Salt-losing Tubulopathy Research Articles

AUP1 Regulates the Endoplasmic Reticulum-Associated Degradation and Polyubiquitination of NKCC2.

Inactivating mutations of kidney Na-K-2Cl cotransporter NKCC2 lead to antenatal Bartter syndrome (BS) type 1, a life-threatening salt-losing tubulopathy. We previously reported that this serious inherited renal disease is linked to the endoplasmic reticulum-associated degradation (ERAD) pathway. The purpose of this work is to characterize further the ERAD machinery of NKCC2. Here, we report the identification of ancient ubiquitous protein 1 (AUP1) as a novel interactor of NKCC2 ER-resident form in renal cells. AUP1 is also an interactor of the ER lectin OS9, a key player in the ERAD of NKCC2. Similar to OS9, AUP1 co-expression decreased the amount of total NKCC2 protein by enhancing the ER retention and associated protein degradation of the cotransporter. Blocking the ERAD pathway with the proteasome inhibitor MG132 or the α-mannosidase inhibitor kifunensine fully abolished the AUP1 effect on NKCC2. Importantly, AUP1 knock-down or inhibition by overexpressing its dominant negative form strikingly decreased NKCC2 polyubiquitination and increased the protein level of the cotransporter. Interestingly, AUP1 co-expression produced a more profound impact on NKCC2 folding mutants. Moreover, AUP1 also interacted with the related kidney cotransporter NCC and downregulated its expression, strongly indicating that AUP1 is a common regulator of sodium-dependent chloride cotransporters. In conclusion, our data reveal the presence of an AUP1-mediated pathway enhancing the polyubiquitination and ERAD of NKCC2. The characterization and selective regulation of specific ERAD constituents of NKCC2 and its pathogenic mutants could open new avenues in the therapeutic strategies for type 1 BS treatment.

2559. Antibiotic-induced Bartter-like syndrome - A Systematic Review

Abstract Background Bartter syndrome encompasses salt-losing tubulopathies characterized by hypokalemia, hypochloremic metabolic alkalosis, and hyperreninemic hyperaldosteronism with normal blood pressure. Acquired Bartter or Bartter-like syndrome (BLS) is commonly associated with the use of diuretics and antibiotics such as capreomycin, netilmicin, colistin, and amphotericin B. This systematic review was done to understand the time of occurrence of Bartter syndrome post-inciting drug and gain knowledge regarding its progression and management. Methods We searched PubMed, Google Scholar, Embase, medRxiv, and bioRxiv databases for articles published from 1986 to March 2022 using the keywords "Antibiotic," "Bartter Syndrome," "Pseudobartter Syndrome," and "Bartter-like." Studies that reported cases of BLS in adults or children with confirmation of the inciting drug were included. We excluded studies that did not provide sufficient details of the clinical presentation and outcome. Results We identified 31 studies, comprising 43 cases of BLS. The mean age of the patients was 41.1 years (range: 25–57 years), and 65.1% were female. The most common antimicrobial class associated with BLS was aminoglycosides (65.1%), followed by the polymyxin group. The most common antimicrobial was Gentamicin (41.8%), followed by Colistin (32.5%), Streptomycin, and Capreomycin (6.9% each). Hypokalemia(100%), metabolic alkalosis(97.2%), hypocalcemia(92.5%), and hypomagnesemia(100%) were reported in almost all patients. The median time of onset after exposure to the inciting agent was 10 days (range 7–18 days), and the median time for resolution after stopping the drug was 14 days (range 7–31 days). There was a complete resolution of the symptoms following discontinuation in all cases. Demographic and baseline characteristics of the patients Prevalence of electrolyte, acid-base disorder, and urinary electrolyte abnormalities Laboratory parameters among the reported cases Conclusion BLS is a rare but potentially reversible complication of various antimicrobials, most commonly aminoglycosides. The diagnosis requires a high index of suspicion and a careful review of the drug history. The prognosis is favorable, with a complete reversal of the state after drug discontinuation. Disclosures All Authors: No reported disclosures

