Introduction
Congenital hypothyroidism (CH) is the most frequent endocrine-metabolic disease in infancy, with an incidence of about 1/2000-4000 newborns . With the exception of rare cases due to hypothalamic or pituitary defects, CH is characterized by elevated TSH in response to reduced thyroid hormone levels.
In the majority of cases (80-85%), primary permanent CH is due to alterations occurring during the gland organogenesis, resulting either in a thyroid that is absent (thyroid agenesis or athyreosis) or hypoplastic (thyroid hypoplasia) or located in an unusual position (thyroid ectopy). All these entities are grouped under the term “ thyroid dysgenesis ” (TD) . TD occurs mostly as a sporadic disease, however a genetic cause of the disease has been demonstrated in about 5% of the reported cases. Genes associated with TD include several thyroid transcription factors expressed in the early phases of thyroid organogenesis ( NKX2.1/TITF1 , FOXE1/TITF2 , PAX8 , NKX2.5 ) as well as genes, like the thyrotropin receptor gene ( TSHR ) expressed later during gland morphogenesis.
In the remaining 15-20% of cases, CH is caused by inborn errors in the molecular steps required for the biosynthesis of thyroid hormones, and generally it is characterized by enlargement of the gland (goiter), presumably due to elevated TSH levels . Thyroid dyshormonogenesis shows classical Mendelian recessive inheritance.
Rarely CH has a central origin, as consequence of hypothalamic and/or pituitary diseases, with reduced production and/or effect of the thyrotropin releasing hormone (TRH) or of the thyrotropin hormone (TSH) .
Congenital hypothyroidism: diagnostics and treatment
Clinical manifestations
CH is usually a sporadic disease with a 2:1 female to male ratio. Familial cases occur with a frequency that is 15-fold higher than by chance alone ; the genetic basis of these familial cases has been established in some, but not all pedigrees .
International studies show that the incidence of permanent primary CH is approximately 1 in 3500 newborns (in iodine sufficient areas). There is considerable ethnic variation in incidence, ranging from 1 in 30,000 in the African-American population in the United States to 1 in 900 in Asian populations in the United Kingdom .
In absence of an adequate treatment, severe CH results in serious mental retardation, in motor handicaps as well as in the signs and symptoms of impaired metabolism. Before the introduction of a neonatal screening program, congenital hypothyroidism was one of the most frequent causes of mental retardation.
The clinically detectable consequences of CH strongly depend on severity and duration of thyroid hormone deprivation, but there is also a large individual variability in treatment response. In the first four-six months after birth, only untreated patients with severe CH have clinical manifestations. Milder cases can remain undiscovered for years. The only characteristic sign of CH is goiter, but this is present only in the few patients with a defect of the hormonogenesis (Table 1). Thus, the most common feature in young infants with CH is the absence of specific signs.
Table 1. Clinical picture of the forms of congenital hypothyroidism with a genetic origin
| Thyroid alteration | Thyroid morphology | Gene | Clinical manifestations |
| Central hypothyroidism | No goiter | LHX3 and LHX4 | Hypothyroidism, combined pituitary hormone deficiency, short stature, metabolic disorders, reproductive system deficits, nervous system developmental abnormalities |
| HESX1 | Hypothyroidism, septo-optic dysplasia (SOD): hypoplasia of the optic nerves, various types of forebrain defects, multiple pituitary hormone deficiencies | ||
| TRH and TRHR | Hypothyroidism, short stature | ||
| Thyroid dysgenesis | Athyreosis | PAX8 | No goiter, severe hypthyroidism |
| NKX2-5 | No goiter, severe hypothyroidism, no cardiac alterations | ||
| FOXE1 | Severe hypothyroidism, Bamforth-Lazarus syndrome | ||
| Thyroid ectopy | NKX2-5 | No goiter, hypothyroidism, no cardiac alterations | |
| FOXE1 | Hypothyroidism, Bamforth-Lazarus syndrome | ||
| Thyroid hypoplasia | NKX2-1 | No goter, variable hypothyroidism (mild to severe), choreoathetosis, pulmonary alterations | |
| TSHR | Reistance to TSH: no goiter, variable hypothyroidism (mild to severe) | ||
| PAX8 | No goiter, variable hypothyroidism (moderate to severe) | ||
| Dysormonogenesis | Goiter | NIS | Variable hypothyroidism (moderate to severe) |
| TPO | Variable hypothyroidism (moderate to severe) | ||
| DUOX1 and DUOX2 | Permanent hypothyroidism (mild to severe), transient and moderate hypothyroidism | ||
| DUOXA2 | Variable hypothyroidism (mild to severe) | ||
| PDS | Moderate hypothyroidism and deafness; | ||
| TG | Variable hypothyroidism (from moderate to severe) | ||
| DHEAL1 | Variable hypothyroidism (mild to severe) |
I nfants with CH appear to be at increased risk of other congenital anomalies, mostly cardiac (approximately 10% of infants with CH, compared with 3% in the general population) .
