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Nephronophthisis is one of the leading genetic causes of end-stage renal disease in childhood. Early diagnostics and prognostics for nephronophthisis are currently limited. We aimed to identify non-invasive protein biomarkers for nephronophthisis in urinary extracellular vesicles. Extracellular vesicles were isolated from urine of 12 patients with a nephronophthisis-related ciliopathy and 12 age- and gender-matched controls, followed by in-depth label-free LC-MS/MS proteomics analysis of gel fractionated extracellular vesicle proteins. Supervised cluster analysis of proteomic profiles separated patients from controls. We identified 156 differentially expressed proteins with fold change ≥4 in patients compared to controls (P

BackgroundObesity is a global and severe health problem. Due to genetic heterogeneity, the identification of genetic defects in patients with obesity can be time consuming and costly. Therefore, we developed a custom diagnostic targeted next-generation sequencing (NGS)-based analysis to simultaneously identify mutations in 52 obesity-related genes. The aim of this study was to assess the diagnostic yield of this approach in patients with suspected genetic obesity.MethodsDNA of 1230 patients with obesity (median BMI adults 43.6 kg/m2; median body mass index-SD children +3.4 SD) was analysed in the genome diagnostics section of the Department of Genetics of the UMC Utrecht (The Netherlands) by targeted analysis of 52 obesity-related genes.ResultsIn 48 patients pathogenic mutations confirming the clinical diagnosis were detected. The majority of these were observed in theMC4Rgene (18/48). In an additional 67 patients a probable pathogenic mutation was identified, necessitating further analysis to confirm the clinical relevance.ConclusionsNGS-based gene panel analysis in patients with obesity led to a definitive diagnosis of a genetic obesity disorder in 3.9% of obese probands, and a possible diagnosis in an additional 5.4% of obese probands. The highest yield was achieved in a selected paediatric subgroup, establishing a definitive diagnosis in 12 out of 164 children with severe early onset obesity (7.3%). These findings give a realistic insight in the diagnostic yield of genetic testing for patients with obesity and could help these patients to receive (future) personalised treatment.

Dravet syndrome is a severe genetic encephalopathy, caused by pathogenic variants inDeep sequencing ofSequence coverage of at least 100× at the location of the corresponding pathogenic variant was reached for 80 parent couples. The variant ratio was significantly higher than the established noise ratio in eight parent couples, of which four (5%) were regarded as true mosaics, based on ddPCR results. The false positive rate of smMIP analysis without ddPCR was therefore 50%. Three of these variants had previously been considered de novo in the proband by Sanger sequencing.smMIP technology combined withnext generation sequencing (NGS) performs better than Sanger sequencing in the detection of parental mosaicism. Because parental mosaicism has important implications for genetic counselling and recurrence risks, we stress the importance of implementing high-sensitivity NGS-based assays in standard diagnostics.

ObjectivePathogenic variants in SCN1A can give rise to extremely variable disease severities that may be indistinguishable at their first presentation. We aim to find clinical features that can help predict the evolution of seizures into Dravet syndrome and clinical features that predict cognitive outcome in Dravet syndrome. We specifically investigate the role of contraindicated medication (CIM) as a possible modifier of cognitive decline.MethodsA cohort of 164 Dutch participants with SCN1A-related seizures was evaluated. Clinical data were collected from medical records and semistructured telephone interviews. Cognitive function was classified by a child neurologist, neuropsychologist, and clinical geneticist. Several clinical variables, including duration of CIM use in the first 5years of disease, were evaluated in univariate and multivariate analyses.ResultsA longer duration of CIM use in the first 5years after seizure onset was significantly associated with a worse cognitive outcome at time of inclusion, and with lower interpolated intelligence quotient/developmental quotient scores after the first 5years of disease in Dravet syndrome patients. CIM use remained a significant predictor for cognitive outcome in a multivariate regression model, as did age at the first observation of developmental delay and age at first afebrile seizure. Age at first afebrile seizure was the most accurate predictor for evolution of seizures into Dravet syndrome for the complete cohort.SignificanceOur data suggest that a longer CIM use in the first 5years of disease can have negative effects on cognitive outcome in Dravet syndrome. An early diagnosis is essential to avoid these drugs. Furthermore, we identified age at first afebrile seizure as an important predictor for evolution of seizures into Dravet syndrome and for the severity of Dravet syndrome, which can be used to counsel parents of young patients with SCN1A-related seizures.

Objective: Phenotypes caused by de novo SCN1A pathogenic variants are very variable, ranging from severely affected patients with Dravet syndrome to much milder genetic epilepsy febrile seizures plus cases. The most important determinant of disease severity is the type of variant, with variants that cause a complete loss of function of the SCN1A protein (α-subunit of the neuronal sodium channel Nav1.1) being detected almost exclusively in Dravet syndrome patients. However, even within Dravet syndrome disease severity ranges greatly, and consequently other disease modifiers must exist. A better prediction of disease severity is very much needed in daily practice to improve counseling, stressing the importance of identifying modifying factors in this patient group. We evaluated 128 participants with de novo, pathogenic SCN1A variants to investigate whether mosaicism, caused by postzygotic mutation, is a major modifier in SCN1A-related epilepsy. Methods: Mosaicism was investigated by reanalysis of the pathogenic SCN1A variants using single molecule molecular inversion probes and next generation sequencing with high coverage. Allelic ratios of pathogenic variants were used to determine whether mosaicism was likely. Selected mosaic variants were confirmed by droplet digital polymerase chain reaction and sequencing of different tissues. Developmental outcome was classified based on available data on intelligence quotient and school functioning/education. Results: Mosaicism was present for 7.5% of de novo pathogenic SCN1A variants in symptomatic patients. Mosaic participants were less severely affected than nonmosaic participants if only participants with truncating variants are considered (distribution of developmental outcome scores, Mann-Whitney U, P =.023). Significance: Postzygotic mutation is a common phenomenon in SCN1A-related epilepsies. Participants with mosaicism have on average milder phenotypes, suggesting that mosaicism can be a major modifier of SCN1A-related diseases. Detection of mosaicism has important implications for genetic counseling and can be achieved by deep sequencing of unique reads.

