Alteration of thyroid gland morphogenesis (thyroid dysgenesis) is a frequent human malformation. Among the one in three to four thousand newborns in which congenital hypothyroidism is detected, 80% have either an ectopic, small and sublingual thyroid, or have no thyroid tissue. Most of these cases appear sporadically, although a few cases of recurring familial thyroid dysgenesis have been described. The lack of evidence for hereditary thyroid dysgenesis may be due to the severity of the hypothyroid phenotype. Neonatal screening and early thyroid hormone therapy have eliminated most of the clinical consequences of hypothyroidism such that the heritability of this condition may become apparent in the near future. We have recently cloned cDNA encoding a forkhead domain-containing transcription factor, TTF-2, and have located the position of the gene, designated Titf2, to mouse chromosome 4 (ref. 3). Titf2 is expressed in the developing thyroid, in most of the foregut endoderm and in craniopharyngeal ectoderm, including Rathke's pouch. Expression of Titf2 in thyroid cell precursors is down-regulated as they cease migration, suggesting that this factor is involved in the process of thyroid gland morphogenesis. Here we show that Titf2-null mutant mice exhibit cleft palate and either a sublingual or completely absent thyroid gland. Thus, mutation of Titf2-/- results in neonatal hypothyroidism that shows similarity to thyroid dysgenesis in humans.

Congenital hypothyroidism with thyroid dysgenesis (TD) is a frequent human condition characterized by elevated levels of TSH in response to reduced thyroid hormone levels. Congenital hypothyroidism is a genetically heterogeneous disease. In the majority of cases studied, no causative mutations have been identified and very often the disease does not show a Mendelian transmission. However, in approximately 5% of cases, it can be a consequence of mutations in genes encoding the TSH receptor or the transcription factors TITF1, FOXE1, or PAX8. We report here that in mouse models, the combination of partial deficiencies in the Titf1 and Pax8 genes results in an overt TD phenotype that is absent in either of the singly deficient, heterozygous mice. The disease is characterized by a small thyroid gland, elevated levels of TSH, reduced thyroglobulin biosynthesis, and high occurrence of hemiagenesis. The observed phenotype is strain specific, and the pattern of transmission indicates that at least two other genes, in addition to Titf1 and Pax8, are necessary to generate the condition. These results show that TD can be of multigenic origin in mice and strongly suggest that a similar pathogenic mechanism may be observed in humans.

The development and the function of central nervous system depend on thyroid hormones. In humans, the lack of thyroid hormones causes cretinism, a syndrome of severe mental deficiency. It is assumed that thyroid hormones affect the normal development and function of the brain by activating or suppressing target gene expression because several genes expressed in the brain have been shown to be under thyroid hormone control. Among these, the Rhes gene, encoding a small GTP-binding protein, is predominantly expressed in the striatal region of the brain. To clarify the role of Rhes in vivo, we disrupted the Rhes gene by homologous recombination in embryonic stem cells and generated mice homozygous for the Rhes null mutation (Rhes ؊/؊ ). Rhes ؊/؊ mice were viable but weighed less than wild-type mice. Furthermore, they showed behavioral abnormalities, displaying a gender-dependent increase in anxiety levels and a clear motor coordination deficit but no learning or memory impairment. These results suggest that Rhes disruption affects selected behavioral competencies.The thyroid hormones thyroxine (T 4 ) and triiodothyronine (T 3 ) have many physiological effects. They exert their actions in all tissues examined and affect many metabolic pathways. Some of the most prominent effects of thyroid hormones occur during fetal development and in early childhood. In humans, the lack of adequate levels of thyroid hormones in the first trimester of life, such as in iodine deficiency (endemic cretinism) (8, 9), or in developmental abnormalities of the thyroid gland (congenital hypothyroidism) (22, 28, 55) results in cretinism, a syndrome of severe mental deficiency, which may be accompanied by retarded growth and/or neurological deficits, such as spastic diplegia. Many of these developmental effects are not reversed by later treatment with hormone, indicating that thyroid hormone acts in a specific developmental window. Therefore, adequate levels of thyroid hormone are required for normal central nervous system development.To date, several specific central nervous system genes whose expression is controlled by thyroid hormone have been identified. The expression of these genes may be decreased (2, 5) or increased (1, 18) in hypothyroidism. Furthermore, the total or partial absence of thyroid hormones may also affect either mRNA stability (43, 54) or the mRNA translational process (43,57,60). The identification of thyroid hormone target genes in the central nervous system and the understanding of their function in central nervous system development are important to understanding the pathogenesis of neurological cretinism at the molecular level.In order to understand the molecular basis of neurological cretinism, we studied the Rhes (Ras homolog enriched in striatum) gene (24). Rhes is predominantly expressed in the striatum, and its expression is controlled by thyroid hormones (59). Interestingly, several lines of evidence indicate that in neurological cretinism, there is damage of striatum, which determines a striatopallidal syndr...