Neonatal screening
Screening programs for CH were initially developed in Quebec, Canada, and Pittsburgh, Pennsylvania, in 1974 , and have now been established in Western Europe, North America, Japan, Australia, and parts of Eastern Europe, Asia, South America, and Central America . Since the introduction of the screening, the apparent overall incidence of CH has increased considerably as a consequence of the detection of mild disorders that previously remained undetected or were not recognized as congenital problems.
The population-based newborn screening measures TSH or TSH and total T4 in dried blood spots obtained in the first 3 days of life. In newborns with a screening result suspicious for hypothyroidism, the diagnosis of primary CH is confirmed when serum TSH levels are above and free T 4 levels are below the age-related reference ranges. Hypothalamic-pituitary hypothyroidism is more difficult to diagnose. Most infants with this diagnosis are missed in screening programs unless T 4 and TSH or TSH, T 4 and thyroxine binding globulin (TBG) are simultaneously measured.
If hypothyroidism is confirmed by laboratory analysis, imaging studies should be performed, but it is not acceptable to delay hormone replacement therapy if imaging studies are not readily available .
Diagnosis
Tests commonly used to determine the underlying cause of congenital hypothyroidism are presented in Table 2.
| Table 2. T ests used to complete the diagnosis of CH |
| 1. Imaging studies (to determine thyroid location and size)a. Scintigraphy (99mTc or 123I)b. Ultrasonography |
| 2. Functional studiesa. 123I uptakeb. Serum thyroglobulin |
| 3. Suspected inborn errors of thyroid hormone synthesisa. 123I uptake and perchlorate dischargeb. Serum/salivary/urine iodine studies |
| 4. Suspected autoimmune thyroid diseasea. Maternal and neonatal serum thyroid-antibodies determination |
| 5. Suspected iodine exposure (or deficiency)a. Urinary iodine measurement |
Imaging studies, will be useful to establish the presence of thyroid morphogenesis alterations, which are the most common cause of CH. Thyroid scintigraphy, with 99m technetium or 123 I, is the most informative diagnostic procedure in patients with thyroid dysgenesis . Scintigraphy should be performed immediately at birth, if this will not delay the start of thyroxine (L-T4) treatment, or around the age of 4, when L-T4 therapy can be interrupted for 4 weeks without consequences for the child development. Very recently it has been reported that 123 I - uptake studies can be performed during L-thyroxine treatment in adult CH patients after intramuscular injections with recombinant human TSH.
Although thyroid ultrasonography is useful in demonstrating enlarged or absent glands, it is less accurate than scintigraphy in showing ectopic glands .
Assay of serum thyroglobulin (Tg) will be usefull in to estabilish the presence of some thyroid tissue, while 123 I - will provide information about the thyroidal uptake of iodide.
More specialized tests, such as perchlorate discharge (see chapter. 6e), evaluation of serum, salivary, and urinary radioiodine , and measurement of serum T 4 precursors (see chapter 6e), may be necessary to delineate specific inborn errors of thyroid hormone biosynthesis .
The measurement of the total urinary iodine excretion differentiates inborn errors from acquired transient forms of hypothyroidism due to iodine deficiency or iodine excess.
A small number of infants with abnormal screening values will have transient hypothyroidism as demonstrated by normal serum T 4 and TSH concentrations at the confirmatory laboratory tests. Transient hypothyroidism is more frequent in iodine-deficient areas and it is much more common in preterm infants. CH can also be the consequence of intrauterine exposure to maternal antithyroid drugs, maternal TSHR-blocking antibodies (TSHRBAb), as well as heterozygous DUOX1 and DUOX2 or TSHR germ-line mutations .
Because the transient nature of the hypothyroidism will not be recognized clinically or through laboratory tests, initial treatment will be similar to that of the infant with permanent CH, however at a later age interruption of therapy allows to distinguish from transient to permanent hypothyroidism .
