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Contains fulltext : 125691.pdf (Publisher’s version ) (Closed access) OBJECTIVE: Retinitis pigmentosa (RP) is a clinically and genetically heterogeneous disorder characterized by night blindness and peripheral vision loss, and in many cases leads to blindness. Despite extensive knowledge about genes involved in the pathogenesis of RP, the genetic cause remains elusive in many patients. In this study, we aimed to identify novel genes that are involved in the cause of RP. DESIGN: We present a case series with mutations in the mevalonate kinase (MVK) gene. PARTICIPANTS: A total of 769 patients with nonsyndromic RP and 174 Dutch control individuals participated in this study. METHODS: Exome sequencing analysis was performed in a proband of Dutch origin who was initially diagnosed with nonsyndromic autosomal recessive RP. Mutations in MVK were identified and subsequently tested for segregation within the patient's family and screened in a large cohort of patients with genetically unsolved RP. Patients with mutations underwent extensive clinical reexamination. MAIN OUTCOME MEASURES: Digital fundus photography, spectral-domain optical coherence tomography (OCT), and fundus autofluorescence analysis were performed in patients with MVK mutations. Mevalonate kinase (MK) enzyme activity was analyzed in cultured lymphoblastoid cells, and mevalonic acid levels were measured in urine samples. RESULTS: Exome variant filtering and prioritization led to the identification of compound heterozygous mutations in MVK (p.I268T and p.A334T) in the proband and her affected brother. Screening of our nonsyndromic RP patient cohort revealed an additional individual who was homozygous for the p.A334T alteration. Clinical reevaluation of all 3 patients showed a classic form of RP with variable extraocular symptoms, such as history of recurrent childhood febrile crises in 2 patients, mild ataxia in 1, and renal failure in 1. All 3 affected individuals showed a significantly decreased MK activity and highly elevated levels of urinary mevalonic acid. CONCLUSIONS: Although the MK activity in cells and mevalonic acid concentrations in urine are strongly aberrant and comparable to that in patients with systemic mevalonate kinase deficiency (MKD), only mild clinical symptoms related to this syndrome were observed in our patients. In the current article, we add another phenotype to the spectrum of diverging disorders associated with mutations in MVK. FINANCIAL DISCLOSURE(S): The author(s) have no proprietary or commercial interest in any materials discussed in this article.

Contains fulltext : 125275.pdf (Publisher’s version ) (Closed access) Through whole-genome sequencing of 2,230 Icelanders, we detected a rare nonsynonymous SNP (minor allele frequency = 0.55%) in the C3 gene encoding a p.Lys155Gln substitution in complement factor 3, which, following imputation into a set of Icelandic cases with age-related macular degeneration (AMD) and controls, associated with disease (odds ratio (OR) = 3.45; P = 1.1 x 10(-7)). This signal is independent of the previously reported common SNPs in C3 encoding p.Pro314Leu and p.Arg102Gly that associate with AMD. The association of p.Lys155Gln was replicated in AMD case-control samples of European ancestry with OR = 4.22 and P = 1.6 x 10(-10), resulting in OR = 3.65 and P = 8.8 x 10(-16) for all studies combined. In vitro studies have suggested that the p.Lys155Gln substitution reduces C3b binding to complement factor H, potentially creating resistance to inhibition by this factor. This resistance to inhibition in turn is predicted to result in enhanced complement activation.

Item does not contain fulltext Turner syndrome (TS) is the result of (partial) X chromosome monosomy. In general, the diagnosis is based on karyotyping of 30 blood lymphocytes. This technique, however, does not rule out tissue mosaicism or low grade mosaicism in the blood. Because of the associated risk of gonadoblastoma, mosaicism is especially important in case this involves a Y chromosome. We investigated different approaches to improve the detection of mosaicisms in 162 adult women with TS (mean age 29.9 +/- 10.3). Standard karyotyping identified 75 patients (46.3%) with a non-mosaic monosomy 45,X. Of these 75 patients, 63 underwent additional investigations including FISH on buccal cells with X- and Y-specific probes and PCR-Y on blood. FISH analysis of buccal cells revealed a mosaicism in 19 of the 63 patients (30.2%). In five patients the additional cell lines contained a (derivative) Y chromosome. With sensitive real-time PCR we confirmed the presence of this Y chromosome in blood in three of the five cases. Although Y chromosome material was established in ovarian tissue in two patients, no gonadoblastoma was found. Our results confirm the notion that TS patients with 45,X on conventional karyotyping often have tissue specific mosaicisms, some of which include a Y chromosome. Although further investigations are needed to estimate the risk of gonadoblastoma in patients with Y chromosome material in buccal cells, we conclude that FISH or real-time PCR on buccal cells should be considered in TS patients with 45,X on standard karyotyping.

Item does not contain fulltext OBJECTIVES: While urine sampling is necessary in the diagnosis of urinary tract infection and electrolyte disturbances, the collection of urine in neonates and non-toilet-trained children is often difficult. A universal urine collection method providing representative urinalyses results is needed. The objective of this study is to evaluate the applicability of the currently used urine collection pads (gauze compresses) and a new urine collection device (Peespot). DESIGN AND METHODS: We tested the reliability of routine (semi-)quantitative urinalysis results with these two different kinds of urine collection methods in a laboratory model. Although important in clinical diagnosis, we did not evaluate the effects on cellular and other components such as casts in the urinary sediment. RESULTS: Most semi-quantitative variables determined by urine stick (pH, blood, protein, leukocytes, nitrite, glucose, ketones, bilirubin and urobilinogen) gave concordant results for both methods compared with native urine. Using the Peespot urine collection device, reliable quantitative results were obtained for calcium, chloride, glucose, magnesium, phosphate, potassium, sodium, osmolality, urea nitrogen and urate compared with native urine. Data were concordant only for chloride, phosphate, glucose, sodium and urea nitrogen by use of gauze compresses. CONCLUSIONS: Urine collection pads are non-invasive methods useful in the collection of urine in non-toilet-trained children. Because of better practical standardisation and more reliable (semi-) quantitative urinalysis results, the Peespot urine collection device is preferred for the collection of urine.

Contains fulltext : 119120.pdf (Publisher’s version ) (Closed access) OBJECTIVE: To identify the underlying genetic defect in a patient with intellectual disability, seizures, ataxia, macrothrombocytopenia, renal and cardiac involvement, and abnormal protein glycosylation. METHODS: Genetic studies involved homozygosity mapping by 250K single nucleotide polymorphism array and SLC35A1 sequencing. Functional studies included biochemical assays for N-glycosylation and mucin-type O-glycosylation and SLC35A1-encoded cytidine 5'-monophosphosialic acid (CMP-sialic acid) transport after heterologous expression in yeast. RESULTS: We performed biochemical analysis and found combined N- and O-glycosylation abnormalities and specific reduction in sialylation in this patient. Homozygosity mapping revealed homozygosity for the CMP-sialic acid transporter SLC35A1. Mutation analysis identified a homozygous c.303G>C (p.Gln101His) missense mutation that was heterozygous in both parents. Functional analysis of mutant SLC35A1 showed normal Golgi localization but 50% reduction in transport activity of CMP-sialic acid in vitro. CONCLUSION: We confirm an autosomal recessive, generalized sialylation defect due to mutations in SLC35A1. The primary neurologic presentation consisting of ataxia, intellectual disability, and seizures, in combination with bleeding diathesis and proteinuria, is discriminative from a previous case described with deficient sialic acid transporter. Our study underlines the importance of sialylation for normal CNS development and regular organ function.