Background Nephronophthisis is an autosomal recessive ciliopathy and important cause of end-stage renal disease (ESRD) in children and young adults. Diagnostic delay is frequent. This study investigates clinical characteristics, initial symptoms, and genetic defects in a cohort with nephronophthisis-related ciliopathy, to improve early detection and genetic counseling. Methods Forty patients from 36 families with nephronophthisis-related ciliopathy were recruited at university medical centers and online. Comprehensive clinical and genotypic data were recorded. Patients without molecular diagnosis were offered genetic analysis. Results Of 40 patients, 45% had isolated nephronophthisis, 48% syndromic diagnosis, and 7% nephronophthisis with extrarenal features not constituting a recognizable syndrome. Patients developed ESRD at median 13 years (range 5–47). Median age of symptom onset was 9 years in both isolated and syndromic forms (range 5–26 vs. 5–33). Common presenting symptoms were fatigue (42%), polydipsia/polyuria (33%), and hypertension (21%). Renal ultrasound showed small-to-normal-sized kidneys, increased echogenicity (65%), cysts (43%), and abnormal corticomedullary differentiation (32%). Renal biopsies in eight patients showed nonspecific signs of chronic kidney disease (CKD). Twenty-three patients (58%) had genetic diagnosis upon inclusion. Thirteen of those without a genetic diagnosis gave consent for genetic testing, and a cause was identified in five (38%). Conclusions Nephronophthisis is genetically and phenotypically heterogeneous and should be considered in children and young adults presenting with persistent fatigue and polyuria, and in all patients with unexplained CKD. As symptom onset can occur into adulthood, presymptomatic monitoring of kidney function in syndromic ciliopathy patients should continue until at least age 30. Electronic supplementary material The online version of this article (10.1007/s00467-018-3958-7) contains supplementary material, which is available to authorized users.

Background: Autosomal dominant tubulointerstitial kidney disease subtype HNF1B (ADTKD-HNF1B) is caused by a mutation in hepatocyte nuclear factor 1 homeobox beta (HNF1B). Although 50-60% of ADTKD-HNF1B patients develop hypomagnesemia, HNF1B mutations are mainly identified in patients with structural kidney defects or diabetes. Cases: The current case series describes 3 patients in whom hypomagnesemia proved to be the first clinical manifestation of ADTKD-HNF1B. All patients presented with hypomagnesemia with a high fractional excretion of Mg2+ and hypocalciuria. Exome sequencing performed for analysis of known and candidate hypomagnesaemia genes and subsequent multiplex ligation-dependent probe amplification analysis revealed a large deletion at the chromosome 17q12. Follow-up analysis showed increased blood glucose concentrations in all 3 patients and high hemoglobin A1c levels in 2 out of 3 patients, indicating diabetes mellitus. Although all patients suffered from mild renal insufficiency, only 1 of the 3 patients was shown to have renal cysts on CT. Conclusion: The prevalence of HNF1B mutations and the relative contribution of hypomagnesemia to its symptoms are underestimated. Therefore, patients with primary renal magnesium wasting should be tested for HNF1B mutations to ensure early detection and optimal management of ADTKD-HNF1B.

Type 2 collagen disorders encompass a diverse group of skeletal dysplasias that are commonly associated with orthopedic, ocular, and hearing problems. However, the frequency of many clinical features has never been determined. We retrospectively investigated the clinical, radiological, and genotypic data in a group of 93 patients with molecularly confirmed SEDC or a related disorder. The majority of the patients (80/93) had short stature, with radiological features of SEDC (n=64), others having SEMD (n=5), Kniest dysplasia (n=7), spondyloperipheral dysplasia (n=2), or Torrance-like dysplasia (n=2). The remaining 13 patients had normal stature with mild SED, Stickler-like syndrome or multiple epiphyseal dysplasia. Over 50% of the patients had undergone orthopedic surgery, usually for scoliosis, femoral osteotomy or hip replacement. Odontoid hypoplasia was present in 56% (95% CI 38-74) and a correlation between odontoid hypoplasia and short stature was observed. Atlanto-axial instability, was observed in 5 of the 18 patients (28%, 95% CI 10-54) in whom flexion-extension films of the cervical spine were available; however, it was rarely accompanied by myelopathy. Myopia was found in 45% (95% CI 35-56), and retinal detachment had occurred in 12% (95% CI 6-21; median age 14 years; youngest age 3.5 years). Thirty-two patients complained of hearing loss (37%, 95% CI 27-48) of whom 17 required hearing aids. The ophthalmological features and possibly also hearing loss are often relatively frequent and severe in patients with splicing mutations. Based on clinical findings, age at onset and genotype-phenotype correlations in this cohort, we propose guidelines for the management and follow-up in this group of disorders. (c) 2015 Wiley Periodicals, Inc.

Introduction:: The research into the question ‘why does my child have this anomaly?’, or the aetiology of paediatric urology anomalies, started in 2002. For this purpose a project plan was submitted to the Internal Review Board on Human Studies to collect blood and additional information with a time schedule of 25 years. These data are currently stored in the Radboudumc AGORA data- en biobank. Methods:: The basic collection of the Radboudumc AGORA data- and biobank data consists of the clinical diagnostic information, questionnaires and genetic material. To collect, use and distribute these data and materials an information brochure, informed consent forms, a collaboration contract, a study request form and a material transfer agreement (MTA) are part of the project. The AGORA program is a collaboration between clinical departments, epidemiology and genetics. Results:: In 2014 there is a collection of biobank data of 1,836 urological and 616 uro-nefrological patients. Most research has been performed into the anomaly hypospadias and recently an international collaboration concerning these patients was established. Furthermore studies were performed concerning undescended testis, vesico-ureteral reflux and bladder exstrophy. A summary of the different publications is hereby presented. Conclusion:: Different studies have been performed concerning the etiological aspects of hypospadias from which known and new findings were discovered. Currently, the aetiology cannot be totally explained by the environmental and genetic factors found, but step by step new associations are being found. In small studies and within (inter)national collaborations new results were also found for undescended testis, vesico-ureteral reflux and bladder exstrophy. For this kind of studies a pleasant and productive collaboration with all people involved, clear appointments and contracts, and good logistic procedures play an essential role.

Objective:To study the effect of vaccination-associated seizure onset on disease course and estimate the risk of subsequent seizures after infant pertussis combination and measles, mumps, and rubella (MMR) vaccinations in Dravet syndrome (DS).Methods:We retrospectively analyzed data from hospital medical files, child health clinics, and the vaccination register for children with DS and pathogenic SCN1A mutations. Seizures within 24 hours after infant whole-cell, acellular, or nonpertussis combination vaccination or within 5 to 12 days after MMR vaccination were defined as "vaccination-associated." Risks of vaccination-associated seizures for the different vaccines were analyzed in univariable and in multivariable logistic regression for pertussis combination vaccines and by a self-controlled case series analysis using parental seizure registries for MMR vaccines. Disease courses of children with and without vaccination-associated seizure onset were compared.Results:Children who had DS (n = 77) with and without vaccination-associated seizure onset (21% and 79%, respectively) differed in age at first seizure (median 3.7 vs 6.1 months, p Conclusions:Our results suggest that vaccination-associated earlier seizure onset does not alter disease course in DS, while the risk of subsequent vaccination-associated seizures is probably vaccine-specific.