Hypokalemia as a risk factor for prolonged QT interval and arrhythmia in inherited salt-losing tubulopathy

Purpose: To analyze electrocardiograms (ECGs) of patients with a salt-losing tubulopathy (SLT) and to determine the frequency and risk factors for long QT and arrhythmia.Methods: A total of 203 patients aged <19 years with SLT, specifically Bartter syndrome and Gitelman syndrome, who had a 12-lead ECG were included in this retrospective study. We analyzed the presence of an arrhythmia or prolonged corrected QT (QTc) on ECGs obtained for these patients. Demographic and laboratory data were compared between patients with abnormal and normal ECG findings.Results: Out of the 203 SLT patients, 38 (18.7%) underwent electrocardiography and 10 (40.0%) of 25 patients with inherited SLT had abnormal ECG findings, including prolonged QTc and arrhythmias. The abnormal ECG group had significantly lower serum potassium levels than the normal group (median [interquartile range]: 2.50 mmol/L [2.20–2.83] vs. 2.90 mmol/L [2.70–3.30], <i>p</i>=0.036), whereas other serum chemistry values did not show significant differences. The cutoff level for a significant difference in QTc interval was serum potassium level <2.50 mmol/L. One cardiac event occurred in a 13-year-old boy, who developed paroxysmal supraventricular tachycardia and underwent cardiac ablation. No sudden cardiac deaths occurred in this cohort.Conclusions: The incidence of ECG abnormalities in patients with inherited SLT was 40.0%, whereas the ECG screening rate was relatively low (18.7%). Therefore, we recommend ECG screening in patients with inherited SLT, especially in those with serum potassium level <2.50 mmol/L.

#2568 LONG-READ SEQUENCING IDENTIFIES NOVEL PATHOGENIC INTRONIC VARIANTS IN GITELMAN SYNDROME

Abstract Background and Aims Gitelman syndrome is a salt-losing tubulopathy characterized by hypokalemic alkalosis and hypomagnesemia. It is caused by homozygous recessive or compound heterozygous pathogenic variants in SLC12A3, which encodes the Na+-Cl− cotransporter (NCC). In up to 10% of patients with Gitelman syndrome, current genetic techniques detect only one specific pathogenic variant. This study aimed to identify a second pathogenic variant in introns, splice sites, or promoters to increase the diagnostic yield. Method Long-read sequencing of SLC12A3 was performed in 67 DNA samples from individuals with suspected Gitelman syndrome in whom a single likely pathogenic or pathogenic variant was previously detected. In addition, we sequenced DNA samples from 28 individuals with one variant of uncertain significance or no candidate variant. Midigene splice assays assessed the pathogenicity of novel intronic variants. Results A second likely pathogenic/pathogenic variant was identified in 45 (67%) patients. Those with two likely pathogenic/pathogenic variants had a more severe electrolyte phenotype than other patients. Of the 45 patients, 16 had intronic variants outside of canonic splice sites (nine variants, mostly deep intronic, six novel), whereas 29 patients had an exonic variant or canonic splice site variant. Midigene splice assays of the previously known c.1670-191C>T variant and intronic candidate variants demonstrated aberrant splicing patterns. Conclusion Intronic pathogenic variants explain an important part of the missing heritability in Gitelman syndrome. Long-read sequencing should be considered in diagnostic workflows for Gitelman syndrome, especially in patients with one pathogenic variant.

