) and free T4 levels (~ 50% lower than control), and a remarkable increase of TSH levels (125% higher than control) (Wu F et al. 2012). Additional studies with quantitative data for this KER are also described in Empirical Support for Linkage. However, further studies are needed in order to drive global conclusions about the magnitude of iodide uptake inhibition required to impact TH synthesis.

High Mixed

High

During brain development

High High Not Specified

Empirical evidence comes from in vivo studies in rats (Wu F et al., 2012; Davidson et al., 1978), in vitro studies using thyroid follicular rat cells (Spitzweg et al., 1999; Sue et al., 2012) and porcine thyroid follicles (Sugawara et al., 1999), and human epidemiological studies (Steinmaus et al., 2016b; Horton et al., 2015; Brechner et al., 2000)

Brechner RJ, Parkhurst GD, Humble WO, Brown MB, Herman WH. (2000). Ammonium perchlorate contamination of Colorado River drinking water is associated with abnormal thyroid function in newborns in Arizona. J Occup Environ Med. Aug;42(8):777-82. Caldwell KL, Jones R, Hollowell JG. (2005). Urinary iodine concentration: United States National Health and Nutrition Examination Survey 2001-2002. Thyroid., 15:692–699. Charnley G. (2008) Perchlorate: overview of risks and regulation. Food Chem Toxicol. 46(7):2307-15 (Review). Charatcharoenwitthaya N, Ongphiphadhanakul B, Pearce EN, Somprasit C, Chanthasenanont A, He X, Chailurkit L, Braverman LE (2014). The association between perchlorate and thiocyanate exposure and thyroid function in first-trimester pregnant Thai women. J Clin Endocrinol Metab. Jul;99(7):2365-71. Dai G, Levy O, Carrasco N. (1996). Cloning and characterization of the thyroid iodide transporter. Nature;379:458–460. Davidson, B., Soodak, M., Neary, J.T., Strout, H.V., and Kieffer, J.D. (1978). The irreversible inactivation of thyroid peroxidase by methylmercaptoimidazole, thiouracil, and propylthiouracil in vitro and its relationship to in vivo findings. Endocrinology 103:871–882. de Benoist B, Andersson M, Takkouche B, et al. (2003).Prevalence of iodine deficiency worldwide. Lancet,  362:1859–1860. De Groef B, Decallonne BR, Van der Geyten S, Darras VM, Bouillon R. (2006). Perchlorate versus other environmental sodium/iodide symporter inhibitors: potential thyroid-related health effects. Europ J Endocr. 155:17-25. Gilbert ME, McLanahan ED, Hedge J, Crofton KM, Fisher JW, Valentín-Blasini L, Blount BC (2011). Marginal iodide deficiency and thyroid function: dose-response analysis for quantitative pharmacokinetic modeling. Toxicology. Apr 28;283(1):41-8. Hollowell JG, Staehling NW, Hannon WH, et al.(1998).  Iodine nutrition in the United States. Trend and public health implications: iodine excretion data from National Health and Nutrition Examination Survey I and III (1971-1974 and 1988-1994). The Journal of Clinical Endocrinology and Metabolism.,  83:3401–3408. Horton MK, Blount BC, Valentin-Blasini L, Wapner R, Whyatt R, Gennings C, Factor-Litvak P. (2015). CO-occurring exposure to perchlorate, nitrate and thiocyanate alters thyroid function in healthy pregnant women. Environ Res. Nov;143(Pt A):1-9. Jones PA, Pendlington RU, Earl LK, Sharma RK, Barrat MD. (1996). In vitro investigations of the direct effects of complex anions on thyroidal iodide uptake: identification of novel inhibitors. Toxicol. In Vitro. 10: 149-160. Kleiman DA, Buitrago D, Crowley MJ, Beninato T, Veach AJ, Zanzonico PB, Jin M, Fahey TJ 3rd, Zarnegar R. (2013). Thyroid stimulating hormone increases iodine uptake by thyroid cancer cells during BRAF silencing. J Surg Res. Jun 1;182(1):85-93. Lazarus JH, Delange F. (2004). Prevalence of iodine deficiency worldwide. Lancet, 363:901-910. Lumen A, George NI (2017). Evaluation of the risk of perchlorate exposure in a population of late-gestation pregnant women in the United States: Application of probabilistic biologically-based dose response modeling. Toxicol Appl Pharmacol. 