OXGR1 is a candidate disease gene for human calcium oxalate nephrolithiasis.

PURPOSE: Nephrolithiasis (NL) affects 1 in 11 individuals worldwide, leading to significant patient morbidity. NL is associated with nephrocalcinosis (NC), a risk factor for chronic kidney disease. Causative genetic variants are detected in 11% to 28% of NL and/or NC, suggesting that additional NL/NC-associated genetic loci await discovery. Therefore, we employed genomic approaches to discover novel genetic forms of NL/NC. METHODS: Exome sequencing and directed sequencing of the OXGR1 locus were performed in a worldwide NL/NC cohort. Putatively deleterious, rare OXGR1 variants were functionally characterized. RESULTS: Exome sequencing revealed a heterozygous OXGR1 missense variant (c.371T>G, p.L124R) cosegregating with calcium oxalate NL and/or NC disease in an autosomal dominant inheritance pattern within a multigenerational family with 5 affected individuals. OXGR1 encodes 2-oxoglutarate (α-ketoglutarate [AKG]) receptor 1 in the distal nephron. In response to its ligand AKG, OXGR1 stimulates the chloride-bicarbonate exchanger, pendrin, which also regulates transepithelial calcium transport in cortical connecting tubules. Strong amino acid conservation in orthologs and paralogs, severe in silico prediction scores, and extreme rarity in exome population databases suggested that the variant was deleterious. Interrogation of the OXGR1 locus in 1107 additional NL/NC families identified 5 additional deleterious dominant variants in 5 families with calcium oxalate NL/NC. Rare, potentially deleterious OXGR1 variants were enriched in patients with NL/NC compared with Exome Aggregation Consortium controls (χ2 = 7.117, P = .0076). Wild-type OXGR1-expressing Xenopus oocytes exhibited AKG-responsive Ca2+ uptake. Of 5 NL/NC-associated missense variants, 5 revealed impaired AKG-dependent Ca2+ uptake, demonstrating loss of function. CONCLUSION: Rare, dominant loss-of-function OXGR1 variants are associated with recurrent calcium oxalate NL/NC disease.

Genetic stratification reveals COL4A variants and spontaneous remission in Egyptian children with proteinuria in the first 2 years of life.

AIM: The earlier the onset of proteinuria, the higher the incidence of genetic forms. Therefore, we aimed to analyse the spectrum of monogenic proteinuria in Egyptian children presenting at age <2 years. METHODS: The results of 27-gene panel or whole-exome sequencing were correlated with phenotype and treatment outcomes in 54 patients from 45 families. RESULTS: Disease-causing variants were identified in 29/45 (64.4%) families. Mutations often occurred in three podocytopathy genes: NPHS1, NPHS2 and PLCE1 (19 families). Some showed extrarenal manifestations. Additionally, mutations were detected in 10 other genes, including novel variants of OSGEP, SGPL1 and SYNPO2. COL4A variants phenocopied isolated steroid-resistant nephrotic syndrome (2/29 families, 6.9%). NPHS2 M1L was the single most common genetic finding beyond the age of 3 months (4/18 families, 22.2%). Biopsy results did not correlate with genotypes (n = 30). On renin-angiotensin-aldosterone system antagonists alone, partial and complete remission occurred in 3/24 (12.5%) patients with monogenic proteinuria each, whereas 6.3% (1/16) achieved complete remission on immunosuppression. CONCLUSION: Genotyping is mandatory to avoid biopsies and immunosuppression when proteinuria presents at age <2 years. Even with such a presentation, COL4A genes should be included. NPHS2 M1L was prevalent in Egyptian children (4 months-2 years) with proteinuria, demonstrating precision diagnostic utility.

Reverse phenotyping facilitates disease allele calling in exome sequencing of patients with CAKUT.