Purpose: Pheochromocytomas and paragangliomas (PGL) are neuroendocrine tumors of sympathetic and parasympathetic paraganglia. This study investigated the relationships between genotype-specific differences in mitochondrial function and catecholamine content in PGL tumors. Experimental Design: Respiratory chain enzyme assays and 1H-nuclear magnetic resonance (NMR) spectroscopy at 500 MHz were conducted on homogenates of 35 sporadic PGLs and 59 PGLs from patients with hereditary mutations in succinate dehydrogenase subunits B and D (SDHB, SDHD), succinate dehydrogenase assembly factor 2, von Hippel-Lindau (VHL), rearranged during transfection (RET), neurofibromatosis type 1 (NF1), and myc-associated factor X. Results: In SDHx-related PGLs, a significant decrease in complex II activity (P < 0.0001) and a significant increase in complex I, III, and IV enzyme activities were observed when compared to sporadic, RET, and NF1 tumors. Also, a significant increase in citrate synthase (P < 0.0001) enzyme activity was observed in SDHx-related PGLs when compared to sporadic-, VHL-, RET-, and NF1-related tumors. An increase in succinate accumulation (P < 0.001) and decrease in ATP/ADP/AMP accumulation (P < 0.001) was observed when compared to sporadic PGLs and PGLs of other genotypes. Positive correlations (P < 0.01) were observed between respiratory chain complex II activity and total catecholamine content and ATP/ADP/AMP and total catecholamine contents in tumor tissues. Conclusions: This study for the first time establishes a relationship between determinants of energy metabolism, like activity of respiratory chain enzyme complex II, ATP/ADP/AMP content, and catecholamine content in PGL tumors. Also, this study for the first time successfully uses NMR spectroscopy to detect catecholamines in PGL tumors and provides ex vivo evidence for the accumulation of succinate in PGL tumors with an SDHx mutation. Clin Cancer Res; 19(14); 3787–95. ©2013 AACR.

Contains fulltext : 118864.pdf (Publisher’s version ) (Closed access) BackgroundFocal segmental glomerulosclerosis (FSGS) is a leading cause of steroid-resistant nephrotic syndrome. Hereditary FSGS is frequently caused by mutations in important structural podocyte proteins, including the slit diaphragm-associated transient receptor potential channel C6 (TRPC6).MethodsIn five patients with biopsy-proven autosomal-dominant FSGS from five different Dutch families, all 13 exons of TRPC6 were sequenced. Upon identification of a novel TRPC6 sequence variant, the resultant amino acid change was introduced in the wild-type TRPC6 protein and functionally tested using patch-clamp analyses and cell-surface biotinylation experiments.ResultsNone of the previously described TRPC6 mutations were found in our cohort. In one family, we identified a novel c.524G>A sequence variant resulting in a p.Arg175Gln (R175Q) substitution in the TRPC6 protein. This sequence variant was absent in 449 control subjects and from public SNP databases. The mutation was located in the third ankyrin repeat domain (ANK3) in the cytoplasmic N-tail of TRPC6, important for protein-protein interaction and regulation of ion channel activity. Patch-clamp analyses of the mutant channel indeed showed an increased TRPC6 channel-mediated current. However, cell-surface expression of the mutant channel was not increased.ConclusionsWe identified a novel TRPC6 p.Arg175Gln gain-of-function mutation that shows increased TRPC6-mediated current, which is not due to altered cell-surface expression. This is the first mutation identified in ANK3 of the TRPC6 N-tail and is most likely responsible for the late-onset autosomal dominant FSGS in this family.

Background— Loss-of-function mutations in CHD7 cause Coloboma, Heart Disease, Atresia of Choanae, Retardation of Growth and/or Development, Genital Hypoplasia, and Ear Abnormalities With or Without Deafness (CHARGE) syndrome, a variable combination of multiple congenital malformations including heart defects. Heart defects are reported in 70% to 92% of patients with a CHD7 mutation, but most studies are small and do not provide a detailed classification of the defects. We present the first, detailed, descriptive study on the cardiac phenotype of 299 patients with a CHD7 mutation and discuss the role of CHD7 in cardiac development. Methods and Results— We collected information on congenital heart defects in 299 patients with a pathogenic CHD7 mutation, of whom 220 (74%) had a congenital heart defect. Detailed information on the heart defects was available for 202 of these patients. We classified the heart defects based on embryonic cardiac development and compared the distribution to 1007 equally classified nonsyndromic heart defects of patients registered by EUROCAT, a European Registry of Congenital Anomalies. Heart defects are highly variable in patients with CHD7 mutations, but atrioventricular septal defects and conotruncal heart defects are over-represented. Sex did not have an effect on the presence of heart defects, but truncating CHD7 mutations resulted in a heart defect significantly more often than missense or splice-site mutations (χ 2 , P Conclusions— CHD7 plays an important role in cardiac development, given that we found a wide range of heart defects in 74% of a large cohort of patients with a CHD7 mutation. Conotruncal defects and atrioventricular septal defects are over-represented in patients with CHD7 mutations compared with patients with nonsyndromic heart defects.

International audience; The M-type receptor for phospholipase A2 (PLA2R1) is the major target antigen in idiopathic membranous nephropathy (iMN). Our recent genome-wide association study showed that genetic variants in an HLA-DQA1 and phospholipase A2 receptor (PLA2R1) allele associate most significantly with biopsy-proven iMN, suggesting that rare genetic variants within the coding region of the PLA2R1 gene may contribute to antibody formation. Here, we sequenced PLA2R1 in a cohort of 95 white patients with biopsy-proven iMN and assessed all 30 exons of PLA2R1, including canonical (GT-AG) splice sites, by Sanger sequencing. Sixty patients had anti-PLA2R1 in serum or detectable PLA2R1 antigen in kidney tissue. We identified 18 sequence variants, comprising 2 not previously described, 7 reported as rare variants (

Item does not contain fulltext Leigh syndrome (LS) is a progressive neurodegenerative disease caused by either mitochondrial or nuclear DNA mutations resulting in dysfunctional mitochondrial energy metabolism. Mutations in genes encoding for subunits of the respiratory chain or assembly factors of respiratory chain complexes are often documented in LS cases. Nicotinamide adenine dinucleotide (NADH):ubiquinone oxidoreductase (complex I) enzyme deficiencies account for a significant proportion of mitochondrial disorders, including LS. In an attempt to expand the repertoire of known mutations accounting for LS, we describe the clinical, radiological, biochemical and molecular data of six patients with LS found to have novel mutations in two complex I subunits (NDUFV1 and NDUFS2). Two siblings were homozygous for the previously undescribed R386C mutation in NDUFV1, one patient was a compound heterozygote for the R386C mutation in NDUFV1 and a frameshift mutation in the same gene, one patient was a compound heterozygote for the R88G and R199P mutations in NDUFV1, and two siblings were compound heterozygotes for an undescribed E104A mutation in NDUFS2. After the novel mutations were identified, we employed prediction models using protein conservation analysis (SIFT, PolyPhen and UCSC genome browser) to determine pathogenicity. The R386C, R88G, R199P, and E104A mutations were found to be likely pathogenic, and thus presumably account for the LS phenotype. This case series broadens our understanding of the etiology of LS by identifying new molecular defects that can result in complex I deficiency and may assist in targeted diagnostics and/or prenatal diagnosis of LS in the future.

Background/Purpose: In fetuses with gastroschisis, toxic products in the amniotic fluid and constriction at the defect of the abdominal wall are considered causative of damage to the eviscerated bowel. The aim of this study was to cover the eviscerated bowel in gastroschisis with a collagen scaffold to protect the bowel and induce cell growth into the scaffold, which could lead to skin or abdominal wall formation replacing the scaffold.Methods: In 12 fetal lambs gastroschisis was surgically created at 79 days gestation. A dual-layer type I collagen scaffold was sutured into the skin of the abdominal wall around the defect covering the eviscerated bowel. Lambs were examined after caesarean section at 140 days' gestation.Results: Survival was 67%. In 7 of 8 surviving lambs the bowel was found to be covered after birth. One scaffold had ruptured. The bowel was found repositioned in the abdominal cavity in 5 lambs. In 2 lambs it was still partially outside. Only minor adherence of bowel loops and no fibrous peel formation were seen. Connective tissue and skin tissue replaced the scaffold.Conclusions: Prenatal coverage of the bowel in experimental gastroschisis with a collagen scaffold is feasible in fetal lambs, significantly diminished damage to the bowel wall, and skin and connective tissue replaced the scaffold. This technique may be promising in the care of fetuses with this congenital anomaly. (C) 2013 Elsevier Inc. All rights reserved.

