Structure and thiazide inhibition mechanism of the human Na-Cl cotransporter.

The sodium-chloride cotransporter (NCC) is critical for kidney physiology1. The NCC has a major role in salt reabsorption in the distal convoluted tubule of the nephron2,3, and mutations in the NCC cause the salt-wasting disease Gitelman syndrome4. As a key player in salt handling, the NCC regulates blood pressure and is the target of thiazide diuretics, which have been widely prescribed as first-line medications to treat hypertension for more than 60 years5-7. Here we determined the structures of human NCC alone and in complex with a commonly used thiazide diuretic using cryo-electron microscopy. These structures, together with functional studies, reveal major conformational states of the NCC and an intriguing regulatory mechanism. They also illuminate how thiazide diuretics specifically interact with the NCC and inhibit its transport function. Our results provide critical insights for understanding the Na-Cl cotransport mechanism of the NCC, and they establish a framework for future drug design and for interpreting disease-related mutations.

Structure and mechanism of the cation-chloride cotransporter NKCC1.

Cation-chloride cotransporters (CCCs) mediate the electroneutral transport of chloride, potassium and/or sodium across the membrane. They have critical roles in regulating cell volume, controlling ion absorption and secretion across epithelia, and maintaining intracellular chloride homeostasis. These transporters are primary targets for some of the most commonly prescribed drugs. Here we determined the cryo-electron microscopy structure of the Na-K-Cl cotransporter NKCC1, an extensively studied member of the CCC family, from Danio rerio. The structure defines the architecture of this protein family and reveals how cytosolic and transmembrane domains are strategically positioned for communication. Structural analyses, functional characterizations and computational studies reveal the ion-translocation pathway, ion-binding sites and key residues for transport activity. These results provide insights into ion selectivity, coupling and translocation, and establish a framework for understanding the physiological functions of CCCs and interpreting disease-related mutations.

Paradoxes in magnesium transport in type 1 Bartter's syndrome and Gitelman's syndrome: a modeling analysis.

Type 1 Bartter's syndrome and Gitelman's syndrome are characterized by mutations in two key renal Na+ transporters, Na-K-2Cl cotransporter (NKCC2) and Na-Cl cotransporter (NCC). Since these two transporters play an important role in regulating magnesium (Mg2+) and calcium (Ca2+) transport in the kidney, significant alterations in the transport of these two electrolytes are observed in type 1 Bartter's syndrome and Gitelman's syndrome. In this study, we used our sex-specific computational models of renal electrolyte transport in rats to understand the complex compensatory mechanisms, in terms of alterations in tubular dimensions and ion transporter activities, that lead to Mg2+ and Ca2+ preservation or wasting in these two genetic disorders. Given the sexual dimorphism in renal transporter patterns, we also assessed how the magnitude of these alterations may differ between males and females. Model simulations showed that in type 1 Bartter's syndrome, nephron adaptations prevent salt wasting and favor Mg2+ preservation but not Ca2+, whereas in Gitelman's syndrome, those adaptations favor Ca2+ preservation over Mg2+. In addition, our models predicted that the compensatory alterations in tubular dimensions and ion transporter activities are stronger in females than in males.NEW & NOTEWORTHY Although changes in Ca2+ excretion in type 1 Bartter's syndrome and Gitelman's syndrome are well understood, Mg2+ excretion displays an interesting paradox. This computational modeling study provides insights into how renal adaptations in these two disorders impact Ca2+ and Mg2+ transport along different nephron segments. Model simulations showed that nephron adaptations favor Mg2+ preservation over Ca2+ in Bartter's syndrome and Ca2+ preservation over Mg2+ in Gitelman's syndrome and are stronger in females than in males.

Clinical Features and Unusual Heterozygous Mutations in Patients with Renal Hypokalemia.

