A recognizable type of syndromic short stature with arthrogryposis caused by bi-allelic SEMA3A loss-of-function variants.

The semaphorins constitute a large family of secreted and membrane-associated proteins that regulate many developmental processes, including neural circuit assembly, bone formation and angiogenesis. Recently, bi-allelic loss-of-function variants in SEMA3A (semaphorin 3A) were identified in a single patient with a particular pattern of multiple congenital anomalies (MCA). Using homozygosity mapping combined with exome sequencing, we identified a homozygous SEMA3A variant causing a premature stop codon in an 8 year old boy with the same pattern of MCA. The phenotype of these patients is characterized by postnatal short stature, skeletal anomalies of the thorax, a minor congenital heart or vascular defect, camptodactyly, micropenis, and variable additional anomalies. Motor development is delayed in both patients, and intellectual development is delayed in one patient. Our observation of a second case supports the notion that bi-allelic mutations in SEMA3A cause an autosomal recessive type of syndromic short stature.

Expanding CEP290 mutational spectrum in ciliopathies.

Ciliopathies are an expanding group of rare conditions characterized by multiorgan involvement, that are caused by mutations in genes encoding for proteins of the primary cilium or its apparatus. Among these genes, CEP290 bears an intriguing allelic spectrum, being commonly mutated in Joubert syndrome and related disorders (JSRD), Meckel syndrome (MKS), Senior-Loken syndrome and isolated Leber congenital amaurosis (LCA). Although these conditions are recessively inherited, in a subset of patients only one CEP290 mutation could be detected. To assess whether genomic rearrangements involving the CEP290 gene could represent a possible mutational mechanism in these cases, exon dosage analysis on genomic DNA was performed in two groups of CEP290 heterozygous patients, including five JSRD/MKS cases and four LCA, respectively. In one JSRD patient, we identified a large heterozygous deletion encompassing CEP290 C-terminus that resulted in marked reduction of mRNA expression. No copy number alterations were identified in the remaining probands. The present work expands the CEP290 genotypic spectrum to include multiexon deletions. Although this mechanism does not appear to be frequent, screening for genomic rearrangements should be considered in patients in whom a single CEP290 mutated allele was identified.

Clinical spectrum of SIX3-associated mutations in holoprosencephaly: correlation between genotype, phenotype and function.

BACKGROUND: Holoprosencephaly (HPE) is the most common structural malformation of the human forebrain. There are several important HPE mutational target genes, including the transcription factor SIX3, which encodes an early regulator of Shh, Wnt, Bmp and Nodal signalling expressed in the developing forebrain and eyes of all vertebrates. OBJECTIVE: To characterise genetic and clinical findings in patients with SIX3 mutations. METHODS: Patients with HPE and their family members were tested for mutations in HPE-associated genes and the genetic and clinical findings, including those for additional cases found in the literature, were analysed. The results were correlated with a mutation-specific functional assay in zebrafish. RESULTS: In a cohort of patients (n = 800) with HPE, SIX3 mutations were found in 4.7% of probands and additional cases were found through testing of relatives. In total, 138 cases of HPE were identified, 59 of whom had not previously been clinically presented. Mutations in SIX3 result in more severe HPE than in other cases of non-chromosomal, non-syndromic HPE. An over-representation of severe HPE was found in patients whose mutations confer greater loss of function, as measured by the functional zebrafish assay. The gender ratio in this combined set of patients was 1.5:1 (F:M) and maternal inheritance was almost twice as common as paternal. About 14% of SIX3 mutations in probands occur de novo. There is a wide intrafamilial clinical range of features and classical penetrance is estimated to be at least 62%. CONCLUSIONS: Our data suggest that SIX3 mutations result in relatively severe HPE and that there is a genotype-phenotype correlation, as shown by functional studies using animal models.

Two novel mutations in the GDAP1 and PRX genes in early onset Charcot-Marie-Tooth syndrome.