Item does not contain fulltext BACKGROUND: Marinesco-Sjogren syndrome is an autosomal recessive cerebellar ataxia, characterised by cerebellar ataxia, myopathy, cataracts and intellectual disability, due to mutations in the SIL1 gene. METHODS: The clinical features and two novel SIL1 mutations of four Dutch patients with Marinesco-Sjogren syndrome are described and compared to the literature on genetically proven Marinesco-Sjogren patients. RESULTS: The core phenotype of this syndrome appears homogeneous, but: [1] cataract can develop later than the motor and cognitive signs; [2] myopathy is an early feature that seems progressive during the course of the disease; [3] serum creatine kinase is normal or only mildly elevated; [4] peripheral neuropathy is absent; and [5] a variable degree of intellectual disability is present in most Marinesco-Sjogren patients. CONCLUSIONS: Because the late appearance of some hallmarks and the uncertainty as to whether incomplete phenotypes occur, SIL1 mutation analysis is helpful early in the diagnostic work-up of children with suspected inherited ataxias.

Item does not contain fulltext Survival rates in esophageal atresia (EA) patients have reached 90%. In long-term follow-up studies the focus has shifted from purely surgical or gastrointestinal evaluation to a multidisciplinary approach. We evaluated the long-term morbidity in adolescent and adult EA patients and discussed mainly nonsurgical issues. Dysphagia is common and reported in up to 85% of patients. In young adults gastroesophageal reflux disease occurs frequently with development of Barrett esophagus in 6% reported in different series. It is difficult to estimate respiratory morbidity from the literature because many different definitions, questionnaires, and study designs have been used. However, many patients seem to suffer from respiratory problems even into adulthood. In conclusion, morbidity is not only restricted to surgical problems but many different domains are involved. These are all related and together determine to a large extent the quality of life of EA patients and also of their families. We assume that a multidisciplinary care approach seems best to address their special needs.

Contains fulltext : 118302.pdf (Publisher’s version ) (Closed access) Creatine (Cr) plays an important role in muscle energy homeostasis by its participation in the ATP-phosphocreatine phosphoryl exchange reaction mediated by creatine kinase. Given that the consequences of Cr depletion are incompletely understood, we assessed the morphological, metabolic and functional consequences of systemic depletion on skeletal muscle in a mouse model with deficiency of l-arginine:glycine amidinotransferase (AGAT(-/-)), which catalyses the first step of Cr biosynthesis. In vivo magnetic resonance spectroscopy showed a near-complete absence of Cr and phosphocreatine in resting hindlimb muscle of AGAT(-/-) mice. Compared with wild-type, the inorganic phosphate/beta-ATP ratio was increased fourfold, while ATP levels were reduced by nearly half. Activities of proton-pumping respiratory chain enzymes were reduced, whereas F(1)F(0)-ATPase activity and overall mitochondrial content were increased. The Cr-deficient AGAT(-/-) mice had a reduced grip strength and suffered from severe muscle atrophy. Electron microscopy revealed increased amounts of intramyocellular lipid droplets and crystal formation within mitochondria of AGAT(-/-) muscle fibres. Ischaemia resulted in exacerbation of the decrease of pH and increased glycolytic ATP synthesis. Oral Cr administration led to rapid accumulation in skeletal muscle (faster than in brain) and reversed all the muscle abnormalities, revealing that the condition of the AGAT(-/-) mice can be switched between Cr deficient and normal simply by dietary manipulation. Systemic creatine depletion results in mitochondrial dysfunction and intracellular energy deficiency, as well as structural and physiological abnormalities. The consequences of AGAT deficiency are more pronounced than those of muscle-specific creatine kinase deficiency, which suggests a multifaceted involvement of creatine in muscle energy homeostasis in addition to its role in the phosphocreatine-creatine kinase system.

Item does not contain fulltext The clinical and molecular characterizations of two patients with a 1.4 Mb overlapping deletion in the 6p25.1p24.3 region are reported. In addition to the mild intellectual disability, they shared feeding problems in infancy and several dysmorphic facial features including a prominent forehead, almond-shaped eyes, a short philtrum, and low-set ears with square helices. The overlapping deleted region harbors six genes (RREB1, NRN1, CAGE1, LY86, SSR1, and F13A1), of which NRN1 and RREB1 are considered as candidate genes for the intellectual disability and the overlapping dysmorphism, respectively.

Item does not contain fulltext The majority of the genetic causes of autosomal-recessive (ar) cone-rod dystrophy (CRD) are currently unknown. A combined approach of homozygosity mapping and exome sequencing revealed a homozygous nonsense mutation (c.565C>T [p.Glu189*]) in RAB28 in a German family with three siblings with arCRD. Another homozygous nonsense mutation (c.409C>T [p.Arg137*]) was identified in a family of Moroccan Jewish descent with two siblings affected by arCRD. All five affected individuals presented with hyperpigmentation in the macula, progressive loss of the visual acuity, atrophy of the retinal pigment epithelium, and severely reduced cone and rod responses on the electroretinogram. RAB28 encodes a member of the Rab subfamily of the RAS-related small GTPases. Alternative RNA splicing yields three predicted protein isoforms with alternative C-termini, which are all truncated by the nonsense mutations identified in the arCRD families in this report. Opposed to other Rab GTPases that are generally geranylgeranylated, RAB28 is predicted to be farnesylated. Staining of rat retina showed localization of RAB28 to the basal body and the ciliary rootlet of the photoreceptors. Analogous to the function of other RAB family members, RAB28 might be involved in ciliary transport in photoreceptor cells. This study reveals a crucial role for RAB28 in photoreceptor function and suggests that mutations in other Rab proteins may also be associated with retinal dystrophies.

Item does not contain fulltext Large defects in congenital diaphragmatic hernia are closed by patch repair, which is associated with a high complication risk and reherniation rate. New treatment modalities are warranted. We evaluated the feasibility of using an acellular biodegradable collagen bioscaffold for a regenerative medicine approach to close a surgically created diaphragmatic defect in a rat model. Scaffold degradation, cellular ingrowth and regeneration of the diaphragm were studied. In 25 rats, a subcostal incision was made and one third of the right hemidiaphragm was resected. Crosslinked porous type I collagen scaffolds (O ~ 14 mm) were sutured into the lesion. Rats were sacrificed at 2, 4, 8, 12 or 24 weeks after scaffold implantation. Implants were evaluated macroscopically and (immuno)histologically. Survival after surgery was 88% with no evidence of reherniation. Histological examination showed that the collagen scaffold degraded slowly and new collagen, elastin and mesothelium were deposited. Blood vessels were observed primarily at the outer borders of the scaffold; their number gradually increased in time. Muscle fibres were found on the scaffold covering up to 10% of the defect. Macroscopically, adhesion of the scaffold to the liver was observed. Use of a collagen scaffold to close a surgically created diaphragmatic defect is feasible, with evidence of new tissue formation. The use of crosslinked collagen scaffolds allows targeted modification; e.g. addition of growth factors to further stimulate growth of muscle cells. Copyright (c) 2012 John Wiley & Sons, Ltd.

Item does not contain fulltext Inherited retinal diseases display a very high degree of clinical and genetic heterogeneity, which poses challenges in identifying the underlying defects in known genes and in identifying novel retinal disease genes. Here, we outline the state-of-the-art techniques to find the causative DNA variants, with special attention for next-generation sequencing which can combine molecular diagnostics and retinal disease gene identification.