2017 May 1;322:9-14. McMullen J, Ghassabian A, Kohn B, Trasande L (2017). Identifying Subpopulations Vulnerable to the Thyroid-Blocking Effects of Perchlorate and Thiocyanate. J Clin Endocrinol Metab. Jul 1;102(7):2637-2645. Moog N.K., Entringer S., Heim Ch., Wadhwa PD., Kathmann N., Buss C. (2017).  Influence of maternal thyroid hormones during gestation on fetal  brain development. Neuroscience, 342: 68–100. Pearce EN, Lazarus JH, Smyth PP, He X, Dall'amico D, Parkes AB, Burns R, Smith DF, Maina A, Bestwick JP, Jooman M, Leung AM, Braverman LE. (2010). Perchlorate and thiocyanate exposure and thyroid function in first-trimester pregnant women. J Clin Endocrinol Metab. Jul;95(7):3207-15. Riesco-Eizaguirre G, Rodríguez I, De la Vieja A, Costamagna E, Carrasco N, Nistal M, Santisteban P. (2009). The BRAFV600E oncogene induces transforming growth factor beta secretion leading to sodium iodide symporter repression and increased malignancy in thyroid cancer. Cancer Res. Nov 1;69(21):8317-25. Spitzweg C, Morris JC. (2010). Genetics and phenomics of hypothyroidism and goiter due to NIS mutations. Mol Cell Endocrinol. 322: 56-63. Spitzweg C, Joba W, Morris JC, Heufelder AE. (1999). Regulation of sodium iodide symporter gene expression in FRTL-5 rat thyroid cells. Thyroid. Aug;9(8):821-30. Steinmaus CM. (2016a). Perchlorate in Water Supplies: Sources, Exposures, and Health Effects. Curr Environ Health Rep. Jun;3(2):136-43. Steinmaus C, Pearl M, Kharrazi M, Blount BC, Miller MD, Pearce EN, Valentin-Blasini L, DeLorenze G, Hoofnagle AN, Liaw J. (2016b). Thyroid Hormones and Moderate Exposure to Perchlorate during Pregnancy in Women in Southern California. Environ Health Perspect. Jun;124(6):861-7. Sue M, Akama T, Kawashima A, Nakamura H, Hara T, Tanigawa K, Wu H, Yoshihara A, Ishido Y, Hiroi N, Yoshino G, Kohn LD, Ishii N, Suzuki K.(2012). Propylthiouracil increases sodium/iodide symporter gene expression and iodide uptake in rat thyroid cells in the absence of TSH. Thyroid. 2012 Aug;22(8):844-52. Sugawara M, Sugawara Y, Wen K. (1999). Methimazole and propylthiouracil increase cellular thyroid peroxidase activity and thyroid peroxidase mRNA in cultured porcine thyroid follicles. Thyroid. May;9(5):513-8. Szkudlinski MW, Fremont V, Ronin C, Weintraub BD. (2002). Thyroid-stimulating hormone and thyroid-stimulating hormone receptor structure-function relationships. Physiological Reviews. 82 (2): 473–502. Tarone RE, Lipworth L, McLaughlin JK. (2010). The epidemiology of environmental perchlorate exposure and thyroid function: a comprehensive review. J Occup Environ Med. Jun;52(6):653-60. Taurog A. 2005. Hormone synthesis. In: Werner and Ingbar’s The Thyroid: A Fundamental and Clinical Text (Braverman LE, Utiger RD, eds). Philadelphia:Lippincott, Williams and Wilkins, 47–81. Téllez Téllez R, Michaud Chacón P, Reyes Abarca C, Blount BC, Van Landingham CB, Crump KS, Gibbs JP. (2005). Long-term environmental exposure to perchlorate through drinking water and thyroid function during pregnancy and the neonatal period. Thyroid, Sep;15(9):963-75. Tonacchera M, Pinchera A, Dimida A, Ferrarini E, Agretti P, Vitti P, Santini F, Crump K, Gibbs J. (2004). Relative potencies and additivity of perchlorate, thiocyanate, nitrate, and iodide on the inhibition of radioactive iodide uptake by the human sodium iodide symporter. Thyroid, Dec;14(12):1012-9. Waltz F, Pillette L, Ambroise Y. (2010). A nonradioactive iodide uptake assay for sodium iodide symporter function. Anal Biochem. 396:91-95. Wu F, Zhou X, Zhang R, Pan M, Peng KL. (2012). The effects of ammonium perchlorate on thyroid homeostasis and thyroid-specific gene expression in rat. Environ Toxicol. Aug;27(8):445-52. Wu Y, Beland FA1, Fang JL. (2016). Effect of triclosan, triclocarban, 2,2',4,4'-tetrabromodiphenyl ether, and bisphenol A on the iodide uptake, thyroid peroxidase activity, and expression of genes involved in thyroid hormone synthesis. Toxicol In Vitro. Apr;32:310-9. AOPWiki 2016-11-29T18:41:35

