Genetic testing facilitates prepubertal diagnosis of congenital hypogonadotropic hypogonadism.

Neonatal micropenis and cryptorchidism raise the suspicion of congenital hypogonadotropic hypogonadism (CHH), a rare genetic disorder caused by gonadotropin-releasing hormone deficiency. Low plasma testosterone levels and low gonadotropins during minipuberty provide a clinical diagnostic clue, yet these tests are seldomly performed in general practice. We report a male neonate with no family history of reproductive disorders who was born with micropenis and cryptorchidism. Hormonal testing at age 2.5 months showed low testosterone (0.3 nmol/L) and undetectable gonadotropins (luteinizing hormone and follicle-stimulating hormone both <0.5 U/L), suggestive of CHH. Genetic testing identified a de novo, heterozygous mutation in fibroblast growth factor receptor 1 (FGFR1 p.L630P). L630 resides on the ATP binding cleft of the FGFR1 tyrosine kinase domain, and L630P is predicted to cause a complete loss of receptor function. Cell-based assays confirmed that L630P abolishes FGF8 signaling activity. Identification of a loss-of-function de novo FGFR1 mutation in this patient confirms the diagnosis of CHH, allowing for a timely hormonal treatment to induce pubertal development. Therefore, genetic testing can complement clinical and hormonal assessment for a timely diagnosis of CHH in childhood.

Clinical, genetic and functional characteristics of three novel CYP17A1 mutations causing combined 17alpha-hydroxylase/17,20-lyase deficiency.

BACKGROUND: P450c17 has two distinct activities: 17alpha-hydroxylase activity and 17,20-lyase activity. Combined 17alpha-hydroxylase/17,20-lyase deficiency leads to a severe defect in the production of cortisol and sex steroids. In affected males this results in impaired masculinization with ambiguous or female external genitalia. Female patients have normal genitalia but show a lack of pubertal development in adolescence. An increased production of mineralocorticoids often leads to hypertension and hypokalemia in both sexes. METHODS: To better understand the mechanisms of P450c17 deficiency, we studied 2 patients (both 46,XY) with combined 17alpha-hydroxylase/17,20-lyase deficiency of different severity: one with complete lack of masculinization and one with ambiguous genitalia. RESULTS: Four mutations were identified by sequencing of the CYP17A1 gene: I332T and A355T in the less severely affected patient; G111S and R440H in the patient with complete lack of masculinization. The three novel mutations were expressed in COS1 cells and all mutant proteins except I332T showed a complete loss of both enzymatic activities. I332T retained some residual 17alpha-hydroxylase as well as 17,20-lyase activity. CONCLUSION: We identified 2 patients with the phenotypical spectrum of P450c17 deficiency. Three novel mutations in the CYP17A1 gene were identified and their functional characterization provided a good phenotype-genotype correlation. The location of the mutated residues in the three-dimensional model of P450c17 gave some additional insights into its structure-function relationship.

P450c17 deficiency: clinical and molecular characterization of six patients.

CONTEXT: The characteristics of P450c17 deficiency include 46,XY disorder of sex development, hypertension, hypokalemia, and lack of pubertal development. OBJECTIVE: To better understand this rare enzymatic deficiency, we analyzed the CYP17A1 gene in six affected patients. DESIGN AND PATIENTS: We examined six patients, five 46,XY, and one 46,XX (age 9-29 yr) with complete lack of masculinization (female infantile external genitalia, no uterus) and delayed puberty, respectively, and different degrees of hypertension. MAIN OUTCOME MEASUREMENTS: Genotype-phenotype correlation was measured. RESULTS: Four homozygote mutations were identified by direct sequencing of the CYP17A1 gene corresponding to an alanin 302-proline (A302P) exchange; the loss of lysine 327 (K327del); the deletion of glutamate 331 (E331del); and the replacement of arginine 416 with a histidine (R416H). Both P450c17 activities were abolished in all the mutant proteins, except one, when expressed in COS1 cells. The E331del-mutated P450c17 retained 17alpha-hydroxylase activity. The mutant proteins were normally expressed, suggesting that the loss of enzymatic activity is not due to defects of synthesis, stability, or localization of P450c17 proteins. CONCLUSION: These studies confirm lack of masculinization in 46,XY individuals as the pathognomic sign of the complete P450c17 deficiency. In XX individuals P450c17 deficiency should be considered in cases of delayed puberty. Age of onset and the severity of hypertension do not seem to be constant. Careful examination of long-term follow-ups in two of our patients suggested to us that estrogen treatment in P450c17-deficient patients might worsen the enzymatic defect, leading to aggravation of the hypertension.