Genetic classification of congental thyroid diseases
1. Central hypothyroidism
Central hypothyroidism is the less frequent form of CH. It occurs with an incidence of 1 in 50000 newborn, and is generally associated to alterations in hypothalamus or pituitary development.
Most patients with central CH are mildly to moderately hypothyroid. The accompanying pituitary hormonal deficiencies, especially the lack of cortisol, may be responsible for high morbidity and mortality.
Developmental defects of the pituitary.
The pituitary gland is formed from an invagination of the floor of the third ventricle and from Rathke’s pouch, developing into the thyrotropic cell lineage and the four other neuroendocrine cell types, each defined by the hormone produced: TSH, growth hormone (GH), prolactin, gonadotropins (luteinizing hormone [LH] and follicle-stimulating hormone [FSH]), and adrenocorticotropic hormone (ACTH).
The ontogeny of the pituitary gland depends on numerous developmental genes that guide differentiation and proliferation. These genes are highly conserved among species, suggesting crucial evolutionary roles for the proteins (PIT1 and PRPO1, HESX1, LHX3, LHX4 and SOX3).
Lhx3 and Lhx4 belong to the LIM family of homeobox genes that are expressed early in Rathke ’ s pouch. In Lhx3 knockout mice the thyrotropes, somatotropes, lactotropes, and gonadotropes cell lineages are depleted, whereas the adrenocorticotropic cell lineage fails to proliferate. This murine knock out model shows that pituitary organ fate commitment depends on Lhx3. Lhx4 null mutants show Rathke ’ s pouch formation with expression of a glycoprotein subunit , TSH-beta, GH and Pit1 transcripts, although cell numbers are reduced.
Recent studies have identified a variety of mutations in the LHX3 and LHX4 genes in patients with combined pituitary hormone deficiency diseases . These patients have complex and variable syndromes involving short stature, metabolic disorders, reproductive system deficits, and nervous system developmental abnormalities .
Hesx1 (also called Rpx ), a member of the paired-like class of homeobox genes, is one of the earliest markers of the pituitary primordium . Extinction of Hesx1 is important for activation of downstream genes such as Prop1 , suggesting that both proteins act as opposing transcription factors . Targeted disruption of Hesx1 in the mouse revealed a reduction in the prospective forebrain tissue, absent optic vesicles, markedly decreased head size, and severe microphthalmia. A similar phenotype it has been observed in patients with the syndrome of septo-optic dysplasia (SOD).
SOD is a rare heterogeneous hypoplasia of the optic nerves, various types of forebrain defects, and a variety of pituitary hormone deficiencies. Endocrine dysfunction ranges from isolated GH deficiency to complete pituitary hormonal deficiency. The human HESX1 gene maps to chromosome 3p21.1 – 3p21.2, and its coding region spans 1.7 Kb, with a highly conserved genomic organization consisting of four coding exons. The first homozygous missense mutation (Arg160Cys) was found in the homeobox of HESX1 in two siblings with SOD . Subsequently several other homozygous and heterozygous mutations have been shown to present with different phenotypes characterized by pituitary hormone deficiency and SOD .
Defects in the TRH and TRH receptor.
In mice, homozygous deletion of the TRH gene produced a phenotype characterized by hypothyroidism and hyperglycemia . Only few patients with reduced TRH production have been described in the literature , but no human mutations have been identified so far.
Similarly, mice lacking the TRH receptor appear almost normal, with some growth retardation, and decreased serum T 3 , T 4 , and prolactin (PRL) levels but normal serum TSH . So far only one family with a compound heterozygous and one family with homozygous loss of function mutation of TRH receptor have been described.
Defects in Thyroid-Stimulating Hormone (TSH) synthesis
The TSH is synthesized in the pituitary gland under the control of local thyroid hormone and TRH. TSH consists of two different subunits (alpha and beta) noncovalently linked. The TSH alpha-subunit is in common with LH, FSH, and chorionic gonadotropin, while the beta-subunit is unique for TSH. The beta-subunit (gene map locus 1p13) synthesis is under the control of several transcription factors, including POU1F1 and PROP1.