Item does not contain fulltext The acronym VATER/VACTERL association describes the combination of at least three of the following congenital anomalies: vertebral defects (V), anorectal malformations (A), cardiac defects (C), tracheoesophageal fistula with or without esophageal atresia (TE), renal malformations (R), and limb defects (L). We aimed to identify highly penetrant de novo copy number variations (CNVs) that contribute to VATER/VACTERL association. Array-based molecular karyotyping was performed in a cohort of 41 patients with VATER/VACTERL association and 6 patients with VATER/VACTERL-like phenotype including all of the patients' parents. Three de novo CNVs were identified involving chromosomal regions 1q41, 2q37.3, and 8q24.3 comprising one (SPATA17), two (CAPN10, GPR35), and three (EPPK1, PLEC, PARP10) genes, respectively. Pre-existing data from the literature prompted us to choose GPR35 and EPPK1 for mouse expression studies. Based on these studies, we prioritized GPR35 for sequencing analysis in an extended cohort of 192 patients with VATER/VACTERL association and VATER/VACTERL-like phenotype. Although no disease-causing mutation was identified, our mouse expression studies suggest GPR35 to be involved in the development of the VATER/VACTERL phenotype. Follow-up of GPR35 and the other genes comprising the identified duplications is warranted.

Item does not contain fulltext There is growing evidence that laminopathies, diseases associated with mutations in the LMNA gene, are caused by a combination of mechanical and gene regulatory distortions. Strikingly, there is a large variability in disease symptoms between individual patients carrying an identical LMNA mutation. This is why classical genetic screens for mutations appear to have limited predictive value for disease development. Recently, the widespread occurrence of repetitive nuclear ruptures has been described in fibroblast cultures from various laminopathy patients. Since this phenomenon was strongly correlated with disease severity, the identification of biomarkers that report on these rupture events could have diagnostic relevance. One such candidate marker is the PML nuclear body, a structure that is normally confined to the nuclear interior, but leaks out of the nucleus upon nuclear rupture. Here, we show that a variety of laminopathies shows the presence of these cytoplasmic PML particles (PML CPs), and that the amount of these protein aggregates increases with severity of the disease. In addition, between clinically healthy individuals, carrying LMNA mutations, significant differences can be found. Therefore, we postulate that detection of PML CPs in patient fibroblasts could become a valuable marker for diagnosis of disease development.

Item does not contain fulltext Mutations in the skeletal muscle ryanodine receptor (RYR1) gene are a common cause of inherited neuromuscular disorders and have been associated with a wide clinical spectrum, ranging from various congenital myopathies to the malignant hyperthermia susceptibility (MHS) trait without any associated weakness. RYR1-related myopathies are usually of early-childhood onset. Here we present 11 patients from 8 families with a late-onset axial myopathy associated with RYR1 variants. Patients presented between the third and seventh decade of life to neuromuscular centres in Norway, the Netherlands and the United Kingdom with predominant axial muscle involvement, comprising variable degrees of lumbar hyperlordosis, scapular winging and/or camptocormia. Marked myalgia was commonly associated. Serum creatine kinase levels were normal or moderately elevated. Muscle imaging showed consistent involvement of the lower paravertebral muscles and the posterior thigh. Muscle biopsy findings were often discrete, featuring variability in fibre size, increased internal nuclei and unevenness of oxidative enzyme staining, but only rarely overt cores. RYR1 sequencing revealed heterozygous missense variants, either previously associated with the MHS trait or localizing to known MHS mutational hotspots. These findings indicate that MHS-related RYR1 mutations may present later in life with prominent axial weakness but not always typical histopathological features. We propose a combined effect of RyR1 dysfunction, aging and particular vulnerability of axial muscle groups as a possible pathogenic mechanism. RYR1 is a candidate for cases with "idiopathic" camptocormia or bent spine syndrome (BSS).

Item does not contain fulltext Aortic aneurysm, including both abdominal aortic aneurysm and thoracic aortic aneurysm, is the cause of death of 1% to 2% of the Western population. This review focuses only on thoracic aortic aneurysms and dissections. During the past decade, the genetic contribution to the pathogenesis of thoracic aortic aneurysms and dissections has revealed perturbed extracellular matrix signaling cascade interactions and deficient intracellular components of the smooth muscle contractile apparatus as the key mechanisms. Based on the study of different Marfan mouse models and the discovery of several novel thoracic aortic aneurysm genes, the involvement of the transforming growth factor-beta signaling pathway has opened unexpected new avenues. Overall, these discoveries have 3 important consequences. First, the pathogenesis of thoracic aortic aneurysms and dissections is better understood, although some controversy still exists. Second, the management strategies for the medical and surgical treatment of thoracic aortic aneurysms and dissections are becoming increasingly gene-tailored. Third, the pathogenetic insights have delivered new treatment options that are currently being investigated in large clinical trials.

Item does not contain fulltext OBJECTIVE: To identify the mutated gene in a group of patients with an unclassified heritable white matter disorder sharing the same, distinct MRI pattern. METHODS: We used MRI pattern recognition analysis to select a group of patients with a similar, characteristic MRI pattern. We performed whole-exome sequencing to identify the mutated gene. We examined patients' fibroblasts for biochemical consequences of the mutant protein. RESULTS: We identified 6 patients from 5 unrelated families with a similar MRI pattern showing predominant abnormalities of the cerebellar cortex, deep cerebral white matter, and corpus callosum. The 4 tested patients had a respiratory chain complex capital I, Ukrainian deficiency. Exome sequencing revealed mutations in NUBPL, encoding an iron-sulfur cluster assembly factor for complex capital I, Ukrainian, in all patients. Upon identification of the mutated gene, we analyzed the MRI of a previously published case with NUBPL mutations and found exactly the same pattern. A strongly decreased amount of NUBPL protein and fully assembled complex I was found in patients' fibroblasts. Analysis of the effect of mutated NUBPL on the assembly of the peripheral arm of complex I indicated that NUBPL is involved in assembly of iron-sulfur clusters early in the complex I assembly pathway. CONCLUSION: Our data show that NUBPL mutations are associated with a unique, consistent, and recognizable MRI pattern, which facilitates fast diagnosis and obviates the need for other tests, including assessment of mitochondrial complex activities in muscle or fibroblasts.

Item does not contain fulltext OBJECTIVE: To evaluate developmental and social-emotional outcomes at 8 years of age for children with congenital diaphragmatic hernia (CDH), treated with or without neonatal extracorporeal membrane oxygenation (ECMO) between January 1999 and December 2003. DESIGN: Cohort study with structural prospective follow-up. SETTING: Level III University Hospital. PATIENTS: 35 children (ECMO: n=16; non-ECMO: n=19) were assessed at 8 years of age. INTERVENTIONS: None. MAIN OUTCOME MEASURES: Intelligence and motor function. Concentration, behaviour, school performance, competence and health status were also analysed. RESULTS: Mean (SD) intelligence for the ECMO group was 91.7 (19.5) versus 111.6 (20.9) for the non-ECMO group (p=0.015). Motor problems were apparent in 16% of all participants and differed significantly from the norm (p=0.015) without differences between treatment groups. For all participants, problems with concentration (68%, p

Item does not contain fulltext BACKGROUND: /st> Asthma is a common disease in children and often develops early in life. This multicentre retrospective case series describe the use and effectiveness of sevoflurane inhalation therapy in a series of children with severe asthma in the paediatric intensive care unit (PICU). METHODS: /st> Seven children ranging from 4 to 13 yr of age admitted to the PICU of two tertiary care hospitals in the Netherlands were included. They all were admitted with the diag-nosis of severe asthma requiring invasive mechanical ventilation and were treated with sevoflurane inhalation therapy. RESULTS: /st> The median (range) Pco2 level at the start, after 2 h, and at the end of sevoflurane treatment were 14 (5.1-24.8), 9.8 (5.4-17.0), and 6.2 (4.5-11.4) kPa (P=0.05) while the median (range) pH was 7.02 (6.97-7.36), 7.18 (7.04-7.35), and 7.43 (7.15-7.47) kPa (P=0.01), respectively. The median (range) peak pressure values declined from 30 (23-56) to 20.4 (14-33) cm H2O (P=0.03). No severe adverse effects besides hypotension, with sufficient response to norepinephrine treatment, were seen. CONCLUSIONS: /st> Sevoflurane inhalation corrects high levels of Pco2 and provides clinical improvement in mechanically ventilated children with life-threatening asthma who fail to respond to conventional treatment. 01 december 2013