PURPOSE: Congenital anomalies of the kidneys and urinary tract (CAKUT) constitute the leading cause of chronic kidney disease in children. In total, 174 monogenic causes of isolated or syndromic CAKUT are known. However, syndromic features may be overlooked when the initial clinical diagnosis of CAKUT is made. We hypothesized that the yield of a molecular genetic diagnosis by exome sequencing (ES) can be increased by applying reverse phenotyping, by re-examining the case for signs/symptoms of the suspected clinical syndrome that results from the genetic variant detected by ES. METHODS: We conducted ES in an international cohort of 731 unrelated families with CAKUT. We evaluated ES data for variants in 174 genes, in which variants are known to cause isolated or syndromic CAKUT. In cases in which ES suggested a previously unreported syndromic phenotype, we conducted reverse phenotyping. RESULTS: In 83 of 731 (11.4%) families, we detected a likely CAKUT-causing genetic variant consistent with an isolated or syndromic CAKUT phenotype. In 19 of these 83 families (22.9%), reverse phenotyping yielded syndromic clinical findings, thereby strengthening the genotype-phenotype correlation. CONCLUSION: We conclude that employing reverse phenotyping in the evaluation of syndromic CAKUT genes by ES provides an important tool to facilitate molecular genetic diagnostics in CAKUT.

Clinical performance of resin-matrix ceramic partial coverage restorations: a systematic review.

OBJECTIVE: To evaluate clinical performance of the new CAD/CAM resin-matrix ceramics and compare it with ceramic partial coverage restorations. MATERIALS AND METHODS: An electronic search of 3 databases (The National Library of Medicine (MEDLINE/PubMed), Scopus, and the Cochrane Central Register of Controlled Trials) was conducted. English clinical studies published between 2005 and September 2020 that evaluated the clinical performance of CAD/CAM resin-matrix ceramics inlays, onlays, or overlays were selected. The primary clinical question was applied according to PICOS strategy (Population, Intervention, Comparison, Outcome, Study design). The included studies were individually evaluated for risk of bias according to the modified Cochrane Collaboration tool criteria. RESULTS: A total of 7 studies were included according to the established inclusion and exclusion criteria. From the included studies, 6 were randomized clinical trials while one study was longitudinal observational study without control group. According to the results of the included studies, the success rate of CAD/CAM resin-based composite ranged from 85.7 to 100% whereas the success rate reported for ceramic partial coverage restorations ranged from 93.3 to 100%. Fractures and debondings are found to be the most common cause of restorations failure. CONCLUSION: CAD/CAM resin-based composite can be considered a reliable material for partial coverage restorations with clinical performance similar to glass ceramic restorations. However, this result needs to be confirmed in long-term evaluations. CLINICAL RELEVANCE: CAD/CAM resin-based composites provide a potential alternative to ceramic indirect restorations. However, clinicians must be aware of the lake of knowledge regarding long-term outcome.

Effect of different surface treatments on resin-matrix CAD/CAM ceramics bonding to dentin: in vitro study.

BACKGROUND: Evaluating the effect of different surface treatment methods on the micro-tensile bond strength (µTBS) of two different resin-matrix computer-aided design/computer-aided manufacturing (CAD/CAM) ceramics (RMCs). METHODS: A standardized inlay preparations were performed on 100 intact maxillary premolars. According to the type of the restorative material, the teeth were randomly divided into two equally sized groups (n = 50): (polymer-infiltrated ceramic (Vita Enamic) and resin-based composites (Lava Ultimate)). The inlays were fabricated using CAD/CAM technology. In each group, the specimens were randomly assigned to five subgroups (n = 10) according to the surface treatment method: group 1 used was the control group (no surface treatment); group 2, was treated with air abrasion with 50 µm Al2O3 (A) and universal adhesive (UA); group 3, was treated with air abrasion with 50 µm Al2O3 (A) and silane coupling agent (S); group 4, was treated with hydrofluoric acid (HF) and universal adhesive (UA) and group 5, was treated with Hydrofluoric acid (HF) + silane coupling agent (S). The inlays were then cemented to their respective preparations using dual-cure self-adhesive resin cement (RelyX U200, 3 M ESPE) according to the manufacturer's instructions. The µTBS test was conducted in all groups, and stereomicroscope and scanning electron microscope were used to inspect the failure mode. The data were statistically analyzed using a two-way analysis of variance (ANOVA) and Tukey's post-hoc multiple comparison tests at a significance level of p < 0.05. RESULTS: Surface treatments significantly increased the µTBS of the materials compared to the control group (p < 0.05). For CAD/CAM RBCs, the µTBS value highest in group 2 whereas, for PICN, the µTBS value was highest in group 3. Cohesive failure of CAD/CAM restorative material was the most predominant mode of failure in all treated groups, whereas adhesive failure at restoration-cement interface was the most predominant failure mode in the control group. CONCLUSION: Surface treatments increase the µTBS of resin-matrix CAD/CAM ceramics to tooth structure. Air abrasion followed by universal adhesive and hydrofluoric acid followed by silane application appears to be the best strategies for optimizing the bond strength of CAD/CAM RBCs and PICN respectively.