BACKGROUND: Renal hypokalemia is associated with mutation. This study aimed to investigate the clinical features and pathogenic mutations in patients with renal hypokalemia. METHODS: The patients with hypokalemia were enrolled, and the renal function, thyroid function, renin-aldosterone system, urinary potassium excretion, and exome sequencing were performed. The correlation between the clinical phenotypes and causative genes was assessed. RESULTS: Five patients with hypokalemia were enrolled and diagnosed as tubular hypokalemia. The patients with common clinical manifestations were difficult to differentiate based on atypical laboratory findings. The results of the genetic analysis were as follows: both patient 1 and patient 2 were heterozygous for the c.C625T mutation of the KCNJ1 gene, which is responsible for Bartter syndrome. Patient 3 was heterozygous for the c.G298A mutation of the ATP6V1B1 gene, which is responsible for renal tubular acidosis. Patient 4 had a compound heterozygous mutation of c.G893A of the BSND gene, responsible for Bartter syndrome, and c.1029+5G>A, the ATP6V0A4 gene responsible for distal renal tubular acidosis. Patient 5 had Gitelman syndrome and carried the compound heterozygous mutations c.C1963T and c.G2029A of the SLC12A3 gene. All the above loci were known heterozygous mutations. CONCLUSIONS: The unusual heterozygous mutations were identified in five renal hypokalemia patients. Molecular diagnosis of tubular hypokalemia was conducive to accurate diagnosis and treatment.

Genotype-phenotype correlations in children with Gitelman syndrome.

BACKGROUND: This study aimed to analyze genotype-phenotype correlations in children with Gitelman syndrome (GS). METHODS: This multicenter retrospective study included 50 Korean children diagnosed with SLC12A3 variants in one or both alleles and the typical laboratory findings of GS. Genetic testing was performed using the Sanger sequencing except for one patient. RESULTS: The median age at the diagnosis was 10.5 years (interquartile range, 6.8;14.1), and 41 patients were followed up for a median duration of 5.4 years (interquartile range, 4.1;9.6). A total of 30 different SLC12A3 variants were identified. Of the patients, 34 (68%) had biallelic variants, and 16 (32%) had monoallelic variants on examination. Among the patients with biallelic variants, those (n = 12) with the truncating variants in one or both alleles had lower serum chloride levels (92.2 ± 3.2 vs. 96.5 ± 3.8 mMol/L, P = 0.002) at onset, as well as lower serum potassium levels (3.0 ± 0.4 vs. 3.4 ± 0.3 mMol/L, P = 0.016), and lower serum chloride levels (96.1 ± 1.9 vs. 98.3 ± 3.0 mMol/L, P = 0.049) during follow-up than those without truncating variants (n = 22). Patients with monoallelic variants on examination showed similar phenotypes and treatment responsiveness to those with biallelic variants. CONCLUSIONS: Patients with GS who had truncating variants in one or both alleles had more severe electrolyte abnormalities than those without truncating variants. Patients with GS who had monoallelic SLC12A3 variants on examination had almost the same phenotypes, response to treatment, and long-term prognosis as those with biallelic variants.

Adult classic Bartter syndrome: a case report with 5-year follow-up and literature review.

Bartter syndrome (BS) is a rare, inherited salt-losing renal tubular disorder characterized by secondary hyperaldosteronism, hypokalemia, hypochloremia, metabolic alkalosis, and low-to-normal blood pressure. Classic BS, or BS Type 3, the most common subtype in the Asian population, is caused by a molecular defect in ClC-Kb, a voltage-gated chloride channel in renal tubules, due to CLCNKB gene mutation. Because the onset of BS is more common in children than in adults, the diagnosis, treatment outcomes, genotype/phenotype association, and follow-up of adult-onset BS Type 3 are limited. This case report describes the findings in a 20-year-old man who was admitted with hypokalemic paralysis, with clinical manifestations were similar to those of Gitelman syndrome (GS); however, the patient was later diagnosed to have BS Type 3 through genetic testing (NM_000085.4 (CLCNKB): c.1052G>T). A literature review showed that no homozygous mutations have been reported to date. After 5 years of treatment and follow-up, we found that this genotype requires high levels of potassium and is prone to urinary protein and metabolic syndrome. Distinguishing adult-onset BS from GS is challenging in clinical practice. However, genetic diagnosis can help solve this problem effectively, and genotypes play a guiding role in treatment planning.

Interpreting epigenetic causes of recurrent hypokalemia and seizures: Gitelman syndrome co-exist with pseudohypoparathyroidism type 1B.