2018-06-04T06:11:03

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Not Specified

AOPWiki 2020-08-25T16:43:49

2020-08-25T16:43:49

<upstream-id>9bedfcac-14d1-49a0-b56e-8eb86d7db03f</upstream-id> <downstream-id>fa76c581-3528-4c8a-b62c-86b29dac93fb</downstream-id> Thyroid hormones (THs), thyroxine (T4) and triiodothyronine (T3) are synthesized by NIS and TPO in the thyroid gland as iodinated thyroglobulin (Tg) and stored in the colloid of thyroid follicles across vertebrates. Secretion from the follicle into serum is a multi-step process. The first involves thyroid stimulating hormone (TSH) stimulation of the separation of the peptide linkage between Tg and TH. The next steps involve endocytosis of colloid, fusion of the endosome with the basolateral membrane of the thyrocyte, and finally release of TH into blood. More detailed descriptions of this process can be found in reviews by Braverman and Utiger (2012) and Zoeller et al. (2007).

The weight of evidence linking these two KEs of decreased TH synthesis and decreased T4 in serum is strong. It is commonly accepted dogma that decreased synthesis in the thyroid gland will result in decreased circulating TH (serum T4).

The biological relationship between two KEs in this KER is well understood and documented fact within the scientific community.

It is widely accepted that TPO inhibition leads to declines in serum T4 levels in adult mammals. This is due to the fact that the sole source for circulating T4 derives from hormone synthesis in the thyroid gland. Indeed, it has been known for decades that insufficient dietary iodine will lead to decreased serum TH concentrations due to inadequate synthesis. Strong qualitative and quantitative relationships exist between reduced TH synthesis and reduced serum T4 (Ekerot et al., 2013; Degon et al., 2008; Cooper et al., 1982; 1983; Leonard et al., 2016; Zoeller and Tan, 2007).  There is more limited evidence supporting the relationship between decreased TH synthesis and lowered circulating hormone levels during development.  Lu and Anderson (1994) followed the time course of TH synthesis, measured as thyroxine secretion rate, in non-treated pregnant rats and correlated it with serum T4 levels. Modeling of TH in the rat fetus demonstrates the quantitative relationship between TH synthesis and serum T4 concentrations (Hassan et al., 2017, 2020; Handa et al., 2021). Furthermore, a wide variety of drugs and chemicals that inhibit TPO are known to result in decreased release of TH from the thyroid gland, as well as decreased circulating TH concentrations. This is evidenced by a very large number of studies that employed a wide variety of techniques, including thyroid gland explant cultures, tracing organification of 131-I and in vivo treatment of a variety of animal species with known TPO inhibitors (King and May,1984; Atterwill et al., 1990; Brown et al., 1986; Brucker-Davis, 1998; Haselman et al., 2020; Hornung et al., 2010; Hurley et al., 1998; Kohrle, 2008; Tietge et al., 2010). Additionally, evidence is available from studies investigating responses to TPO inhibitors in fish. For example, Stinckens et al. (2020) showed reduced whole body T4 concentrations in zebrafish larvae exposed to 50 or 100 mg/L methimazole, a potent TPO inhibitor, from immediately after fertilization until 21 or 32 days of age. Exposure to 37 or 111 mg/L propylthiouracil also reduced T4 levels after exposure up to 14, 21 and 32 days in the same study. Walter et al. (2019) showed that propylthiouracil had no effect on T4 levels in 24h old zebrafish, but decreased T4 levels of 72h old zebrafish. This difference is probably due to the onset of embryonic TH production between the age of 24 and 72 hours (Opitz et al., 2011). Stinckens et al. (2016) showed that exposure to 2-mercaptobenzothiazole (MBT), an environmentally relevant TPO inhibitor, decreased whole body T4 levels in continuously exposed 5 and 32 day old zebrafish larvae. Several other studies have also shown that chemically induced Inhibition of TPO results in reduced TH synthesis in zebrafish (Van der Ven et al., 2006; Raldua and Babin, 2009; Liu et al., 2011; Thienpont et al., 2011; Rehberger et al., 2018). A high concentration of MBT also decreased whole body T4 levels in 6 day old fathead minnows, but recovery was observed at the age of 21 days although the fish were kept in the exposure medium (Nelson et al., 2016). Crane et al. (2006) showed decreased T4 levels in 28 day old fathead minnows continuously exposed to 32 or 100 µg/L methimazole. Temporal Evidence: In mammals, the temporal nature of this KER is applicable to all life stages, including development (Seed et al., 2005).  There are currently no studies that measured both TPO synthesis and TH production during development. However, the impact of decreased TH synthesis on serum hormones is similar across all ages in mammals. Good evidence for the temporal relationship comes from thyroid system modeling of the impacts of iodine deficiency and NIS inhibition (e.g., Degon et al., 2008; Fisher et al., 2013). In addition, recovery experiments have demonstrated that serum thyroid hormones recovered in athyroid mice following grafting of in-vitro derived follicles (Antonica et al., 2012). In Xenopus, it has been shown that depression of TH synthesis in the thyroid gland precedes depression of circulating TH within 7 days of exposure during pro-metamorphosis (Haselman et al., 2020).    In oviparous fish such as zebrafish and fathead minnow, the nature of this KER depends on the life stage since the earliest stages of embryonic development rely on maternal THs transferred to the eggs. Embryonic TH synthesis is activated later during embryo-larval development. (See Domain of applicability) Dose-response Evidence: Dose-response data is lacking from studies that include concurrent measures of both TH synthesis and serum TH concentrations. However, data is available demonstrating correlations between thyroidal TH and serum TH concentrations during gestation and lactation during development (Gilbert et al., 2013). This data was used to develop a rat quantitative biologically-based dose-response model for iodine deficiency (Fisher et al., 2013). In Xenopus, dose-responses were demonstrated in both thyroidal T4 and circulating T4 following exposure to three TPO inhibitors (Haselman et al., 2020).