Association of childhood type 1 diabetes mellitus with a variant of PAX4: possible link to beta cell regenerative capacity.

AIMS/HYPOTHESIS: Loss of pancreatic beta cells is the crucial event in the development of type 1 diabetes. It is the result of an imbalance between autoimmune destruction and insufficient regeneration of islet cells. To study the role of islet cell regeneration in the pathogenesis of type 1 diabetes, we focused on PAX4, a paired homeodomain transcriptional repressor that is involved in islet cell growth. METHODS: The study included 379 diabetic children and 1,070 controls from two distinct populations, and a cohort of children who had not developed type 1 diabetes, despite the presence of islet cell antibodies. Genomic DNA analysis of PAX4 was carried out via direct sequencing of PCR-amplified fragments and allelic discrimination. We compared the transrepression potential of the PAX4 variants in betaTC3 cells and analysed their influence on beta cell growth. RESULTS: The type 1 diabetic subjects are different from the normal individuals in terms of the genotype distribution of the A1168C single nucleotide polymorphism in PAX4. The C/C genotype is frequent among type 1 diabetic children (73%) and rare among the control population (32%). Conversely, the A/C genotype is prevalent among control subjects (62%) and antibody-positive children without type 1 diabetes (73.6%), but uncommon among subjects with type 1 diabetes (17.5%). The combination of PAX4A and PAX4C is functionally more active than PAX4C alone (the "diabetic" variant). Beta cells expressing PAX4A and PAX4C efficiently proliferate when stimulated with glucose, whereas cells expressing the PAX4C variant alone do not. CONCLUSIONS/INTERPRETATION: We have identified a link between beta cell regenerative capacity and susceptibility to type 1 diabetes. This finding could explain the fact that not all of the individuals who develop autoimmunity against beta cells actually contract the disease. The C/C genotype of the A1168C polymorphism in PAX4 can be viewed as a predisposition marker that can help to detect individuals prone to develop type 1 diabetes.

Incidence of enteropathy-associated T-cell lymphoma in celiac disease: implications for children and adolescents with type 1 diabetes.

The long-term consequences of screening for celiac disease in diabetic children are not known. Routine screening is not practiced in our pediatric diabetic population. This study of the incidence of the most severe and specific long-term complication of untreated celiac disease, i.e., enteropathy-associated T-cell lymphoma (EATCL) and its association with diabetes, is done in order to justify our strategy not to practice routine screening. In the first phase of this study, a questionnaire was sent to all Swiss pathologists. The second phase consisted of a search in the cancer registry of the canton of Zurich. The incidence of EATCL in the general population of a Swiss region and the theoretical risk for a diabetic patient to develop this type of lymphoma were calculated. Ten cases of EATCL were found. Five had a long history of malabsorption, three of them since childhood. The mean age of the patients was 61.9 yr. None suffered from diabetes mellitus. The incidence of EATCL was 0.07/100,000 inhabitants/year. The expected risk for EATCL in patients with type 1 diabetes is 12.4/100,000 diabetic patients over a period of 60 yr. The data suggest that the risk for EATCL is small in diabetic patients. Therefore, we restrict the investigation for celiac disease to patients with typical and atypical symptoms, but do not perform routine screening.

Epidemiology of type I diabetes mellitus in Switzerland: steep rise in incidence in under 5 year old children in the past decade.