Machine Learning to Identify Genetic Salt-Losing Tubulopathies in Hypokalemic Patients

Clinically distinguishing patients with the inherited salt-losing tubulopathies (SLTs), Gitelman or Bartter syndrome (GS or BS) from other causes of hypokalemia (LK) patients is difficult, and genotyping is costly. We decided to identify clinical characteristics that differentiate SLTs from LK. A total of 66 hypokalemic patients with possible SLTs were recruited to a prospective observational cohort study at the University College London Renal Tubular Clinic, London. All patients were genotyped for pathogenic variants in genes which cause SLTs; 39 patients had pathogenic variants in genes causing SLTs. We obtained similar data sets from cohorts in Taipei and Kobe, as follows: the combined data set comprised 419 patients; 291 had genetically confirmed SLT. London and Taipei data sets were combined to train machine learning (ML) algorithms, which were then tested on the Kobe data set. Single biochemical variables (e.g., plasma renin) were significantly, but inconsistently, different between SLTs and LK in all cohorts. A decision table algorithm using serum bicarbonate and urinary sodium excretion (FENa) achieved a classification accuracy of 74%. This was superior to all the single biochemical variables identified previously. ML algorithms can differentiate true SLT in the context of a specialist clinic with some accuracy. However, based on routine biochemistry, the accuracy is insufficient to make genotyping redundant.

Gitelman syndrome diagnosed in the first trimester of pregnancy: a case report and literature review

Abstract Objectives Gitelman syndrome is a rare salt-losing tubulopathy caused by inactivating mutations in the SLC12A3 gene, which is expressed in the distal convoluted tubule and accounts for 5–10% of renal sodium reabsorption. Atypical symptoms and insidious conditions generally delay diagnosis until childhood or even adulthood. Here, we report the case of a 22-year-old Chinese woman who was admitted to our endocrinology department for severe hypokalemia during pregnancy. Case presentation The patient had no specific symptoms but exhibited hypokalemia, metabolic alkalosis, hypomagnesemia, hypocalciuria, hyperreninemia, hyperaldosteronism, and normal blood pressure. Together, these symptoms indicated the clinical diagnosis of Gitelman syndrome, which was confirmed by genetic analysis. Many drugs have limited safety data during early pregnancy, and optimum potassium and magnesium levels are necessary for a successful pregnancy. Conclusions Diagnosis and management of Gitelman syndrome are crucial during pregnancy to ensure the safety of the mother and fetus, especially during the first trimester.

Long-Read Sequencing Identifies Novel Pathogenic Intronic Variants in Gitelman Syndrome.

Gitelman syndrome is a salt-losing tubulopathy characterized by hypokalemic alkalosis and hypomagnesemia. It is caused by homozygous recessive or compound heterozygous pathogenic variants in SLC12A3 , which encodes the Na + -Cl - cotransporter (NCC). In up to 10% of patients with Gitelman syndrome, current genetic techniques detect only one specific pathogenic variant. This study aimed to identify a second pathogenic variant in introns, splice sites, or promoters to increase the diagnostic yield. Long-read sequencing of SLC12A3 was performed in 67 DNA samples from individuals with suspected Gitelman syndrome in whom a single likely pathogenic or pathogenic variant was previously detected. In addition, we sequenced DNA samples from 28 individuals with one variant of uncertain significance or no candidate variant. Midigene splice assays assessed the pathogenicity of novel intronic variants. A second likely pathogenic/pathogenic variant was identified in 45 (67%) patients. Those with two likely pathogenic/pathogenic variants had a more severe electrolyte phenotype than other patients. Of the 45 patients, 16 had intronic variants outside of canonic splice sites (nine variants, mostly deep intronic, six novel), whereas 29 patients had an exonic variant or canonic splice site variant. Midigene splice assays of the previously known c.1670-191C>T variant and intronic candidate variants demonstrated aberrant splicing patterns. Intronic pathogenic variants explain an important part of the missing heritability in Gitelman syndrome. Long-read sequencing should be considered in diagnostic workflows for Gitelman syndrome.

