Jasmine Parma

Familial congenital hypothyroidism due to inactivating mutation of the thyrotropin receptor causing profound hypoplasia of the thyroid gland.

Thyroid gland agenesis is the most common cause of congenital hypothyroidism and is usually sporadic. We investigated a brother and sister from consanguineous parents, ascertained through systematic newborn screening, and initially diagnosed with thyroid agenesis. Careful cervical ultrasonography in both patients revealed a very hypoplastic thyroid gland. By direct sequencing of the thyrotropin receptor gene, we identified the substitution of threonine in place of a highly conserved alanine at position 553, in the fourth predicted transmembrane domain. The mutation was found homozygous in the affected siblings, and heterozygous in both parents and two unaffected siblings. Functional analysis in transfected COS-7 cells showed that it resulted in extremely low expression at the cell surface as compared with the wild-type receptor, in spite of an apparently normal intracellular synthesis. The small amount of mutated receptor expressed at the surface of transfected cells bound thyrotropin with normal affinity and responded in terms of cAMP production, but the in vivo significance of these data from overexpressed receptor in transfected cells is unclear. Of note, blood thyroglobulin was unexpectedly elevated in the patients at the time of diagnosis, a finding that might prove useful in refining etiologies of congenital hypothyroidism.

Constitutive activation of the TSH receptor by spontaneous mutations affecting the N-terminal extracellular domain.

Activating mutations of the TSH receptor gene have been found in toxic adenomas and hereditary toxic thyroid hyperplasia. Up to now, all mutations have been located in the serpentine portion of the receptor. We now describe two additional mutations affecting Ser‐281 (Ser‐281‐Thr and Ser‐281‐Asn) in the ectodomain of the receptor. After transfection in COS cells, both mutants displayed increased constitutive activity for cAMP generation despite expression at a lower level than the wild type. The mutants were responsive to TSH. The present results are compatible with a model in which the activity of the unliganded receptor is kept at a low level by an inhibitory interaction between the N‐terminal domain and the serpentine portion of the receptor.

Nephrogenic Syndrome of Inappropriate Antidiuresis in Adults: High Phenotypic Variability in Men and Women from a Large Pedigree

Nephrogenic syndrome of inappropriate antidiuresis (NSIAD) is a recently described genetic cause of hyponatremia in male infants. Whether this X-linked condition could be detected in the adult or also could affect women is unknown. A large five-generation family was identified in which the recently described arginine-vasopressin receptor type 2 (AVPR2) mutation that is responsible for NSIAD was segregated. The proband was a 74-yr-old patient who had a syndrome of inappropriate antidiuresis and whose hyponatremia resisted administration of two AVPR2 antagonists. The phenotype of family members who carry the mutation was investigated. Patients with normal serum sodium were subjected to a water-load test. The previously reported activating missense R137C mutation in the AVPR2 gene in three hemizygous male and four heterozygous female individuals was identified. Except in one woman, spontaneous episodes of hyponatremia or abnormal water-load test were identified in all patients with the mutation, whether male or female. Skewed X inactivation was evidenced in the blood of the asymptomatic woman, which is compatible with preferential inactivation of her mutated allele. NSIAD is not limited to male infants. The diagnosis also should be considered in both male and female adults.

TSH receptor and disease

The TSH, LH/CG and FSH receptors belong to a subfamily of G protein-coupled receptors. As such, their primary structures deduced from the sequence of the corresponding cDNA (Parmentieret al., 1989; Nagayama et al., 1989; Libertet al., 1989; Minegishiet al., 1990; 1991), predict the existence of seven segments with hydropathy compatible with a transmembrane location. The glycoprotein hormone receptor subfamily (TSH, LH/CG, FSH) share characteristics that distinguish them from the other G protein-coupled receptors. They contain a signal peptide (20 amino acids for the TSH receptor) and they have a long extracellular aminoterminal domain (398 aminoacids for the TSH receptor) comprising the loose repetition of a motif of 25 residues rich in leucine (Parmentieret al., 1989; McFarlandet al., 1989). Similar leucine-rich motifs are also found in a number of widely different proteins (Roth, 1991), in which they are believed to confer the ability to interact with other proteins. The threedimensional structure of one such protein, the ribonuclease inhibitor, has been determined (Kobe & Deisenhofer, 1993), and provides a basis for the modelling of other leucine-rich protein segments. Site-directed mutagenesis studies involving chimeric receptors have clearly shown that the binding specificity and the effector properties of the glycoprotein hormone receptors are encoded in separate domains of the proteins (Xie et al., 1990; Braunet al., 1991; Nagayamaet al., 1991; Vassart & Dumont, 1992); the extracellular domain mediates the binding specificities and the ‘serpentine’ portion with the seven transmembrane segments displays the effector properties triggering G protein activation. This duality is reflected at the genomic level where a single exon encodes the serpentine portion of the receptors and many exons (nine for the TSH receptor) encode the extracellular domain (Gross et al., 1991). When aligned, the three glycoprotein hormone receptors show stronger conservation in the serpentine domain (approximately 70% similarity) than in the extracellular domain (approximately 40% similarity). A peculiarity of the TSH receptor with no counterpart in the FSH or LH/CG receptors is a 50-residue insert upstream from the hinge between the aminoterminal extracellular portion and the first transmembrane segment. A first model for the three-dimensional structure of the aminoterminal extracellular segment of the thyrotrophin receptor has been recently proposed (Kajavaet al., 1995) (Fig. 1). It is based on the known structure of the ribonuclease inhibitor (Kobe & Deisenhofer, 1993).