There are no inconsistencies in this KER, but there are some uncertainties. The first uncertainty stems from the paucity of data for quantitative modeling of the relationship between the degree of synthesis decrease and resulting changes in circulating T4 concentrations. In addition, most of the data supporting this KER comes from inhibition of TPO, and there are a number of other processes (e.g., endocytosis, lysosomal fusion, basolateral fusion and release) that are not as well studied. For example, Kim et al. (2015) investigated the adverse effects of Triphenyl phosphate (TPP), a substance that disrupts the thyroid system. Therefore, Rat pituitary (GH3) and thyroid follicular cell lines (FRTL-5) were studied. In the GH3 cells, TPP led to an upregulation of the expression of important thyroid genes (tsh, tr alpha and tr beta) while T3, a positive control, downregulated the expression of these genes. In FRTL-5 cells, the expression of nis and tpo genes was significantly upregulated, suggesting that TPP stimulates TH synthesis in the thyroid gland. In zebrafish larvae at the age of 7 days post-fertilisation (dpf), TPP exposure resulted in a significant increase in T3 and T4 concentrations and the expression of genes involved in thyroid hormone synthesis. Exposure to TPP also significantly regulated the expression of genes involved in the metabolism (dio1), transport (ttr) and excretion (ugt1ab) of THs. The down-regulation of the crh and tsh genes in the zebrafish larvae suggests the activation of a central regulatory feedback mechanism that is triggered by the increased T3 levels in vivo. Taken together, these observations indicate that TPP increases TH concentrations in early life stages of zebrafish by disrupting central regulatory and hormone synthesis pathways.

During Xenopus metamorphosis, circulating T4 steadily increases to peak levels at metamorphic climax. Therefore, during Xenopus metamorphosis, this KER is operable at an increased rate as compared to a system that is maintaining steady circulating T4 levels through homeostatic control. In this case, developmental status is a modulating factor for the rates and trajectories of these KEs.

In rats, Hassan et al. (2020) demonstrated in vitro: ex vivo correlations of TPO inhibition using PTU and MMI and constructed a quantitative model relating level of TPO inhibition with changes in circulating T4 levels. They determined that 30% inhibition of TPO was sufficient to decrease circulating T4 levels by 20%. This is further supported by studies of Hassan et al. (2017) and Handa et al. (2021) In Xenopus, Haselman et al. (2020) collected temporal and dose-response data for both thyroidal and circulating T4 which showed strong qualitative concordance of the response-response relationship. A quantitative relationship exists there in, but is yet to be demonstrated mathematically in this species.

Fisher et al. (2013) published a quantitative biologically-based dose-response model for iodine deficiency in the rat. This model provides quantitative relationships for thyroidal T4 synthesis (iodine organification) and predictions of serum T4 concentrations in developing rats. There are other computational models that include thyroid hormone synthesis. Ekerot et al. (2012) modeled TPO, T3, T4 and TSH in dogs and humans based on exposure to myeloperoxidase inhibitors that also inhibit TPO.  This model was recently adapted for rats(Leonard et al., 2016) and Hassan et al (2017) have extended it to include the pregnant rat dam in response to TPO inhibition induced by PTU. While the original model predicted serum TH and TSH levels as a function of oral dose, it was not used to explicitly predict the relationship between serum hormones and TPO inhibition, or TH synthesis. Leonard et al. (2016) recently incorporated TPO inhibition into the model. Degon et al (2008) developed a human thyroid model that includes TPO, but does not make quantitative prediction of organification changes due to inhibition of the TPO enzyme. Further empirical support for the response-response relationship has been demonstrated in the amphibian model, Xenopus laevis, exposed to TPO inhibitors during pro-metamorphosis (Haselman et al., 2020) wherein temporal profiles were measured for both thyroidal and circulating T4.

Given that the thyroid gland contains follicular lumen space filled with stored thyroglobulin/T4, complete inhibition of thyroid hormone synthesis at a given point in time will not result in an instantaneous decrease in circulating T4. The system will be capable of maintaining sufficient circulating T4 levels until the gland stores are depleted. The time it takes to deplete stored hormone will greatly depend on species, developmental status and numerous other factors. In Xenopus, Haselman et al. (2020) demonstrated an approximately 5 day difference between a significant decrease in thyroidal T4 preceding a significant decrease in circulating T4 while exposed to a potent TPO inhibitor (MMI) continuously during pro-metamorphosis.

This KER is entirely influenced by the feedback loop between circulating T4 originating from the thyroid gland and circulating TSH originating from the pituitary. Intermediate biochemical processes exist within the hypothalamus to affirm feedback and coordinately release TSH from the pituitary. However, quantitative representations of these feedback processes are limited to models discussed previously. In Xenopus, circulating levels of T4 increase through pro-metamorphosis indicating a "release" of feedback to allow circulating levels of T4 to increase and drive metamorphic changes (Sternberg et al., 2011). This provides evidence that homeostatic control of feedback can be developmentally dependent, and likely species dependent.