Systemic lupus erythematosus children in Egypt: Homeland spectrum amid the global situation.

OBJECTIVES: This study aims to present the manifestations of juvenile systemic lupus erythematosus (JSLE) across Egypt, to focus on age at onset and gender-driven influence on disease characteristics, and to compare findings to other countries. METHODS: The study included 404 Egyptian children with systemic lupus erythematosus (SLE) presenting to one of the specialized rheumatology centers corresponding to 13 major governorates. Juvenile cases age was ≤ 16°years at the time of recruitment. The SLE Disease Activity Index (SLEDAI) and damage index (DI) were assessed. RESULTS: The mean age was 13.2 ± 2.4°years; 355 females and 49 males (7.2:1), and the disease duration was 2.3 ± 1.6 years, while age at disease onset was 11.1 ± 2.5°years. Their SLEDAI was 13.5 ± 12.3, and DI, 0.36 ± 0.78. The overall estimated prevalence of childhood-SLE patients in the recruited cohort in Egypt was 1/100,000 population (0.24/100000 males and 1.8/100000 females). 7.4% developed pre-pubertal SLE (≤ 7 years); 73.3%, peri-pubertal; and 19.3% during early adolescence. The differences according to age group were equal for gender and clinical manifestations except skin lesions present in 59.3% of pre-pubertal onset, 74.6% of peri-pubertal, and 84.2% of adolescents (p = 0.029), and renal involvement in 73.8% of peripubertal, 62.1% of pre-pubertal and 58.9% of adolescents (p = 0.03). Laboratory investigations, SLEDAI, and DI were similar among age categories. Lupus nephritis was more common in Egypt compared to JSLE from other countries. CONCLUSION: Our large multicenter study identified that female gender influenced disease characteristics with more frequent skin involvement. Skin lesions were significantly higher in adolescents, while renal involvement in peri-pubertal children.

Whole exome sequencing identified ATP6V1C2 as a novel candidate gene for recessive distal renal tubular acidosis.

Distal renal tubular acidosis is a rare renal tubular disorder characterized by hyperchloremic metabolic acidosis and impaired urinary acidification. Mutations in three genes (ATP6V0A4, ATP6V1B1 and SLC4A1) constitute a monogenic causation in 58-70% of familial cases of distal renal tubular acidosis. Recently, mutations in FOXI1 have been identified as an additional cause. Therefore, we hypothesized that further monogenic causes of distal renal tubular acidosis remain to be discovered. Panel sequencing and/or whole exome sequencing was performed in a cohort of 17 families with 19 affected individuals with pediatric onset distal renal tubular acidosis. A causative mutation was detected in one of the three "classical" known distal renal tubular acidosis genes in 10 of 17 families. The seven unsolved families were then subjected to candidate whole exome sequencing analysis. Potential disease causing mutations in three genes were detected: ATP6V1C2, which encodes another kidney specific subunit of the V-type proton ATPase (1 family); WDR72 (2 families), previously implicated in V-ATPase trafficking in cells; and SLC4A2 (1 family), a paralog of the known distal renal tubular acidosis gene SLC4A1. Two of these mutations were assessed for deleteriousness through functional studies. Yeast growth assays for ATP6V1C2 revealed loss-of-function for the patient mutation, strongly supporting ATP6V1C2 as a novel distal renal tubular acidosis gene. Thus, we provided a molecular diagnosis in a known distal renal tubular acidosis gene in 10 of 17 families (59%) with this disease, identified mutations in ATP6V1C2 as a novel human candidate gene, and provided further evidence for phenotypic expansion in WDR72 mutations from amelogenesis imperfecta to distal renal tubular acidosis.