Isodisomy of chromosome 6 in a newborn with methylmalonic acidemia and agenesis of pancreatic beta cells causing diabetes mellitus.

Isodisomy (ID) is a genetic anomaly defined as the inheritance of two copies of the same genetic material from one parent. ID in an offspring is a rare cause of recessive genetic diseases via inheritance of two copies of a mutated gene from one carrier parent. We studied a newborn female with a mut(o) of methylmalonic acidemia and complete absence of insulin-producing beta cells in otherwise normal-appearing pancreatic islets, causing insulin-dependent diabetes mellitus. The patient died 2 wk after birth. Serotyping of the HLA antigens, DNA typing of HLA-B and HLA class II loci, study of polymorphic DNA markers of chromosome 6, and cytogenetic analysis demonstrated paternal ID, involving at least a 25-centiMorgan portion of the chromosome pair that encompasses the MHC. ID probably caused methylmalonic acidemia by duplication of a mutated allele of the corresponding gene on the chromosome 6 inherited from the father. It is also very likely that ID was etiologically related to the agenesis of beta cells and consequent insulin-dependent diabetes mellitus in our patient. We thus speculate on the existence of a gene on chromosome 6 involved in beta cell differentiation.

Relative contribution of Ki-ras gene analysis and brush cytology during ercp for the diagnosis of biliary and pancreatic diseases

BACKGROUND Ki-ras mutation analysis from material collected during ERCP has been claimed to improve the diagnosis of pancreatic and bile duct carcinomas as compared with conventional cytology. Our aim was to study the relative contribution of both Ki-ras analysis and brush cytology in patients with a significant stricture at ERCP. METHODS Brushings were collected in duplicate for both analyses in 142 patients in whom a definitive diagnosis was obtained by histology or a minimal follow-up of 6 months. RESULTS For pancreatic strictures, sensitivity, specificity, and accuracy of Ki-ras analysis vs. cytology in detecting malignancy were 81% vs. 66%, 72% vs. 100%, and 70% vs. 74%, respectively. For biliary strictures, they were 25% vs. 42%, 100% vs. 100%, and 35% vs. 43%, respectively. The combination of the two methods only marginally increased their sensitivity and accuracy in both types of strictures. CONCLUSION Ki-ras analysis is a sensitive method for diagnosing pancreatic but not biliary carcinoma. However, its specificity is lowered by a high frequency of Ki-ras mutations in patients with chronic pancreatitis (25%) who did not manifest cancer development within a 6-month follow-up period. In pancreatic duct strictures, brush cytology appears to be more specific in detecting malignancy; specificity for Ki-ras and cytology are equivalent for the diagnosis of malignant bile duct strictures. Therefore, making a clinical decision on the sole basis of Ki-ras analysis is probably not justified in the majority of the cases.

Lethal respiratory failure and mild primary hypothyroidism in a term girl with a de novo heterozygous mutation in the TITF1/NKX2.1 gene.

CONTEXT Thyroid transcription factor 1 (TITF1/NKX2.1) is expressed in the thyroid, lung, ventral forebrain, and pituitary. In the lung, TITF1/NKX2.1 activates the expression of genes critical for lung development and function. Titf/Nkx2.1(-/-) mice have pituitary and thyroid aplasia but also impairment of pulmonary branching. Humans with heterozygous TITF1/NKX2.1 mutations present with various combinations of primary hypothyroidism, respiratory distress, and neurological disorders. OBJECTIVE The objective of the study was to report clinical and molecular studies of the first patient with lethal neonatal respiratory distress from a novel heterozygous TITF1/NKX2.1 mutation. PARTICIPANT This girl, the first child of healthy nonconsanguineous French-Canadian parents, was born at 41 wk. Birth weight was 3,460 g and Apgar scores were normal. Soon after birth, she developed acute respiratory failure with pulmonary hypertension. At neonatal screening on the second day of life, TSH was 31 mU/liter (N <15) and total T(4) 245 nmol/liter (N = 120-350). Despite mechanical ventilation, thyroxine, surfactant, and pulmonary vasodilators, the patient died on the 40th day. RESULTS Histopathology revealed pulmonary tissue with low alveolar counts. The thyroid was normal. Sequencing of the patients lymphocyte DNA revealed a novel heterozygous TITF1/NKX2.1 mutation (I207F). This mutation was not found in either parent. In vitro, the mutant TITF-1 had reduced DNA binding and transactivation capacity. CONCLUSION This is the first reported case of a heterozygous TITF1/NKX2.1 mutation leading to neonatal death from respiratory failure. The association of severe unexplained respiratory distress in a term neonate with mild primary hypothyroidism is the clue that led to the diagnosis.