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Alexander EK, Pearce EN, Brent GA, Brown RS, Chen H, Dosiou C, Grobman WA, Laurberg P, Lazarus JH, Mandel SJ, Peeters RP, Sullivan S.  2017 Guidelines of the American Thyroid Association for the Diagnosis and Management of Thyroid Disease During Pregnancy and the Postpartum. Thyroid. 2017 Mar;27(3):315-389. Antonica F, Kasprzyk DF, Opitz R, Iacovino M, Liao XH, Dumitrescu AM, Refetoff S, Peremans K, Manto M, Kyba M, Costagliola S.  Generation of functional thyroid from embryonic stem cells.  Nature. 2012 491(7422):66-71. Atterwill CK, Fowler KF. A comparison of cultured rat FRTL-5 and porcine thyroid cells for predicting the thyroid toxicity of xenobiotics. Toxicol In Vitro. 1990. 4(4-5):369-74. Braverman, L.E. and Utiger, R.D. (2012). Werner and Ingbar's The Thyroid: A Fundamental and Clinical Text (10 ed.). Philadelphia, PA: Lippincott Williams & Wilkins. pp. 775-786. ISBN 978-1451120639. Brown CG, Fowler KL, Nicholls PJ, Atterwill C. Assessment of thyrotoxicity using in vitro cell culture systems. Food Chem Toxicol. 1986 24(6-7):557-62. Brucker-Davis F. Effects of environmental synthetic chemicals on thyroid function. Thyroid. 1998 8(9):827-56. Chang J, Wang M, Gui W, Zhao Y, Yu L, Zhu G. 2012. Changes in thyroid hormone levels during zebrafish development. Zoological Science. 29(3):181-184. Cooper DS, Kieffer JD, Halpern R, Saxe V, Mover H, Maloof F, Ridgway EC (1983) Propylthiouracil (PTU) pharmacology in the rat. II. Effects of PTU on thyroid function. Endocrinology 113:921-928. Cooper DS, Saxe VC, Meskell M, Maloof F, Ridgway EC.Acute effects of propylthiouracil (PTU) on thyroidal iodide organification and peripheral iodothyronine deiodination: correlation with serum PTU levels measured by radioimmunoassay. J Clin Endocrinol Metab. 1982 54(1):101-7. Crane HM, Pickford DB, Hutchinson TH, Brown JA. 2006. The effects of methimazole on development of the fathead minnow, pimephales promelas, from embryo to adult. Toxicological Sciences. 93(2):278-285. Degon, M., Chipkin, S.R., Hollot, C.V., Zoeller, R.T., and Chait, Y. (2008). A computational model of the human thyroid. Mathematical Biosciences 212, 22–53 Ekerot P, Ferguson D, Glämsta EL, Nilsson LB, Andersson H, Rosqvist S, Visser SA. Systems pharmacology modeling of drug-induced modulation of thyroid hormones in dogs and translation to human. Pharm Res. 2013 30(6):1513-24. Fisher JW, Li S, Crofton K, Zoeller RT, McLanahan ED, Lumen A, Gilbert ME.  Evaluation of iodide deficiency in the lactating rat and pup using a biologically based dose-response model. Toxicol Sci. 2013 132(1):75-86. Gilbert ME, Hedge JM, Valentín-Blasini L, Blount BC, Kannan K, Tietge J, Zoeller RT, Crofton KM, Jarrett JM, Fisher JW.  An animal model of marginal iodine deficiency during development: the thyroid axis and neurodevelopmental outcome.  Toxicol Sci. 2013 132(1):177-95. Handa S, Hassan I, Gilbert M, El-Masri H. 2021. Mechanistic Computational Model for Extrapolating In Vitro Thyroid Peroxidase (TPO) Inhibition Data to Predict Serum Thyroid Hormone Levels in Rats. Toxicological Sciences 183(1):36-48. Haselman, J.T., Olker, J.H., Kosian, P.A., Korte, J.J., Swintek, J.A., Denny, J.S., Nichols, J.W., Tietge, J.E., Hornung, M.W. and Degitz, S.J., 2020. Targeted pathway-based in vivo testing using thyroperoxidase inhibition to evaluate plasma thyroxine as a surrogate metric of metamorphic success in model amphibian Xenopus laevis. Toxicological Sciences, 175(2), pp.236-250. Hassan, I, El-Masri, H., Kosian, PA, Ford, J, Degitz, SJ and Gilbert, ME. Quantitative Adverse Outcome Pathway for Neurodevelopmental Effects of Thyroid Peroxidase-Induced Thyroid Hormone Synthesis Inhibition. Toxicol Sci. 2017 Nov 1;160(1):57-73 Hassan, I., El-Masri, H., Ford, J., Brennan, A., Handa, S., Paul Friedman, K. and Gilbert, M.E., 2020. Extrapolating in vitro screening assay data for thyroperoxidase inhibition to predict serum thyroid hormones in the rat. Toxicological Sciences, 173(2), pp.280-292. Hernandez-Mariano JA, Torres-Sanchez L, Bassol-Mayagoitia S, Escamilla-Nunez M, Cebrian ME, Villeda-Gutierrez EA, Lopez-Rodriguez G, Felix-Arellano EE, Blanco-Munoz J. 2017. Effect of exposure to p,p '-dde during the first half of pregnancy in the maternal thyroid profile of female residents in a mexican floriculture area. Environmental Research. 156:597-604. Hornung MW, Degitz SJ, Korte LM, Olson JM, Kosian PA, Linnum AL, Tietge JE. Inhibition of thyroid hormone release from cultured amphibian thyroid glands by methimazole, 6-propylthiouracil, and perchlorate. Toxicol Sci. 2010 118(1):42-51. Hurley PM. Mode of carcinogenic action of pesticides inducing thyroid follicular cell tumors in rodents. Environ Health Perspect. 1998 106(8):437-45. Kim, S., Jung, J., Lee, I., Jung, D., Youn, H., & Choi, K. (2015). Thyroid disruption by triphenyl phosphate, an organophosphate flame retardant, in zebrafish (Danio rerio) embryos/larvae, and in GH3 and FRTL-5 cell lines. Aquatic Toxicology, 160, 188–196. https://doi.org/10.1016/j.aquatox.2015.01.016 King DB, May JD. Thyroidal influence on body growth. J Exp Zool. 1984 Dec;232(3):453-60. Köhrle J. Environment and endocrinology: the case of thyroidology. Ann Endocrinol (Paris). 