RF30 | PSAT136 Gitelman Syndrome and Primary Hyperparathyroidism, a Case Report

Abstract Introduction Gitelman syndrome, also known as familial hypokalemia-hypomagnesemia, is a rare autosomal recessive disorder that impairs the sodium chloride cotransporter (NCC) and magnesium channels in the renal distal convoluted tubule, causing salt-losing tubulopathy. It is characterized by hypokalemia, hypomagnesemia, hypocalciuria, and secondary hyperaldosteronism. Although hypocalciuria is a distinctive characteristic of Gitelman syndrome, it seldom can cause any change in the total plasma calcium level. Thus, the occurrence of hypercalcemia would need further investigation. Case presentation We reported a case of a 36-year-old normotensive man with a past medical history significant for bipolar depression disorder and a history of chronic atrial fibrillation secondary to chronic hypokalemia, hypomagnesemia, and mild hypercalcemia. He was diagnosed with Gitelman syndrome. However, he was noncompliant with spironolactone, potassium, and magnesium supplementation. He suffered a motor vehicle accident, and on presentation to the emergency room, he was found with a serum potassium of 2.5 mEq/L (reference value 3.5-5.1), serum magnesium of 0.8 mEq/L (reference value 1.6-2.6), total serum calcium of 11.3 mEq/L (reference value 8.4-10.2), and serum phosphorus of 2.7 (reference value 2.5-4.5 mg/dl). Random urine potassium was 122.1 mEq/L, and intact parathyroid hormone was 144 pg/mL (reference value 14-65). His CT neck scan showed a 1.5 cm nodule inferior to the right thyroid lobe pole consistent with an inferior parathyroid adenoma. However, a parathyroid radionuclide scan using technetium-99m MIBI showed no radiopharmaceutical localization for parathyroid adenoma. Conclusion Hypercalcemia is extremely rare in Gitelman syndrome, offset by the action of concomitant hypomagnesemia. Our case shows the importance of investigating other causes of hypercalcemia, such as primary hyperparathyroidism in patients with Gitelman syndrome. To our knowledge, Gitelman syndrome and primary hyperparathyroidism are an extremely rare association that is present in our case. Presentation: Saturday, June 11, 2022 1:00 p.m. - 3:00 p.m., Monday, June 13, 2022 12:48 p.m. - 12:53 p.m.

Abnormal Electrocardiogram Incidence and Risk Factors in Hypokalemic Salt-Losing Tubulopathy

Abnormal Electrocardiogram Incidence and Risk Factors in Hypokalemic Salt-Losing Tubulopathy

A case of Gitelman syndrome with membranous nephropathy

BackgroundGitelman syndrome (GS) is a rare autosomal recessive inherited salt-losing tubulopathy (SLT). Here, we report, for the first time, a case of GS overlapping nephrotic syndrome (NS) related to PLA2R-associated membranous nephropathy (MN).Case presentationWe described a male patient had a 4-year history of recurrent fatigue. Serum biochemistry revealed hypokalemia with renal potassium wasting, hypomagnesemia, metabolic alkalosis, hyperreninemia, hypocalciuria, as well as nephrotic-range proteinuria, hypoalbuminemia, and elevated serum anti-phospholipase A2 receptor (PLA2R) antibody. Gene sequencing identified compound heterozygous mutations in SLC12A3 [c.536T > A(p.V179D) and c.1456G > A(p.D486N)]. The unusual association of SLTs and nephrotic-range glomerular proteinuria prompted us to perform a renal biopsy. Renal biopsy showed idiopathic MN. Due to the potential to activate the sodium-chloride co-transporter (NCC) and cause hyperkalemia, tacrolimus was selected to treat NS. Following treatment with potassium chloride, magnesium oxide, low-dose glucocorticoid combined with tacrolimus, the fatigue significantly improved, and concurrently hypokalemia, hypomagnesemia were corrected and NS was remitted.ConclusionsRenal biopsy should be warranted for GS patients with moderate to nephrotic-range proteinuria. Tacrolimus was preferred to the management of GS patients with NS.

Detecting pathogenic deep intronic variants in Gitelman syndrome.