Item does not contain fulltext A regenerative medicine approach to restore the morphology and function of the diaphragm in congenital diaphragmatic hernia is especially challenging because of the position and flat nature of this organ, allowing cell ingrowth primarily from the perimeter. Use of porous collagen scaffolds for the closure of surgically created diaphragmatic defects in rats has been shown feasible, but better ingrowth of cells, specifically blood vessels and muscle cells, is warranted. To stimulate this process, heparin, a glycosaminoglycan involved in growth factor binding, was covalently bound to porous collagenous scaffolds (14%), with or without vascular endothelial growth factor (VEGF; 0.4 microg/mg scaffold), hepatocyte growth factor (HGF; 0.5 microg/mg scaffold) or a combination of VEGF + HGF (0.2 + 0.5 microg/mg scaffold). All components were located primarily at the outside of scaffolds. Scaffolds were implanted in the diaphragm of rats and evaluated after 2 and 12 weeks. No herniations or eventrations were observed, and in several cases, growth factor-substituted scaffolds showed macroscopically visible blood vessels at the lung site. The addition of heparin led to an accelerated ingrowth of blood vessels at 2 weeks. In all scaffold types, giant cells and immune cells were present primarily at the liver side of the scaffold, and immune cells and individual macrophages at the lung side; these cell types decreased in number from week 2 to week 12. The addition of growth factors did not influence cellular response to the scaffolds, indicating that further optimization with respect to dosage and release profile is needed.

Contains fulltext : 115395.pdf (Publisher’s version ) (Closed access) Pre/perinatal risk factors and body growth abnormalities have been studied frequently as early risk markers in autism spectrum disorder (ASD), yet their interrelatedness in ASD has received very little research attention. This is surprising, given that pre/perinatal risk factors can have a substantial impact on growth trajectories in the first years of life. We aimed to determine which pre/perinatal factors were more prevalent in ASD children and if these factors differentially influenced body growth in ASD and control children. A total of 96 ASD and 163 control children matched for gender participated. Data of growth of head size and body length during the first 13 months of life were collected. Data on pre/perinatal risk factors were retrospectively collected through standardized questionnaires. Results indicated that after matching for SES, prematurity/low birth weight and being first born were more prevalent in the ASD versus the control group. In addition, with increasing age children with ASD tended to have a proportionally smaller head circumference compared to their height. However, the effect of prematurity/low birth weight on head growth corrected for height was significantly different in ASD and control children: premature/low birth weight control children showed a disproportionate larger head circumference in relation to height during their first year of life, whereas this effect was absent in premature/low birth weight ASD children. This may suggest that the etiology of abnormal growth is potentially different in ASD and control children: where abnormal growth in control children is related to suboptimal conditions in the uterus, abnormal growth in ASD may be more strongly related to the causal factors that also increase the risk for ASD. However, prospective studies measuring growth and ASD characteristics in both premature/low birth weight and a terme children are necessary to support this conclusion.

The diagnosis of respiratory chain deficiencies (RCDs) is complicated and the need for a diagnostic biomarker or biosignature has been widely expressed. In this study, the metabolic profile of a selected group of 29 RCD patients, with a predominantly muscle disease phenotype, and 22 controls were investigated using targeted and untargeted analyses of three sub-sections of the human metabolome, including urinary organic acids and amino acids [measured by gas chromatography–mass spectrometry (GC–MS)], as well as acylcarnitines (measured by electrospray ionization tandem MS). Although MS technologies are highly sensitive and selective, they are restrictive by being applied only to sub-sections of the metabolome; an untargeted nuclear magnetic resonance (NMR) spectroscopy approach was therefore also included. After data reduction and pre-treatment, a biosignature comprising six organic acids (lactic, succinic, 2-hydroxyglutaric, 3-hydroxyisobutyric, 3-hydroxyisovaleric and 3-hydroxy-3-methylglutaric acids), six amino acids (alanine, glycine, glutamic acid, serine, tyrosine and α-aminoadipic acid) and creatine, was constructed from uni- and multivariate statistical analyses and verified by cross-validation. The results presented here provide the first proof-of-concept that the metabolomics approach is capable of defining a biosignature for RCDs. We postulate that the composite of organic acids ≈ amino acids > creatine > betaine > carnitines represents the basic biosignature for RCDs. Validated through a prospective study, this could offer an improved ability to assign individual patients to a group with defined RCD characteristics and improve case selection for biopsy procedures, especially in infants and children.

Contains fulltext : 108770.pdf (Publisher’s version ) (Closed access) We report on four families affected by a clinical presentation of complex hereditary spastic paraplegia (HSP) due to recessive mutations in DDHD2, encoding one of the three mammalian intracellular phospholipases A(1) (iPLA(1)). The core phenotype of this HSP syndrome consists of very early-onset (

Item does not contain fulltext OBJECTIVE: Congenital disorders of glycosylation (CDG) are a group of metabolic diseases due to defects in protein and lipid glycosylation. We searched for the primary defect in 3 children from 2 families with a severe neurological phenotype, including profound developmental delay, intractable epilepsy, progressive microcephaly, severe hypotonia with elevated blood creatine kinase levels, and early fatal outcome. There was clinical evidence of a muscular dystrophy-dystroglycanopathy syndrome, supported by deficient O-mannosylation by muscle immunohistochemistry. METHODS: Biochemical and molecular methods were combined to pinpoint the defect in the glycosylation pathway in the endoplasmic reticulum. RESULTS: Metabolic investigations revealed CDG-I, pointing to a defect in protein N-glycosylation in the endoplasmic reticulum. Analysis of lipid-linked oligosaccharides in fibroblasts showed accumulation of Dol-PP-GlcNAc(2) -Man(5) . DNA analysis revealed mutations in DPM2, 1 of the subunits of the dolichol-phosphate-mannose (DPM) synthase; the patient in the first family is compound heterozygous for 2 mutations (c.68A>G, predicting a missense mutation p.Y23C and c.4-1G>C, a splice mutation), whereas the patients in the second family are homozygous for the same missense mutation (c.68A>G, p.Y23C). INTERPRETATION: We describe a new CDG, due to a deficiency of DPM2. Hence, mutations have now been described in the genes for the 3 subunits of DPM: DPM1, DPM2, and DPM3, whereby DPM2-CDG links the congenital disorders of glycosylation to the congenital muscular dystrophies. ANN NEUROL 2012;72:550-558.

Although the combined presence of ataxia and pyramidal features has a long differential, the presence of a true spastic-ataxia as the predominant clinical syndrome has a rather limited differential diagnosis. Autosomal recessive ataxia of Charlevoix-Saguenay, late-onset Friedreich ataxia, and hereditary spastic paraplegia type 7 are examples of genetic diseases with such a prominent spastic-ataxic syndrome as the clinical hallmark. We review the various causes of spastic-ataxic syndromes with a focus on the genetic disorders, and provide a clinical framework, based on age at onset, mode of inheritance, and additional clinical features and neuroimaging signs, that could serve the diagnostic workup. Neurology (R) 2012;79:1507-1514

Item does not contain fulltext Uterine leiomyomas are benign solid tumors of mesenchymal origin which occur with an estimated incidence of up to 77% of all women of reproductive age. The majority of these tumors remains symptomless, but in about a quarter of cases they cause leiomyoma-associated symptoms including chronic pelvic pain, menorrhagia-induced anemia, and impaired fertility. As a consequence, they are the most common indication for pre-menopausal hysterectomy in the USA and Japan and annually translate into a multibillion dollar healthcare problem. Approximately 40% of these neoplasms present with recurring structural cytogenetic anomalies, including del(7)(q22), t(12;14)(q15;q24), t(1;2)(p36;p24), and anomalies affecting 6p21 and/or 10q22. Using positional cloning strategies, we and others previously identified HMGA1, HMGA2, RAD51L1, MORF, and, more recently, NCOA1 as primary target (fusion) genes associated with tumor initiation in four of these distinct cytogenetic subgroups. Despite the fact that the del(7)(q22) subgroup is the largest among leiomyomas, and was first described more than twenty years ago, the 7q22 leiomyoma target gene still awaits unequivocal identification. We here describe a positional cloning effort from two independent uterine leiomyomas, containing respectively a pericentric and a paracentric chromosomal inversion, both affecting band 7q22. We found that both chromosomal inversions target the cut-like homeobox 1 (CUX1) gene on chromosomal band 7q22.1 in a way which is functionally equivalent to the more frequently observed del(7q) cases, and which is compatible with a mono-allelic knock-out scenario, similar as was previously described for the cytogenetic subgroup showing chromosome 14q involvement.