AIMS/HYPOTHESIS: In this nationwide prospective study we wanted to verify the trend of increasing diabetes incidence data from our earlier retrospective analysis of the military registry of Swiss men. SUBJECTS AND METHODS: The data collection of newly diagnosed children in Switzerland at an age younger than 15 years started in 1991. The countrywide survey used a small questionnaire which was sent back to the study centre. The questionnaire was anonymous and contained: hospital of diagnosis, initials, sex, birth date, date of diagnosis, residence, country of citizenship, and responsible physician. General data on the population were taken from publications of the Swiss Federal Statistical Office. RESULTS: A total of 941 children below the age of 15 years with newly diagnosed Type I (insulin-dependent) diabetes mellitus were collected (434 girls, 507 boys). The incidence in children aged 0 to 14 years rose significantly between 1991 and 1999 with a yearly average increase of 5.1%. In the age group 0 to 4 years a more than four-fold increase in incidence from 2.4/100,000 per year to 10.5/100,000 per year (p = 0.0002) was recorded, whereas the age-specific incidence in the 5 to 9-year-old and 10 to 14-year-old children did not change during the data collection period. The incidence was significantly higher in boys than in girls, whereas no difference was found between rural and urban populations. CONCLUSION/INTERPRETATION: The incidence of Type I diabetes is rising in children living in Switzerland but only the youngest age group of under 5 years of age is affected showing a large annual average increase of 23.8%.

A novel mutation in the anti-müllerian hormone gene as cause of persistent müllerian duct syndrome.

UNLABELLED: Persistent müllerian duct syndrome is a relatively rare inherited defect of sexual differentiation characterised by failure of regression of the müllerian ducts in males. In affected individuals, uterus and tubes are present because of defects of synthesis or action of anti-müllerian hormone (AMH), normally produced by the Sertoli cells of the testis. Patients are normally virilised, although mono- or bilateral cryptorchidism may be present. We observed two brothers (chromosomes 46 XY), aged 11 years and 2 months and 8 years and 3 months respectively, with bilateral cryptorchidism. The diagnosis of persistent müllerian duct syndrome was made on the basis of laparoscopic evidence of uterus and tubes, undetectable plasma levels of AMH and a 23 base pair duplicative insertion in exon 5 of the AMH gene, causing the introduction of a premature stop codon, homozygous in the two brothers. The surgical correction of the genital abnormalities was successfully carried out by laparoscopic orchidopexy according to Fowler-Stephens. CONCLUSION: Persistent müllerian duct syndrome should be taken into consideration in all cases of bilateral cryptorchidism. Laparoscopy is the elective procedure for diagnosis of this disease and laparoscopic surgery for orchidopexy of intra-abdominal testes. Mutation analysis of the anti-müllerian hormone gene in these patients helps to understand the structure-function relationship of the anti-müllerian hormone protein, although it is not clear at present whether anti-müllerian hormone is necessary to maintain normal testicular function.

Epidermolysis bullosa junctionalis progressiva in three siblings.

Three siblings of Swiss origin with epidermolysis bullosa junctionalis progressiva are described. The following clinical features were present from school age: dystrophy of the nails, non-scarring blistering of the skin, mild skin atrophy, hypodontia and dental caries. Light microscopy showed subepidermal blistering. Direct immunofluorescence was negative. On indirect immunofluorescence staining of a fresh spontaneous blister, bullous pemphigoid antigen and laminin were localized to the blister roof, and collagen IV and collagen VII to the blister base, indicating junctional splitting. Electron microscopy revealed a normal dermo-epidermal junction zone, including normal hemidesmosomes. There were no deposits of electron-dense amorphous material.

A physiological mode of puberty induction in hypogonadal girls by low dose transdermal 17 beta-oestradiol.

Transdermal 17 beta-oestradiol administration (17 beta-E2), used mainly in menopausal women, allows a continuous 17 beta-E2 delivery through the skin into the systemic circulation, avoiding intestinal and hepatic passage. In order to explore whether transdermal 17 beta-E2 could be used for the induction of puberty, 17 beta-E2 patches with low dose delivery were administered in nine prepubertal girls with Turner syndrome (bone age greater than 10.5 years) for a mean period of 2.2 years. Treatment schedule: 5 micrograms/day for 6-9 months, 10 micrograms/day for 6-9 months, 25 micrograms/day for long-term substitution; addition of cyclic gestagen p.o. after 18-24 months. Breast development started within 3 months of therapy and menstruation occurred after 2 years. Growth rate increased from 3.2 to 5.0 cm/year during the 1st year of therapy, height prediction did not change. Serum oestradiol (E2) and urinary E2 conjugates increased proportionally with 17 beta-E2 doses, serum oestrone (E1) rose much less. The possibility to imitate time course, clinical events and hormonal changes of normal puberty, the absence of adverse drug reactions and the excellent acceptance and easy mode of application suggest that transdermal 17 beta-E2 is optimally suited for hormonal substitution in girls with hypogonadism.