Gitelman syndrome (GS) is a rare, autosomal recessive, salt-losing tubulopathy caused by loss of function in the SLC12A3 gene (NM_000339.2), which encodes the natrium chloride cotransporter. The detection of homozygous or compound heterozygous SLC12A3 variants is expected in GS, but 18%-40% of patients with clinical GS carry only one mutant allele. Previous reports identified some pathogenic deep intronic variants in SLC12A3. Here, we report the screening of SLC12A3 deep intronic variants in 13 patients with suspected GS carrying one mutated SLC12A3 allele. Variant screening used the HaloPlex Target Enrichment System Kit capturing whole introns and the promotor region of SLC12A3, followed by SureCall variant analysis. Rare intronic variants (<1% frequency) were identified, and pathogenicity evaluated by the minigene system. Deep intronic variant screening detected seven rare SLC12A3 variants from six patients. Only one variant showed pathogenicity in the minigene system (c.602-16G>A, intron 4) through activation of a cryptic acceptor site. No variants were detected in the promotor region. Deep intronic screening identified only one pathogenic variant in patients with suspected GS carrying monoallelic SLC12A3 variants. Our results suggest that deep intronic variants partially explain the cause of monoallelic variants in patients with GS.

Gitelman‐like syndrome caused by pathogenic variants in mitochondrial DNA

Gitelman syndrome is the most frequent hereditary salt‐losing tubulopathy characterized by hypokalemic alkalosis and hypomagnesemia. Gitelman syndrome is caused by biallelic pathogenic variants in SLC12A3, encoding the Na+‐Cl‐ cotransporter (NCC) expressed in the distal convoluted tubule. Pathogenic variants in CLCNKB, HNF1B, FXYD2 or KCNJ10 may result in renal phenocopies of Gitelman syndrome, as they can lead to reduced NCC activity. Nevertheless, ±10% of patients with a Gitelman syndrome phenotype remain genetically unsolved.After identification of mitochondrial DNA (mtDNA) variants in three families with Gitelman syndrome‐like electrolyte abnormalities, 156 families were investigated for variants in MT‐TI and MT‐TF, encoding the transfer RNAs for phenylalanine and isoleucine. Mitochondrial respiratory chain function was assessed in patient fibroblasts. In NCC‐expressing HEK293 cells, mitochondrial dysfunction was induced to assess the effect on thiazide‐sensitive 22Na+ transport.Genetic investigations revealed four mtDNA variants in 13 families: m.591C&gt;T (n=7), m.616T&gt;C (n=1), m.643A&gt;G (n=1) (all in MT‐TF) and m.4291T&gt;C (n=4, in MT‐TI). Variants were near homoplasmic in affected individuals. Importantly, affected members of six families with an MT‐TF variant additionally suffered from progressive chronic kidney disease. Maximal mitochondrial respiratory capacity was reduced in patient fibroblasts, caused by dysfunction of oxidative phosphorylation complex IV. In vitro pharmacological inhibition of complex IV, mimicking the effect of the mtDNA variants, demonstrated an inhibitory effect on NCC phosphorylation and NCC‐mediated sodium uptake.Pathogenic mtDNA variants in MT‐TF and MT‐TI can cause a Gitelman syndrome‐like syndrome. Genetic investigation of mtDNA should be considered in patients with unexplained Gitelman syndrome‐like tubulopathies.

Parathyroid hormone and phosphate homeostasis in patients with Bartter and Gitelman syndrome: an international cross-sectional study.

Small cohort studies have reported high parathyroid hormone (PTH) levels in patients with Bartter syndrome and lower serum phosphate levels have anecdotally been reported in patients with Gitelman syndrome. In this cross-sectional study, we assessed PTH and phosphate homeostasis in a large cohort of patients with salt-losing tubulopathies. Clinical and laboratory data of 589 patients with Bartter and Gitelman syndrome were provided by members of the European Rare Kidney Diseases Reference Network (ERKNet) and the European Society for Paediatric Nephrology (ESPN). A total of 285 patients with Bartter syndrome and 304 patients with Gitelman syndrome were included for analysis. Patients with Bartter syndrome type I and II had the highest median PTH level (7.5pmol/L) and 56% had hyperparathyroidism (PTH>7.0pmol/L).Serum calcium was slightly lower in Bartter syndrome type I and II patients with hyperparathyroidism (2.42versus 2.49mmol/L; P=.038) compared to those with normal PTH levels and correlated inversely with PTH (rs -0.253; P=.009). Serum phosphate and urinary phosphate excretion did not correlate with PTH. Overall, 22% of patients had low serum phosphate levels (phosphate-standard deviation score < -2), with the highest prevalence in patients with Bartter syndrome type III (32%). Serum phosphate correlated with tubular maximum reabsorption of phosphate/glomerular filtration rate (TmP/GFR) (rs 0.699; P <.001), suggesting renal phosphate wasting. Hyperparathyroidism is frequent in patients with Bartter syndrome type I and II. Low serum phosphate is observed in a significant number of patients with Bartter and Gitelman syndrome and appears associated with renal phosphate wasting.