Pathogenic variants in the PCDH19 gene are associated with epilepsy, intellectual disability (ID) and behavioural disturbances. Only heterozygous females and mosaic males are affected, likely due to a disease mechanism named cellular interference. Until now, only four affected mosaic male patients have been described in literature. Here, we report five additional male patients, of which four are older than the oldest patient reported so far. All reported patients were selected for genetic testing because of developmental delay and/or epilepsy. Custom-targeted next generation sequencing gene panels for epilepsy genes were used. Clinical data were collected from medical records. All patients were mosaic in blood for likely pathogenic variants in the PCDH19 gene. In most, clinical features were very similar to the female phenotype, with normal development before seizure onset, which occurred between 5 and 10 months of age, clustering of seizures and sensitivity to fever. Four out of five patients had mild to severe ID and behavioural problems. We reaffirm the similarity between male and female PCDH19-related phenotypes, now also in a later phase of the disorder (ages 10–14 years). Electronic supplementary material The online version of this article (doi:10.1007/s10048-017-0517-5) contains supplementary material, which is available to authorized users.

Rare kidney diseases encompass at least 150 different conditions, most of which are inherited. Although individual rare kidney diseases raise specific issues, as a group these rare diseases can have overlapping challenges in diagnosis and treatment. These challenges include small numbers of affected patients, unidentified causes of disease, lack of biomarkers for monitoring disease progression, and need for complex care. To address common clinical and patient issues among rare kidney diseases, the KDIGO Controversies Conference entitled, Common Elements in Rare Kidney Diseases, brought together a panel of multidisciplinary clinical providers and patient advocates to address five central issues for rare kidney diseases. These issues encompassed diagnostic challenges, management of kidney functional decline and progression of chronic kidney disease, challenges in clinical study design, translation of advances in research to clinical care, and provision of practical and integrated patient support. Thus, by a process of consensus, guidance for addressing these challenges was developed and is presented here.

Congenital anomalies of the kidney and urinary tract (CAKUT) constitute one of the most common developmental diseases in humans; however, the cause for most patients remains unknown. Efforts to identify novel genetic causes for CAKUT through next-generation sequencing techniques have led to the discovery of new genes and risk factors. Concomitantly, these same efforts have generated large gene candidate lists requiring individual functional characterization. Appropriate model systems are needed to assess the functionality of genes and pathogenicity of genetic variants discovered in CAKUT patients. In this review, we discuss how cellular, animal, and personal (human) models are being used to study CAKUT candidate genes and what their major advantages and disadvantages are with respect to relevance and throughput. (C) 2014 S. Karger AG, Basel

The advent of next-generation sequencing technologies has enabled genetic nephrology research to move beyond single gene analysis to the simultaneous investigation of hundreds of genes and entire pathways. These new sequencing approaches have been used to identify and characterize causal factors that underlie inherited heterogeneous kidney diseases such as nephronophthisis and congenital anomalies of the kidney and urinary tract. In this Review, we describe the development of next-generation sequencing in basic and clinical research and discuss the implementation of this novel technology in routine patient management. Widespread use of targeted and nontargeted approaches for gene identification in clinical practice will require consistent phenotyping, appropriate disease modelling and collaborative efforts to combine and integrate data analyses. Next-generation sequencing is an exceptionally promising technique that has the potential to improve the management of patients with inherited kidney diseases. However, identifying the molecular mechanisms that lead to renal developmental disorders and ciliopathies is difficult. A major challenge in the near future will be how best to integrate data obtained using next-generation sequencing with personalized medicine, including use of high-throughput disease modelling as a tool to support the clinical diagnosis of kidney diseases.

Following initial glucocorticoid treatment, the clinical course in children with nephrotic syndrome is highly variable. Intrinsic sensitivity to glucocorticoids might be a determinant of this variability. Functional polymorphisms of the glucocorticoid receptor gene NR3C1 have been associated with either relatively impaired (GR-9β) or increased (BclI) glucocorticoid sensitivity. Here, in a prospective, well-defined cohort of children with nephrotic syndrome, we evaluated both carriage of GR-9β+TthIII-1 and BclI haplotypes in 113 children and a dexamethasone suppression test in 90 children in relation to their clinical outcome over a median follow-up of 4.4 years. Carriers of GR-9β+TthIII-1 had a significantly higher incidence of steroid dependence 13/25 (52%) compared with noncarriers 19/75 (25%) with a hazard ratio adjusted for gender, age, and descent of 3.04 with 95% confidence interval 1.37-6.74. Both first and frequent relapses happened significantly more often in GR-9β+TthIII-1 carriers than in noncarriers. There were no significant differences in therapeutic outcomes between carriers and noncarriers of the BclI haplotype. Results of the dexamethasone test showed no associations with clinical outcome. Thus, the GR-9β+TthIII-1 haplotype of the glucocorticoid receptor gene offers new insights into the clinical course of children with nephrotic syndrome.

Bartter’s syndrome is a rare, genetically heterogeneous disorder characterized by renal salt wasting, hypokalemic metabolic alkalosis, and secondary hyperaldosteronism with normal to low blood pressure. Two distinct presentations of the syndrome exist: antenatal Bartter’s syndrome and classical Bartter’s syndrome. Both forms are inherited as autosomal recessive traits. Patients with antenatal Bartter’s syndrome may carry loss-of-function mutations in the genes encoding the furosemide-sensitive sodium–potassium–chloride cotransporter NKCC2, the inwardly rectifying potassium channel ROMK, or the chloride channel β-subunit barttin.1 The concerted action of these transporters in the renal thick ascending limb of the loop of Henle ensures transcellular sodium chloride reabsorption in . . .

X-linked sideroblastic anemia (XLSA) is the most common form of congenital sideroblastic anemia. In affected males, it is uniformly associated with partial loss-of-function missense mutations in the erythroid-specific heme biosynthesis protein 5-aminolevulinate synthase 2 (ALAS2). Here, we report five families with XLSA owing to mutations in a GATA transcription factor binding site located in a transcriptional enhancer element in intron 1 of the ALAS2 gene. As such, this study defines a new class of mutations that should be evaluated in patients undergoing genetic testing for a suspected diagnosis of XLSA.