Lupus nephritis: correlation of immunohistochemical expression of C4d, CD163-positive M2c-like macrophages and Foxp3-expressing regulatory T cells with disease activity and chronicity.

BACKGROUND: C4d, which is a serum complement cleavage product of the activated complement component C4, was found to be an accurate indicator of lupus activity compared to complement levels. Recently, macrophages have been considered to be pivotal members in the pathogenesis of lupus nephritis (LN). M2c-like macrophages have anti-inflammatory functions and promote fibrosis. Multiple studies have detected that LN is associated with an imbalance between the regulatory T cell (Treg) population and the inflammatory T helper subtypes. METHODS: We evaluated and scored the immunohistochemical expression of C4d, CD163-positive M2C-macrophages and Foxp3-expressing Tregs in 53 renal biopsies of LN. Their expression was scored and correlated with clinical and histological disease activity and chronicity. RESULTS: Class IV was the most prevalent class (50.9%), followed by class III (17%). PTC-C4d intensity score, CD163% of positive M2c macrophages and FOXP3% of positive Tregs were significantly correlated with chronicity index (rs = 0.292, p = 0.034; rs = 0.407, p = 0.003; and rs = 0.296, p = 0.031, respectively). Also, FOXP3% of positive Tregs was significantly correlated with LN class (rs = 0.31, p = 0.024). CONCLUSION: C4d-PTC, CD163-positive M2c macrophages and FOXP3-positive Tregs are markers that significantly correlated with chronicity in LN. Further studies are needed to evaluate their prognostic value.

Whole-Exome Sequencing Identifies Causative Mutations in Families with Congenital Anomalies of the Kidney and Urinary Tract.

BACKGROUND: Congenital anomalies of the kidney and urinary tract (CAKUT) are the most prevalent cause of kidney disease in the first three decades of life. Previous gene panel studies showed monogenic causation in up to 12% of patients with CAKUT. METHODS: We applied whole-exome sequencing to analyze the genotypes of individuals from 232 families with CAKUT, evaluating for mutations in single genes known to cause human CAKUT and genes known to cause CAKUT in mice. In consanguineous or multiplex families, we additionally performed a search for novel monogenic causes of CAKUT. RESULTS: In 29 families (13%), we detected a causative mutation in a known gene for isolated or syndromic CAKUT that sufficiently explained the patient's CAKUT phenotype. In three families (1%), we detected a mutation in a gene reported to cause a phenocopy of CAKUT. In 15 of 155 families with isolated CAKUT, we detected deleterious mutations in syndromic CAKUT genes. Our additional search for novel monogenic causes of CAKUT in consanguineous and multiplex families revealed a potential single, novel monogenic CAKUT gene in 19 of 232 families (8%). CONCLUSIONS: We identified monogenic mutations in a known human CAKUT gene or CAKUT phenocopy gene as the cause of disease in 14% of the CAKUT families in this study. Whole-exome sequencing provides an etiologic diagnosis in a high fraction of patients with CAKUT and will provide a new basis for the mechanistic understanding of CAKUT.

Whole exome sequencing frequently detects a monogenic cause in early onset nephrolithiasis and nephrocalcinosis.

