Mutation of BSND causes Bartter syndrome with sensorineural deafness and kidney failure.

Antenatal Bartter syndrome (aBS) comprises a heterogeneous group of autosomal recessive salt-losing nephropathies. Identification of three genes that code for renal transporters and channels as responsible for aBS has resulted in new insights into renal salt handling, diuretic action and blood-pressure regulation. A gene locus of a fourth variant of aBS called BSND, which in contrast to the other forms is associated with sensorineural deafness (SND) and renal failure, has been mapped to chromosome 1p. We report here the identification by positional cloning, in a region not covered by the human genome sequencing projects, of a new gene, BSND, as the cause of BSND. We examined ten families with BSND and detected seven different mutations in BSND that probably result in loss of function. In accordance with the phenotype, BSND is expressed in the thin limb and the thick ascending limb of the loop of Henle in the kidney and in the dark cells of the inner ear. The gene encodes a hitherto unknown protein with two putative transmembrane alpha-helices and thus might function as a regulator for ion-transport proteins involved in aBS, or else as a new transporter or channel itself.

Germline mosaicism in keratitis-ichthyosis-deafness syndrome: pre-natal diagnosis in a familial lethal form.

Keratitis-ichthyosis-deafness (KID) syndrome is an autosomal dominant congenital ectodermal defect characterized by the association of skin lesions, hearing loss and keratitis. Most of the cases appear to be sporadic. KID syndrome is mostly related to mutations of GJB2 gene encoding connexin-26. Recently, a lethal form of the disease during the first year of life has been reported in two unrelated Caucasian patients. This rare lethal form is caused by the G45E mutation of GJB2 gene. We here report the first pre-natal molecular genetic diagnosis of the lethal form of KID syndrome relating to a G45E mutation. In the same family, the occurrence of this condition in three other siblings born to African non-consanguineous healthy parents lead to perform pre-natal diagnosis for this last pregnancy. Molecular analysis confirms the diagnosis of the lethal form of KID for the fetus. These results establish the role of germline mosaicism in KID syndrome and warrant careful genetic counseling. Furthermore, analysis of our cases and the literature allowed us to define a characteristic severe neonatal phenotype including facial dysmorphy, severe cornification with massive focal hyperkeratosis of the skin with erythroderma, dystrophic nails, complete atrichia and absence of foreskin.

New surfactant protein C gene mutations associated with diffuse lung disease.

BACKGROUND: Mutations in the surfactant protein C gene (SFTPC) have been recently associated with the development of diffuse lung disease, particularly sporadic and familial interstitial lung disease (ILD). OBJECTIVE: We have investigated the prevalence and the spectrum of SFTPC mutations in a large cohort of infants and children with diffuse lung disease and suspected with surfactant dysfunction. METHOD AND RESULTS: 121 children were first screened for the common SFTPC mutation, p.Ile73Thr (I73T). Ten unrelated patients were shown to carry this mutation. The I73T mutation was inherited in six cases, and appeared de novo in four. The 111 patients without the I73T mutation were screened for the entire coding sequence of SFTPC. Of these, eight (seven unrelated) subjects were shown to carry a novel mutant allele of SFTPC. All these seven new mutations are located in the BRICHOS domain except the p.Val39Ala (V39A) mutation, which is in the surfactant protein C (SP-C) mature peptide. CONCLUSIONS: Our results confirm that SFTPC mutations are a frequent cause of diffuse lung disease, and that I73T is the most frequent SFTPC mutation associated with diffuse lung disease.

The very low penetrance of cystic fibrosis for the R117H mutation: a reappraisal for genetic counselling and newborn screening.