We describe a unique case of 27-year-old male with Gitelman syndrome (GS) co-exist with pseudohypoparathyroidism type 1B (PHP1B). The patient presented with a 5-year history of seizures, tetany, and numbness of the extremities. Further examinations showed recurrent hypokalemia, inappropriate kaliuresis, hypocalcemia, hyperphosphatemia, and elevated PTH levels. A novel variant of autosomal recessive GS (p.Val287Met SLC12A3) and a novel 492.3Kb deletion containing the whole of STX16, were discovered by a whole-exome sequencing. Following the diagnosis, calcitriol, calcium, and potassium supplements were started. Hematuria calcium and phosphorus levels, as well as blood potassium levels, have recovered and remained within normal ranges after 3 years of follow-up. Our findings have important consequences for supporting the idea that heterozygosity for variants have effects on the patients' clinical performance with autosomal recessive inheritance disorders. Further study is need for the putative effects of the variant. Likewise, further investigation with regards to the gene-gene interaction relations between GS and other electrolyte imbalance disorders is warranted.

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

BACKGROUND: 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. METHODS: 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.

Untangling the Uncertain Role of Overactivation of the Renin-Angiotensin-Aldosterone System with the Aging Process Based on Sodium Wasting Human Models.

Every individual at some point encounters the progressive biological process of aging, which is considered one of the major risk factors for common diseases. The main drivers of aging are oxidative stress, senescence, and reactive oxygen species (ROS). The renin-angiotensin-aldosterone system (RAAS) includes several systematic processes for the regulation of blood pressure, which is caused by an imbalance of electrolytes. During activation of the RAAS, binding of angiotensin II (ANG II) to angiotensin II type 1 receptor (AGTR1) activates intracellular nicotinamide adenine dinucleotide phosphate (NADPH) oxidase to generate superoxide anions and promote uncoupling of endothelial nitric oxide (NO) synthase, which in turn decreases NO availability and increases ROS production. Promoting oxidative stress and DNA damage mediated by ANG II is tightly regulated. Individuals with sodium deficiency-associated diseases such as Gitelman syndrome (GS) and Bartter syndrome (BS) show downregulation of inflammation-related processes and have reduced oxidative stress and ROS. Additionally, the histone deacetylase sirtuin-1 (SIRT1) has a significant impact on the aging process, with reduced activity with age. However, GS/BS patients generally sustain higher levels of sirtuin-1 (SIRT1) activity than age-matched healthy individuals. SIRT1 expression in GS/BS patients tends to be higher than in healthy age-matched individuals; therefore, it can be assumed that there will be a trend towards healthy aging in these patients. In this review, we highlight the importance of the hallmarks of aging, inflammation, and the RAAS system in GS/BS patients and how this might impact healthy aging. We further propose future research directions for studying the etiology of GS/BS at the molecular level using patient-derived renal stem cells and induced pluripotent stem cells.

[Genetic analysis of two patients with Gitelman syndrome].

OBJECTIVE: To explore the genetic etiology of two patients with Gitelman syndrome (GS). METHODS: Two patients who had presented at the Linyi People's Hospital in January and June 2022 respectively were selected as the study subjects. Peripheral blood samples of them were collected and subjected to whole exome sequencing (WES). Electrolyte levels in their serum and urine were detected. Candidate variants were verified by Sanger sequencing. PyMOL software was used to predict the impact of the variants on the protein structure. RESULTS: Patient 1 was a 27-year-old female with decreased serum levels of sodium, potassium, chloride and magnesium, along with decreased urine chloride and calcium. WES revealed that she has harbored compound heterozygous variants of the SLC12A3 gene, namely c.1456G>A (p.D486N) and c.179C>T (p.T60M). The former was inherited from her mother and known to be pathogenic. Patient 2 was a 4-year-old male with lower serum sodium, chloride and magnesium levels, and his serum potassium level was found to be critically low. He was found to harbor compound heterozygous variants of c.602-16G>A and c.805_806insTTGGCGTGGTCTCGGTCA (p.V268_T269insIGVVSV) of the SLC12A3 gene, which were inherited from his mother and father, respectively. Based on the guidelines from the American College of Medical Genetics and Genomics, both variants were predicted to be pathogenic (PVS1+PM2_Supporting+PP3; PVS1+PM2_Supporting+PM4). CONCLUSION: The above heterozygous variants of the SLC12A3 gene probably underlay the GS in these patients.