Autosomal recessive Charcot-Marie-Tooth syndrome (AR-CMT) is often characterised by an infantile disease onset and a severe phenotype. Mutations in the ganglioside-induced differentiation-associated protein 1 (GDAP1) gene are thought to be a common cause of AR-CMT. Mutations in the periaxin (PRX) gene are rare. They are associated with severe demyelination of the peripheral nerves and sometimes lead to prominent sensory disturbances. To evaluate the frequency of GDAP1 and PRX mutations in early onset CMT, we examined seven AR-CMT families and 12 sporadic CMT patients, all presenting with progressive distal muscle weakness and wasting. In one family also prominent sensory abnormalities and sensory ataxia were apparent from early childhood. In three families we detected four GDAP1 mutations (L58LfsX4, R191X, L239F and P153L), one of which is novel and is predicted to cause a loss of protein function. In one additional family with prominent sensory abnormalities a novel homozygous PRX mutation was found (A700PfsX17). No mutations were identified in 12 sporadic cases. This study suggests that mutations in the GDAP1 gene are a common cause of early-onset AR-CMT. In patients with early-onset demyelinating AR-CMT and severe sensory loss PRX is one of the genes to be tested.

[Van-der-Woude Syndrome]. / Van-der-Woude-Syndrom.

We report on two families with different expression of a Van-der-Woude-Syndrome (VWS) and with proven mutation of the IRF6- gene. The Van-der-Woude syndrome is a rare disease, typically consisting of congenital pits of the lower lip in combination with cleft lip or cleft palate or both. The Van-der-Woude syndrome is an autosomal dominant syndrome with variable expression. The penetrance is between 0,89 and 0,99. It is important to establish the correct diagnosis by careful investigation of patients with cleft lip or cleft palate and their parents. Genetic counselling is recommended in such cases.

Histopathological and neuroradiological features of Usher syndrome type II.

OBJECTIVE: To study the histopathological and neuroradiological features of Usher syndrome (USH), with particular focus on USH type II, an inherited disorder characterized by moderate to severe congenital hearing impairment and retinitis pigmentosa with onset in the late teens. METHODOLOGY: A report of four cases and literature review. RESULTS AND CONCLUSION: Rare examples of histopathological and neuroradiological findings from four USH type II cases are presented. More studies like these are encouraged so that correlation studies between the morphological and clinical findings can be performed on the path to elucidate the pathogenesis of this heterogeneous disorder.

Novel C59T leader peptide mutation in the MPZ gene associated with late-onset, axonal, sensorimotor polyneuropathy.

The objective of this study was to report a novel exon-1 mutation in the myelin protein zero (MPZ) gene, resulting in axonal Charcot-Marie-Tooth neuropathy with recurrent hyper-CK-emia. In a 64-year-old woman slowly progressive distal lower limb weakness, muscle cramps in the lower limb muscles, and stocking-type numbness had developed from the age of 61. Neurologic examination revealed discrete hip flexor weakness, weakness for foot extension, diffuse wasting of the distal lower limb muscles, reduced patella tendon reflexes, and absent Achilles tendon reflexes. There was recurrently elevated creatine kinase with a maximum of 607 U/l (n, <145 U/l). Stimulation of the peroneal and tibial nerves did not evoke a muscular response. Electromyography was neurogenic. Biopsy of the right sural nerve showed diffuse axonal degeneration and loss of axons of all diameters. Muscle biopsy showed increased fiber-size variability, angulated fibers, internalized nuclei, accumulations of nuclei, grouped atrophic muscle fibers, and fiber splitting. Molecular genetic analysis by PCR and direct nucleotide sequencing revealed the heterozygous C59T exon-1 MPZ gene mutation, resulting in the amino acid exchange S20F of the MPZ signal protein domain (leader peptide). The novel C59T mutation in the leader peptide of the MPZ gene is pathogenic and manifests as severe, late-onset, axonal, symmetric sensorimotor polyneuropathy (CMT2) and hyper-CK-emia.

Novel rhodopsin mutations and genotype-phenotype correlation in patients with autosomal dominant retinitis pigmentosa.

AIM: To identify novel or rare rhodopsin gene mutations in patients with autosomal dominant retinitis pigmentosa and description of their clinical phenotype. METHODS: The complete rhodopsin gene was screened for mutations by DNA sequencing in index patients. Mutation specific assays were used for segregation analysis and screening for controls. Eight patients from five families and their relatives were diagnosed with autosomal dominant retinitis pigmentosa (adRP) by means of clinical evaluation. RESULTS: Mutation screening identified five different rhodopsin mutations including three novel mutations: Ser176Phe, Arg314fs16, and Val20Gly and two missense mutations, Pro215Leu and Thr289Pro, that were only reported once in a mutation report. Electrophysiological and psychophysical testings provide evidence of an impaired rod system with additionally affected cone system in subjects from each genotype group. Visual function tended to be less affected in subjects with the Arg314fs16 and Val20Gly mutations than in the Ser176Phe phenotype. In contrast, Pro215Leu and Thr289Pro mutations caused a remarkably severe phenotype. CONCLUSION: The ophthalmic findings support a correlation between disease expression and structural alteration: (1) extracellular/intradiscal Val20Gly and cytoplasmic Arg314fs16 mutation-mild adRP phenotype; (2) Ser176Phe mutation-"mostly type 1" disease; (3) predicted alteration of transmembrane domains TM V and TM VII induced by Pro215Leu and Thr289Pro-severe phenotype. However, variation of phenotype expression in identical genotypes may still be a typical feature of RHO mutations.