BACKGROUND: Cystic fibrosis (CF) is caused by compound heterozygosity or homozygosity of CF transmembrane conductance regulator gene (CFTR) mutations. Phenotypic variability associated with certain mutations makes genetic counselling difficult, notably for R117H, whose disease phenotype varies from asymptomatic to classical CF. The high frequency of R117H observed in CF newborn screening has also introduced diagnostic dilemmas. The aim of this study was to evaluate the disease penetrance for R117H in order to improve clinical practice. METHODS: The phenotypes in all individuals identified in France as compound heterozygous for R117H and F508del, the most frequent CF mutation, were described. The allelic prevalences of R117H (p(R117H)), on either intron 8 T5 or T7 background, and F508del (p(F508del)) were determined in the French population, to permit an evaluation of the penetrance of CF for the [R117H]+[F508del] genotype. RESULTS: Clinical details were documented for 184 [R117H]+[F508del] individuals, including 72 newborns. The disease phenotype was predominantly mild; one child had classical CF, and three adults' severe pulmonary symptoms. In 5245 healthy adults, p(F508del) was 1.06%, p(R117H;T7) 0.27% and p(R117H;T5)<0.01%. The theoretical number of [R117H;T7]+[F508del] individuals in the French population was estimated at 3650, whereas only 112 were known with CF related symptoms (3.1%). The penetrance of classical CF for [R117H;T7]+[F508del] was estimated at 0.03% and that of severe CF in adulthood at 0.06%. CONCLUSIONS: These results suggest that R117H should be withdrawn from CF mutation panels used for screening programmes. The real impact of so-called disease mutations should be assessed before including them in newborn or preconceptional carrier screening programmes.

Phenotype and genotype in females with POU3F4 mutations.

X-linked deafness is a rare cause of hereditary isolated hearing impairment estimated as at least 1% or 2% of the non-syndromic hearing loss. To date, four loci for DFN have been identified and only one gene, POU3F4 responsible for DFN3, has been cloned. In males, DFN3 is characterized by a progressive deafness associated with perilymphatic gusher at stapes surgery and with a characteristic inner ear malformation. The phenotype of eight independent females carrying POU3F4 anomalies is defined, and a late-onset hearing loss is found in three patients. Only one has an inner ear malformation. No genotype/phenotype correlation is identified.

[Sweat testing: review of technical requirements]. / Recommandations pour l'exécution et l'interprétation du test de la sueur.

The sweat test, a quantitative measurement of chloride in sweat, remains a key laboratory test to support the diagnosis of cystic fibrosis. However, because of its delicate execution, sweat test result should be interpreted with biological, clinical and genetic arguments. The following guidelines which we propose, were established in order to harmonize the practices of the sweat test. They are elaborated in a consensual way by biologists from cystic fibrosis reference centers and/or from the working group "Sweat Testing" of the National College of Biochemistry Hospital praticiens, according to the current state of knowledge on the subject, the experiment of the biologists and the recommendations established in the United States and in the United Kingdom.

[Lung diseases associated with inherited disorders of surfactant metabolism]. / Pathologies respiratoires associées à des anomalies héréditaires du métabolisme du surfactant.

Lung diseases associated with surfactant-metabolism disorders are a heterogeneous group of rare diseases. Intra-alveolar accumulation of protein related to surfactant dysfunction leads to cough, hypoxemia, and radiological-diffuse infiltration. Inherited deficiency of pulmonary surfactant protein B (SP-B) was initially described in infants who develop respiratory failure at birth. More recently, mutations in other constitutive surfactant proteins, such as surfactant protein C or implied in its metabolism, such as ATP-binding cassette, subfamily A, member 3 (ABCA3) and thyroid transcription factor 1 (TTF-1) were identified in newborns with respiratory distress as well as in children with chronic-infiltrative pneumonia. The aim of this review is therefore to summarize the current state of our knowledge in this area.

[The mutation 35delG of the gene of the connexin 26 is a frequent cause of autosomal-recessive non-syndromic hearing loss in Morocco]. / La mutation 35delG du gène de la connexine 26, une cause fréquente des surdités non syndromiques autosomiques récessives au Maroc.