2008 69(2):116-22. Leonard JA, Tan YM, Gilbert M, Isaacs K, El-Masri H. Estimating margin of exposure to thyroid peroxidase inhibitors using high-throughput in vitro data, high-throughput exposure modeling, and physiologically based pharmacokinetic/pharmacodynamic modeling. Toxicol Sci. 2016 151(1):57-70. Liu CS, Zhang XW, Deng J, Hecker M, Al-Khedhairy A, Giesy JP, Zhou BS. 2011. Effects of prochloraz or propylthiouracil on the cross-talk between the hpg, hpa, and hpt axes in zebrafish. Environmental Science & Technology. 45(2):769-775. Liu XS, Cai Y, Wang Y, Xu SH, Ji K, Choi K. 2019. Effects of tris(1,3-dichloro-2-propyl) phosphate (tdcpp) and triphenyl phosphate (tpp) on sex-dependent alterations of thyroid hormones in adult zebrafish. Ecotoxicology and Environmental Safety. 170:25-32. Lu, M-H, and Anderson, RR. Thyroxine secretion rats during pregnancy in the rat.  Endo Res. 1994. 20(4):343-364. Nelson K, Schroeder A, Ankley G, Blackwell B, Blanksma C, Degitz S, Flynn K, Jensen K, Johnson R, Kahl M et al. 2016. Impaired anterior swim bladder inflation following exposure to the thyroid peroxidase inhibitor 2-mercaptobenzothiazole part i: Fathead minnow. Aquatic Toxicology. 173:192-203. Opitz R, Maquet E, Zoenen M, Dadhich R, Costagliola S. 2011. Tsh receptor function is required for normal thyroid differentiation in zebrafish. Molecular Endocrinology. 25(9):1579-1599. Paul KB, Hedge JM, Macherla C, Filer DL, Burgess E, Simmons SO, Crofton KM, Hornung MW. Cross-species analysis of thyroperoxidase inhibition by xenobiotics demonstrates conservation of response between pig and rat. Toxicology. 2013. 312:97-107. Power DM, Llewellyn L, Faustino M, Nowell MA, Bjornsson BT, Einarsdottir IE, Canario AV, Sweeney GE. 2001. Thyroid hormones in growth and development of fish. Comp Biochem Physiol C Toxicol Pharmacol. 130(4):447-459. Raldua D, Babin PJ. 2009. Simple, rapid zebrafish larva bioassay for assessing the potential of chemical pollutants and drugs to disrupt thyroid gland function. Environmental Science & Technology. 43(17):6844-6850. Rehberger K, Baumann L, Hecker M, Braunbeck T. 2018. Intrafollicular thyroid hormone staining in whole-mount zebrafish (danio rerio) embryos for the detection of thyroid hormone synthesis disruption. Fish Physiology and Biochemistry. 44(3):997-1010. Sternberg, R.M., Thoemke, K.R., Korte, J.J., Moen, S.M., Olson, J.M., Korte, L., Tietge, J.E. and Degitz Jr, S.J., 2011. Control of pituitary thyroid-stimulating hormone synthesis and secretion by thyroid hormones during Xenopus metamorphosis. General and comparative endocrinology, 173(3), pp.428-437. Stinckens E, Vergauwen L, Blackwell BR, Anldey GT, Villeneuve DL, Knapen D. 2020. Effect of thyroperoxidase and deiodinase inhibition on anterior swim bladder inflation in the zebrafish. Environmental Science & Technology. 54(10):6213-6223. Stinckens E, Vergauwen L, Schroeder A, Maho W, Blackwell B, Witters H, Blust R, Ankley G, Covaci A, Villeneuve D et al. 2016. Impaired anterior swim bladder inflation following exposure to the thyroid peroxidase inhibitor 2-mercaptobenzothiazole part ii: Zebrafish. Aquatic Toxicology. 173:204-217. Thienpont B, Tingaud-Sequeira A, Prats E, Barata C, Babin PJ, Raldua D. 2011. Zebrafish eleutheroembryos provide a suitable vertebrate model for screening chemicals that impair thyroid hormone synthesis. Environmental Science & Technology. 45(17):7525-7532. Tietge, J.E., Butterworth, B.C., Haselman, J.T., Holcombe, G.W., Hornung, M.W., Korte, J.J., Kosian, P.A., Wolfe, M. and Degitz, S.J., 2010. Early temporal effects of three thyroid hormone synthesis inhibitors in Xenopus laevis. Aquatic Toxicology, 98(1), pp.44-50. van der Ven LTM, van den Brandhof EJ, Vos JH, Power DM, Wester PW. 2006. Effects of the antithyroid agent propylthiouracil in a partial life cycle assay with zebrafish. Environmental Science & Technology. 40(1):74-81. Van Herck SL, Geysens S, Delbaere J, Darras VM.  Regulators of thyroid hormone availability and action in embryonic chicken brain development. Gen Comp Endocrinol. 2013. 190:96-104. Wabukebunoti MAN, Firling CE. 1983. The prehatching development of the thyroid-gland of the fathead minnow, pimephales-promelas (rafinesque). General and Comparative Endocrinology. 49(2):320-331. Walter KM, Miller GW, Chen XP, Yaghoobi B, Puschner B, Lein PJ. 2019. Effects of thyroid hormone disruption on the ontogenetic expression of thyroid hormone signaling genes in developing zebrafish (danio rerio). General and Comparative Endocrinology. 272:20-32. Webster GM, Venners SA, Mattman A, Martin JW. 2014. Associations between perfluoroalkyl acids (pfass) and maternal thyroid hormones in early pregnancy: A population-based cohort study. Environmental Research. 133:338-347. Zoeller, R. T., Tan, S. W., and Tyl, R. W. (2007). General background on the hypothalamic-pituitary-thyroid (HPT) axis. Critical reviews in toxicology 37(1-2), 11-53. AOPWiki 2016-11-29T18:41:33