Pit1/POU1F1
Pit1 (called POU1F1 in humans) is a pituitary-specific transcription factor belonging to the POU homeodomain family. The human POUIFI maps to chromosome 3p11 and consists of six exons spanning 17 Kb encoding for a 291 aminoacid protein. After the initial report , several heterozygous, compound heterozygous, and homozygous POU1F1 deletions and missense and nonsense mutations have been reported to cause this type of hereditary CH . Deficiency of GH, prolactin and TSH is generally severe in patients harbouring mutations in POU1F1 .
PROP1
Prop1 (Prophet of Pit1) is a pituitary-specific paired-like homeodomain transcription factor required for the expression of Pit1 , and also important in regulating the Hesx1 expression. Dwarf mice, harboring a homozygous missense mutation in Prop1 , exhibit GH, TSH and prolactin deficiency, and an anterior pituitary gland reduced in size by about 50%. Additionally, these mice have reduced gonadotropin expression .
The human PROP1 maps to chromosome 5q. The gene consists of three exons encoding for a 226 aminoacids protein. After the first report of mutations in PROP1 in four unrelated pedigrees with GH, TSH, prolactin, LH and FSH deficiencies , several distinct mutations have been identified in over 170 patients , suggesting that PROP1 mutations account for 29.5-50% of familial multiple pituitary hormone deficiency. Affected individuals exhibit recessive inheritance . The timing of initiation and the severity of hormonal deficiency in patients with PROP1 mutations is highly variable: diagnosis of GH deficiency preceded that of TSH deficiency in 80%. Following the deficiencies in GH and TSH, there is a reduced fertility due to gonadotropin insufficiency. Although most patients fail to enter puberty spontaneously, some start puberty before deficiencies in LH and FSH evolve. ACTH deficiency is a relatively late manifestation of PROP1 mutation, often evolving several decades after birth. The degree of prolactin deficiency and pituitary morphological alterations are variable .
Structural Thyroid-Stimulating Hormone defects
Mutation in the TSH-beta gene are a rare cause of congenital hypothyroidism, and in all the reported cases, the mutations were homozygous or compound heterozygous. Available data have been recently reviewed by Miyai . The phenotype is very variable and it may range from a very mild hypothyroidism to severe forms associated with mental retardation in case of delayed treatment. Patients with mutation in the TSH-beta are characterized by the presence of low levels of circulating TSH that will not be stimulated by TRH . Finally, cases of immunologycally reactive but biologically inactive TSH have also been reported .
2. Alterations of thyroid morphogenesis (thyroid dysgenesis)
In the majority of cases (80-85%), primary permanent CH is due to alterations occurring during the gland organogenesis, resulting either in a thyroid that is absent (thyroid agenesis or athyreosis) or hypoplastic (thyroid hypoplasia) or located in an unusual position (thyroid ectopy). All these entities are grouped under the term “ thyroid dysgenesis ” (TD). TD occurs mostly as a sporadic disease, however a genetic cause of the disease has been demonstrated in about 5% of the reported cases. Genes associated with TD (Table 3) include several thyroid transcription factors expressed in the early phases of thyroid organogenesis ( NKX2.1/TITF1 , FOXE1/TITF2 , PAX8 , NKX2.5 ) as well as genes, like the thyrotropin receptor gene ( TSHR ) expressed later during gland morphogenesis.
Table 3. Genes involved in thyroid development: chromosomal localization and molecular features of the gene product
| Gene | Chromosome | Features of the gene product | |
| Mouse | Human | ||
| Titf1/Nkx2-1 | 12 C1-C3 | 14q13 | Homeodomain transcription factor |
| Pax8 | 2 | 2q12-14 | Paired domain transcription factor |
| Foxe1 | 4 | 9q22 | Forkhead domain transcription factor |
| Hhex | 19 | 10 | Homeodomain transcription factor |
| Nkx2-5 | 17 | 5q34 | Homeodomain transcription factor |
| Tshr | 12 | 14q31 | G protein coupled receptor |
Athyreosis
The absence of thyroid follicular cells is called athyreosis or agenesis of the thyroid: the term agenesis should be used to define the absence of the gland due to a defective initiation of thyroid morphogenesis, while athyreosis indicates a dysgenesis characterized by the disappearance of the thyroid following any step after the thyroid anlage specification. Athyreosis accounts for 22-44% of the cases of primitive permanent CH ( Figure 1 ). So far, the absence of thyroid was reported in patients with CH associated with FOXE1 gene defects (Bamforth-Lazarus syndrome) , in one subject carrying a mutation in PAX8 , in one patient with NKX2-5 mutation and in one patient with both a heterozygous NKX2-5 mutation and a heterozygous mutation in the PAX8 promoter region (Table 4).