Background/Aims: Mutations in the inwardly-rectifying K+-channel KCNJ10/Kir4.1 cause autosomal recessive EAST syndrome (epilepsy, ataxia, sensorineural deafness and tubulopathy). KCNJ10 is expressed in the distal convoluted tubule of the kidney, stria vascularis of the inner ear and brain glial cells. Patients diagnosed clinically with EAST syndrome were genotyped and mutations in KCNJ10 were studied functionally. Methods: Patient DNA was amplified and sequenced, and new mutations were identified. Mutant and wild-type KCNJ10 constructs were cloned and heterologously expressed in Xenopus oocytes. Whole-cell K+ currents were measured by 2-electrode voltage clamping and channel expression was analysed by Western blotting. Results: We identified 3 homozygous mutations in KCNJ10 (p.F75C, p.A167V and p.V91fs197X), with mutation p.A167V previously reported in a compound heterozygous state. Oocytes expressing wild-type human KCNJ10 showed inwardly rectified currents, which were significantly reduced in all of the mutants (p < 0.001). Specific inhibition of KCNJ10 currents by Ba2+ demonstrated a large residual function in p.A167V only, which was not compatible with causing disease. However, co-expression with KCNJ16 abolished function in these heteromeric channels almost completely. Conclusion: This study provides an explanation for the pathophysiology of the p.A167V KCNJ10 mutation, which had previously not been considered pathogenic on its own. These findings provide evidence for the functional cooperation of KCNJ10 and KCNJ16. Thus, in vitro ascertainment of KCNJ10 function may necessitate co-expression with KCNJ16.

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.

Ciliopathies are a group of clinically and genetically overlapping disorders whose etiologies lie in defective cilia. These are antenna-like organelles on the apical surface of numerous cell types in a variety of tissues and organs, the kidney included. Cilia play essential roles during development and tissue homeostasis, and their dysfunction in the kidney has been associated with renal cyst formation and renal failure. Recently, the term “renal ciliopathies” was coined for those human genetic disorders that are characterized by nephronophthisis, cystic kidneys or renal cystic dysplasia. This review focuses on renal ciliopathies from a human genetics perspective. We survey the newest insights with respect to gene identification and genotype–phenotype correlations, and we reflect on candidate ciliopathies. The opportunities and challenges of next-generation sequencing (NGS) for genetic renal research and clinical DNA diagnostics are also reviewed, and we discuss the contribution of NGS to the development of personalized therapy for patients with renal ciliopathies.

International audience; Mutations in SCN2A, a gene encoding the voltage-gated sodium channel Nav1.2, have been associated with a spectrum of epilepsies and neurodevelopmental disorders. Here, we report the phenotypes of 71 patients and review 130 previously reported patients. We found that (i) encephalopathies with infantile/childhood onset epilepsies (≥3 months of age) occur almost as often as those with an early infantile onset (

What's known on the subject? and What does the study add? The various phenotypes of hypospadias may result from disturbances of dissimilar embryonic processes in different time windows, suggesting aetiological heterogeneity; however, only a few studies have investigated the risk factors for the different hypospadias phenotypes. The study confirmed that genetic predisposition possibly plays a role in anterior and middle hypospadias, as shown by the large effect estimates for familial occurrence of these forms of hypospadias. By contrast, the posterior phenotype was more often associated with pregnancy-related factors, such as primiparity, preterm delivery, and being small for gestational age. New findings were that hormone-containing contraceptive use after conception increased the risk of middle and posterior hypospadias, while multiple pregnancies were associated with the posterior form in particular. Objective To identify specific risk factors for different phenotypes of hypospadias that may arise as a result of dissimilar embryonic processes in different time windows. Patients and Methods A total of 405 hypospadias cases and 627 male controls were included in a Dutch case–control study. Medical records of cases were reviewed to determine the anatomical location of the urethral opening, while demographic, lifestyle and pregnancy-related risk factor data were obtained from self-administered questionnaires. Multivariable and multinomial logistic regression analyses were used to calculate effect estimates for the group containing all cases of hypospadias and for the different hypospadias phenotypes. Results Cases were subdivided into anterior (glandular and coronal; 59%), middle (penile; 29%) and posterior (penoscrotal, scrotal and perineal; 12%) hypospadias. Being a twin/triplet, primiparity, preterm delivery, and being small for gestational age were associated with hypospadias, particularly in posterior cases. Family history of hypospadias increased the risk of hypospadias, an effect that seemed to be more predominant in anterior and middle forms. Maternal obesity seemed to increase the risk of hypospadias in general, and hormone-containing contraceptive use during pregnancy especially increased the risk of middle and posterior hypospadias. Conclusions Our study provides some indications for aetiological heterogeneity of hypospadias, separating anterior and middle phenotypes from posterior hypospadias. Future research should continue to try to establish which specific risk factors and mechanisms may differ according to hypospadias phenotype.

In the last decade, an overwhelming number of genetic aberrations have been discovered and linked to the development of treatment for cancer. With the rapid advancement of next-generation sequencing (NGS) techniques, it is expected that large-scale DNA analyses will increasingly be used to select patients for treatment with specific anticancer agents. Personalizing cancer treatment has many advantages, but sequencing germline DNA as reference material for interpreting cancer genetics may have consequences that extend beyond providing cancer care for an individual patient. In sequencing germline DNA, mutations may be encountered that are associated with increased susceptibility not only to hereditary cancer syndromes but also to other diseases; in those cases, disclosing germline data could be clinically relevant and even lifesaving. In the context of personal autonomy, it is necessary to develop an ethical and legal framework for how to deal with identified hereditary disease susceptibilities and how to return the data to patients and their families. Because clear legislation is lacking, we need to establish guidelines on disclosure of genetic information and, in the process, we need to balance privacy issues with the potential advantages and drawbacks of sharing genetic data with patients and their relatives. Importantly, a strong partnership with patients is critical for understanding how to maximize the translation of genetic information for the benefit of patients with cancer. This review discusses the ethical, legal, and counseling issues surrounding disclosure of genetic information generated by NGS to patients with cancer and their relatives. We also provide a framework for returning these genetic results by proposing a design for a qualified disclosure policy.