2022-10-10T08:56:38

Sodium Iodide Symporter (NIS) Inhibition leading to altered amphibian metamorphosis

NIS inhib alters metamorphosis

Jonathan Haselman

Jonathan T. Haselman, Center for Computational Toxicology and Exposure, US EPA, Duluth, MN, USA <haselman.jon@epa.gov> Sigmund J. Degitz, Center for Computational Toxicology and Exposure, US EPA, Duluth, MN, USA <degitz.sigmund@epa.gov> Michael W. Hornung, Center for Computational Toxicology and Exposure, US EPA, Duluth, MN, USA <hornung.michael@epa.gov>

BY-SA

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This AOP describes how intracellular iodine deficits in thyroid follicular cells via chemical inhibition of sodium-iodide symporter (NIS) decrease thyroid hormone (TH) synthesis and cause delayed amphibian metamorphosis, or in extreme cases, arrests development. Amphibian metamorphosis is mediated by TH and successful completion of metamorphosis is generally required for organism survival. NIS is a critical transport protein that mediates iodine uptake into thyroid follicular cells making it available for thyroperoxidase (see TPO AOP) to catalyze its covalent bonding to tyrosine residues of thyroglobulin. TPO subsequently couples the iodinated tyrosines to form thyroxine (T4). Conversion of T4 to the active hormone, triiodothyronine (T3), is catalyzed by type I or II deiodinase enzymes (see DIO1 and DIO2 pAOPs) located within the peripheral organs and tissues, which then binds to thyroid receptor (TR). Activated TR then stimulates gene expression that drives the anatomical and physiological changes encompassed by the metamorphic process including limb emergence and development, lung development, gill and tail resorption, gut remodeling, metabolic profile changes in the liver, skin keratinization, etc. The model NIS inhibitor, perchlorate, has been tested in amphibian model species Xenopus laevis using in vivo study designs aiming to characterize temporal profiles of glandular hormone levels in addition to serum hormone levels and associated thyroid gland histopathology. Although there are only a few studies in amphibians that directly address NIS inhibition, these studies provide a strong weight of evidence supporting the specificity and essentiality of NIS inhibition leading to well-supported essential key events downstream.