Gitelman-Like Syndrome Caused by Pathogenic Variants in mtDNA.

Gitelman syndrome is the most frequent hereditary salt-losing tubulopathy characterized by hypokalemic alkalosis and hypomagnesemia. Gitelman syndrome is caused by biallelic pathogenic variants in SLC12A3, encoding the Na+-Cl- cotransporter (NCC) expressed in the distal convoluted tubule. Pathogenic variants of CLCNKB, HNF1B, FXYD2, or KCNJ10 may result in the same renal phenotype of Gitelman syndrome, as they can lead to reduced NCC activity. For approximately 10 percent of patients with a Gitelman syndrome phenotype, the genotype is unknown. We identified mitochondrial DNA (mtDNA) variants in three families with Gitelman-like electrolyte abnormalities, then investigated 156 families for variants in MT-TI and MT-TF, which encode the transfer RNAs for phenylalanine and isoleucine. Mitochondrial respiratory chain function was assessed in patient fibroblasts. Mitochondrial dysfunction was induced in NCC-expressing HEK293 cells to assess the effect on thiazide-sensitive 22Na+ transport. Genetic investigations revealed four mtDNA variants in 13 families: m.591C>T (n=7), m.616T>C (n=1), m.643A>G (n=1) (all in MT-TF), and m.4291T>C (n=4, in MT-TI). Variants were near homoplasmic in affected individuals. All variants were classified as pathogenic, except for m.643A>G, which was classified as a variant of uncertain significance. Importantly, affected members of six families with an MT-TF variant additionally suffered from progressive chronic kidney disease. Dysfunction of oxidative phosphorylation complex IV and reduced maximal mitochondrial respiratory capacity were found in patient fibroblasts. In vitro pharmacological inhibition of complex IV, mimicking the effect of the mtDNA variants, inhibited NCC phosphorylation and NCC-mediated sodium uptake. Pathogenic mtDNA variants in MT-TF and MT-TI can cause a Gitelman-like syndrome. Genetic investigation of mtDNA should be considered in patients with unexplained Gitelman syndrome-like tubulopathies.

Gitelman syndrome: A first published clinical association with chronic pancreatitis, a case report and review of literature.

Introduction and importanceGitelman syndrome (GS) is an autosomal recessive, salt-losing tubulopathy, also referred to as familial hypokalemia-hypomagnesemia, caused by mutation of genes encoding the sodium chloride cotransporter (NCCT) and magnesium transporters in the thiazide sensitive segments of the distal convoluted tubule (DCT) of the nephron. Patients may present with a spectrum of clinical presentations and associations.Case presentationHere, we report a case of a 39-year-old female with Gitelman syndrome and chronic pancreatitis in the absence of well-known causes of CP. Her clinical and radiographic profile constituted an indication for surgical intervention, namely pancreatic head and body coring and pancreaticojejunostomy (Frey's procedure) (FP). On follow up 3 month later, the patient is pain-free and is satisfied. To the best of our knowledge and based on literature review, this is the first reported case of GS with CP.ConclusionThe purpose of this paper is to describe a case of CP in association with established GS as a first published clinical association, raising a possibility of another possible clinical manifestation of GS. Further observational studies are encouraged to support this association.

