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.

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.

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.

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.

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.

Mutations in multiple components of the nuclear pore complex cause nephrotic syndrome.

Steroid-resistant nephrotic syndrome (SRNS) almost invariably progresses to end-stage renal disease. Although more than 50 monogenic causes of SRNS have been described, a large proportion of SRNS remains unexplained. Recently, it was discovered that mutations of NUP93 and NUP205, encoding 2 proteins of the inner ring subunit of the nuclear pore complex (NPC), cause SRNS. Here, we describe mutations in genes encoding 4 components of the outer rings of the NPC, namely NUP107, NUP85, NUP133, and NUP160, in 13 families with SRNS. Using coimmunoprecipitation experiments, we showed that certain pathogenic alleles weakened the interaction between neighboring NPC subunits. We demonstrated that morpholino knockdown of nup107, nup85, or nup133 in Xenopus disrupted glomerulogenesis. Re-expression of WT mRNA, but not of mRNA reflecting mutations from SRNS patients, mitigated this phenotype. We furthermore found that CRISPR/Cas9 knockout of NUP107, NUP85, or NUP133 in podocytes activated Cdc42, an important effector of SRNS pathogenesis. CRISPR/Cas9 knockout of nup107 or nup85 in zebrafish caused developmental anomalies and early lethality. In contrast, an in-frame mutation of nup107 did not affect survival, thus mimicking the allelic effects seen in humans. In conclusion, we discovered here that mutations in 4 genes encoding components of the outer ring subunits of the NPC cause SRNS and thereby provide further evidence that specific hypomorphic mutations in these essential genes cause a distinct, organ-specific phenotype.

Tumor necrosis factor-α -308 A/G gene polymorphism in children with juvenile idiopathic arthritis: relation to disease activity, damage, and functional status.

The study aims to evaluate the clinical significance of serum levels of tumor necrosis factor alpha (TNF-α) and -308 A/G promoter polymorphism in juvenile idiopathic arthritis (JIA) patients and find any association to the subsets, clinical and laboratory features, disease activity, and damage as well as functional disability. Forty-eight JIA children and 30 controls were included in the present study. Juvenile arthritis disease activity score in 27 joints (JADAS-27) was calculated, juvenile arthritis damage index (JADI) was assessed, and Childhood Health Assessment Questionnaire (CHAQ) measured the functional status. Serum TNF-α was assayed by ELISA and gene (-308) promoter polymorphism was determined by polymerase chain reaction. The 48 JIA children (mean age 11.5 ± 2.8 years) were 13 systemic, 17 oligoarticular, and 18 polyarticular onset. The serum TNF-α was significantly higher in patients (90.4 ± 6.3 ng/ml) compared to control (3.5 ± 2.6 ng/ml) (p < 0.0001) with a tendency to be higher in the polyarticular subtype. All controls had TNF-α -308 GG alleles. The frequency of GG genotype tended to be higher in systemic onset compared to oligoarticular and polyarticular subtypes. The serum TNF-α significantly correlated with JADAS-27 (r = 0.32, p = 0.03) and CHAQ (r = 0.37, p = 0.01) and negatively with the presence of GG alleles (r = -0.48, p = 0.001). The GG alleles were significantly negatively associated with C-reactive protein (r = -0.32, p = 0.03) with a tendency to negatively correlate with JADAS-27, CHAQ, and JADI-extrarticular (r = -0.28, p = 0.06; r = -0.25, p = 0.09 and r = -0.25, p = 0.09, respectively). There is evidence of a possible influence of the -308 SNP promoter position on the production of TNF-α, the severity of JIA which may consequently influence the response to anti-TNF-α treatment.

ARHGDIA mutations cause nephrotic syndrome via defective RHO GTPase signaling.