The incidence of nephrolithiasis continues to rise. Previously, we showed that a monogenic cause could be detected in 11.4% of individuals with adult-onset nephrolithiasis or nephrocalcinosis and in 16.7-20.8% of individuals with onset before 18 years of age, using gene panel sequencing of 30 genes known to cause nephrolithiasis/nephrocalcinosis. To overcome the limitations of panel sequencing, we utilized whole exome sequencing in 51 families, who presented before age 25 years with at least one renal stone or with a renal ultrasound finding of nephrocalcinosis to identify the underlying molecular genetic cause of disease. In 15 of 51 families, we detected a monogenic causative mutation by whole exome sequencing. A mutation in seven recessive genes (AGXT, ATP6V1B1, CLDN16, CLDN19, GRHPR, SLC3A1, SLC12A1), in one dominant gene (SLC9A3R1), and in one gene (SLC34A1) with both recessive and dominant inheritance was detected. Seven of the 19 different mutations were not previously described as disease-causing. In one family, a causative mutation in one of 117 genes that may represent phenocopies of nephrolithiasis-causing genes was detected. In nine of 15 families, the genetic diagnosis may have specific implications for stone management and prevention. Several factors that correlated with the higher detection rate in our cohort were younger age at onset of nephrolithiasis/nephrocalcinosis, presence of multiple affected members in a family, and presence of consanguinity. Thus, we established whole exome sequencing as an efficient approach toward a molecular genetic diagnosis in individuals with nephrolithiasis/nephrocalcinosis who manifest before age 25 years.

IFT81, encoding an IFT-B core protein, as a very rare cause of a ciliopathy phenotype.

BACKGROUND: Bidirectional intraflagellar transport (IFT) consists of two major protein complexes, IFT-A and IFT-B. In contrast to the IFT-B complex, all components of IFT-A have recently been linked to human ciliopathies when defective. We therefore hypothesised that mutations in additional IFT-B encoding genes can be found in patients with multisystemic ciliopathies. METHODS: We screened 1628 individuals with reno-ocular ciliopathies by targeted next-generation sequencing of ciliary candidate genes, including all IFT-B encoding genes. RESULTS: Consequently, we identified a homozygous mutation in IFT81 affecting an obligatory donor splice site in an individual with nephronophthisis and polydactyly. Further, we detected a loss-of-stop mutation with extension of the deduced protein by 10 amino acids in an individual with neuronal ceroid lipofuscinosis-1. This proband presented with retinal dystrophy and brain lesions including cerebellar atrophy, a phenotype to which the IFT81 variant might contribute. Cultured fibroblasts of this latter affected individual showed a significant decrease in ciliated cell abundance compared with controls and increased expression of the transcription factor GLI2 suggesting deranged sonic hedgehog signalling. CONCLUSIONS: This work describes identification of mutations of IFT81 in individuals with symptoms consistent with the clinical spectrum of ciliopathies. It might represent the rare case of a core IFT-B complex protein found associated with human disease. Our data further suggest that defects in the IFT-B core are an exceedingly rare finding, probably due to its indispensable role for ciliary assembly in development.

A single-gene cause in 29.5% of cases of steroid-resistant nephrotic syndrome.

Steroid-resistant nephrotic syndrome (SRNS) is the second most frequent cause of ESRD in the first two decades of life. Effective treatment is lacking. First insights into disease mechanisms came from identification of single-gene causes of SRNS. However, the frequency of single-gene causation and its age distribution in large cohorts are unknown. We performed exon sequencing of NPHS2 and WT1 for 1783 unrelated, international families with SRNS. We then examined all patients by microfluidic multiplex PCR and next-generation sequencing for all 27 genes known to cause SRNS if mutated. We detected a single-gene cause in 29.5% (526 of 1783) of families with SRNS that manifested before 25 years of age. The fraction of families in whom a single-gene cause was identified inversely correlated with age of onset. Within clinically relevant age groups, the fraction of families with detection of the single-gene cause was as follows: onset in the first 3 months of life (69.4%), between 4 and 12 months old (49.7%), between 1 and 6 years old (25.3%), between 7 and 12 years old (17.8%), and between 13 and 18 years old (10.8%). For PLCE1, specific mutations correlated with age of onset. Notably, 1% of individuals carried mutations in genes that function within the coenzyme Q10 biosynthesis pathway, suggesting that SRNS may be treatable in these individuals. Our study results should facilitate molecular genetic diagnostics of SRNS, etiologic classification for therapeutic studies, generation of genotype-phenotype correlations, and the identification of individuals in whom a targeted treatment for SRNS may be available.