Item does not contain fulltext This review article gives a state-of-the-art synopsis of current pathophysiological concepts in Sjogren-Larsson syndrome (SLS) mainly based upon original research data of the authors in one of the world's largest clinical SLS study cohorts. Clinical features are discussed in order of appearance, and diagnostic tests are set out to guide the clinician toward the diagnosis SLS. Furthermore, current and future treatment strategies are discussed to render a comprehensive review of the topic.

Item does not contain fulltext Nemaline myopathy and myofibrillar myopathy are heterogeneous myopathies that both comprise early-onset forms. We present two sisters from a consanguineous Iraqi Kurdish family with predominant axial and limb girdle weakness. Muscle biopsies showed features of both nemaline myopathy and myofibrillar myopathy. We performed homozygosity mapping in both siblings using an Affymetrix 250K Nspl SNP array. One of the overlapping homozygous regions harbored the gene CFL2. Because a mutation in CFL2 was identified in a family with nemaline myopathy, we performed sequence analysis of the gene and a novel homozygous missense mutation in exon 2 (c.19G>A, p.Val7Met) of CFL2 was identified in both siblings. CFL2 encodes the protein cofilin-2, which plays an important role in regulation of sarcomeric actin filaments. To our knowledge, this is the second family in which a mutation in CFL2 causes an autosomal recessive form of congenital myopathy with features of both nemaline and myofibrillar myopathy. Given the clinical variability and the multitude of histological features of congenital myopathies, CFL2 sequence analysis should be considered in patients presenting with an autosomal recessive form of congenital myopathy. 01 juli 2012

Item does not contain fulltext Recent studies have identified PRDM9, a zinc finger (ZF) protein, as a key regulator of meiotic recombination. As both recurrent genomic disorders and chromosomal non-disjunction are known to be associated with specific unusual patterns of recombination, we hypothesized a possible link between PRDM9 ZF variation and susceptibility to microdeletion syndromes and/or trisomy. We sequenced the PRDM9 ZF domain in 271 parents of patients with de novo microdeletions of known parental origin (velocardiofacial syndrome, the 17q21.31 microdeletion syndrome, Prader-Willi/Angelman syndrome and Williams-Beuren syndrome), and in 61 parents of individuals with a supernumerary X chromosome. We compared PRDM9 ZF genotype frequencies between parents in whose germ line the de novo rearrangement occurred and their spouses. We observed a significantly increased frequency (p = 0.006) of PRDM9 variants in parents who transmitted de novo 7q11.23 deletions to their offspring. These data suggest that certain PRDM9 alleles may be associated with an increased susceptibility to recurrent 7q11.23 microdeletions that cause Williams-Beuren syndrome. However, as the majority of parents who transmitted a de novo microdeletion/supernumerary X chromosome to their offspring have the common AA genotype, we conclude that none of the rearrangements we have studied are dependent on specific non-A PRDM9 alleles.

Item does not contain fulltext Creatine transporter deficiency is an X-linked disorder caused by mutations in the SLC6A8 gene. Currently, 38 pathogenic, including 15 missense variants, are reported. In this study, we report 33 novel, including 6 missense variants. To classify all known missense variants, we transfected creatine deficient fibroblasts with the SLC6A8 ORF containing one of the unique variants and tested their ability to restore creatine uptake. This resulted in the definitive classification of 2 non-disease associated and 19 pathogenic variants of which 3 have residual activity. Furthermore, we report the development and validation of a novel DHPLC method for the detection of heterozygous SLC6A8 variants. The method was validated by analysis of DNAs that in total contained 67 unique variants of which 66 could be detected. Therefore, this rapid screening method may prove valuable for the analysis of large cohorts of females with mild intellectual disability of unknown etiology, since in this group heterozygous SLC6A8 mutations may be detected. DHPLC proved also to be important for the detection of somatic mosaicism in mothers of patients who have a pathogenic mutation in SLC6A8. All variants reported in the present and previous studies are included in the Leiden Open Source Variant Database (LOVD) of SLC6A8 (www.LOVD.nl/SLC6A8).

Item does not contain fulltext Revertant mosaicism is an infrequently observed phenomenon caused by spontaneous correction of a pathogenic allele. We have observed such reversions caused by mitotic recombination of mutant TERC (telomerase RNA component) alleles in six patients from four families affected by dyskeratosis congenita (DC). DC is a multisystem disorder characterized by mucocutaneous abnormalities, dystrophic nails, bone-marrow failure, lung fibrosis, liver cirrhosis, and cancer. We identified a 4 nt deletion in TERC in a family with an autosomal-dominant form of DC. In two affected brothers without bone-marrow failure, sequence analysis revealed pronounced overrepresentation of the wild-type allele in blood cells, whereas no such skewing was observed in the other tissues tested. These observations suggest that this mosaic pattern might have resulted from somatic reversion of the mutated allele to the normal allele in blood-forming cells. SNP-microarray analysis on blood DNA from the two brothers indeed showed independent events of acquired segmental isodisomy of chromosome 3q, including TERC, indicating that the reversions must have resulted from mitotic recombination events. Subsequently, after developing a highly sensitive method of detecting mosaic homozygosity, we have found four additional cases with a mosaic-reversion pattern in blood cells; these four cases are part of a cohort of 17 individuals with germline TERC mutations. This shows that revertant mosaicism is a recurrent event in DC. This finding has important implications for improving diagnostic testing and understanding the variable phenotype of DC.

Item does not contain fulltext CONTEXT: Newborn screening for cystic fibrosis (CF) is included in many routine programmes but current strategies have considerable drawbacks, such as false-positive tests, equivocal diagnosis and detection of carriers. OBJECTIVE: To assess the test performance of two newborn screening strategies for CF. DESIGN, SETTING AND PARTICIPANTS: In 2008 and 2009, CF screening was added to the routine screening programme as a prospective study in part of The Netherlands. INTERVENTIONS: Two strategies were performed in all newborns. In the first strategy, concentrations of immunoreactive trypsinogen (IRT) and pancreatitis-associated protein (PAP) were measured. In the second method, samples with IRT >/=60 mug/litre were analysed for 36 CFTR mutations, followed by sequencing when a single mutation was detected. Tests were positive only with two identified CFTR mutations. MAIN OUTCOME: Sensitivity, specificity and positive predictive value (PPV) of both screening strategies. RESULTS: 145,499 infants were screened. The IRT/PAP approach showed a sensitivity of 95.0%, a specificity of 99.897% and a PPV of 12.3%. Test properties for the IRT/DNA/sequencing strategy were respectively 100%, 100% and 64.9%. Combining both strategies (IRT/PAP/DNA/sequencing) led to a sensitivity of 95.0%, a specificity of 100% and a PPV of 87.5%. CONCLUSION: In conclusion, all strategies performed well. Although there was no statistically significant difference in test performance, the IRT/DNA/sequencing strategy detected one infant that was missed by IRT/PAP (/DNA/sequencing). IRT/PAP may be the optimal choice if the use of DNA technology must be avoided. If identification of carriers and equivocal diagnosis is considered an important disadvantage, IRT/PAP/DNA/sequencing may be the best choice. 01 april 2012

