Whole-exome sequencing in diagnosing 2 cases of Gitelman syndrome / 中南大学学报(医学版)

Two patients with Gitelman syndrome were admitted to the Department of Endocrinology, Third Xiangya Hospital of Central South University. The genomic DNA from the patients' peripheral blood was extracted and the whole-exome sequencing was performed to detect the possible mutations. The function of the mutation sites was analyzed by bioinformatics software. Through whole-exome sequencing and Sanger sequencing, we have found that 2 patients with Gitelman syndrome carried compound heterozygous mutations of SLC12A3 gene, which were c.486_490delTACGGinsA, p.R943W, p.D486N, and p.R928C. Among them, c.486_490delTACGGinsA insertion deletion mutation causes frame shift and protein truncation. The p.R943W, p.D486N, and p.R928C of SLC12A3 gene were predicted to be pathogenic mutations by SIFT, PolyPhen2, and Mutation Taster. These 4 mutations were all reported, but p.R943W was first reported in Chinese population. Gitelman syndrome is rare in clinic and the rate of missed diagnosis is high. Early genetic analysis in patients with Gitelman syndrome is helpful to determine the etiology and guide the treatment.

The mechanism of blood pressure regulation by high potassium diet in the kidney / 生理学报

Hypertension is one of the strongest risk factors for cardiovascular diseases, cerebral stroke, and kidney failure. Lifestyle and nutrition are important factors that modulate blood pressure. Hypertension can be controlled by increasing physical activity, decreasing alcohol and sodium intake, and stopping tobacco smoking. Chronic kidney disease patients often have increased blood pressure, which indicates that kidney is one of the major organs responsible for blood pressure homeostasis. The decrease of renal sodium reabsorption and increase of diuresis induced by high potassium intake is critical for the blood pressure reduction. The beneficial effect of a high potassium diet on hypertension could be explained by decreased salt reabsorption by sodium-chloride cotransporter (NCC) in the distal convoluted tubule (DCT). In DCT cells, NCC activity is controlled by with-no-lysine kinases (WNKs) and its down-stream target kinases, Ste20-related proline-alanine-rich kinase (SPAK) and oxidative stress-responsive 1 (OSR1). The kinase activity of WNKs is inhibited by intracellular chloride ([Cl-]i) and WNK4 is known to be the major WNK positively regulating NCC. Based on our previous studies, high potassium intake reduces the basolateral potassium conductance, decreases the negativity of DCT basolateral membrane (depolarization), and increases [Cl-]i. High [Cl-]i inhibits WNK4-SPAK/OSR1 pathway, and thereby decreases NCC phosphorylation. In this review, we discuss the role of DCT in the blood pressure regulation by dietary potassium intake, which is the mechanism that has been best dissected so far.

Analysis of clinical features and genetic variants among 12 children with Gitelman syndrome / 中华医学遗传学杂志

OBJECTIVE@#To summarize clinical manifestations and results of genetic testing in 12 children with Gitelman syndrome (GS).@*METHODS@#Clinical data of the children was collected. Whole exome sequencing(WES) was carried out to screen potential variants of genomic DNA. Candidate variants were verified by Sanger sequencing.@*RESULTS@#The patients have included 10 boys and 2 girls, whom were diagnosed at between 2.8 to 15.0 year old. Six patients were due to infections, 5 were due to short stature, and 1 was due to lower limb weakness. All patients were found to carry variants of SLC12A3 gene, which included 11 with compound heterozygous variants and 1 with homozygous variant. All of the 19 alleles of the SLC12A3 gene carried by the patients were delineated, which included 15 missense variants, 2 frameshift variants and 2 splice region variants. These variants were unreported previously, which included c.578_582dupCCACC (p.Asn195Profs*109), c.251C>T (p.Pro84Leu) and c.2843G>A (p.Trp948X).@*CONCLUSION@#The clinical symptoms of GS in children are atypical and often seen in older children. For children with occasional hypokalemia associated with growth failure, GS should be suspected. The majority of GS children carry two pathogenic variants of the SLC12A3 gene, mainly compound heterozygotes, among which p.Thr60Met is the most common one. The discovery of new variants has enriched the spectrum of SLC12A3 gene variants.