The oral-facial-digital (OFD) syndromes comprise a group of related disorders with a combination of oral, facial and digital anomalies. Variants in several ciliary genes have been associated with subtypes of OFD syndrome, yet in most OFD patients the underlying cause remains unknown. We investigated the molecular basis of disease in two brothers with OFD type II, Mohr syndrome, by performing single-nucleotide polymorphism (SNP)-array analysis on the brothers and their healthy parents to identify homozygous regions and candidate genes. Subsequently, we performed whole-exome sequencing (WES) on the family. Using WES, we identified compound heterozygous variants c.[464G>C];[1226G>A] in NIMA (Never in Mitosis Gene A)-Related Kinase 1 (NEK1). The novel variant c.464G>C disturbs normal splicing in an essential region of the kinase domain. The nonsense variant c.1226G>A, p.(Trp409*), results in nonsense-associated alternative splicing, removing the first coiled-coil domain of NEK1. Candidate variants were confirmed with Sanger sequencing and alternative splicing assessed with cDNA analysis. Immunocytochemistry was used to assess cilia number and length. Patient-derived fibroblasts showed severely reduced ciliation compared with control fibroblasts (18.0 vs 48.9%, P

Next-generation sequencing (NGS) has led to the identification of previously unrecognized phenotypes associated with classic kidney disease genes. In addition to improving diagnostics for genetically heterogeneous diseases and enabling a faster rate of gene discovery, NGS has enabled an expansion and redefinition of nephrogenetic disease categories. Findings from these studies raise the question of whether disease diagnoses should be made on clinical grounds, on genetic evidence or a combination thereof. Here, we discuss the major kidney disease-associated genes and gene categories for which NGS has expanded the phenotypic spectrum. For example, COL4A3-5 genes, which are classically associated with Alport syndrome, are now understood to also be involved in the aetiology of focal segmental glomerulosclerosis. DGKE, which is associated with nephrotic syndrome, is also mutated in patients with atypical haemolytic uraemic syndrome. We examine how a shared genetic background between diverse clinical phenotypes can provide insight into the function of genes and novel links with essential pathophysiological mechanisms. In addition, we consider genetic and epigenetic factors that contribute to the observed phenotypic heterogeneity of kidney diseases and discuss the challenges in the interpretation of genetic data. Finally, we discuss the implications of the expanding phenotypic spectra associated with kidney disease genes for clinical practice, genetic counselling and personalized care, and present our recommendations for the use of NGS-based tests in routine nephrology practice.

Parents' preferences for unsolicited findings (UFs) from diagnostic whole-exome sequencing (WES) for their children remain largely unexplored. Our aim was to gain insight into parental considerations favoring acceptance/decline of UFs pertaining to their child. We conducted 20 qualitative, semistructured interviews with parents (n=34) of children with a developmental delay, aged

Item does not contain fulltext We report an 11-year-old girl with mild intellectual disability, skeletal anomalies, congenital heart defect, myopia, and facial dysmorphisms including an extra incisor, cup-shaped ears, and a preauricular skin tag. Array comparative genomic hybridization analysis identified a de novo 4.5-Mb microdeletion on chromosome 14q24.2q24.3. The deleted region and phenotype partially overlap with previously reported patients. Here, we provide an overview of the literature on 14q24 microdeletions and further delineate the associated phenotype. We performed exome sequencing to examine other causes for the phenotype and queried genes present in the 14q24.2q24.3 microdeletion that are associated with recessive disease for variants in the non-deleted allele. The deleted region contains 65 protein-coding genes, including the ciliary gene IFT43. Although Sanger and exome sequencing did not identify variants in the second IFT43 allele or in other IFT complex A-protein-encoding genes, immunocytochemistry showed increased accumulation of IFT-B proteins at the ciliary tip in patient-derived fibroblasts compared to control cells, demonstrating defective retrograde ciliary transport. This could suggest a ciliary defect in the pathogenesis of this disorder. (c) 2016 Wiley Periodicals, Inc.

We identifiedde novononsense variants inKIDINS220/ARMSin three unrelated patients with spastic paraplegia, intellectual deficit, nystagmus, and obesity. KIDINS220 is an essential scaffold protein coordinating neurotrophin signal pathways in neurites and is spatially and temporally regulated in the brain. Molecular analysis of patients' variants confirmed expression and translation of truncated transcripts similar to recently characterized alternative terminal exon splice isoforms ofKIDINS220KIDINS220undergoes extensive alternative splicing in specific neuronal populations and developmental time points, reflecting its complex role in neuronal maturation. In mice and humans,KIDINS220is alternative spliced in the middle region as well as in the last exon. These full-length andKIDINS220splice variants occur at precise moments in cortical, hippocampal, and motor neuron development, with splice variants similar to the variants seen in our patients and lacking the last exon ofKIDINS220occurring in adult rather than in embryonic brain. We conducted tissue-specific expression studies in zebrafish that resulted in spasms, confirming a functional link with disruption of the KIDINS220 levels in developing neurites. This work reveals a crucial physiological role ofKIDINS220in development and provides insight into how perturbation of the complex interplay ofKIDINS220isoforms and their relative expression can affect neuron control and human metabolism. Altogether, we here show thatde novoprotein-truncatingKIDINS220variants cause a new syndrome, SINO, characterized by spastic paraplegia, intellectual deficit, nystagmus, and obesity. This is the first report ofKIDINS220variants causing a human disease.

Joubert syndrome (JBS) is a rare neurodevelopmental disorder belonging to the group of ciliary diseases. JBS is genetically heterogeneous, with >20 causative genes identified to date. A molecular diagnosis of JBS is essential for prediction of disease progression and genetic counseling. We developed a targeted next-generation sequencing (NGS) approach for parallel sequencing of 22 known JBS genes plus 599 additional ciliary genes. This method was used to genotype a cohort of 51 well-phenotyped Northern European JBS cases (in some of the cases, Sanger sequencing of individual JBS genes had been performed previously). Altogether, 21 of the 51 cases (41%) harbored biallelic pathogenic mutations in known JBS genes, including 14 mutations not previously described. Mutations in C5orf42 (12%), TMEM67 (10%), and AHI1 (8%) were the most prevalent. C5orf42 mutations result in a purely neurological Joubert phenotype, in one case associated with postaxial polydactyly. Our study represents a population-based cohort of JBS patients not enriched for consanguinity, providing insight into the relative importance of the different JBS genes in a Northern European population. Mutations in C5orf42 are relatively frequent (possibly due to a Dutch founder mutation) and mutations in CEP290 are underrepresented compared with international cohorts. Furthermore, we report a case with heterozygous mutations in CC2D2A and B9D1, a gene associated with the more severe Meckel-Gruber syndrome that was recently published as a potential new JBS gene, and discuss the significance of this finding.