MTOR-Activating Mutations in RRAGD Are Causative for Kidney Tubulopathy and Cardiomyopathy.

Over the last decade, advances in genetic techniques have resulted in the identification of rare hereditary disorders of renal magnesium and salt handling. Nevertheless, approximately 20% of all patients with tubulopathy lack a genetic diagnosis. We performed whole-exome and -genome sequencing of a patient cohort with a novel, inherited, salt-losing tubulopathy; hypomagnesemia; and dilated cardiomyopathy. We also conducted subsequent in vitro functional analyses of identified variants of RRAGD, a gene that encodes a small Rag guanosine triphosphatase (GTPase). In eight children from unrelated families with a tubulopathy characterized by hypomagnesemia, hypokalemia, salt wasting, and nephrocalcinosis, we identified heterozygous missense variants in RRAGD that mostly occurred de novo. Six of these patients also had dilated cardiomyopathy and three underwent heart transplantation. We identified a heterozygous variant in RRAGD that segregated with the phenotype in eight members of a large family with similar kidney manifestations. The GTPase RagD, encoded by RRAGD, plays a role in mediating amino acid signaling to the mechanistic target of rapamycin complex 1 (mTORC1). RagD expression along the mammalian nephron included the thick ascending limb and the distal convoluted tubule. The identified RRAGD variants were shown to induce a constitutive activation of mTOR signaling in vitro. Our findings establish a novel disease, which we call autosomal dominant kidney hypomagnesemia (ADKH-RRAGD), that combines an electrolyte-losing tubulopathy and dilated cardiomyopathy. The condition is caused by variants in the RRAGD gene, which encodes Rag GTPase D; these variants lead to an activation of mTOR signaling, suggesting a critical role of Rag GTPase D for renal electrolyte handling and cardiac function.

MTOR-activating mutations of RRAGD cause a novel syndrome combining kidney tubulopathy and cardiomyopathy

Advances in genetic techniques have resulted in the identification of rare inherited disorders of renal magnesium and salt handling. Nevertheless, ∼20% of all patients with primary kidney tubulopathy have no diagnosis. We explore a large multicentric cohort presenting with a novel inherited salt-losing tubulopathy mostly characterized by hypomagnesemia, combined with early-onset dilated cardiomyopathy (DCM). Whole exome and genome sequencings were performed with various subsequent functional analyses of identified RRAGD variants in vitro . In 8 children from unrelated families with a tubulopathy characterized by hypomagnesemia, hypokalemia, salt-wasting, and nephrocalcinosis, we identified heterozygous missense variants in RRAGD that mostly occurred de novo . Six of these patients additionally suffered from DCM, and a heart transplantation was performed in 3 of them before the age of 25y. A dominant variant in RRAGD was simultaneously identified in 8 members of a large family, which segregated with a similar renal magnesium-losing phenotype. No DCM was detected in this family. RRAGD encodes Rag GTPase D mediating amino acid signaling to the mechanistic target of rapamycin complex 1 (mTORC1). RagD expression along the mammalian nephron includes the thick ascending limb and the distal convoluted tubule. The identified RRAGD variants were shown to induce a constitutive activation of mTOR signaling in vitro . Our findings establish a novel diseased phenotype combining kidney tubulopathy and cardiomyopathy caused by an activation of mTOR signaling suggesting a critical role of Rag GTPase D for renal electrolyte handling and cardiac function.