Correlation between genotypes with metabolic markers and microstructure of bones in children with Gitelman syndrome / 中华医学遗传学杂志

OBJECTIVE@#To explore the correlation between the genotypes and metabolic markers and microstructure of bones in children with Gitelman syndrome (GS).@*METHODS@#For 15 children with GS and 10 healthy individuals, baseline data and bone metabolic markers including parathyroid hormone, alkaline phosphatase, osteocalcin, N-terminal propeptide of type I procollagen, beta isomer of the C-terminal telopeptide of type I collagen and 25-hydroxyvitamin D, high-resolution peripheral quantitative computed tomography indicators (volumetric bone mineral density, bone microstructure indicators) were collected. Genetic testing was carried out to determine their genotypes.@*RESULTS@#The volumetric bone mineral density, bone geometry and bone microstructure parameters of the GS group were better than those of the healthy controls (P<0.05). Variants of the SLC12A3 gene were identified in 9 of the 15 patients but none of the 10 healthy controls.@*CONCLUSION@#The phenotype of GS children is influenced by the interaction of genetic variants, though the phenotype associated with high frequency mutations showed no specificity. There is also a correlation between their genotype and the bone microstructure.

Identification of pathological variants of SLC12A3 gene in a pedigree affected with Gitelman syndrome / 中华医学遗传学杂志

OBJECTIVE@#To detect pathological variants of the SLC12A3 gene in a Chinese pedigree affected with Gitelman syndrome (GS).@*METHODS@#Clinical data and peripheral blood samples of the proband and his family members were collected. All exons of the SLC12A3 gene were amplified by PCR and subjected to Sanger sequencing.@*RESULTS@#Sanger sequencing has revealed that the proband has carried a c.486_489 delTACG (p.Ile162Met fs*8) deletion and a heterozygous c.2890C>T (p.Arg964Trp) missense variant in the SLC12A3 gene. Neither variant was reported previously and was not found among healthy controls.@*CONCLUSION@#The c.486_489delTACG (p.Ile162Met fs*8) and c.2890C>T (p.Arg964Trp) variants of the SLC12A3 gene probably underlay the GS in the proband. Above discovery has enriched the variant spectrum of GS.

Rheumatoid arthritis accompanied by Gitelman syndrome / 영남의대학술지

Gitelman syndrome is a condition caused by a mutation of the thiazide sensitive Na-Cl cotransporter gene on the distal convoluted tubule. It results in a variety of clinical features, including hypokalemia, hypomagnesemia, hypocalciuria, and metabolic alkalosis. It is often diagnosed in asymptomatic adults presented with unexplained hypokalemia; however, it is sometimes associated with muscular cramps, numbness, fatigue, weakness, or paralysis. We experienced a case of rheumatoid arthritis accompanied by Gitelman syndrome, presented with hand tremor. We diagnosed her using renal clearance study and genetic analysis. Here, we report our experiences regarding this case along with a literature review.

Rheumatoid arthritis accompanied by Gitelman syndrome / 영남의대학술지

Gitelman syndrome is a condition caused by a mutation of the thiazide sensitive Na-Cl cotransporter gene on the distal convoluted tubule. It results in a variety of clinical features, including hypokalemia, hypomagnesemia, hypocalciuria, and metabolic alkalosis. It is often diagnosed in asymptomatic adults presented with unexplained hypokalemia; however, it is sometimes associated with muscular cramps, numbness, fatigue, weakness, or paralysis. We experienced a case of rheumatoid arthritis accompanied by Gitelman syndrome, presented with hand tremor. We diagnosed her using renal clearance study and genetic analysis. Here, we report our experiences regarding this case along with a literature review.

A novel homozygous mutation in the solute carrier family 12 member 3 gene in a Chinese family with Gitelman syndrome

Loss of function of mutated solute carrier family 12 member 3 (SLC12A3) gene is the most frequent etiology for Gitelman syndrome (GS), which is mainly manifested by hypokalemia, hypomagnesemia and hypocalciuria. We report the genetic characteristics of one suspicious Chinese GS pedigree by gene sequencing. Complete sequencing analysis of the SLC12A3 gene revealed that both the proband and his elder sister had a novel homozygous SLC12A3 mutation: c.2099T>C and p.Leu700Pro. Moreover, the SLC12A3 genes of his mother and daughter encoded the same mutated heterozygote. It was noted that in this pedigree, only the proband complained about recurrent episodes of bilateral lower limb weakness over 8 years, while his elder sister, mother and daughter did not present symptoms. The inconsistent clinical features of this pedigree implied that besides diverse phenotypes possibly originated from the same genotype, gender difference may also dominate the variant GS phenotypes. Further genetic and proteomic research are needed to investigate the precise mechanisms of GS, including the study of specific ethnicities.