The anti-diuretic hormone arginine vasopressin (AVP) is released from the pituitary upon hypovolemia or hypernatremia, and regulates water reabsorption in the renal collecting duct principal cells. Binding of AVP to the arginine vasopressin receptor type 2 (AVPR2) in the basolateral membrane leads to translocation of aquaporin 2 (AQP2) water channels to the apical membrane of the collecting duct principal cells, inducing water permeability of the membrane. This results in water reabsorption from the pro-urine into the medullary interstitium following an osmotic gradient. Congenital nephrogenic diabetes insipidus (NDI) is a disorder associated with mutations in either the AVPR2 or AQP2 gene, causing the inability of patients to concentrate their pro-urine, which leads to a high risk of dehydration. This review focuses on the current knowledge regarding the cell biological aspects of congenital X-linked, autosomal-recessive and autosomal-dominant NDI while specifically addressing the latest developments in the field. Based on deepened mechanistic understanding, new therapeutic strategies are currently being explored, which we also discuss here.

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

Contains fulltext : 108791.pdf (Author’s version postprint ) (Open Access) BACKGROUND: Hypospadias is a common congenital malformation of the male external genitalia. Most cases have an unknown aetiology, which is probably a mix of monogenic and multifactorial forms, implicating both genes and environmental factors. This review summarizes current knowledge about the aetiology of hypospadias. METHODS: Pubmed was used to identify studies on hypospadias aetiology published between January 1995 and February 2011. Reference lists of the selected manuscripts were also searched to identify additional studies, including those published before 1995. RESULTS: The search provided 922 articles and 169 articles were selected for this review. Studies screening groups of patients with hypospadias for single gene defects found mutations in WT1, SF1, BMP4, BMP7, HOXA4, HOXB6, FGF8, FGFR2, AR, HSD3B2, SRD5A2, ATF3, MAMLD1, MID1 and BNC2. However, most investigators are convinced that single mutations do not cause the majority of isolated hypospadias cases. Indeed, associations were found with polymorphisms in FGF8, FGFR2, AR, HSD17B3, SRD5A2, ESR1, ESR2, ATF3, MAMLD1, DGKK, MID1, CYP1A1, GSTM1 and GSTT1. In addition, gene expression studies indentified CTGF, CYR61 and EGF as candidate genes. Environmental factors consistently implicated in hypospadias are low birthweight, maternal hypertension and pre-eclampsia, suggesting that placental insufficiency may play an important role in hypospadias aetiology. Exogenous endocrine-disrupting chemicals have the potential to induce hypospadias but it is unclear whether human exposure is high enough to exert this effect. Other environmental factors have also been associated with hypospadias but, for most, the results are inconsistent. CONCLUSIONS: Although a number of contributors to the aetiology of hypospadias have been identified, the majority of risk factors remain unknown.

Contains fulltext : 97141.pdf (Publisher’s version ) (Closed access) Meier-Gorlin syndrome (ear, patella and short-stature syndrome) is an autosomal recessive primordial dwarfism syndrome characterized by absent or hypoplastic patellae and markedly small ears(1)(3). Both pre- and post-natal growth are impaired in this disorder, and although microcephaly is often evident, intellect is usually normal in this syndrome. We report here that individuals with this disorder show marked locus heterogeneity, and we identify mutations in five separate genes: ORC1, ORC4, ORC6, CDT1 and CDC6. All of these genes encode components of the pre-replication complex, implicating defects in replication licensing as the cause of a genetic syndrome with distinct developmental abnormalities.

Purpose: Differentiating between Dravet syndrome and non-Dravet SCN1A-related phenotypes is important for prognosis regarding epilepsy severity, cognitive development, and comorbidities. When a child is diagnosed with genetic epilepsy with febrile seizures plus (GEFS+) or febrile seizures (FS), accurate prognostic information is essential as well, but detailed information on seizure course, seizure freedom, medication use, and comorbidities is lacking for this milder patient group. In this cross-sectional study, we explore disease characteristics in milder SCNTA-related phenotypes and the nature, occurrence, and relationships of SCN1A-related comorbidities in both patients with Dravet and non-Dravet syndromes.Methods: A cohort of 164 Dutch participants with SCN1A-related seizures was evaluated, consisting of 116 patients with Dravet syndrome and 48 patients with either GEFS+, febrile seizures plus (FS+), or FS. Clinical data were collected from medical records, semi-structured telephone interviews, and three questionnaires: the Functional Mobility Scale (FMS), the Pediatric Quality of Life Inventory (PedsQL) Measurement Model, and the Child or Adult Behavior Checklists (CBCL/ABCL).Results: Walking disabilities and severe behavioral problems affect 71% and 43% of patients with Dravet syndrome respectively and are almost never present in patients with non-Dravet syndromes. These comorbidities are strongly correlated to lower quality-of-life (QoL) scores. Less severe comorbidities occur in patients with non-Dravet syndromes: learning problems and psychological/behavioral problems are reported for 27% and 38% respectively. The average QoL score of the non-Dravet group was comparable with that of the general population. The majority of patients with non-Dravet syndromes becomes seizure-free after 10 years of age (85%).Conclusions: Severe behavioral problems and walking disabilities are common in patients with Dravet syndrome and should receive specific attention during clinical management. Although the epilepsy course of patients with non-Dravet syndromes is much more favorable, milder comorbidities frequently occur in this group as well. Our results may be of great value for clinical care and informing newly diagnosed patient and their parents about prognosis. (C) 2018 The Authors. Published by Elsevier Inc.

Item does not contain fulltext A subset of ciliopathies, including Sensenbrenner, Jeune, and short-rib polydactyly syndromes are characterized by skeletal anomalies accompanied by multiorgan defects such as chronic renal failure and retinitis pigmentosa. Through exome sequencing we identified compound heterozygous mutations in WDR19 in a Norwegian family with Sensenbrenner syndrome. In a Dutch family with the clinically overlapping Jeune syndrome, a homozygous missense mutation in the same gene was found. Both families displayed a nephronophthisis-like nephropathy. Independently, we also identified compound heterozygous WDR19 mutations by exome sequencing in a Moroccan family with isolated nephronophthisis. WDR19 encodes IFT144, a member of the intraflagellar transport (IFT) complex A that drives retrograde ciliary transport. We show that IFT144 is absent from the cilia of fibroblasts from one of the Sensenbrenner patients and that ciliary abundance and morphology is perturbed, demonstrating the ciliary pathogenesis. Our results suggest that isolated nephronophthisis, Jeune, and Sensenbrenner syndromes are clinically overlapping disorders that can result from a similar molecular cause.