Table 4. Genetic basis of thyroid dysgenesis
| Thyroid alteration | Genes | Clinical manifestations | References |
| Athyreosis | PAX8FOXE1NKX2-5 | HypothyroidismBamforth-Lazarus syndromeAthyreosis, no cardiac alterations | |
| Thyroid Ectopy | NKX2-5FOXE1 | Ectopy, no cardiac alterationsBamforth-Lazarus syndrome | |
| Thyroid hypoplasia | NKX2-1TSHRPAX8 | Choreoathetosis, hypothyroidism, and pulmonary alterationsResistance to TSHHypothyroidism |
The Bamfort-Lazarus syndrome is a clinical entity characterized by cleft palate, bilateral choanal atresia, spiky hair and athyreosis. Mutations in FOXE1 gene have been described in two pairs of siblings affected by this syndrome and in one patient with syndromic congenital hypothyroidism but not athyreosis . All affected members carry homozygous missense mutations in conserved aminoacids within the FOXE1 forkhead domain. The mutant proteins were tested in vitro and have shown a reduction in both DNA binding and transcriptional activity.
Ectopic thyroid
The ectopic thyroid is due to a failure in the descent of the developing thyroid from the thyroid anlage region to its definitive location in front of the trachea, therefore an ectopic thyroid can be found in any location along the path of migration from the foramen caecum to the mediastinum.
In humans more than 50% of TD cases are associated with an ectopic thyroid (Figure 1); however, up to now, only three heterozygous mutations in the NKX2-5 gene and one mutation in FOXE1 have been associated to the human ectopic thyroid (Table 4). The functional studies of the mutant NKX2-5, demonstrated a significant functional impairment with reduction of transactivation properties and a dominant negative effect. The patients described were all heterozygous and the mutations were inherited from one of the parents, suggesting that NKX2-5 mutations have variable penetrance and clinical significance.
Figure 1. Prevalence of the various causes of primitive congenital hypothyroidism.
Recently gene expression, genome-wide methylation, and structural genome variations have been compared between normal and ectopic thyroid tissue. Ectopic thyroids show a differential gene expression compared to normal thyroids, although molecular basis could not be properly defined probably as consequence of the small number of samples examined or of the different gene expression pattern between adult and embryonic gland .
Hypoplasia
The presence of hypoplastic thyroid has been reported in 24-36% of cases of CH (Figure 1). Thyroid hypoplasia is a genetically heterogeneous dysgenesis, since mutations in NKX2-1, PAX8 or TSHR gene have been reported in patients with thyroid hypoplasia (Table 4).
Patients with NKX2-1 loss of-function mutations are affected by choreoathetosis, hypothyroidism, and pulmonary alterations with incomplete penetrance and the variability of the phenotype . So far, several loss of function mutations in the NKX2-1 gene have been identified in patients with this clinical picture . The unfavorable outcome in the case of impaired NKX2.1 expression, regardless of early T 4 supplementation, is most likely caused by defects in the central nervous system rather than fetal hypothyroidism.
The involvement of PAX8 has been described in sporadic and familial cases of CH with TD . In vitro transfection assays demonstrated that the mutated proteins are unable to bind DNA and to drive transcription of the TPO promoter. All affected individuals are heterozygous for the mutations and in the familial cases transmission is autosomal dominant with a variable penetrance and expressivity.
The human TSHR gene maps to chromosome 14q31 and is encoded by ten exons producing a 1.8 Kb mRNA. The TSHR belongs to the superfamily of G protein – coupled receptors. It contains an extracellular N-terminal domain with a repetitive leu-rich motif, seven transmembrane helices, three intracellular and three extracellular loops, and an intracellular C-terminal part. The TSHR is responsible for mediating TSH action on thyroid follicular cell growth, metabolism and function, ultimately resulting in TH synthesis and secretion.
The role of TSHR gene in CH with TSH unresponsiveness and absence of goiter was hypothesized almost forty years ago. Identification of hyt/hyt mice, affected by primary hypothyroidism with elevated TSH and hypoplastic thyroid, with a loss-of-function mutation in the Tshr gene , and the production of Tshr -/- mice offered useful models for this autosomal recessive form of CH.