Terra firma-forme dermatosis: Case Series and dermoscopic features.

Terra firma-forme dermatosis (TFFD) is characterized by dirt-like skin lesions that disappear after rubbing with alcohol. We describe the dermoscopic features of TFFD before and after alcohol swabbing in six patients. All patients showed similar dermoscopic appearance with large polygonal plate-like brown scales arranged together giving a mosaic pattern. These features disappear completely after isopropyl alcohol swabbing of the lesions. In conclusion dermoscopy can assist in the evaluation of terra firma-forme dermatosis and the dermoscopic evaluation of other dirty dermatoses is recommended in the future to compare findings with TFFD.

Whole-exome resequencing distinguishes cystic kidney diseases from phenocopies in renal ciliopathies.

Rare single-gene disorders cause chronic disease. However, half of the 6000 recessive single gene causes of disease are still unknown. Because recessive disease genes can illuminate, at least in part, disease pathomechanism, their identification offers direct opportunities for improved clinical management and potentially treatment. Rare diseases comprise the majority of chronic kidney disease (CKD) in children but are notoriously difficult to diagnose. Whole-exome resequencing facilitates identification of recessive disease genes. However, its utility is impeded by the large number of genetic variants detected. We here overcome this limitation by combining homozygosity mapping with whole-exome resequencing in 10 sib pairs with a nephronophthisis-related ciliopathy, which represents the most frequent genetic cause of CKD in the first three decades of life. In 7 of 10 sibships with a histologic or ultrasonographic diagnosis of nephronophthisis-related ciliopathy, we detect the causative gene. In six sibships, we identify mutations of known nephronophthisis-related ciliopathy genes, while in two additional sibships we found mutations in the known CKD-causing genes SLC4A1 and AGXT as phenocopies of nephronophthisis-related ciliopathy. Thus, whole-exome resequencing establishes an efficient, noninvasive approach towards early detection and causation-based diagnosis of rare kidney diseases. This approach can be extended to other rare recessive disorders, thereby providing accurate diagnosis and facilitating the study of disease mechanisms.

Exome sequencing reveals cubilin mutation as a single-gene cause of proteinuria.

In two siblings of consanguineous parents with intermittent nephrotic-range proteinuria, we identified a homozygous deleterious frameshift mutation in the gene CUBN, which encodes cubulin, using exome capture and massively parallel re-sequencing. The mutation segregated with affected members of this family and was absent from 92 healthy individuals, thereby identifying a recessive mutation in CUBN as the single-gene cause of proteinuria in this sibship. Cubulin mutations cause a hereditary form of megaloblastic anemia secondary to vitamin B(12) deficiency, and proteinuria occurs in 50% of cases since cubilin is coreceptor for both the intestinal vitamin B(12)-intrinsic factor complex and the tubular reabsorption of protein in the proximal tubule. In summary, we report successful use of exome capture and massively parallel re-sequencing to identify a rare, single-gene cause of nephropathy.

Dermoscopic criteria of discoid lupus erythematosus: An observational cross-sectional study of 28 patients.