Spectrum of variants in a large Chinese Gitelman syndrome cohort.

Gitelman syndrome (GS) is caused by SLC12A3 biallelic variants. A previous study showed that large rearrangements (LRGs) of SLC12A3 accounted for the low sensitivity of genetic testing. However, a systematic screening for LRGs in Chinese GS patients is lacking. Massively parallel sequencing (MPS) and multiplex ligation-dependent probe amplification (MLPA) were performed to sequence the genomic DNA of patients with clinically diagnosed GS. Of 165 index cases, MPS identified 151 cases with two or more affected alleles and 14 cases with one variant allele. LRGs were detected by MLPA in 20 out of 27 cases, including 15 cases with suspected LRGs by MPS. Among these 20 cases with LRGs, the results of MPS and MLPA were identical in only 8 cases. Additional LRGs in 6 cases were detected by MLPA alone. In 6 cases, E4_E6del was identified by MPS, while E4_E5del and Intron6del were identified by MLPA. Among the 102 distinct variants, 30 are novel. LRGs were found in 20 cases (12.1%). LRGs were found in 12.1% of our Chinese GS patients cohort. We show that MPS and MLPA are two complementary techniques with the ability to improve the diagnostic yield of GS.

Novel SLC12A3 gene mutations and clinical characteristics in two pedigrees with Gitelman syndrome.

OBJECTIVE: Gitelman syndrome (GS) is an autosomal recessive tubulopathy resulting from inactivating mutations in the SLC12A3 gene that encodes the thiazide-sensitive sodium-chloride cotransporter (NCC). To date, more than 500 mutations have been identified in the SLC12A3 gene. In this study, we identified two new mutations in the SLC12A3 gene in two Chinese GS pedigrees. DESIGN, PATIENTS AND MEASUREMENTS: The clinical characteristics and laboratory examination of two suspected GS patients in our hospital were analyzed. In addition, two pedigrees including 11 members and 2 patients underwent SLC12A3 gene analysis. RESULTS: Both patients were middle-aged women with characteristics of hypokalemic metabolic alkalosis, hypomagnesemia, low level of urinary calcium and the elevated levels of renin-angiotensin-aldosterone system. So, they were clinically diagnosed as GS. Patient 2 also had type 2 diabetes and Graves' disease. Both patients were found to carry two mutations of SLC12A3 gene by Sanger direct sequencing, which were all compound heterozygous mutations. We identified three mutations in these two Chinese GS pedigrees, one of which was c.179C>T (Thr60Met). The novel c.2159G>T (p. Gly720Val) and c.2675T>C (p. Leu892Pro) mutations were strongly predicted to be pathogenic using four network programs-Polyphen-2, SIFT, Mutation Taster and LRT. CONCLUSIONS: We identified two novel SLC12A3 genetic variant [c.2159G>T (p.Gly720Val) and c.2675T>C (p.Leu892Pro)] in two Chinese GS pedigrees. The discovery of new mutations has enriched the spectrum of SLC12A3 genotypes.

Gitelman syndrome with Graves' disease leading to rhabdomyolysis: a case report and literature review.

A 14-year-old male patient who suffered from limb numbness, fatigue, and hypokalemia was considered Graves' disease (GD) complicated with thyrotoxic periodic paralysis (TPP) at the first diagnosis. Although with the treatment of antithyroid drugs, he developed severe hypokalemia and rhabdomyolysis (RM). Further laboratory tests revealed hypomagnesemia, hypocalciuria, metabolic alkalosis, hyperrenin, and hyperaldosteronemia. Genetic testing revealed compound heterozygous mutations in the SLC12A3 gene (c.506-1G > A, c.1456G > A) encoding the thiazide-sensitive sodium-chloride cotransporter, which presented a definitive diagnosis of Gitelman syndrome (GS). Moreover, gene analysis revealed his mother diagnosed with subclinical hypothyroidism due to Hashimoto's thyroiditis carried the c.506-1G > A heterozygous mutation in the SLC12A3 gene and his father carried the c.1456G > A heterozygous mutation in the SLC12A3 gene. His younger sister who had hypokalemia and hypomagnesemia carried the same compound heterozygous mutations as the proband and was diagnosed with GS as well, but with a much milder clinical presentation and better treatment outcome. This case suggested the potential relationship between GS and GD, clinicians should strengthen the differential diagnosis to avoid missed diagnosis.