TSHR mutations in humans were identified for the first time in three siblings with CH associated with high serum TSH and normal thyroid hormone . The siblings were compound heterozygous, carrying a different mutation in each of the two alleles. After this report other mutations in TSHR gene have been identified in several patients with thyroid hypoplasia and increased TSH secretion. All the affected individuals are homozygous or compound heterozygous for loss-of-function mutations, and consistently, in the familial forms, the disease is inherited as an autosomal recessive trait. This form of CH is characterized by a “ small ” thyroid gland in normal position. In the case of total failure of the TSHR function, the patient is severely hypothyroid because the complete lack of TSH stimulation represses almost completely the metabolic activity of the thyroid gland . When the TSHR has a diminished affinity to its ligand, the effect may largely be compensated by high plasma TSH concentrations.
Hemiagenesis
Thyroid hemiagenesis is a dysgenesis in which one thyroid lobe fails to develop. The prevalence of this morphological abnormality ranges from 0.05% to 0.2% in healthy children, with the absence of the left lobe in almost all the cases. In these subjects thyroid function tests are within the normal range .
The molecular mechanisms leading to the formation of the two thyroid symmetrical lobes are still unclear and in humans, candidate genes responsible for the hemiagenesis of the thyroid have not yet been described. Indeed, Shh -/- mice embryos can display either a non lobulated gland or hemiagenesis of thyroid , and hemiagenesis of the thyroid is also frequent in mice double heterozygous Titf1 +/- , Pax8 +/- .
Very recently the role of the polyalanine tract in the FOXE1-gene has been investigated in patients with thyroid hemiagenesis, suggesting that FOXE1-poly-alanine tract expansion may contribute to the molecular background of familial but not sporadic forms of TH .
3. Defects in thyroid hormone synthesis (dyshormonogenesis)
As mentioned before, in about 15% of cases, CH is due to hormonogenesis defects ( Figure 1 ) caused by mutations in genes involved in thyroid hormone synthesis, secretion or recycling. These cases are clinically characterized by the presence of goiter, and the molecular mechanisms in most of these forms have been well defined (Table 5).
Table 5. Gene causing defects in thyroid hormone synthesis
| Gene | Protein function | Inheritance | Human phenotype |
| Sodiun-Iodide symporter ( NIS ) | Transports iodine across basal membrane | AR | CH ( moderate to severe);Euthyroid goiter |
| Thyroperoxidase ( TPO ) | Catalyses the oxidation, organification, and coupling reactions | AR | Goiter and CH due to a total iodide organification defect |
| Dual oxidases( DUOX1 and DUOX2 ) | H 2 O 2 generation in the follicle | AD and AR | Permanent hypo (from mild to severe);Transient and moderate hypo |
| Dual oxidase maturation factor 2 ( DUOXA2 ) | Required to express DUOX2 enzymatic activity | AR | Goiter and CH due to partial iodide organification defect |
| Pendrin ( PDS ) | Transport iodine across apical membrane | AR | Goiter, moderate hypothyroidism and deafness; |
| Thyroglobulin ( TG ) | Support for thyroid hormone synthesis | AD and AR | Goiter and CH (from moderate to severe) |
| Iodotyrosine deiodinase ( DHEAL1 ) | Nitroreductase-related enzyme capable of deiodinating iodotyrosines | AR | Hypothyroidism with variable age of diagnosis |
In thyroid follicular cells, iodide is actively transported and concentrated by the sodium iodide symporter present in the baso-lateral membrane. Subsequently it is oxidised by hydrogen peroxide generation system (thyroperoxidase, Pendrin) and bound to tyrosine residues in thyroglobulin to form iodotyrosine (iodide organification). Some of these iodotyrosine residues (monoiodotyrosine and diiodotyrosine) are coupled to form the hormonally active iodothyronines T 4 and triiodotironine (T 3 ), and, when needed, thyroglobulin is hydrolyzed and hormones are released in the blood. A small part of the iodotyronines are hydrolyzed into the gland, and iodine is recovered by the action of specific enzymes, namely the intrathyroidal dehalogenases.
Defects in any of these steps lead to reduced circulating thyroid hormone, resulting in congenital hypothyroidism and goiter. With the exception of rare cases, all mutations in these genes appear to be inherited in autosomal recessive fashion .