UNLABELLED: Mutations of the connexin 26 gene, GJB2, are the most common cause of non syndromic autosomal-recessive hearing loss. One of the GJB2 mutations, the 35delG, is recurrent in European and Mediterranean populations with allelic frequency of at least 70% in patients with hearing loss caused by GJB2 impairment. OBJECTIVES: To determine the prevalence of the 35delG mutation in non-syndromic autosomal-recessive deafness in Morocco. PATIENTS AND METHODS: We looked for the 35delG mutation among 25 non-related Moroccan children suffering from an autosomal recessive hearing loss. A screening for GJB2 mutations, and then a search for GJB6 deletions were carried out among patients who do not bear the 35delG. RESULTS: Twelve patients were homozygous for the 35delG mutation. This mutation was responsible for almost half of the hearing loss among our patients (48%). There was no other GJB2 or GJB6 mutation among 13 patients. CONCLUSION: This study underlines the advantages of a systematic search for this mutation among deaf children when environmental causes are considered irrelevant. The identification of this genetic anomaly signs the etiologic diagnosis of deafness, which allows a relevant genetic advice, and a better treatment of patients.

Results of cochlear implantation in two children with mutations in the OTOF gene.

OBJECTIVE: The purpose of the study is to present the results of cochlear implantation in case of deafness involving mutations in the OTOF gene. This form of deafness is characterized by the presence of transient evoked otoacoustic emissions (TEOAE). In cases of profound deafness with preserved TEOAE, two main etiologies should be considered: either an auditory neuropathy (a retrocochlear lesion) or an endocochlear lesion. It is essential to differentiate these two entities with regards to therapy and screening. PATIENTS: We report two children who presented with profound prelingual deafness, confirmed by the absence of detectable responses to auditory evoked potentials (AEP), associated with the presence of bilateral TEOAE. Genetic testing revealed mutations in OTOF, confirming DFNB9 deafness. Both patients have been successfully implanted (with a follow-up of 18 and 36 months, respectively). MAIN OUTCOME MEASURES: Clinical (oral production, closed and open-set words and sentences list, meaningful auditory integration scale), audiometric evaluation (TEOAE, AEP) before and after implantation, and neural response telemetry (NRT). RESULTS: Both patients present a good quality of clinical responses and electrophysiological tests after implantation, indicating satisfactory functioning of the auditory nerve. This confirms the endocochlear origin of DFNB9 and suggests that these mutations in OTOF lead to functional alteration of inner hair cells. CONCLUSION: In the absence of a context of neurological syndrome, the combination of absent AEP and positive TEOAE should lead to a genetic screening for mutations in OTOF, in order to undertake the appropriate management.

Auditory neuropathy or endocochlear hearing loss?

AIMS: The purpose of the study was to define boundaries between endocochlear hearing loss and auditory neuropathy in children with congenital profound hearing loss and positive otoacoustic emissions. PATIENT: A child presented with bilateral profound hearing loss, which was confirmed by the absence of evoked auditory potentials at 110 dB and with conserved otoacoustic emissions. The lack of any relevant medical history, a normal neurologic pediatric examination, and the improvement obtained with powerful hearing aids suggested an endocochlear cause. Genetic testing identified mutations in OTOF, responsible for the DFNB9 recessive form of hearing loss. RESULTS: In recent years, cases of children with hearing loss associated with positive otoacoustic emissions have been labeled as "auditory neuropathy." Classically, this form of hearing loss is refractory to the use of hearing aids and cochlear implants. Mutations in OTOF lead to inner hair cells dysfunction, whereas the outer hair cells are initially functionally preserved. As this form of endocochlear hearing loss can be detected at a molecular level, genetic testing can be proposed for cases of nonsyndromic auditory neuropathy, as those children could benefit from cochlear implantation. CONCLUSION: It is advisable to reserve the term "auditory neuropathy" for patients who present hearing loss and conserved otoacoustic emissions in the context of a neurologic syndrome or for children with suggestive perinatal history. In other cases, genetic testing for mutations in OTOF should be carried out.

Genotype analysis and phenotypic manifestations of children with intermediate sweat chloride test results.