Gitelman-Like Syndrome Caused by Pathogenic Variants in mtDNA.

BACKGROUND: 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. METHODS: 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. RESULTS: 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. CONCLUSION: 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.

[Regulation of kidney on potassium balance and its clinical significance].

Virtually all of the dietary potassium intake is absorbed in the intestine, over 90% of which is excreted by the kidneys regarded as the most important organ of potassium excretion in the body. The renal excretion of potassium results primarily from the secretion of potassium by the principal cells in the aldosterone-sensitive distal nephron (ASDN), which is coupled to the reabsorption of Na+ by the epithelial Na+ channel (ENaC) located at the apical membrane of principal cells. When Na+ is transferred from the lumen into the cell by ENaC, the negativity in the lumen is relatively increased. K+ efflux, H+ efflux, and Cl- influx are the 3 pathways that respond to Na+ influx, that is, all these 3 pathways are coupled to Na+ influx. In general, Na+ influx is equal to the sum of K+ efflux, H+ efflux, and Cl- influx. Therefore, any alteration in Na+ influx, H+ efflux, or Cl- influx can affect K+ efflux, thereby affecting the renal K+ excretion. Firstly, Na+ influx is affected by the expression level of ENaC, which is mainly regulated by the aldosterone-mineralocorticoid receptor (MR) pathway. ENaC gain-of-function mutations (Liddle syndrome, also known as pseudohyperaldosteronism), MR gain-of-function mutations (Geller syndrome), increased aldosterone levels (primary/secondary hyperaldosteronism), and increased cortisol (Cushing syndrome) or deoxycorticosterone (hypercortisolism) which also activate MR, can lead to up-regulation of ENaC expression, and increased Na+ reabsorption, K+ excretion, as well as H+ excretion, clinically manifested as hypertension, hypokalemia and alkalosis. Conversely, ENaC inactivating mutations (pseudohypoaldosteronism type 1b), MR inactivating mutations (pseudohypoaldosteronism type 1a), or decreased aldosterone levels (hypoaldosteronism) can cause decreased reabsorption of Na+ and decreased excretion of both K+ and H+, clinically manifested as hypotension, hyperkalemia, and acidosis. The ENaC inhibitors amiloride and Triamterene can cause manifestations resembling pseudohypoaldosteronism type 1b; MR antagonist spironolactone causes manifestations similar to pseudohypoaldosteronism type 1a. Secondly, Na+ influx is regulated by the distal delivery of water and sodium. Therefore, when loss-of-function mutations in Na+-K+-2Cl- cotransporter (NKCC) expressed in the thick ascending limb of the loop and in Na+-Cl- cotransporter (NCC) expressed in the distal convoluted tubule (Bartter syndrome and Gitelman syndrome, respectively) occur, the distal delivery of water and sodium increases, followed by an increase in the reabsorption of Na+ by ENaC at the collecting duct, as well as increased excretion of K+ and H+, clinically manifested as hypokalemia and alkalosis. Loop diuretics acting as NKCC inhibitors and thiazide diuretics acting as NCC inhibitors can cause manifestations resembling Bartter syndrome and Gitelman syndrome, respectively. Conversely, when the distal delivery of water and sodium is reduced (e.g., Gordon syndrome, also known as pseudohypoaldosteronism type 2), it is manifested as hypertension, hyperkalemia, and acidosis. Finally, when the distal delivery of non-chloride anions increases (e.g., proximal renal tubular acidosis and congenital chloride-losing diarrhea), the influx of Cl- in the collecting duct decreases; or when the excretion of hydrogen ions by collecting duct intercalated cells is impaired (e.g., distal renal tubular acidosis), the efflux of H+ decreases. Both above conditions can lead to increased K+ secretion and hypokalemia. In this review, we focus on the regulatory mechanisms of renal potassium excretion and the corresponding diseases arising from dysregulation.

[Clinical and genetic analysis of a case of Gitelman syndrome with comorbid Graves disease and adrenocortical adenoma].