Value of Chloride Clearance Test in Differential Diagnosis of Gitelman Syndrome / 中国医学科学院学报

Objective To investigate the value of chloride clearance test in differential diagnosis of Gitelman syndrome (GS). Methods For patients with hypokalemic metabolic alkalosis and highly suspected GS,clinical data were documented and SLC12A3 gene screening was performed as gold standard to diagnose GS. Hydrochlorothiazide (HCT) test and furosemide (FUR) test were performed according to the standard process. Baseline and maximal increasement of chloride excretion fraction (FECl,the net and relative increase measured as εFECl) were compared between patients and controls to evaluated the reaction to the corresponding diuretics. Receiver operating characteristic (ROC) curve was used to evaluate the sensitivity and specificity of HCT test in GS diagnosis. Results Totally 27 patients and 20 health controls received HCT test. Among those patients,23 were diagnosed with GS genetically. When using the net and relative εFECl to diagnose GS,the areas under the ROC curve were 0.987 (95% CI:0.963～1.000,P<0.001) and 0.984 (95%CI:0.950～1.000,P<0.001),respectively. When a reasonable cutoff value for εFECl was selected,the sensitivity and specificity were both higher than 95%. Eight patients received both HCT test and FUR test. Five of them showed decreased reaction to HCT(net εFECl≤2.86% or relative εFECl≤223%),while normal reaction to FUR.SLC12A3 mutations confirmed their GS. Three patients with blunt reaction to FUR showed normal reaction to HCT,finally they were diagnosed as BS clinically because no SLC12A3 gene mutation was detected. Conclusion Comprehensive application of HCT test and FUR test to evaluate the diuretic reaction can effectively differentiate GS and BS.

Clinical Study of a Newly Diagnosed Case of Gitelman Syndrome in a Patient Monitored for Liddle Syndrome

A 55-year-old man who had been monitored for Liddle syndrome in the nephrology division for 15 years visited again Inje University Sanggye Paik Hospital for a newly developed electrolyte disorder. Because his blood pressure was normal and he showed hypomagnesemia and hypokalemia, a renal clearance test and renal biopsy were conducted for suspected Gitelman syndrome. The patient was diagnosed with Gitelman syndrome, which has been previously reported 12 cases in South Korea. The renal clearance test revealed a disorder of the Na-Cl cotransporter (NCCT) in the distal tubule, while the renal biopsy revealed partial expression of NCCT, typical of Gitelman syndrome. Currently, the patient is being monitored, and is receiving oral administration of calcium and magnesium.

Mutations in SLC12A3 and CLCNKB and Their Correlation with Clinical Phenotype in Patients with Gitelman and Gitelman-like Syndrome

Gitelman's syndrome (GS) is caused by loss-of-function mutations in SLC12A3 and characterized by hypokalemic metabolic alkalosis, hypocalciuria, and hypomagnesemia. Long-term prognosis and the role of gene diagnosis in GS are still unclear. To investigate genotype-phenotype correlation in GS and Gitelman-like syndrome, we enrolled 34 patients who showed hypokalemic metabolic alkalosis without secondary causes. Mutation analysis of SLC12A3 and CLCNKB was performed. Thirty-one patients had mutations in SLC12A3, 5 patients in CLCNKB, and 2 patients in both genes. There was no significant difference between male and female in clinical manifestations at the time of presentation, except for early onset of symptoms in males and more profound hypokalemia in females. We identified 10 novel mutations in SLC12A3 and 4 in CLCNKB. Compared with those with CLCNKB mutations, patients with SLC12A3 mutations were characterized by more consistent hypocalciuria and hypomagnesemia. Patients with 2 mutant SLC12A3 alleles, compared with those with 1 mutant allele, did not have more severe clinical and laboratory findings except for lower plasma magnesium concentrations. Male and female patients did not differ in their requirement for electrolyte replacements. Two patients with concomitant SLC12A3 and CLCNKB mutations had early-onset severe symptoms and showed different response to treatment. Hypocalciuria and hypomagnesemia are useful markers in differentiation of GS and classical Bartter's syndrome. Gender, genotypes or the number of SLC12A3 mutant alleles cannot predict the severity of disease or response to treatment.