Ocular manifestations of keratitis-ichthyosis-deafness (KID) syndrome.

OBJECTIVE: Keratitis-ichthyosis-deafness (KID) syndrome is a rare congenital ectodermal dysplasia characterized by the association of hyperkeratotic skin lesions, moderate to profound sensorineural hearing loss and vascularizing keratitis. Mutations in the GJB2 gene coding for connexin 26, a component of gap junctions in epithelial cells, have been observed in several KID patients. Variable ocular manifestations of the disease in 3 patients with molecular genetically confirmed KID syndrome are reported. DESIGN: Retrospective case series. METHODS: Clinical examination and molecular genetic analysis for mutations in the GJB2 gene were performed in 3 patients with KID syndrome ages 5, 13, and 41 years. RESULTS: Visual acuity ranged from normal to severe visual loss. The ocular signs included loss of eyebrows and lashes, thickened and keratinized lids, trichiasis, recurrent corneal epithelial defects, superficial and deep corneal stromal vascularization with scarring, keratoconjunctivitis sicca, and, in one patient, presumed limbal insufficiency. Whereas ocular surface integrity could be maintained with artificial tears in one patient, and an epithelial defect healed under conservative treatment in the second patient, multiple surgical procedures including superficial keratectomies, limbal allograft transplantation with systemic immunosuppression, amniotic membrane transplantation, lateral tarsorrhaphies, and lamellar keratoplasty could not preserve useful vision in the third patient. CONCLUSIONS: KID syndrome may affect the ocular adnexae and surface with variable severity independent of the age of the patient. Lid abnormalities, corneal surface instability, limbal stem cell deficiency with resulting corneal complications, and dry eye are the main ocular manifestations.

Mutation spectrum of type I glycogen storage disease in Hungary.

We performed mutation analysis in 12 Hungarian type I glycogen storage disease (GSD I) patients in order to determine the mutation spectrum. All patients were clinically classified as GSD Ia. Nine patients carried biallelic G6PC mutations (p.Q27fsX35, p.D38V, p.W70X, p.K76N, p.W77R, p.R83C, p.E110Q, p.G222R), with E110Q reported only in Hungary. However, three patients displayed two common G6PT1 (SLC37A4) mutations (p.L348fsX400, p.C183R) which were originally described in association with GSD Inon-a. Review of the literature and our data show that G6PT1 mutations are not associated with neutropenia and related clinical findings in approximately 10% of these cases. Homozygosity for the truncating G6PT1 mutation p.L348fsX400 can be observed with and without neutropenia, indicating that one or more modifiers of the action of G6PT1 exist. Our data are suitable to provide DNA-based and thus noninvasive confirmation of diagnosis in Hungarian patients with this disorder.

p63 Gene mutations in eec syndrome, limb-mammary syndrome, and isolated split hand-split foot malformation suggest a genotype-phenotype correlation.