OBJECTIVE: To report the clinical and genetic characteristics of a rare case of Gitelman syndrome with comorbid Graves disease and ACTH-independent adrenocortical adenoma. METHODS: A patient who had presented at the Nanchong Central Hospital on December 21, 2020 was selected as the study subject. Clinical data of the patient was collected. Whole-exome sequencing was carried out on DNA extracted from peripheral venous blood samples from the patient and her family members. RESULTS: The patient, a 45-year-old woman, was found to have Graves disease, ACTH-independent Cushing syndrome, hypokalemia and hypomagnesemia following the discovery of an adrenal incidentaloma. MRI scan had revealed a 3.8 cm × 3.2 cm mass in the left adrenal gland. The mass was removed by surgery and confirmed as adrenocortical adenoma. DNA sequencing revealed that the patient and her sister have both harbored compound heterozygous variants of the SLC12A3 gene, namely c.1444-10(IVS11)G>A and c.179(exon1)C>T (p.T60M), which were respectively inherited from their father and mother. Based on the guidelines from the American College of Medical Genetics and Genomics (ACMG), the c.1444-10(IVS11)G>A and c.179(exon1)C>T (p.T60M) were respectively classified as a variant of uncertain significance (PM2_Supporting+PP3) and a likely pathogenic variant (PM3_Strong+PM1+PP3). CONCLUSION: The conjunction of Gitelman syndrome with Graves disease and adrenal cortex adenoma is rather rare. The newly discovered c.1444-10(IVS11)G>A variant of the SLC12A3 gene, together with the heterozygous variant of c.179(exon1)C>T (p.T60M), probably underlay the pathogenesis in this patient.

The genetic spectrum of Gitelman(-like) syndromes.

PURPOSE OF REVIEW: Gitelman syndrome is a recessive salt-wasting disorder characterized by hypomagnesemia, hypokalemia, metabolic alkalosis and hypocalciuria. The majority of patients are explained by mutations and deletions in the SLC12A3 gene, encoding the Na+-Cl--co-transporter (NCC). Recently, additional genetic causes of Gitelman-like syndromes have been identified that should be considered in genetic screening. This review aims to provide a comprehensive overview of the clinical, genetic and mechanistic aspects of Gitelman(-like) syndromes. RECENT FINDINGS: Disturbed Na+ reabsorption in the distal convoluted tubule (DCT) is associated with hypomagnesemia and hypokalemic alkalosis. In Gitelman syndrome, loss-of-function mutations in SLC12A3 cause impaired NCC-mediated Na+ reabsorption. In addition, patients with mutations in CLCKNB, KCNJ10, FXYD2 or HNF1B may present with a similar phenotype, as these mutations indirectly reduce NCC activity. Furthermore, genetic investigations of patients with Na+-wasting tubulopathy have resulted in the identification of pathogenic variants in MT-TI, MT-TF, KCNJ16 and ATP1A1. These novel findings highlight the importance of cell metabolism and basolateral membrane potential for Na+ reabsorption in the DCT. SUMMARY: Altogether, these findings extend the genetic spectrum of Gitelman-like electrolyte alterations. Genetic testing of patients with hypomagnesemia and hypokalemia should cover a panel of genes involved in Gitelman-like syndromes, including the mitochondrial genome.

Calcium pyrophosphate crystal deposition in a cohort of 57 patients with Gitelman syndrome.

OBJECTIVE: Gitelman syndrome (GS) is the most frequent salt-wasting genetic tubulopathy and a source of hypokalaemia and hypomagnesemia. Chondrocalcinosis (CC) is a frequent feature of GS. The aim of our study was to determine the prevalence, distribution patterns, clinical phenotypes and risk factors for CC in GS. METHODS: This prospective study of a cohort of 57 patients with GS included a systematic screening for CC by peripheral joint radiography, cervical spine CT and joint US. The prevalence of cervical C1-C2 CC by CT was compared between 33 GS patients and sex- and age-matched controls. Clinical and biochemical features were analysed to identify factors associated with CC. RESULTS: Mean (s.d.) age of patients was 46.5 (12.4) years, 66.7% were women and 93.0% carried SLC12A3 mutations. Mean serum magnesium level was 0.60 (0.30) mmol/l. CC was observed in 79% of patients, with the highest prevalence at the cervical spine (81.8%) followed by the knee (52.6%), wrist (50.9%), ankle (38.6%), TM joint (36.4%), shoulder (33.3%), hip (22.8%), elbow (14.0%) and sclerochoroid (12.1%). Prevalence of CC at the C1-C2 level was higher in the GS cohort than control group (72.7% vs 9.1%) (adjusted odds ratio 21.0, 95% CI 2.8, 156.1, P = 0.003). Independent factors associated with CC were low serum magnesium level and age. CONCLUSION: GS was associated with widespread CC, favoured by aging and hypomagnesemia. The C1-C2 level was the most affected site. Follow-up of this unique cohort will help understanding the clinical consequences of CC, especially the precise characterization of pyrophosphate arthropathy.