A Familial Case of Congenital Hypothyroidism Caused by a Homozygous Mutation of the Thyrotropin Receptor Gene

Most of the time congenital hypothyroidism appears as a sporadic disease. In addition to the rare defects in hormonosynthesis associated with goiters, the causes of congenital hypothyroidism include agenesis and ectopy of the thyroid gland. The study of some familial cases has allowed the identification of a few genes responsible for congenital hypothyroidism. We report here a familial case of congenital hypothyroidism, transmitted as a recessive trait, and caused by a homozygous mutation in the thyrotropin receptor (TSH-R). The initial diagnosis of thyroid agenesis, based on the absence of tracer uptake on scintiscan, was incorrect, because ultrasound examination identified severely hypoplastic thyroid tissue in the cervical region.

A quantitative study of peripheral blood stem cell contamination in diffuse large-cell non-Hodgkin's lymphoma: one-half of patients significantly mobilize malignant cells.

Autologous transplantation using peripheral blood stem cells (PBSCs) collected after chemotherapy, followed by growth factor administration (ASCT), is increasingly used in the treatment of non‐Hodgkins lymphoma (NHL). However, quantitative data regarding contaminating malignant cells in the harvests are still scarce. We prospectively investigated 37 diffuse large‐cell lymphomas (DLCLs) in complete remission (CR) that were treated according to multicentric protocols at our centre. DNA was extracted from the diagnostic lymph node. The complementarity‐determining region (CDR) III was sequenced and a patient‐specific oligomer synthesized. Contamination was evaluated semiquantitatively by polymerase chain reaction (PCR) and was confirmed by a limiting dilution analysis. PBSCs collected at regeneration after administration of granulocyte colony‐stimulating factor (G‐CSF), steady‐state bone marrow (BM) and peripheral blood samples at CR were compared. DNA was available in 37 patients, from which 22 rearrangements could be sequenced. Patients (n = 15) who had both the required follow‐up samples and a suitable clonal marker were investigated. In two cases, the patient‐specific PCR assay set up at diagnosis later gave false‐negative results in samples in which clonal DNA was still detectable by other sets of primers. PBSC contamination was highly variable: 7 out of 15 patients showed a PBSC/BM ratio of NHL cells greater than 1 log, whereas 8 out of 15 patients showed no difference and could vary from one apheresis to another. Eight ASCTs were performed, five of which used highly contaminated PBSCs: four patients relapsed early, three with disseminated lymphoma. Thus, 50% of DLCLs in CR seem to mobilize significantly malignant cells at regeneration under G‐CSF. Considering the higher numbers of cells reinfused, this translates into a much higher number of lymphoma cells reinfused when compared with autologous bone marrow transplantation (ABMT). However, their clonogenic potential remains unknown and, despite concerning observations in certain cases, it is still unclear whether this has an impact upon the outcome of ASCT.

Long-Term Follow-Up of an Infant with Thyrotoxicosis due to Germline Mutation of the TSH Receptor Gene (Met453Thr)

A 18-year clinical follow-up period in a male patient with a germline TSH-R gene mutation (Met453Thr) is described. Nonautoimmune thyrotoxicosis was diagnosed at the age of 7 months. The patient had exophthalmus, failure to thrive, advanced bone age and no goiter. Long-term antithyroid drug treatment (ATD) was necessary during childhood. At the age of 7 years he developed a goiter. Subtotal thyroidectomy was performed at the age of 9 years, followed by repeated ablative radiotherapy at the age of 9.5–13 years due to a toxic multinodular goiter. After 13 years ATD could be discontinued and the patient was euthyroid until 16 years of age, where L-thyroxine substitution had to be started. The exophthalmus diminished, and had disappeared at the age of 18 years, when CT scan of the orbit was performed. Conclusion: TSH-R mutation must be considered in early nonautoimmune thyrotoxicosis. A very agressive treatment strategy is necessary.