Thyroid Disrupting Chemicals (TDCs) are defined as the xenobiotics that interfere with the thyroid axis with different outcomes for the organism. A very well-studied mechanism of action of the TDCs is the reduction of the circulating levels of THs by inhibiting hormone synthesis in the thyroid gland. For example, perchlorate is a very potent inhibitor of iodide uptake through the sodium/iodide symporter (Tonacchera et al., 2004). Perchlorate has been detected in human breast milk ranging from 1.4 to 92.2 mg μl–1 (10.5 μg l–1 mean) in 18 US states (Kirk et al. 2005), and 1.3 to 411 μg l–1 (9.1 μg l–1 median) in the Boston area, United States (Pearce et al. 2007). Perchlorate has also been detected in human colostrum of 46 women in the Boston area (from < 0.05 to 187.2 μmol l–1 (Leung et al. 2009)). The mechanism of perchlorate action is quite simple, as it is believed to be mediated only by the NIS inhibition (Dohan et al., 2007; Wolff, 1998). Additionally, thiocyanate and nitrate are two known inhibitors that have been found to reduce circulating TH levels (Blount et al., 2006; Steinhaus et al., 2007), but they are both less potent than perchlorate (Tonacchera et al., 2004). However, there are also contradictory results from other studies that showed no correlation between thyroid parameters and perchlorate levels in humans (Pearce et al., 2010; Amitai et al., 2007; Tellez et al., 2005). Co-occurrence of perchlorate, nitrate, and thiocyanate can alter thyroid function in pregnant women. Horton et al. (2015) have shown positive associations between the weighted sum of urinary concentrations of these three analytes and increased TSH, with perchlorate showing the largest weight in the index. Interestingly, De Groef et al. 2006 showed that nitrate and thiocyanate, acquired through drinking water or food, account for a much larger proportion of iodine uptake inhibition than perchlorate, suggesting that NIS inhibition and any potential downstream effect by perchlorate are highly dependent on the presence of other environmental NIS inhibitors and iodine intake itself (Leung et al., 2010). In particular, Tonacchera et al. (2004) showed that the relative potency of perchlorate to inhibit radioactive I− uptake by NIS is 15, 30 and 240 times that of thiocyanate, iodide, and nitrate respectively on a molar concentration basis. These data are in line with earlier studies in rats (Alexander and Wolff, 1996; Greer et al. 1966). Contradictory findings in these studies may therefore be a result of the confounding mixtures in the environment, masking the primary effect of perchlorate. Decreased iodine intake can decrease TH production, and therefore exposure to perchlorate might be particularly detrimental in iodine-deficient individuals (Leung et al. 2010). Moreover, biologically based dose-response modeling of the relationships among iodide status (e.g., dietary iodine levels), perchlorate dose, and TH production in pregnant women has shown that iodide intake has a profound effect on the likelihood that exposure to goitrogens will produce hypothyroxinemia (Lewandowski et al. 2015). During pregnancy TH requirements increase, particularly during the first trimester (Alexander et al. 2004; Leung et al. 2010), due to higher concentrations of thyroxine-binding globulin, placental T4 inner-ring deiodination leading to the inactive reverse T3 (rT3), and transfer of small amounts of T4 to the foetus (during the first trimester foetal thyroid function is absent). Moreover, glomerular filtration rate and clearance of proteins and other molecules are both increased during pregnancy, possibly causing increased renal iodide clearance and a decreased of circulating plasma iodine (Glinoer, 1997). Thus, even though the foetal thyroid can trap iodide by about 12 week of gestation (Fisher and Klein, 1981), high concentrations of maternal perchlorate may potentially decrease thyroidal iodine available to the foetus by inhibiting placental NIS (Leung et al. 2010). Consequences of TH deficiency depend on the developmental timing of the deficiency (Zoeller and Rovet, 2004). For instance, if the TH deficiency occurs during early pregnancy, offspring show visual attention, visual processing and gross motor skills deficits, while if it occurs later, offspring may show subnormal visual and visuospatial skills, along with slower response speeds and motor deficits. If TH insufficiency occurs after birth, language and memory skills are most predominantly affected (Zoeller and Rovet, 2004). Along this line, age and developmental stage are crucial in determining sensitivity to NIS inhibitors (e.g., perchlorate, thiocyanate, and nitrate). In this regard, McMullen et al. (2017) have shown that adolescent boys and girls, more than adults, represent vulnerable subpopulations to NIS symporter inhibitors. Altogether these studies indicate that age, gender, developmental stage, and dietary iodine levels can affect the impact of NIS inhibitors. Finally, ten more small simple-structured molecules were identified in a large screening study (Lecat-Guillet et al., 2008b) that could block iodide uptake by specifically disrupting NIS in a dose-dependent manner. These molecules were named Iodide Transport Blockers (ITBs). There are few organic molecules that lead to NIS inhibition but no direct interaction with NIS has been determined (Gerard et al., 1994; Kaminsky et al., 1991). Up to date, only dysidenin, a toxin isolated from the marine sponge Dysidea herbacea, has been reported to specifically inhibit NIS (Van Sande et al., 2003). Finally, the aryltrifluoroborates were found to inhibit iodide uptake with an IC50 value of 0.4 μM on rat-derived thyroid cells (Lecat-Guillet et al., 2008a). The biological activity is rationalized by the presence of the BF3− ion as a minimal binding motif for substrate recognition at the iodide binding site. It has been also shown that many anions, such as ClO3-, SCN-, NO3-, ReO4-, TcO4- and in a lower extent Br- and BF4-, are also acting as NIS substrates and they enter the cell by the same transporter mechanism (Van Sande et al., 2003). It has been also shown that ClO4- is transferred by NIS with high affinity and is considered as one of its most potent inhibitors (Dohan et al., 2007). Most recently, the aryltrifluoroborates were also shown to inhibit NIS function (Lecat-Guillet et al., 2008a). A library of 17,020 compounds was tested by a radioactive screening method with high specificity using transfected mammalian cells (Lecat-Guillet et al., 2008b; 2007) for NIS inhibition evaluation. Further studies with the most powerful inhibitors showed a high diversity in their structure and mode of action (Lindenthal et al., 2009).

Altered metamorphosis is a critical apical endpoint evaluated as part of regulatory test guideline studies (OECD, 2009, 2015; US EPA 2009, 2015). Measurable effects on metamorphic rates can be an indication of endocrine disruption, and more specifically thyroid disruption, due to the requirement of thyroid hormone for amphibians to undergo metamorphosis. Although this outcome is evaluated at the level of the individual organism, delayed or arrested metamorphosis can have implications toward population-level effects; however, significant effects on metamorphic rates are typically considered in a weight-of-evidence evaluation to determine a chemical's potential to cause thyroid disruption.

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Hornung, M.W., Degitz, S.J., Korte, L.M., Olson, J.M., Kosian, P.A., Linnum, A.L. and Tietge, J.E., 2010. Inhibition of thyroid hormone release from cultured amphibian thyroid glands by methimazole, 6-propylthiouracil, and perchlorate. Toxicological Sciences, 118(1), pp.42-51. Tietge, J.E., Holcombe, G.W., Flynn, K.M., Kosian, P.A., Korte, J.J., Anderson, L.E., Wolf, D.C. and Degitz, S.J., 2005. Metamorphic inhibition of Xenopus laevis by sodium perchlorate: effects on development and thyroid histology. Environmental Toxicology and Chemistry, 24(4), pp.926-933. Tietge, J.E., Butterworth, B.C., Haselman, J.T., Holcombe, G.W., Hornung, M.W., Korte, J.J., Kosian, P.A., Wolfe, M. and Degitz, S.J., 2010. Early temporal effects of three thyroid hormone synthesis inhibitors in Xenopus laevis. Aquatic Toxicology, 98(1), pp.44-50. AOPWiki 2016-11-29T18:41:17

2026-01-11T16:56:07