p63 mutations have been associated with EEC syndrome (ectrodactyly, ectodermal dysplasia, and cleft lip/palate), as well as with nonsyndromic split hand-split foot malformation (SHFM). We performed p63 mutation analysis in a sample of 43 individuals and families affected with EEC syndrome, in 35 individuals affected with SHFM, and in three families with the EEC-like condition limb-mammary syndrome (LMS), which is characterized by ectrodactyly, cleft palate, and mammary-gland abnormalities. The results differed for these three conditions. p63 gene mutations were detected in almost all (40/43) individuals affected with EEC syndrome. Apart from a frameshift mutation in exon 13, all other EEC mutations were missense, predominantly involving codons 204, 227, 279, 280, and 304. In contrast, p63 mutations were detected in only a small proportion (4/35) of patients with isolated SHFM. p63 mutations in SHFM included three novel mutations: a missense mutation (K193E), a nonsense mutation (Q634X), and a mutation in the 3' splice site for exon 5. The fourth SHFM mutation (R280H) in this series was also found in a patient with classical EEC syndrome, suggesting partial overlap between the EEC and SHFM mutational spectra. The original family with LMS (van Bokhoven et al. 1999) had no detectable p63 mutation, although it clearly localizes to the p63 locus in 3q27. In two other small kindreds affected with LMS, frameshift mutations were detected in exons 13 and 14, respectively. The combined data show that p63 is the major gene for EEC syndrome, and that it makes a modest contribution to SHFM. There appears to be a genotype-phenotype correlation, in that there is a specific pattern of missense mutations in EEC syndrome that are not generally found in SHFM or LMS.

Molecular genetics of type 1 glycogen storage disease.

Glycogen storage disease type 1 (GSD 1) comprises a group of autosomal recessive inherited metabolic disorders caused by deficiency of the microsomal multicomponent glucose-6-phosphatase system. Of the two known transmembrane proteins of the system, malfunction of the catalytic subunit (G6Pase) characterizes GSD 1a. GSD 1 non-a is characterized by defective microsomal glucose-6-phosphate or pyrophosphate/phosphate transport due to mutations in G6PT (glucose-6-phosphate translocase gene) encoding a microsomal transporter protein. Mutations in G6Pase and G6PT account for approximately 80 and approximately 20% of GSD 1 cases, respectively. G6Pase and G6PT work in concert to maintain glucose homeostasis in gluconeogenic organs. Whereas G6Pase is exclusively expressed in gluconeogenic cells, G6PT is ubiquitously expressed and its deficiency generally causes a more severe phenotype. Rapid confirmation of clinically suspected diagnosis of GSD 1, reliable carrier testing, and prenatal diagnosis are facilitated by mutation analyses of the chromosome 11-bound G6PT gene as well as the chromosome 17-bound G6Pase gene.

De novo mutation of the connexin 26 gene associated with dominant non-syndromic sensorineural hearing loss.

Mutations of the connexin 26 (Cx26) gene cause isolated recessive or dominant hearing loss or both sensorineural hearing impairment and keratoderma. We have identified the first de novo mutation of the Cx26 gene, R75 W, in a sporadic case of isolated profound hearing loss. R75 W has been previously observed in association with hearing impairment and keratoderma in one family and is thus thought to cause both syndromic and non-syndromic hearing loss. This case illustrates the risk of a possible erroneous diagnosis of autosomal recessive hearing loss in a sporadic case.

Sensorineural hearing loss and the incidence of Cx26 mutations in Austria.

A clinical evaluation and Cx26 mutation analysis was performed in 92 consecutive patients with sensorineural hearing loss in order to delineate the spectrum of genetically caused hearing loss. Among patients of Austrian origin, 53% were classified with hereditary hearing loss. Cx26 mutations were found in 26% of NSHL patients (40% of familial vs 18% of sporadic cases). The mutation 35delG accounted for 52.8% of all presumed GJB2 disease alleles. The second most frequent mutation was L90P (16.7%) having been reported with a prevalence of 0.7-3.5% in other populations. Three novel mutations were found. The novel mutation, R143Q, was associated with dominant high-frequency hearing loss. Pseudodominant transmission of NSHL was seen in four families with Cx26 mutations. A mutation 35delG carrier rate of 0.9% was observed among 672 controls from West-Austria. Cx26 mutations were found associated with mild to profound, and with asymmetric hearing impairment.

Congenital sodium diarrhea is an autosomal recessive disorder of sodium/proton exchange but unrelated to known candidate genes.