Nephrotic syndrome (NS) is divided into steroid-sensitive (SSNS) and -resistant (SRNS) variants. SRNS causes end-stage kidney disease, which cannot be cured. While the disease mechanisms of NS are not well understood, genetic mapping studies suggest a multitude of unknown single-gene causes. We combined homozygosity mapping with whole-exome resequencing and identified an ARHGDIA mutation that causes SRNS. We demonstrated that ARHGDIA is in a complex with RHO GTPases and is prominently expressed in podocytes of rat glomeruli. ARHGDIA mutations (R120X and G173V) from individuals with SRNS abrogated interaction with RHO GTPases and increased active GTP-bound RAC1 and CDC42, but not RHOA, indicating that RAC1 and CDC42 are more relevant to the pathogenesis of this SRNS variant than RHOA. Moreover, the mutations enhanced migration of cultured human podocytes; however, enhanced migration was reversed by treatment with RAC1 inhibitors. The nephrotic phenotype was recapitulated in arhgdia-deficient zebrafish. RAC1 inhibitors were partially effective in ameliorating arhgdia-associated defects. These findings identify a single-gene cause of NS and reveal that RHO GTPase signaling is a pathogenic mediator of SRNS.

Prevalence of celiac disease, Helicobacter pylori and gastroesophageal reflux in patients with refractory iron deficiency anemia.

OBJECTIVE: The aim of this article is to determine the prevalence of celiac disease (CD), Helicobacter pylori (H. pylori) and gastroesophageal reflux (GER) in patients with resistant iron deficiency anemia (IDA). PATIENTS: The study included 25 patients <18 years of age with refractory IDA (not responding to iron therapy for 3 months in a dose of 6 mg elemental iron/kg/day). METHODS: All patients included in the study were subjected to careful history taking and thorough clinical examination. Blood sample was taken for analysis of antibodies for CD including: antigliadin antibody (AGA), antiendomysial antibody (EMA), antireticulin antibody (ARA) and antitissue Transglutaminase (tTg) IgG antibody. Anti-H. pylori IgG antibodies and a (13)C-urea breath test (UBT) was done to all patients to diagnose H. pylori. Upper gastrointestinal tract endoscopy was done for all patients to evaluate for the presence of some etiologies of intractable anemia as chronic blood loss. These included: CD, H. pylori infection and GER. The upper gastrointestinal tract endoscopy was also done to evaluate the presence of bleeding spots, ulcers or angiomatous malformations. In addition, gastric antral biopsies were taken for diagnosis of H. pylori infection by the following tests: rapid urease test, histopathological examination and culture. RESULTS: CD was positive in 11 out of 25 patients (44%), H. pylori infection in 12 out of 25 patients (48%), while GER was diagnosed in 11 out of 25 patients (44%). Patients with CD had age of presentation < or =2 years in two patients (18.2%) while the remaining nine patients (81.8%) had age of presentation >2 years and it was statistically significant (p = 0.05*). Also patients with H. pylori had age of presentation < or =4 years in five patients (41.7%) and the remaining seven patients (81.8%) had age of presentation >4 years and it was statistically significant (p = 0.03*). Logistic regression analysis demonstrated that the risk factors for severity of anemia were age of patients and duration of anemia. On the other hand, other parameters have no significant influence on the severity of anemia. Also risk factors of short stature were age of presentation of anemia, degree of anemia and H. pylori infection. AGA had the highest sensitivity (100%) followed by antiendomysium antibody (81.8%) while the tTG antibody had the highest specificity (85.7%) for diagnosis of CD. UBT and histopathology had the highest sensitivity (100%) for diagnosis of H. pylori while rapid urease test, culture, H. pylori stool antigen and anti-H. pylori IgG antibody had the highest specificity (100%). In conclusion, refractory IDA may be due to clinically unapparent H. pylori gastritis and CD. CD is one of the most common causes of intestinal malabsorption during childhood which leads to impairment of iron absorption. Apart from offering them gluten-free diet rich in iron, early detection and treatment of IDA and prophylactic iron and folic acid supplementation will go a long way to optimize their mental and psychological functions. Eradication of H. pylori infection with concomitant iron therapy should correct the anemia.