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Sodium-iodide symporter
The sodium-iodide symporter (NIS) is a member of the sodium/solute symporter family that actively transports iodide across the membrane of the thyroid follicular cells. In 1996, NIS mRNAs from rats and humans were isolated. The human gene ( SLC5A5 ) maps to chromosome 19p13.2-p12. It has 15 exons encoding for a 643-amino acid protein expressed primarily in thyroid, but also in salivary glands, gastric mucosa, small intestinal mucosa, lacrimal gland, nasopharynx, thymus, skin, lung tissue, choroid plexus, ciliary body, uterus, lactating mammary tissue and mammary carcinoma cells, and placenta . Only in thyroid cells is iodide transport regulated by TSH.
The inability of the thyroid gland to accumulate iodine was one of the early known causes of CH, and before the cloning of NIS, a clinical diagnosis of hereditary iodide transport defect had been made on the basis of goitrous hypothyroidism and absent thyroidal radioiodine uptake. To date, several mutations inherited in an autosomal recessive manner have been described, with a clinical picture characterized by hypothyroidism of variable severity (from severe to fully compensated) and goiter . Thyroid morphology is heterogeneous in patients with the same NIS mutation .
In the neonatal period, infants with iodide transport defects are found to have a normal-size or slightly enlarged thyroid gland by ultrasonography and elevated serum thyroglobulin levels . Radioactive iodide uptake is absent. Measurement of the saliva-to-plasma 123 I ratio is around one. The degree of hypothyroidism is variable and ranges from mild to severe, possibly depending on the amount of iodide in the diet. These children are severely hypothyroid if maintained with a normal iodine diet, while addition of high amount of iodide to the diet tends to compensate the iodide transport failure.
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Thyroperoxidase
The most frequent cause of dyshormonogenesis is thyroperoxidase (TPO) deficiency. TPO is the enzyme that catalyses the oxidation, organification, and coupling reactions (see Chapter 74).
Accumulation of iodine in the thyroid gland reaches a steady state between active influx, protein binding, and efflux, resulting in a relatively low free intracellular iodide concentration in normal conditions, while increased in the presence of TPO defects. The kinetics of iodide uptake and release can be traced by administration of radioiodide and iodide re-uptake can be inhibited by anions of similar molecular size and charge, such as perchlorate or thiocyanate. Radioiodide uptake and perchlorate inhibition gives an idea of the intrathyroidal iodide concentration in relation to the circulating iodine. Iodine organification defects can be quantified as total or partial: total iodide organification defects are characterized by discharge of more than 90% of the radioiodide taken up by the gland within 1 hour after administration of sodium perchlorate, usually given 2 hours after radioiodide. A total disappearance of the thyroid image is also observed. Partial iodide organification defects are characterized by discharge of 20% to 90% of the accumulated radioiodine .
The human TPO gene is located on chromosome 2p25 and spans approximately 150 kb; the coding sequence of 3048 bp is divided over 17 exons and encodes for a 933 amino acid, membrane bound, glycated, haem containing protein, located on the apical membranes of the thyroid follicular cell.
Defects in the TPO gene have been reported to cause congenital hypothyroidism by a total iodide organification defect, and mutations have been identified in the all exons of the TPO gene. Most mutations are found in exons 8, 9, or 10, encoding the active center and heme-binding place of the enzyme. Nonsense, splice-site, and frameshift mutations have been also described by several groups .
If untreated, patients with organification defects show variable degrees of mental retardation, very large goiter and hypothyroidism. In some cases with partial defects hypothyroidism appears compensated .
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DUOX1 and DUOX2
The generation of H 2 O 2 is a crucial step in thyroid hormonogenesis. Recently two new proteins involved in the H 2 O 2 generation in the apical membrane of the follicular thyroid cell have been identified . These proteins, initially named THOX1 and THOX2 (for th yroid ox idase), maps on chromosome 15q15.3, only 16kb apart from each other and in opposite transcriptional orientation. In 2001, since these proteins contain two distinct functional domains, it has been suggested to call them DUOX ( du al ox idase).
DUOX1 and DUOX2 are glycoproteins with seven putative transmembrane domains. Their function remained unclear until a factor, named DUOXA2, which allows the transition of DUOX2 from the endoplasmic reticulum to the Golgi was identified . The coexpression of this factor with DUOX2 in HeLa cells is able to reconstitute the H 2 O 2 production in vitro. A similar protein (DUOXA1) is necessary for the complete maturation of the DUOX1. Interestingly, both DUOXA genes maps in the 16kb that separates the DUOX1 and DUOX2 genes on chromosome 15.