Item does not contain fulltext Congenital disorders of glycosylation type I (CDG-I) form a growing group of recessive neurometabolic diseases. Identification of disease genes is compromised by the enormous heterogeneity in clinical symptoms and the large number of potential genes involved. Until now, gene identification included the sequential application of biochemical methods in blood samples and fibroblasts. In genetically unsolved cases, homozygosity mapping has been applied in consanguineous families. Altogether, this time-consuming diagnostic strategy led to the identification of defects in 17 different CDG-I genes. Here, we applied whole-exome sequencing (WES) in combination with the knowledge of the protein N-glycosylation pathway for gene identification in our remaining group of six unsolved CDG-I patients from unrelated non-consanguineous families. Exome variants were prioritized based on a list of 76 potential CDG-I candidate genes, leading to the rapid identification of one known and two novel CDG-I gene defects. These included the first X-linked CDG-I due to a de novo mutation in ALG13, and compound heterozygous mutations in DPAGT1, together the first two steps in dolichol-PP-glycan assembly, and mutations in PGM1 in two cases, involved in nucleotide sugar biosynthesis. The pathogenicity of the mutations was confirmed by showing the deficient activity of the corresponding enzymes in patient fibroblasts. Combined with these results, the gene defect has been identified in 98% of our CDG-I patients. Our results implicate the potential of WES to unravel disease genes in the CDG-I in newly diagnosed singleton families.

Contains fulltext : 108772.pdf (Publisher’s version ) (Open Access) Walker-Warburg syndrome (WWS) is an autosomal recessive multisystem disorder characterized by complex eye and brain abnormalities with congenital muscular dystrophy (CMD) and aberrant a-dystroglycan glycosylation. Here we report mutations in the ISPD gene (encoding isoprenoid synthase domain containing) as the second most common cause of WWS. Bacterial IspD is a nucleotidyl transferase belonging to a large glycosyltransferase family, but the role of the orthologous protein in chordates is obscure to date, as this phylum does not have the corresponding non-mevalonate isoprenoid biosynthesis pathway. Knockdown of ispd in zebrafish recapitulates the human WWS phenotype with hydrocephalus, reduced eye size, muscle degeneration and hypoglycosylated a-dystroglycan. These results implicate ISPD in a-dystroglycan glycosylation in maintaining sarcolemma integrity in vertebrates.

A female patient, nine years of age, is reported with a history characterized by delay of psychomotor and speech development, mild to moderate intellectual disability and persistent sleep disturbances since the age of two. The patient showed facial dysmorphisms, a pectus excavatum and a sandal gap. Apart from lowered intelligence, neuropsychological functioning disclosed impaired attentional capacities and executive control as well as weak motor skills. Genome wide SNP array analysis revealed a 3.57 Mb de novo microdeletion in band q12.3 of chromosome 8. The long lasting sleep disorders turned out to originate from a rare juvenile epilepsy, continuous spike-waves during slow sleep (CSWS) syndrome, that includes the electrical status epilepticus in sleep (ESES) phenomenon. MRI-scanning of the brain showed no abnormalities. To the authors knowledge, this is the first report of a de novo 8q12.3q13.2 microdeletion syndrome that presents with ESES/CSWS. (C) 2011 Elsevier Masson SAS. All rights reserved.

Item does not contain fulltext Aim The long-term outcome of the Dutch pyridoxine-dependent epilepsy cohort and correlations between patient characteristics and follow-up data were retrospectively studied. Method Fourteen patients recruited from a national reference laboratory were included (four males, 10 females, from 11 families; median age at assessment 6y; range 2y 6mo-16y). The following data were retrieved: sex; age at seizure onset; age at the start of pyridoxine therapy; level of urinary alpha-aminoadipic semialdehyde; antiquitin mutations; developmental milestones; evaluation of neurocognitive functioning and school career; magnetic resonance imaging (MRI) and electroencephalography (EEG) assessments. Results Pyridoxine was started antenatally in two children, in the first week of life in five, in the first month of life in three, or after the first month of life (range 2.5-8mo) in four. No child was physically disabled; however, only five walked at 2 years of age. Mental development was delayed in most: median IQ or developmental index was 72 (SD 19). Pyridoxine monotherapy controlled seizures in 10 of 14 children, whereas four needed additional antiepileptic drugs. Seizure persistence, antiepileptic drugs (other than pyridoxine), EEG background, and epileptiform activity were not associated with outcome. On neonatal MRI, structural and white matter abnormalities occurred in five of eight children; on follow-up, the number of abnormal MRIs was increased. Delayed initiation of pyridoxine medication and corpus callosum abnormalities were significantly associated with unfavourable neurodevelopmental outcome, but normal follow-up imaging did not predict a good outcome. Interpretation Outcome of patients with pyridoxine-dependent epilepsy remains poor. Individual outcome cannot be predicted by the evaluated characteristics. We suggest that collaborated research in structured settings could help to improve treatment strategies and outcome for pyridoxine-dependent epilepsy. 01 september 2012

Objectives The purpose of this study was describe the cardiovascular phenotype of the aneurysms-osteoarthritis syndrome (AOS) and to provide clinical recommendations. Background AOS, caused by pathogenic SMAD3 variants, is a recently described autosomal dominant syndrome characterized by aneurysms and arterial tortuosity in combination with osteoarthritis. Methods AOS patients in participating centers underwent extensive cardiovascular evaluation, including imaging, arterial stiffness measurements, and biochemical studies. Results We included 44 AOS patients from 7 families with pathogenic SMAD3 variants (mean age: 42 +/- 17 years). In 71%, an aortic root aneurysm was found. In 33%, aneurysms in other arteries in the thorax and abdomen were diagnosed, and in 48%, arterial tortuosity was diagnosed. In 16 patients, cerebrovascular imaging was performed, and cerebrovascular abnormalities were detected in 56% of them. Fifteen deaths occurred at a mean age of 54 +/- 15 years. The main cause of death was aortic dissection (9 of 15; 60%), which occurred at mildly increased aortic diameters (range: 40 to 63 mm). Furthermore, cardiac abnormalities were diagnosed, such as congenital heart defects (6%), mitral valve abnormalities (51%), left ventricular hypertrophy (19%), and atrial fibrillation (22%). N-terminal brain natriuretic peptide (NT-proBNP) was significantly higher in AOS patients compared with matched controls (p < 0.001). Aortic pulse wave velocity was high-normal (9.2 +/- 2.2 m/s), indicating increased aortic stiffness, which strongly correlated with NT-proBNP (r = 0.731, p = 0.005). Conclusions AOS predisposes patients to aggressive and widespread cardiovascular disease and is associated with high mortality. Dissections can occur at relatively mildly increased aortic diameters; therefore, early elective repair of the ascending aorta should be considered. Moreover, cerebrovascular abnormalities were encountered in most patients. (J Am Coll Cardiol 2012;60:397-403) (C) 2012 by the American College of Cardiology Foundation

Item does not contain fulltext Fibroblast growth factor receptors (FGFRs) play critical roles in craniofacial and skeletal development via multiple signaling pathways including MAPK, PI3K/AKT, and PLC-?. FGFR-mediated signaling is modulated by several regulators. Proteins with leucine-rich repeat (LRR) and/or immunoglobulin (IG) superfamily domains have been suggested to interact with FGFRs. In addition, fibronectin leucine-rich repeat transmembrane protein 3 (FLRT3) has been shown to modulate the FGFR-mediated signaling via the fibronectin type III (FNIII) domain. Therefore proteins with LRR, IG, and FNIII are candidate regulators of the FGFRs. Here we identify leucine-rich repeat, immunoglobulin-like and transmembrane domain 3 (LRIT3) as a regulator of the FGFRs.