A Pedigree with c.179 Cytosine to Threonine Missense Mutation of SLC12A3 Gene Presenting Gitelman's Syndrome

A 42-year-old man came to the hospital presenting chest discomfort and general weakness. He had come to the hospital with the same symptoms 3 months ago and 12 years prior. His laboratory test showed hypokalemia, hypomagnesemia and hypocalciuria. The arterial blood gas analysis showed hypochloremic metabolic alkalosis. He had an ultrasonography guided renal biopsy, the result was normal at light microscopy and immunofluorescence microscopy. However, a special stain for Na-Cl cotransporter was weakly expressed compared with the control. The patient and his family underwent genetic sequencing about the SLC12A3 gene. He had a homozygous mutation in the 179th nucleotide of Exon 1 on the SLC12A3 gene (p.Thr60Met) and his parents and sisters were diagnosed as carrier state of Gitelman's syndrome (GS). GS is an inherited tubular disorder which presents mild hypokalemia, hypomagnesemia and hypocalciuria. Since the symptoms and laboratory results are not severe, it can go unnoticed by physicians. Herein we present a family with GS, diagnosed by genetic sequencing.

Absence of intact thiazide-sensitive sodium-chloride cotransporter in the renal tissue of a Gitelman's syndrome patient / 대한내과학회지

BACKGROUND: Gitelman's syndrome is an autosomal recessive renal tubular disorder characterized by hypokalemic metabolic alkalosis, hypomagnesemia, and hypocalciuria. It is known to be caused by a mutation of SLC12A3 gene coding the sodium-chloride cotransporter (NCCT) in the distal tubule. The defect of NCCT in human renal tissues has not been investigated, and we tested whether the defect of NCCT can be detected in renal tissue of a patient with Gitelman's syndrome by using immunohistochemistry. METHODS: In an adult patient with Gitelman's syndrome, blood and urine samples were collected for measurement of biochemical parameters. Renal clearance study and gene analysis were performed. Immunohistochemistry was performed on the renal tissue of the patient using a rabbit polyclonal antibody directed against a synthetic peptide corresponding to a portion in the amino terminal tail for human NCCT. Normal human renal tissues from surgical nephrectomy due to renal cell carcinoma and renal biopsy tissues from patients with glomerulonephritis but without any electrolyte disturbance were used as controls. RESULTS: The patient had hypokalemic metabolic alkalosis, hypocalciuria and hypomagnesemia. Renal clearance study revealed a decrease in distal fractional chloride reabsorption after the administration of furosemide. SLC12A3 gene mutation (S967F) was found by direct sequencing method. Immunohistochemistry showed the absence of NCCT staining in the renal tissue of the patient. On the other hand, the immunostaining of other transporters was all positive in renal tissues from both Gitelman's syndrome patients and controls. CONCLUSIONS: We report the absence of intact NCCT in the renal tissue of a Gitelman's syndrome patient.

Two Cases of Gitelman's Syndrome Diagnosed by Renal Clearance Study

Gitelman's syndrome is an autosomal recessive disorder characterized by hypokalemic metabolic alkalosis, hypomagnesemia, and hypocalciuria that has recently been reported to be linked to thiazide-sensitive Na-Cl cotransporter gene mutation. In this study, we performed renal clearance studies to differentiate Gitelman's from Bartter's syndrome and to confirm the diagnosis in two patients clinically diagnosed with Gitelman's syndrome. Each patient was hydrated by 20 mL/kg body weight of oral water within 30 minutes, which was followed by intravenous half saline. When urinary flow reached 10 mL/min, samples of urine and serum were obtained to calculate the osmolar clearance, free water clearance, chloride clearance, and distal fractional chloride reabsorption. Subsequently, furosemide or hydrochlorothiazide was administered. Samples were collected and the same parameters were calculated. In our patients, chloride clearance was increased more than 10 times after furosemide administration(2.1:25.7 and 2.2:27.4 mL/min/100 mL GFR), but not increased after hydrochlorothiazide treatment(2.1:1.6 and 2.2:2.6 mL/min/100 mL GFR). And the distal fractional chloride reabsorption was significantly decreased by furosemide injection (73%:15% and 75%:4.6%), whereas hydrochlorothiazide had no effect on it(73%:63% and 75%:78%). These findings indicate that our patients have a defect in thiazide-sensitive Na-Cl cotransporter in the distal tubule, which is compatible with the pathophysiology of Gitelman's syndrome.