STUDY OBJECTIVES: Cystic fibrosis (CF) is one of the most common inherited diseases among whites. Since the cloning of the CF transmembrane conductance regulator (CFTR) gene, a number of studies have focused on associations between the genotype and phenotype in CF. This had led to the progressive identification of new groups of patients, including those who have mild lung disease and those who have normal sweat chloride values (< 60 mEq/L). The aim of the present work was to provide information on the genotype and the phenotypic characteristics of children with intermediate-range sweat chloride test results. PATIENTS AND RESULTS: We focused on children referred to the pulmonary department for various types of pulmonary disease and who had several sweat chloride test results with median values in the range of 40 to 60 mEq/L. Twenty-four patients over a 10-year period were enrolled (mean age, 4.8 years). Respiratory manifestations at initial evaluation included recurrent bronchitis, wheezing, chronic cough, and pneumonia. The duration of the follow-up ranged from 0.5 to 10.5 years. Sputum cultures revealed the presence of Haemophilus influenzae (10 children), Staphylococcus aureus (4 children), and Pseudomonas aeruginosa (3 children). Pancreatic insufficiency was found in two patients. Analysis of the entire coding sequence allowed identification of 16 known mutations in CFTR gene. Fifteen chromosomes (31.2%) carried a mutation in CFTR gene and one allele carried two mutations. Three patients were homozygous or double heterozygous (DeltaF508/DeltaF508, DeltaF508/3849 + 10 kb C-->T, S1235R/G551D). The 5-thymidine allele was identified in four children. CONCLUSION: These results indicate an higher frequency of CFTR gene mutations in patients with borderline sweat chloride test results, compared to data reported in the general population. They lead to the recommendations for complete pulmonary and GI investigations in this group of patients, as well as assiduous care and medical follow-up.

Development of a synovial cyst following anterior cruciate ligament reconstruction.

The use of bone-patellar tendon-bone allograft or autograft in anterior cruciate ligament (ACL) reconstruction is an accepted method of repair of the ACL-deficient knee with few complications. We report an unusual complication associated with this technique, the development of synovial fistula and cyst associated with the use of nonabsorbable suture material.

[Assay of total serum bile acids. Reference values in children]. / Dosage des acides biliaires totaux sériques. Valeurs de référence chez l'enfant.

Serum total bile acids were measured using an enzyme micromethod involving 3-hydroxysteroid dehydrogenase (Sterognost 3 Fluorometry) without prior extraction. The test specimen was 50 microliters of serum, with reading by spectrofluorometry. The degree of accuracy assessed at several concentration levels was acceptable for values greater than 5 mumol/l. The calibration function was linear between 0 and 80 mumol/l. Lower limit of detection was 0.88 mumol/l. Results given by this method were compared with those obtained using the Schwarz technique involving extraction of bile acids and their enzyme estimation. The correlation between the two methods was satisfactory. Absence of interference by bilirubin was confirmed. Total bile acids were measured in the serum of fasting children, aged from 1 day to 4 years. High values were seen at birth (m = 8.33 mumol/l, SD = 3.46, n = 10) as well as during the first month of life. Serum total bile acid concentration decreased up to the age of 10 months (m = 2.13 mumol/l, SD = 1.91, n = 31). After one year, they increased progressively, reaching at the age of 4 values similar to those in the adult (m = 3.80 mumol/l, SD = 2.17, n = 30). Particularly high values were found in children with delayed growth or in premature infants (m = 19.66 mumol/l, SD = 10.4, n = 15). Serum total bile acid levels were lower in umbilical cord blood and in the newborn. The increase in serum bile acids found in the newborn may be due to slight cholestasis or to a reduction in the hepatic clearance, showing the special type of hepatic function in the young child.

[Optimization of the determination in urine of alanine aminopeptidase, gamma-glutamyltransferase and N acetyl-beta-D-glucosaminidase]. / Optimisation de la mesure dans l'urine de l'alanine aminopeptidase, de la gamma-glutamyltransférase et de la N-acétyl-bêta-D-glucosaminidase.