BACKGROUND: Discoid lupus erythematosus (DLE) affects mainly the head and neck and lesions heal with scaring. Early diagnosis of DLE is crucial; dermoscopy may enable early diagnosis and help to assess the prognosis of well-established lesions. AIMS: To describe the dermoscopic features of DLE and to correlate them with the histological findings, site and duration of DLE. MATERIAL AND METHOD: This study included 28 patients diagnosed as DLE based on clinical and histopathological examination. We examined the lesions clinically, dermoscopically and histopathologically. Evaluated dermoscopic variables were based on data in the available literature and on our observations. RESULTS: Whitish scales (89.3%), arborizing blood vessels (85.7%), follicular plugging (82.1%), and pigmentation (82.1%) were the commonest dermoscopic findings. Radial arrangement of arborizing blood vessel in between a radially arranged perifollicular whitish halo (starburst pattern) (39.3%) was noticed for the first time in this study. Rosettes (57.1%) were also seen. There was significant agreement between many dermoscopic and pathological findings with high sensitivity and specificity of many dermoscopic variants in the diagnosis of DLE. Follicular plugging, perifollicular whitish halo, starburst pattern, follicular red dots and rosettes were detected in early stages of the disease but structureless whitish areas and telangiectasia need more time to develop. LIMITATIONS: We examined our patients at the time of presentation only without prospective monitoring and we had a relatively small sample size. CONCLUSION: Dermoscopy helps in the diagnosis of DLE at different body sites.

Copy Number Variation Analysis Facilitates Identification of Genetic Causation in Patients with Congenital Anomalies of the Kidney and Urinary Tract.

Background: Congenital anomalies of the kidneys and urinary tract (CAKUT) are the most common cause of chronic kidney disease among children and adults younger than 30 yr. In our previous study, whole-exome sequencing (WES) identified a known monogenic cause of isolated or syndromic CAKUT in 13% of families with CAKUT. However, WES has limitations and detection of copy number variations (CNV) is technically challenging, and CNVs causative of CAKUT have previously been detected in up to 16% of cases. Objective: To detect CNVs causing CAKUT in this WES cohort and increase the diagnostic yield. Design setting and participants: We performed a genome-wide single nucleotide polymorphism (SNP)-based CNV analysis on the same CAKUT cohort for whom WES was previously conducted. Outcome measurements and statistical analysis: We evaluated and classified the CNVs using previously published predefined criteria. Results and limitations: In a cohort of 170 CAKUT families, we detected a pathogenic CNV known to cause CAKUT in nine families (5.29%, 9/170). There were no competing variants on genome-wide CNV analysis or WES analysis. In addition, we identified novel likely pathogenic CNVs that may cause a CAKUT phenotype in three of the 170 families (1.76%). Conclusions: CNV analysis in this cohort of 170 CAKUT families previously examined via WES increased the rate of diagnosis of genetic causes of CAKUT from 13% on WES to 18% on WES + CNV analysis combined. We also identified three candidate loci that may potentially cause CAKUT. Patient summary: We conducted a genetics study on families with congenital anomalies of the kidney and urinary tract (CAKUT). We identified gene mutations that can explain CAKUT symptoms in 5.29% of the families, which increased the percentage of genetic causes of CAKUT to 18% from a previous study, so roughly one in five of our patients with CAKUT had a genetic cause. These analyses can help patients with CAKUT and their families in identifying a possible genetic cause.

Cystin genetic variants cause autosomal recessive polycystic kidney disease associated with altered Myc expression.

Mutation of the Cys1 gene underlies the renal cystic disease in the Cys1cpk/cpk (cpk) mouse that phenocopies human autosomal recessive polycystic kidney disease (ARPKD). Cystin, the protein product of Cys1, is expressed in the primary apical cilia of renal ductal epithelial cells. In previous studies, we showed that cystin regulates Myc expression via interaction with the tumor suppressor, necdin. Here, we demonstrate rescue of the cpk renal phenotype by kidney-specific expression of a cystin-GFP fusion protein encoded by a transgene integrated into the Rosa26 locus. In addition, we show that expression of the cystin-GFP fusion protein in collecting duct cells down-regulates expression of Myc in cpk kidneys. Finally, we report the first human patient with an ARPKD phenotype due to homozygosity for a deleterious splicing variant in CYS1. These findings suggest that mutations in Cys1/CYS1 cause an ARPKD phenotype in mouse and human, respectively, and that the renal cystic phenotype in the mouse is driven by overexpression of the Myc proto-oncogene.