Contains fulltext : 89067.pdf (Publisher’s version ) (Closed access) Autosomal-recessive cerebellar ataxias comprise a clinically and genetically heterogeneous group of neurodegenerative disorders. In contrast to their dominant counterparts, unraveling the molecular background of these ataxias has proven to be more complicated and the currently known mutations provide incomplete coverage for genotyping of patients. By combining SNP array-based linkage analysis and targeted resequencing of relevant sequences in the linkage interval with the use of next-generation sequencing technology, we identified a mutation in a gene and have shown its association with autosomal-recessive cerebellar ataxia. In a Dutch consanguineous family with three affected siblings a homozygous 12.5 Mb region on chromosome 3 was targeted by array-based sequence capture. Prioritization of all detected sequence variants led to four candidate genes, one of which contained a variant with a high base pair conservation score (phyloP score: 5.26). This variant was a leucine-to-arginine substitution in the DUF 590 domain of a 16K transmembrane protein, a putative calcium-activated chloride channel encoded by anoctamin 10 (ANO10). The analysis of ANO10 by Sanger sequencing revealed three additional mutations: a homozygous mutation (c.1150_1151del [p.Leu384fs]) in a Serbian family and a compound-heterozygous splice-site mutation (c.1476+1G>T) and a frameshift mutation (c.1604del [p.Leu535X]) in a French family. This illustrates the power of using initial homozygosity mapping with next-generation sequencing technology to identify genes involved in autosomal-recessive diseases. Moreover, identifying a putative calcium-dependent chloride channel involved in cerebellar ataxia adds another pathway to the list of pathophysiological mechanisms that may cause cerebellar ataxia.

Contains fulltext : 88664.pdf (Publisher’s version ) (Closed access) Sensenbrenner syndrome/cranioectodermal dysplasia (CED) is an autosomal-recessive disease that is characterized by craniosynostosis and ectodermal and skeletal abnormalities. We sequenced the exomes of two unrelated CED patients and identified compound heterozygous mutations in WDR35 as the cause of the disease in each of the two patients independently, showing that it is possible to find the causative gene by sequencing the exome of a single sporadic patient. With RT-PCR, we demonstrate that a splice-site mutation in exon 2 of WDR35 alters splicing of RNA on the affected allele, introducing a premature stop codon. WDR35 is homologous to TULP4 (from the Tubby superfamily) and has previously been characterized as an intraflagellar transport component, confirming that Sensenbrenner syndrome is a ciliary disorder.

Contains fulltext : 89679.pdf (Publisher’s version ) (Closed access) The renal distal convoluted tubule (DCT) has an essential role in maintaining systemic magnesium (Mg(2+)) concentration. The DCT is the final determinant of plasma Mg(2+) levels, as the more distal nephron segments are largely impermeable to Mg(2+). In the past decade, positional candidate strategies in families with inherited forms of hypomagnesemia have led to the identification of genes involved in Mg(2+) handling. A large fraction of this resides in the DCT, namely, (i) the transient receptor potential channel melastatin subtype 6 (TRPM6), a divalent cation-permeable channel located at the luminal membrane of the DCT, facilitates Mg(2+) entry from the pro-urine into the cell; (ii) the epidermal growth factor is a novel hormone regulating active Mg(2+) transport through TRPM6; (iii) the voltage-gated K(+) channel, Kv1.1, establishes a favorable luminal membrane potential for TRPM6-mediated Mg(2+) transport; (iv) the Na(+)/K(+)-ATPase gamma-subunit (gamma-Na(+)/K(+)-ATPase) was identified as mutated protein in a family with isolated dominant hypomagnesemia. The molecular mechanism by which gamma-Na(+)/K(+)-ATPase is involved in DCT Mg(2+) handling remains unknown; (v) a high percentage of patients with mutations in the renal transcription factor HNF1B (hepatocyte nuclear factor 1 homeobox B) gene develop hypomagnesemia; and (vi) Gitelman and EAST/SeSAME syndrome patients suffer from a similar tubulopathy due to mutations in NCC (NaCl cotransporter) and Kir4.1, respectively. In these patients, decreased expression of TRPM6 is proposed to cause hypomagnesemia. Insights into the molecular mechanisms of the identified genes, as well as the identification of novel genes, will further improve our knowledge about renal Mg(2+) handling. 01 januari 2010

Contains fulltext : 80180.pdf (Publisher’s version ) (Open Access) In this study, we developed and analytically validated a fully automated, robust confirmation sensitive capillary electrophoresis (CSCE) method to perform mutation scanning of the large SACS gene. This method facilitates a rapid and cost-effective molecular diagnosis of autosomal recessive spastic ataxia of Charlevoix-Saguenay. Critical issues addressed during the development of the CSCE system included the position of a DNA variant relative to the primers and the CG-content of the amplicons. The validation was performed in two phases; a retrospective analysis of 32 samples containing 41 different known DNA variants and a prospective analysis of 20 samples of patients clinically suspected of having autosomal recessive spastic ataxia of Charlevoix-Saguenay. These 20 samples appeared to contain 73 DNA variants. In total, in 32 out of the 45 amplicons, a DNA variant was present, which allowed verification of the detection capacity during the validation process. After optimization of the original design, the overall analytical sensitivity of CSCE for the SACS gene was 100%, and the analytical specificity of CSCE was 99.8%. In conclusion, CSCE is a robust technique with a high analytical sensitivity and specificity, and it can readily be used for mutation scanning of the large SACS gene. Furthermore this technique is less time-consuming and less expensive, as compared with standard automated sequencing.

Item does not contain fulltext The kidney plays an important role in ion homeostasis in the human body. Several hereditary disorders characterized by perturbations in renal magnesium reabsorption leading to hypomagnesemia have been described over the past 50 years, with the most important of these being Gitelman syndrome, familial hypomagnesemia with hypercalciuria and nephrocalcinosis, familial hypomagnesemia with secondary hypocalcemia, autosomal dominant hypomagnesemia with hypocalciuria, and autosomal recessive hypomagnesemia. Only recently, following positional cloning strategies in affected families, have mutations in renal ion channels and transporters been identified in these diseases. In this short review, I give an update on these hypomagnesemic disorders. Elucidation of the genetic etiology and, for most of these disorders, also the underlying pathophysiology of the disease, has greatly increased our understanding of the normal physiology of renal magnesium handling. This is yet another example of the importance of studying rare disorders in order to unravel physiological and pathophysiological processes in the human body.