Polypeptide hormones produce a wide range of metabolic and morphologic responses in specific target tissues.1 Clearly the initial interaction of hormone with sensitive tissue must be binding of the hormone to some cellular constituent. To delineate this interaction we have studied the effects of thyroid-stimulating hormone (TSH) on thyroid slices and of insulin on skeletal muscle. Tissues that were exposed to hormone and washed thoroughly in hormone-free medium showed a persistent hormone effect. However, when the washed tissues were subsequently exposed to antibody or to trypsin, this effect was obliterated. Thus, the persistent hormone effect was due to the continued presence of relatively intact hormone at a readily accessible tissue site. .05 M phosphate, pH 7.4, at 500 v at 40 on Whatman 3MM chromatography paper, over 95% of the radioactivity of each of the labeled proteins migrated appropriately as a single peak. Substantially less than 1% of the radioactivity was iodide. At least 80% of the TSH-I131 was bound by TSH antibody. Labeled proteins were stored at -20°and used within 72 hr of preparation.Antibodies were produced in a burro by four intramuscular injections of 6 mg of bovine TSH (4 IU/mg) in complete Freund's adjuvant (Difco). Antibody was prepared from citrated plasma by defibrination with calcium and thrombin and precipitation at 230 with ammonium sulfate at 40% saturation. The guinea pig anti-insulin serum (Suburban Laboratories) was similarly purified. Hereafter antiserum refers to the purified gamma globulin. The anti-TSH serum at a final dilution of 1: 5 neutralized bovine TSH at 50 mU/ml but had little effect on human TSH. The anti-insulin serum at a 1:20 dilution neutralized insulin at 10 mU/ml. Neutralization indicates that a solution of hormone and antibody, allowed to mix for 30 min, failed to stimulate glucose-1-C14 oxidation in thyroid or glucose-i-C14 incorporation into rat diaphragms. An aliquot of burro anti-TSH serum was treated with papain (Worthington) at pH 6 for 5 hr.5 Papain was inactivated by iodoacetamide, and Porter fraction III was precipitated by dialysis in water and removed by centrifugation. Simultaneous with the preparation of each antiserum, serum from an unimmunized animal was treated identically and served as control throughout. All sera were dialyzed against Krebs-Ringer bicarbonate before use.The buffer for in vitro experiments was Krebs-Ringer bicarbonate pH 7.4 containing bovine fraction V (fr. V, Armour) at 5 mg/ml. Routinely, dog thyroid slices weighing 15-30 mg were preincubated with bovine TSH (4 IU/mg) in 2-5 ml of buffer and washed three times by agitation for 1 min in 10 ml of hormone-free buffer at 10. The slices were then blotted, weighed, and incubated at 370 in 1 ml of buffer containing 1 mg of glucose and 0.5 ,uc of glucose-i-C14. The C1402 was collected in hyamine.6 Data are expressed as cpm/mg of thyroid slice per hr of incubation. The thyroid of a single normal dog was used for each experiment.The effect of insulin on glucose utilization in the r...

Ras oncogenes are frequently mutated in thyroid carcinomas. To verify the role played by N-ras in thyroid carcinogenesis, we generated transgenic mice in which a human N-ras(Gln61Lys) oncogene (Tg-N-ras) was expressed in the thyroid follicular cells. Tg-N-ras mice developed thyroid follicular neoplasms; 11% developed follicular adenomas and B40% developed invasive follicular carcinomas, in some cases with a mixed papillary/ follicular morphology. About 25% of the Tg-N-ras carcinomas displayed large, poorly differentiated areas, featuring vascular invasion and forming lung, bone or liver distant metastases. N-ras(Gln61Lys) expression in cultured PC Cl 3 thyrocytes induced thyroid-stimulating hormone-independent proliferation and genomic instability with micronuclei formation and centrosome amplification. These findings support the notion that mutated ras oncogenes could be able to drive the formation of thyroid tumors that can progress to poorly differentiated, metastatic carcinomas.