Contains fulltext : 108685.pdf (Publisher’s version ) (Closed access) We present two new patients with the recently described mitochondrial m.3242G > A mutation. Although the mutation is situated next to the well known m.3243A > G mutation, the most common alteration associated with mitochondrial myopathy, encephalopathy, lactic acidosis, and stroke-like episodes (MELAS) syndrome, the clinical presentation is quite different, but characteristic. All three m.3242G > A patients presented in the neonatal period with hypertrophic and dilated cardiomyopathy, generalized muscle hypotonia and lactic acidosis. Two additionally had creatine kinase elevation, renal tubular acidosis/dysfunction and showed a mild clinical course with a favourable psychomotor development. The third patient had more neurological involvement and died in infancy. The mutation occurred de novo in the two patients where maternal investigations were performed. The combination of hypertrophic cardiomyopathy and renal tubular acidosis/renal tubular dysfunction is clinically distinctive and may represent a separate entity.

Item does not contain fulltext Cone-rod dystrophy (CRD) and retinitis pigmentosa (RP) are clinically and genetically overlapping heterogeneous retinal dystrophies. By using homozygosity mapping in an individual with autosomal-recessive (ar) RP from a consanguineous family, we identified three sizeable homozygous regions, together encompassing 46 Mb. Next-generation sequencing of all exons, flanking intron sequences, microRNAs, and other highly conserved genomic elements in these three regions revealed a homozygous nonsense mutation (c.497T>A [p.Leu166( *)]) in C8orf37, located on chromosome 8q22.1. This mutation was not present in 150 ethnically matched control individuals, single-nucleotide polymorphism databases, or the 1000 Genomes database. Immunohistochemical studies revealed C8orf37 localization at the base of the primary cilium of human retinal pigment epithelium cells and at the base of connecting cilia of mouse photoreceptors. C8orf37 sequence analysis of individuals who had retinal dystrophy and carried conspicuously large homozygous regions encompassing C8orf37 revealed a homozygous splice-site mutation (c.156-2A>G) in two siblings of a consanguineous family and homozygous missense mutations (c.529C>T [p.Arg177Trp]; c.545A>G [p.Gln182Arg]) in siblings of two other consanguineous families. The missense mutations affect highly conserved amino acids, and in silico analyses predicted that both variants are probably pathogenic. Clinical assessment revealed CRD in four individuals and RP with early macular involvement in two individuals. The two CRD siblings with the c.156-2A>G mutation also showed unilateral postaxial polydactyly. These results underline the importance of disrupted ciliary processes in the pathogenesis of retinal dystrophies.

Item does not contain fulltext PURPOSE: Hereditary spastic paraplegias compose a group of neurodegenerative disorders with a large clinical and genetic heterogeneity. Among the autosomal recessive forms, spastic paraplegia type 11 is the most common. METHODS: To better understand the spastic paraplegia type 11 mutation spectrum, we studied a group of 54 patients with hereditary spastic paraplegia. Mutation screening was performed by PCR amplification of SPG11 coding regions and intron boundaries, followed by sequencing. For the detection of large gene rearrangements, we performed multiplex ligation-dependent probe amplification. RESULTS: We report 13 families with spastic paraplegia type 11 carrying either novel or previously identified mutations. We describe a complex entire SPG11 rearrangement and show that large gene rearrangements are frequent among patients with spastic paraplegia type 11. Moreover, we mapped the deletion breakpoints of three different large SPG11 deletions and provide evidence for Alu microhomology-mediated exon deletion. CONCLUSION: Our analysis shows that the high number of repeated elements in SPG11 together with the presence of recombination hotspots and the high intrinsic instability of the 15q locus all contribute toward making this genomic region more prone to large gene rearrangements. These findings enlarge the amount of data relating repeated elements with neurodegenerative disorders and highlight their importance in human disease and genome evolution. 01 januari 2012

Background— Arrhythmogenic ventricular remodeling is hallmarked by both reduced gap junction expression and increased collagen deposition. We hypothesized that reduced connexin43 (Cx43) expression is responsible for enhanced fibrosis in the remodeled heart, resulting in an arrhythmogenic substrate. Therefore, we investigated the effect of normal or reduced Cx43 expression on the formation of fibrosis in a physiological (aging) and pathophysiological (transverse aortic constriction [TAC]) mouse model. Methods and Results— The Cx43 fl/fl and Cx43 CreER(T)/fl mice were aged 18 to 21 months or, at the age of 3 months, either TAC or sham operated and euthanized after 16 weeks. Epicardial activation mapping of the right and left ventricles was performed on Langendorff perfused hearts. Sustained ventricular arrhythmias were induced in 0 of 11 aged Cx43 fl/fl and 10 of 15 Cx43 Cre-ER(T)/fl mice ( P CreER(T)/fl compared with aged Cx43 fl/fl mice, whereas collagen deposition was significantly increased from 1.1±0.2% to 7.4±1.3%. Aged Cx43 CreER(T)/fl mice with arrhythmias had significantly higher levels of fibrosis and conduction heterogeneity than aged Cx43 CreER(T)/fl mice without arrhythmias. The TAC operation significantly increased fibrosis in control compared with sham (4.0±1.2% versus 0.4±0.06%), but this increase was significantly higher in Cx43 CreER(T)/fl mice (10.8±1.4%). Discoidin domain receptor 2 expression was unchanged, but procollagen peptide I and III expression and collagen type 1α2 mRNA levels were higher in TAC–operated Cx43HZ mice. Conclusions— Reduced cellular coupling results in more excessive collagen deposition during aging or pressure overload in mice due to enhanced fibroblast activity, leading to increased conduction in homogeneity and proarrhythmia.

Objective: To describe a novel pattern of magnetic resonance imaging (MRI) abnormalities as well as the associated clinical and laboratory findings. Design: The MRIs of more than 3000 patients with an unclassified leukoencephalopathy were systematically reviewed. Clinical and laboratory data were retrospectively collected. Setting: University hospital. Patients: Seven patients (3 male) shared similar MRI abnormalities and clinical features. Main Outcome Measures: Pattern of MRI abnormalities and clinical and laboratory findings. Results: The MRIs showed signal abnormalities of the deep cerebral white matter, corpus callosum, thalamus, basal ganglia, brainstem, and cerebellar white matter between the ages of 9 months and 2 years. On follow-up, abnormalities gradually improved. Clinical regression occurred in the second half-year of life with spasticity and loss of milestones. From the second year on, clinical improvement occurred. So far, no second episode of regression has happened. Lactate levels were elevated during clinical regression. Conclusion: These patients represent a single novel leukoencephalopathy, probably caused by a mitochondrial defect. Arch Neurol. 2012; 69(6): 718-722. Published online February 6, 2012. doi:10.1001/archneurol.2011.1048

Item does not contain fulltext Background: In order to correctly assess the efficacy of therapy or diet in intervention studies on the activity of cystathionine beta-synthase (CBS) a sensitive analytical method is necessary. Methods: An electrospray LC-MS/MS method preceded by a solid phase extraction step was developed for the measurement of CBS activity in cell extracts. Nonafluoropentanoic acid was used as an ionpair to provide the underivatized cystathionine the desired retention on a C18 column. Results: A detection limit of 50pmol cystathionine/h/mg protein was achieved. In fibroblasts, intra- and inter-assay CVs for the CBS activity were 5.2% and 14.7%, respectively. A K(m) value of 8mumol/L for homocysteine, and 2.5mumol/L for serine was calculated. In fibroblasts wildtype, heterozygous, and homozygous CBS activity ranges measured were 8.5-27.0, 4.2-13.4, 0.0-0.7nmol/hxmg protein, respectively. The method was applied to a study where rats were fed 2 diets. Increase of dietary methionine (7.7 versus 3.8mg/kg methionine) significantly increased the CBS activity in rat liver lysates from a median of 58.0 to a median of 71.5 (P=0.037)nmol/hxmg protein. In a lymphoblasts cell culture experiment, the addition of Hcy to the culture media increased the activity of CBS 3 fold. Conclusion: This LC-MS/MS is able to diagnose CBS deficiency at the enzyme level, and can accurately measure the effect diets or therapy might have on the CBS activity in a variety of cell types.