Several mutations in DUOX genes have been reported in patients with congenital hypothyroidism showing very variable phenotype . In order to produce congenital permanent hypothyroidism a severe alteration of both alleles of DUOX2 gene is required. The presence of some residual activity in one of the alleles may produce a less severe phenotype, whereas monoallelic severe inactivation of the DUOX2 gene is associated with transient CH. In addition, the phenotype of monoallelic inactivation seems to be modulated by other factors, including environmental conditions (such as iodine insufficiency), lifetime events (pregnancy, immediate postnatal life) or alterations in the dual oxidase maturation factor 2 ( DUOXA2 ) .
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Pendrin
In 1896, Vaughan Pendred described a syndrome characterized by congenital neurosensorial deafness and goiter . The disease is transmitted as autosomal recessive disorder. Patients have a moderately enlarged thyroid gland, are usually euthyroid and show only a partial discharge of iodide after the administration of thiocyanate or perchlorate. The impaired hearing is not constant, and is due to a cochlear defect that corresponds to the Mondini ’ s type of developmental abnormality of the cochlea.
In 1997, the PDS gene was cloned and the predicted protein of 780 amino acids (86-kD) was called Pendrin . The PDS gene maps to human chromosome 7q31, contains 21 exons, and it is expressed both in the cochlea and in the thyroid. Pendrin has been localized into the apical membrane of thyroid follicular cell . In thyroid follicular cells, and in transfected oocytes, pendrin is able to transport iodide..
Patients with Pendred ’ s syndrome are subclinically hypothyroid with goiter, and show moderate-to-severe sensineural hearing impairment. Discharge of radioiodide after administration of sodium perchlorate is moderately increased (>20%). The prevalence varies between 1:15,000 and 1:100,000 .
A number of mutations in the PDS gene have been described in patients with Pendred syndrome. Despite the goiter, individuals are likely to be euthyroid and only rarely present congenital hypothyroidism. However, TSH levels are often in the upper limit of the normal range, and hypothyroidism of variable severity may eventually develop .
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Thyroglobulin
Thyroglobulin is a homodimer protein synthesized exclusively in the thyroid. The human gene is located on chromosome 8q24 and the coding sequence, containing 8307 bp , is divided into 42 exons . Following a signal peptide of 19 amino acids, the polypeptide chain is composed of 2750 amino acids containing 66 tyrosine residues. Thyroglobulin is a dimer with identical 330-kDa subunits containing 10% carbohydrate residues.
Patients with disorders of thyroglobulin synthesis are moderately to severely hypothyroid. Usually, plasma thyroglobulin concentration is low, especially in relation to the TSH concentrations, and does not changes after T 4 treatment or injection of TSH. Patients classified in the category “ thyroglobulin synthesis defects ” often have abnormal iodoproteins, mainly iodinated plasma albumin, and they excrete iodopeptides of low molecular weight in the urine .
Several mutations in the thyroglobulin gene have been reported in patients with CH and in animals including Afrikander cattle (p.R697X) , Dutch goats (p.Y296X) , cog/cog mouse (p.L2263P) and rdw rats (p.G2300R) .
Mutations in the human thyroglobulin gene are associated with congenital goiter and with moderate to severe hypothyroidism.
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DEHAL1
In addition to the active transport from the blood due to NIS, iodine in the thyroid follicular cells derives also from the deiodination of monoiodotyrosine and diiodotyrosine . The gene encoding for this enzymatic activity was recently identified and named IYD (or DEHAL1 ) . The human gene maps to chromosome 6q24-q25 and it consists in six exons encoding a protein of 293 amino acids with a nitroreductase-related enzyme capable of deiodinating iodotyrosines.
In the past it was suggested that IYD mutations could be responsible for congenital hypothyroidism, but only very recently four patients with three mutations in the IYD gene have been reported . The disease was transmitted either as autosomal recessive character or dominant pattern of inheritance with incomplete penetration , patients were hypothyroid and goitrous with a high phenotypic variability depending on the time of expression of the disease manifestations. The patients born after the introduction of screening program for CH were not identified by the screening. There is also a variable severity in the clinical picture, and this can derive either form the molecular effects of the mutation (complete absence or partial activity of the protein), or due to environmental factors, such as iodine diet content.
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