Germline loss-of-function mutations of TSH receptor (TSHR) gene have been described in families with partial or complete TSH resistance. Large TSH elevations were generally found in the patients with homozygous or compound heterozygous mutations. In this study, we sequenced the entire TSHR gene in a series of 10 unrelated patients with slight (6.6-14.9 mU/liter) to moderate (24-46 mU/liter) elevations of serum TSH, associated with definitely normal free thyroid hormone concentrations. Thyroid volume was normal in all patients, except two with a modest hypoplasia. Autoimmune thyroid disease was excluded in all patients on the basis of clinical and biochemical parameters. Eight patients had at least one first-degree relative bearing the same biochemical picture. TSHR mutations were detected in 4 of 10 cases by analyzing DNA from peripheral leukocytes. A compound heterozygosity (P162A on maternal allele, and the novel mutation C600R on the paternal one) was found in the patient with the highest TSH levels. Only one TSHR allele was mutated in the remaining three cases, and no alterations in TSHR gene promoter were detected in all of these probands. A novel mutation (L467P) was detected on the maternal allele in one patient and in her monozygotic twin. Previously described inactive mutants, T655Delta and C41S, were detected in the other two cases. When tested on several occasions, circulating TSH values fluctuating above the upper limit of the normal range could be shown in heterozygous subjects of these families. A dominant mode of inheritance of the biochemical alterations was detected in these cases. Mutant TSHRs were studied during transient expression in COS7 and HEK293T cells. Their TSH-independent cAMP accumulation activities were very low or similar to mock-transfected cells, and no increases were seen after maximal hormone stimulation. Flow cytometry experiments showed a poor level of expression of all mutant TSHRs at the cell membrane. In conclusion, we found several loss-of-function mutations of TSHR, including two novel ones, in a series of unrelated patients with slightly elevated TSH levels. Therefore, partial resistance to TSH action is a frequent finding among patients with slight hyperthyrotropinemia of nonautoimmune origin. Germline mutations of TSHR may be associated with serum TSH values fluctuating above the upper limit of the normal range, also in the heterozygous state.

Mutations in the orphan nuclear receptor DAX-1 cause X-linked adrenal hypoplasia congenita. Affected boys usually present with primary adrenal failure in early infancy or childhood. Impaired sexual development because of hypogonadotropic hypogonadism becomes apparent at the time of puberty. We report adult-onset adrenal hypoplasia congenita in a patient who presented with hypogonadism at 28 yr of age. Although he had no clinical evidence of adrenal dysfunction, compensated primary adrenal failure was diagnosed by biochemical testing. Semen analysis showed azoospermia, and he did not achieve fertility after 8 months of treatment with gonadotropins. A novel Y380D DAX-1 missense mutation, which causes partial loss of function in transient gene expression assays, was found in this patient. This case demonstrates that partial loss-of-function mutations in DAX1 can present with hypogonadotropic hypogonadism and covert adrenal failure in adulthood. Further, an important role for DAX-1 in spermatogenesis in humans is confirmed, supporting findings in the Dax1 (Ahch) knockout mouse.

A 29-yr-old woman with pituitary resistance to thyroid hormones (PRTH) was found to harbor a novel point mutation (T337A) on exon 9 of the thyroid hormone receptor beta (TRbeta) gene. She presented with symptoms and signs of hyperthyroidism and was successfully treated with 3,5,3'-triiodothyroacetic acid (TRIAC) until the onset of pregnancy. This therapy was then discontinued in order to prevent TRIAC, a compound that crosses the placental barrier, from exerting adverse effects on normal fetal development. However, as the patient showed a recurrence of thyrotoxic features after TRIAC withdrawal, we sought to verify, by means of genetic analysis and hormone measurements, whether the fetus was also affected by RTH, in order to rapidly reinstitute TRIAC therapy, which could potentially be beneficial to both the mother and fetus. At 17 weeks gestation, fetal DNA was extracted from chorionic villi and was used as a template for PCR and restriction analysis together with direct sequencing of the TRbeta gene. The results indicated that the fetus was also heterozygous for the T337A mutation. Accordingly, TRIAC treatment at a dose of 2.1 mg/day was restarted at 20 weeks gestation. The mother rapidly became euthyroid, and the fetus grew normally up to 24 weeks gestation. At 29 weeks gestation mild growth retardation and fetal goiter were observed, prompting cordocentesis. Circulating fetal TSH was very high (287 mU/L) with a markedly reduced TSH bioactivity (B/I: 1.1 +/- 0.4 vs 12.7 +/- 1.2), while fetal FT4 concentrations were normal (8.7 pmol/L; normal values in age-matched fetuses: 5-22 pmol/L). Fetal FT3 levels were raised (7.1 pmol/L; normal values in age-matched fetuses: <4 pmol/L), as a consequence of 100% cross-reactivity of TRIAC in the FT3 assay method. To reduce the extremely high circulating TSH levels and fetal goiter, the dose of TRIAC was increased to 3.5 mg/day. To monitor the possible intrauterine hypothyroidism, another cordocentesis was performed at 33 weeks gestation, showing that TSH levels were reduced by 50% (from 287 to 144 mU/L). Furthermore, a simultaneous ultrasound examination revealed a clear reduction in fetal goiter. After this latter cordocentesis, acute complications occured, prompting delivery by cesarean section. The female neonate was critically ill, with multiple-organ failure and respiratory distress syndrome. In addition, a small goiter and biochemical features ofhypothyroidism were noted transiently and probably related to the prematurity of the infant. At present, the baby is clinically euthyroid, without goiter, and only exhibits biochemical features of RTH. In summary, although further fetal studies in cases of RTH are necessary to determine whether elevated TSH levels with a markedly reduced bioactivity are a common finding, our data suggest transient biochemical hypothyroidism in RTH during fetal development. Furthermore, we advocate prenatal diagnosis of RTH and adequate treatment of the disease in case of maternal hyperthyroidism, to avoid fetal thyrotrope hyperplasia, reduce ...