BACKGROUND & AIMS: Congenital sodium diarrhea (CSD) is caused by defective sodium/proton exchange with only 6 sporadic cases reported. The genetics of the disease have not been established. We studied 5 infants with secretory diarrhea, identified in a circumscribed rural area in Austria, to define the mode of transmission and the involvement of candidate genes known to encode for sodium/proton exchangers (NHEs). METHODS: We collected clinical and laboratory data from 5 affected patients, analyzed the pedigrees of their families, and performed homozygosity mapping and multipoint linkage analysis studies in 4 candidate regions known to contain NHE genes. RESULTS: The diagnosis of CSD in 4 of 5 patients was based on daily fecal sodium excretion between 98 and 190 mmol/L, hyponatremia, metabolic acidosis, and low-to-normal urinary sodium concentrations. Pedigree analysis of the affected 2 CSD families revealed parental consanguinity and a common single ancestor 5 generations ago. Homozygosity mapping and/or multipoint linkage analysis excluded the NHE1 locus on chromosome 1, NHE2 locus on chromosome 2, NHE3 locus on chromosome 5, and NHE5 locus on chromosome 16 as potential candidate genes for CSD in this pedigree. Results on NHE4 were inconclusive because the precise chromosomal location of this NHE gene in humans is currently unknown. CONCLUSIONS: Our data indicate that CSD is an autosomal recessive disorder but is not related to mutations in the NHE1, NHE2, NHE3, and NHE5 genes encoding for currently known sodium/proton exchangers.

Mutation analysis in glycogen storage disease type 1 non-a.

We report molecular and clinical findings in 13 patients with rare types of glycogen storage disease 1 (GSD1 non-a). Analysis of G6PT encoding a microsomal transporter protein has revealed mutations on both chromosomes in each case, four of which are novel. Diagnosis has been confirmed in three patients suspected of having GSD1 non-a without enzymatic studies involving liver biopsy, thus emphasising the advantage of G6PT mutation analysis for all GSD1 non-a patients.

[Application of indirect calorimetry in monitoring feeding of low birth-weight preterm infants]. / Anwendung der indirekten Kalorimetrie zur Ernährungssteuerung bei kleinen Frühgeborenen.

BACKGROUND: We investigated the practical use of indirect calorimetry for the individual nutritional support of preterm infants in order to answer the question whether it is possible to reliably calculate energy expenditure, fat and carbohydrate oxidation in preterm infants individually by using the results of a timed 6-hour-measurement of oxygen consumption and carbon dioxide production. PATIENTS: Measurements were performed in 20 preterm infants (gestational age 30.2 +/- 0.6 weeks, birth weight 1.09 +/- 0.07; mean +/- SEM) at a mean postnatal age of 25 +/- 4 days and with a body weight of 1.35 +/- 0.06 kg. METHODS: Carbon dioxide production (24 h-VCO2), oxygen consumption (24 h-VO2) and respiratory quotient (24 h-RQ) were measured by indirect calorimetry for 24 hours using the Deltatrac II metabolic monitor (Datex, Helsinki, Finland). Additionally, 6 h-VCO2, 6 h-VO2 and 6 h-RQ were determined by measurement over 6 hours. The patients' energy expenditure, fat and carbohydrate oxidation were calculated from VCO2 and VO2 measured over a 24 hour- and 6 hour-period with or without consideration of urinary nitrogen excretion (NU). RESULTS: If NU was not included in the calculation of energy expenditure, the values differed by maximally 1.1% from the calculations including NU. The correlations between the 24 h-RQ and the calculated 24 h-fat or 24 h-carbohydrate oxidation values were statistically significant (r = -0.99; p = 0.0001 and r = 0.773; p = 0.0002 respectively). However, in individual patients, it was not possible to predict 24 h energy expenditure, fat and carbohydrate oxidation of preterm infants using values determined by 6 h indirect calorimetry. CONCLUSION: The determination of the urine-nitrogen excretion is not necessary for calculation of energy expenditure of preterm infants. It is possible to estimate fat and carbohydrate oxidation of preterm infants by the measured 24 h-RQ, but 6 h indirect calorimetry is not accurate enough for calculating the individual nutritional needs of preterm infants in clinical practice. Indirect calorimetry over 24 h may be helpful in the management of selected patients with nutritional problems.

Molecular diagnosis of type 1c glycogen storage disease.

Glycogen storage disease type 1 (GSD 1) results from deficiency of the microsomal multicomponent glucose-6-phosphatase system. Malfunction of the catalytic subunit characterises GSD 1a. GSD 1b and GSD 1c are characterised by defective microsomal glucose-6-phosphate or pyrophosphate/phosphate transport, respectively. Recently, a gene encoding a microsomal transporter protein has been found to be mutated in GSD 1b and 1c patients. Here, we report the genomic sequence of the transporter gene and the detection of a homozygous 2-bp deletion (1211delCT) and a homozygous donor splice site mutation (317+1G-->T) in two GSD 1c patients, confirming that GSD 1c is allelic to GSD 1b.