The authors describe the optimization of determination of alanine aminopeptidase (AAP), gamma-glutamyltransferase (GGT) and N-acetyl-beta-D-glucosaminidase (NAG) in urine by multivariate analysis. The optimal conditions found are: for AAP at 30 degrees C TRIS HCl buffer 300 mmol/l pH 7.9, L-alanine-4-nitroanilide 5.8 mmol/l, for GGT at 30 degrees C buffer glycylglycine 150 mmol/l pH 8.0, gamma-L-glutamyl-3-carboxy-4-nitroanilide 9.0 mmol/l, for NAG at 37 degrees C citrate buffer 50 mmol/l pH 5.8, m cresolsulfonphtaleinyl-N-acetyl-beta-D-glucosaminide 5.5 mmol/l. These methods are easy to perform, apply to urine without pretreatment through Sephadex: therefore complete automatization is possible. The stability of enzymatic activities in urine is of ten days at +4 degrees C in the presence of sodium azide at neutral pH. Freezing resulted in a considerable loss of activity for AAP and GGT.

[Genetic testing for cystic fibrosis: evaluation of the Elucigene CF20 kit in blood and buccal cells]. / Diagnostic moléculaire de la mucoviscidose: evaluation de la trousse Elucigene CF20 dans le sang et les cellules buccales.

Routine determination of mutations in cystic fibrosis requires accurate, rapid, reliable and low-cost methods, permitting the simultaneous detection of multiple mutations. The Elucigene CF20 kit developped by Cellmark Diagnostics, uses multiplex ARMS, which allows the screening for 20 CFTR gene mutations (deltaF508, G542X, N1303K, 1717-1G>A, G551D, W1282X, R553X, deltaI507, 1078delT, 2183AA>G, 3849+10kbC>T, R1162X, 621+1G>T, R334W, R347P, 3659delC, R117H, S1251N, E60X, A455E ) in a work day without specific instrumentation. The kit distinguishes between homozygotes and heterozygotes for deltaF508, but not for rare mutations. The kit detects from 68 to 92% of defective alleles in Caucasians. We evaluate the kit in a blind study in two independent laboratories. Thirty blood samples and thirty mouthwash samples from CF patients, carriers and unaffected individuals were analysed by the Elucigene CF20 kit. All the samples were previously analysed by denaturing gradient gel electrophoresis and sequencing. The Elucigene CF20 kit consists of three multiplexes. Each mutiplex contains ARMS specific primers for six to eight mutations and two control reactions. The absence of the upper control fragment indicates that a repeat test is required. We demonstrated a first time amplification rate of 98.3%: of the 60 samples tested, one required a reamplification. Results compared with the reference method demonstrated that in all cases where one or more of the 20 mutations detected by the kit were present in the test set, the kit accurately identified them. Reproducibility was assessed by repeating the analysis of a blood and mouthwash sample five times. Cross reactivity between R117C and R117H, R117P and R117H, R347P and R347H, deltaI507 and deltaF508, G551D and R553X were evaluated. Only a cross reactivity between R347P and R347H was observed. The kit is specially useful for first line study of patients and carrier identification.

[Measurement of total bile acids. Comparison of different preparations of 3 alpha-hydroxysteroid dehydrogenase]. / Dosage des acides biliaires totaux. Comparaison de différentes préparations de 3 alpha-hydroxystéroïde déshydrogénase.

Oxidation of free and conjugated biliary acids was carried out with 5 different enzymatic preparations of 3 alpha-hydroxysteroid dehydrogenase. The comparison of the catalytic activities obtained, shows the existence of important variations according to the enzymes. Preparations purified by chromatography have a stronger activity toward primary biliary acids than toward secondary biliary acids, while preparations obtained from bacterial mutants have a low activity toward cholic acid and its conjugates.

[Evaluation of enzymuria as a tracer in nephrotoxicity: results of a multicenter study]. / Evaluation de l'enzymurie comme marqueur de néphrotoxicité: résultats d'une expérience multicentrique.