Epilepsy, ataxia, sensorineural deafness, tubulopathy, and KCNJ10 mutations. N Engl J Med 360: 1960–1970, 2009 Bockenhauer D, Feather S, Stanescu HC, Bandulik S, Zdebik AA, Reichold M, Tobin J, Lieberer E, Sterner C, Landoure G, Arora R, Sirimanna T, Thompson D, Cross JH, van’t Hoff W, Al Masri O, Tullus K, Yeung S, Anikster Y, Klootwijk E, Hubank M, Dillon MJ, Heitzmann D, ArcosBurgos M, Knepper MA, Dobbie A, Gahl WA, Warth R, Sheridan E, Kleta R

Item does not contain fulltext Water reabsorption in the renal collecting duct is regulated by the antidiuretic hormone vasopressin (AVP). When the vasopressin V2 receptor, present on the basolateral site of the renal principal cell, becomes activated by AVP, aquaporin-2 (AQP2) water channels will be inserted in the apical membrane, and in this fashion, water can be reabsorbed from the pro-urine into the interstitium. The essential role of the vasopressin V2 receptor and AQP2 in the maintenance of body water homeostasis became clear when it was shown that mutations in their genes cause nephrogenic diabetes insipidus, a disorder in which the kidney is unable to concentrate urine in response to AVP. This review describes the current knowledge on AQP2 mutations in nephrogenic diabetes insipidus.

Contains fulltext : 69050.pdf (Publisher’s version ) (Open Access) Autosomal recessive spastic ataxia of Charlevoix-Saguenay (ARSACS: MIM 270550) is a neurodegenerative disorder characterized by early-onset cerebellar ataxia with spasticity and peripheral neuropathy. This disorder, considered to be rare, was first described in the late seventies among French Canadians in the isolated Charlevoix-Saguenay region of Quebec. Nowadays, it is known that the disorder is not only limited to this region but occurs worldwide. Our objective was to identify cases of autosomal recessive spastic ataxia of Charlevoix-Saguenay (ARSACS) in Dutch patients with recessive early-onset cerebellar ataxia by sequencing the complete SACS gene. In a Dutch cohort of 43 index patients with ataxia onset before age 25, we identified 16 index patients (total 23 patients) with mutations in the SACS gene. Nine of them had homozygous mutations, and seven of them had compound heterozygous mutations. Retrospectively, the phenotype of patients carrying mutations was remarkably uniform: cerebellar ataxia with onset before age 13 years, lower limb spasticity and sensorimotor axonal neuropathy, and cerebellar (vermis) atrophy on magnetic resonance imaging, consistent with the core ARSACS phenotype previously described. The high rate of mutations (37%) identified in this cohort of Dutch patients suggests that ARSACS is substantially more frequent than previously estimated. We predict that the availability of SACS mutation analysis as well as an increasing awareness of the characteristic ARSACS phenotype will lead to the diagnosis of many additional patients, possibly even at a younger age.

Item does not contain fulltext BACKGROUND: Loss-of-function mutations in SLC12A3 coding for the thiazide-sensitive NaCl cotransporter (NCC) cause Gitelman's syndrome (GS), a recessively inherited salt-losing tubulopathy. Most GS patients are compound heterozygous. However, up to 30% of GS patients carry only a single mutant allele, and a normal SLC12A3 screening is also observed in a small subset of patients. Locus heterogeneity could explain the lack of detection of mutant SLC12A3 alleles in GS patients. The renal phenotype of the parvalbumin knockout mice pointed to PVALB as a candidate gene for GS for SLC12A3-negative cases. METHODS: PCR and direct sequencing of PVALB was performed in 132 GS patients in whom only one or no (N = 79) mutant SLC12A3 allele was found. The possible interference of biallelic SNPs (single nucleotide polymorphisms) on normal transcription or normal splicing was investigated. Genotyping of 110 anonymous blood donors was performed to determine the allelic frequency in the normal population. RESULTS: No sequence variants resulting in amino acid substitution or truncated protein within the PVALB gene were found in the 264 chromosomes tested. Ten biallelic SNPs, including six novel polymorphisms, were identified: five in the 5' UTR, none of them affecting predicted regulatory elements; three in the coding region, without alteration of the consensus splice sites, and two in the 3' UTR. The observed allelic frequencies did not differ significantly between GS patients and controls. CONCLUSION: Our results strongly suggest that mutations in the PVALB gene are not involved in GS patients who harbour a single or no mutant SLC12A3 allele.

The importance of genetics for nephrology is underlined by the fact that more than 200 genes have been associated with inherited renal disease. Information about the genotypes of patients with kidney disease is key to providing an early and accurate diagnosis and determining prognosis. In addition, genetics influences patient management and allows for the development of targeted therapeutics. After briefly discussing major developments in the genetics field, this review focuses on the development of next-generation sequencing (NGS) technology that can be used to analyze numerous genes or even whole genomes in a single experiment. This technology allowed for the rapid identification of novel disease genes for a variety of recessive and dominant renal disorders, varying from congenital anomalies of the kidney and urinary tract to atypical hemolytic uremic syndrome and nephronophthisis to adult-onset focal segmental glomerulosclerosis. This review discusses how to interpret NGS data, summarizes NGS discoveries in the kidney field, highlights challenges with respect to technology and variant interpretation, and underlines the importance of NGS usage in the nephrology clinic. We discuss how sequencing of all or a selection of coding DNA is progressing toward whole-genome analysis, and we touch on how NGS has opened an era of personalized medicine that includes drug screening, gene therapy, and tissue regeneration therapy.

Contains fulltext : 171962.PDF (Publisher’s version ) (Open Access) The Wnts can be considered as candidates for the Congenital Anomaly of Kidney and Urinary Tract, CAKUT diseases since they take part in the control of kidney organogenesis. Of them Wnt5a is expressed in ureteric bud (UB) and its deficiency leads to duplex collecting system (13/90) uni- or bilateral kidney agenesis (10/90), hypoplasia with altered pattern of ureteric tree organization (42/90) and lobularization defects with partly fused ureter trunks (25/90) unlike in controls. The UB had also notably less tips due to Wnt5a deficiency being at E15.5 306 and at E16.5 765 corresponding to 428 and 1022 in control (p