Recessive Mutations in SYNPO2 as a Candidate of Monogenic Nephrotic Syndrome.

INTRODUCTION: Most of the approximately 60 genes that if mutated cause steroid-resistant nephrotic syndrome (SRNS) are highly expressed in the glomerular podocyte, rendering SRNS a "podocytopathy." METHODS: We performed whole-exome sequencing (WES) in 1200 nephrotic syndrome (NS) patients. RESULTS: We discovered homozygous truncating and homozygous missense mutation in SYNPO2 (synaptopodin-2) (p.Lys1124∗ and p.Ala1134Thr) in 2 patients with childhood-onset NS. We found SYNPO2 expression in both podocytes and mesangial cells; however, notably, immunofluorescence staining of adult human and rat kidney cryosections indicated that SYNPO2 is localized mainly in mesangial cells. Subcellular localization studies reveal that in these cells SYNPO2 partially co-localizes with α-actinin and filamin A-containing F-actin filaments. Upon transfection in mesangial cells or podocytes, EGFP-SYNPO2 co-localized with α-actinin-4, which gene is mutated in autosomal dominant SRNS in humans. SYNPO2 overexpression increases mesangial cell migration rate (MMR), whereas shRNA knockdown reduces MMR. Decreased MMR was rescued by transfection of wild-type mouse Synpo2 cDNA but only partially by cDNA representing mutations from the NS patients. The increased mesangial cell migration rate (MMR) by SYNPO2 overexpression was inhibited by ARP complex inhibitor CK666. SYNPO2 shRNA knockdown in podocytes decreased active Rac1, which was rescued by transfection of wild-type SYNPO2 cDNA but not by cDNA representing any of the 2 mutant variants. CONCLUSION: We show that SYNPO2 variants may lead to Rac1-ARP3 dysregulation, and may play a role in the pathogenesis of nephrotic syndrome.

Quantification of circulating endothelial cells in peripheral blood of systemic lupus erythematosus patients: a simple and reproducible method of assessing endothelial injury and repair.

BACKGROUND: Quantification of circulating endothelial cells (CECs) in peripheral blood is developing as a novel and reproducible method of assessing endothelial damage/dysfunction. Accordingly, elevated levels of CECs may be a marker of vascular injury in systemic lupus erythaematosus (SLE). This study was undertaken to assess the blood level of CECs in SLE and to correlate its level with the activity of the disease and to find out the possibility that the presence of increased numbers of CECs can be used as a marker of immune-mediated vessel damage. METHODS: The study included 33 patients with SLE and 20 healthy controls. They were subjected to clinical examination together with laboratory investigations including complete blood count (CBC), erythrocyte sedimentation rate (ESR), urine analysis, renal function test, C3, C4, ANA, anti-ds DNA antibody, antiphospholipid (IgM and IgG) antibodies and quantification of CECs in blood. CECs were calculated using flow cytometry after staining with a mouse anti-human CD45 antibody (pan-leukocyte marker), mouse anti-human CD146 antibody (endothelial cell marker) and 7-amino-actinomycin D (7-AAD) viability marker. CECs were defined as the live cells with 7-AAD negative, CD45 negative and CD146 positive. RESULTS: The number of CECs was significantly higher in patients with SLE compared with those in healthy control (mean +/- SD 38.6 +/- 21.2 versus 7.4 +/- 3.4). Furthermore, CECs were correlated positively with SLE disease activity index (SLEDAI) score, ESR and anti-ds DNA. CECs from patients with vasculitic skin lesions, renal and central nervous system (CNS) manifestation were significantly higher than patients free from the previous signs. CONCLUSIONS: An increased number of CECs observed in patients with SLE was associated with the active phase of the disease and may represent a marker of widespread endothelial injury.