Examination of the predicted prevalence of Gitelman syndrome by ethnicity based on genome databases

Gitelman syndrome is an autosomal recessive inherited salt-losing tubulopathy. It has a prevalence of around 1 in 40,000 people, and heterozygous carriers are estimated at approximately 1%, although the exact prevalence is unknown. We estimated the predicted prevalence of Gitelman syndrome based on multiple genome databases, HGVD and jMorp for the Japanese population and gnomAD for other ethnicities, and included all 274 pathogenic missense or nonsense variants registered in HGMD Professional. The frequencies of all these alleles were summed to calculate the total variant allele frequency in SLC12A3. The carrier frequency and the disease prevalence were assumed to be twice and the square of the total allele frequency, respectively, according to the Hardy–Weinberg principle. In the Japanese population, the total carrier frequencies were 0.0948 (9.5%) and 0.0868 (8.7%) and the calculated prevalence was 0.00225 (2.3 in 1000 people) and 0.00188 (1.9 in 1000 people) in HGVD and jMorp, respectively. Other ethnicities showed a prevalence varying from 0.000012 to 0.00083. These findings indicate that the prevalence of Gitelman syndrome in the Japanese population is higher than expected and that some other ethnicities also have a higher prevalence than has previously been considered.

Defective claudin-10 causes a novel variation of HELIX syndrome through compromised tight junction strand assembly

Formation of claudin-10 based tight junctions (TJs) is paramount to paracellular Na+ transport in multiple epithelia. Sequence variants in CLDN10 have been linked to HELIX syndrome, a salt-losing tubulopathy with altered handling of divalent cations accompanied by dysfunctional salivary, sweat, and lacrimal glands. Here, we investigate molecular basis and phenotypic consequences of a newly identified homozygous CLDN10 variant that translates into a single amino acid substitution within the fourth transmembrane helix of claudin-10. In addition to hypohidrosis (H), electrolyte (E) imbalance with impaired urine concentrating ability, and hypolacrimia (L), phenotypic findings include altered salivary electrolyte composition and amelogenesis imperfecta but neither ichthyosis (I) nor xerostomia (X). Employing cellular TJ reconstitution assays, we demonstrate perturbation of cis- and trans-interactions between mutant claudin-10 proteins. Ultrastructures of reconstituted TJ strands show disturbed continuity and reduced abundance in the mutant case. Throughout, both major isoforms, claudin-10a and claudin-10b, are differentially affected with claudin-10b showing more severe molecular alterations. However, expression of the mutant in renal epithelial cells with endogenous TJs results in wild-type-like ion selectivity and conductivity, indicating that aberrant claudin-10 is generally capable of forming functional paracellular channels. Thus, mutant proteins prove pathogenic by compromising claudin-10 TJ strand assembly. Additional ex vivo investigations indicate their insertion into TJs to occur in a tissue-specific manner.

A Novel Homozygous CLCNKB Mutation of Classic Bartter Syndrome Presenting with Renal Cysts in 6-year-Old Identical Twin Boys : A Case Report

Bartter syndrome is an autosomal recessive hypokalemic salt-losing tubulopathy, and classic Bartter syndrome is associated with mutations in the CLCNKB gene. While chronic hypokalemia is known to induce renal cyst formation in different renal diseases, renal cyst formation in Bartter syndrome is rarely reported. Russian six-year-old identical male twins were referred to our hospital for the evaluation of renal cysts, which were incidentally detected on abdominal sonography due to diarrhea. Both twins had shown symptoms of polydipsia, polyuria, and nocturia since they were one year olds. Vital signs including blood pressure were normal in both twins. Renal sonography revealed nephromegaly, increased echogenicity of renal cortex, and various sized multiple cysts in both kidneys for both twins. Laboratory findings included hyponatremia, hypokalemia, hypochloremia, and metabolic alkalosis. Bartter syndrome with renal cysts were suspected. Genetic analysis for both twins confirmed a homozygous c.1614delC deletion on exon 15 of the CLCNKB gene, which was confirmed as a previously unreported variant to the best of our knowledge. They were managed with potassium chloride, nonsteroidal anti-inflammatory drugs, and angiotensin-converting-enzyme inhibitors. Metabolic alkalosis, hypokalemia, hypochloremia, and polyuria partially improved during the short course of treatment. This is the first report of a homozygous mutation in the CLCNKB gene in an identical twin, presenting with renal cysts. Key words: Bartter syndrome, Type 3, Chloride channels, Cysts, Hypokalemia, Twins, Monozygotic, Case report