Molecular diagnosis of adult patients with clinically unexplained hypokalemia without hypertension demonstrated a diagnostic yield of 30.5.

Hypokalemia is a common disorder in clinical settings; however, nonmolecular diagnostic testing cannot explain some causes of hypokalemia. To determine the etiology of clinically unexplained hypokalemia without hypertension (CUHypoNH) and to obtain a diagnostic yield of monogenic hypokalemia without hypertension in adults (MHNHA), we enrolled 82 patients with CUHypoNH for whole-exome sequencing or targeted gene sequencing of genes associated with 4000 monogenic disorders. Through molecular diagnosis, 25 patients were diagnosed with monogenic hypokalemia, and a diagnostic yield of 30.5% was obtained. Among patients with MHNHA, 18 patients (18/82, 22.0% and 72% of MHNHA) with Gitelman syndrome accounted for the largest proportion. Among the 29 diagnostic variants found, eight mutations have not been reported previously; these include three point mutations, one frameshift mutation, and four exon deletions. Based on the clinical presentation of patients with CUHypoNH, the diagnostic yield of monogenic hypokalemia was the highest for chronic asymptomatic hypokalemia (8/11, 72.7%). Twenty-one patients had concomitant hypomagnesemia, when accompanied with hypocalciuria, the molecular diagnostic yield of Gitelman syndrome increased to 88.2%. Overall, this study on hospitalized adult patients explored the etiology of CUHypoNH using high-throughput sequencing. Molecular diagnosis of CUHypoNH is clinically significant in guiding precision treatment and improving disease prognosis.

Na-Cl Co-transporter (NCC) gene inactivation is associated with improved bone microstructure.

Gitelman syndrome (GS) is the disease model of the inactivation of thiazide-sensitive sodium chloride cotransporter (NCC), which is believed to benefit bone mass and reduce fracture risk. In this study, we found that GS patients have superior bone microarchitecture, which is associated with the disease status. Several decreased bone parameters with aging in healthy controls were reversed in GS patients to a certain extent. PURPOSE: To evaluate the impact of the inactivation of NCC on bone turnover and microarchitecture in Gitelman syndrome patients. METHODS: A cross-sectional study was conducted in 45 GS patients (25 males and 20 females). Serum procollagen type 1 N-terminal propeptide (P1NP), ß-carboxy-terminal crosslinked telopeptide of type 1 collagen (ß-CTX), and osteocalcin were measured. High-resolution peripheral quantitative computed tomography (HR-pQCT) was conducted to evaluate bone microarchitecture in GS patients and age- and sex-matched healthy controls. Areal bone mineral density (aBMD) was measured by dual-energy X-ray absorptiometry (DXA) simultaneously. RESULTS: GS patients had a relatively lower level of ß-CTX. aBMD at several skeletal sites was improved in GS patients. HR-pQCT assessment revealed that GS patients had slightly thinner but significantly more compact trabecular bone (increased trabecular number and decreased thickness), notably decreased cortical porosity, and increased volume BMD (vBMD) at both the radius and tibia compared with controls. The disease severity, represented as the relationship with the minimum level of magnesium during the course and standard base excess, was associated with bone microarchitecture parameters after adjusting for age, sex, and BMI. The decreased vBMD and Tb.BV/TV, and increased Tb.Sp and Ct.Po with aging, were reversed in GS patients to a certain extent. CONCLUSION: GS patients have superior bone microarchitecture, which suggests that the inactivation of NCC might be beneficial for avoiding osteoporosis.