Urinary excretion of three enzymes of different subcellular location in kidney tissue, alanine aminopeptidase (AAP), gammaglutamyltransferase (GGT), N acetyl-beta-D glucosaminidase (NAG), was carried out in 79 healthy adults and 108 healthy children and in 69 adults with various therapies: antibiotics (32 cases), non steroidal anti-inflammatory drugs (NSAIDs) (22 cases), cisplatinum (12 cases) and cyclosporine (3 cases). A circadian rhythm has been shown in children. In patients treated with antibiotics, the importance and duration of the increased enzymes urinary excretion were variable but the excretion of AAP was always higher than that of GGT and NAG. Short term therapies by NSAIDs were without influence on enzymuria but long term therapies produced a moderate increase of NAG excretion. Enzymuria increased immediately after cisplatinum administration and decreased after each daily dose, except in patients with previously high creatininemia. Cyclosporine induced a slight increase in AAP and NAG excretion. Enzymuria, thus, increased early reflecting a toxic effect of the drug at the cellular level whereas creatininemia increase, marker of renal fonctionnal insufficiency, occurs only occasionally and lately.

[Primary molecular changes and secondary biological problems in Bartter and Gitelman syndrome]. / Altérations moléculaires primaires et troubles biologiques secondaires des syndromes de Bartter et de Gitelman.

Bartter syndrome and Gitelman syndrome are primary hereditary diseases characterized by hypokaliemia, alkalosis, hypertrophy of the juxtaglomerular complex with secondary hyperaldoteronism and normal blood pressure. They result from molecular disorders leading to a defect of sodium reabsorption in respectively the Henle's loop and the distal convoluted tubule. Biological adaptations of downstream tubular segments, i.e. distal convoluted tubule and collecting duct, are responsible for hypokaliemia, alkalosis, renin-aldosterone activation, prostaglandins hypersecretion and dysregulation of the urinary excretion of calcium and magnesium, illustrating the close integration of the regulation of different solutes in the distal tubular structures.

[Constitutional deficiency of pulmonary surfactant protein B: clinical presentation, histologic and molecular diagnosis]. / Déficit constitutionnel en protéine B du surfactant pulmonaire: présentation clinique, diagnostic histologique et moléculaire.

We report a female full-term infant with fatal respiratory failure of early onset due to inherited SP-B deficiency. Lung biopsy was performed at 18 days after birth, with histopathological characterization indicating congenital alveolar proteinosis. Immunohistochemical studies of lung tissue revealed the absence of SP-B and the presence of intra-alveolar SP-A normal quantities. Analysis of genomic DNA showed homozygosity for the 121ins2 mutation of the SFTPB gene. The infant died 21 days after birth. Both parents were heterozygotes for the mutation. Chorionic villus sampling was performed at the first trimester of the following pregnancy. Restriction analysis of amplified fetal DNA, studies of microsatellite segregation and direct sequencing led to the diagnosis of homozygosity for the parental wild-type allele. The diagnosis of congenital SP-B deficiency should be suspected whenever an early and acute respiratory failure in a term or near-term infant does not resolve after five days of age: diagnostic confirmation can be easily and rapidly obtained with the analysis of genomic DNA and immunohistochemical characterization of lung tissue.

[Dental caries (DMFT) in adults in Switzerland 1988]. / Zahnkaries (DMFT) bei Erwachsenen in der Schweiz 1988.

In 1988, 864 adults were examined for dental and periodontal conditions in 47 dental offices in Switzerland according to WHO methods. Among the 57 offices selected at random 10 dentists were unwilling to cooperate. Within each office, 36 patients were chosen by a randomizing mechanism. 51 percent of the selected patients presented themselves for the examination whereas 14 percent refused to participate in the study; of the remaining subjects one third could not be reached during the three to five days available for contacting them and two thirds were unable to come to the respective dental office at the preset day of examination. The DMF data obtained were similar to the results of previous local surveys, part of which were based on random samples. Up to age 74, it was concluded that the data of the present survey are fairly representative of the Swiss resident population. In the age group 75 and above, the number of missing teeth (MT) was too low, mainly due to edentulous persons, institutionalized and other, being obviously underrepresented. The Swiss DMFT of 22.3 in the age group 35-44 was almost the highest mean among 26 European countries. However, prevention efforts at school age, already obvious in the age group 30-34, will substantially improve the results at age 35-44 by the year 1998. The number of DT was 1.0 and 1.3 for the ages up to 29 and below 1.0 in the older groups. Up to age 54, FT was the most important component of the DMFT while in the older groups, MT was highest.