A Chinese Girl with Bartter Syndrome Type III due to a Novel Mutation and/or Single Nucleotide Polymorphisms (SNPs) in CLCNKB Gene
An 811/12-year-old Chinese girl was brought to the pediatric endocrinology ward because of hypokalemia found occasionally. The girl showed no progressive muscle weakness of the extremities. Her serum potassium was 2.8 mmol/l estimated two weeks ago in a local hospital. Her birth history revealed full-term by normal spontaneous delivery and normal birth weight of 3,400 g without antenatal polyhydramnios.\nOn admission, her height (129 cm) and body weight (25 kg) were all more than the 40th percentile. Blood pressure was 106/66 mmHg, heart rate 96 beats/min and respiratory rate 22/min. internal jugular vein was flat. There was no diminished muscle strength and no reflex over the lower extremities. Fasciculation, myoclonus and muscular atrophy were not present. Her pure tone audiometry was normal. The remainder of the physical examination was unremarkable.\nComplete blood cells counts showed that hemoglobin was 116g/l, white blood cells 5,600/µl and platelets 314,000/µl. Urinalysis revealed pH 7.5, negative protein, glucose, leukocytes, and red blood cells. The data of biochemical studies are shown in and . The most striking findings were hypokalemia (2.5-3.0 mmol/l), mild metabolic alkalosis (HCO3 －32.6 mmol/l), normal renal function (BUN 36.4 mg/dl, Cr 3.52 mg/dl), and normal aldosterone concentration (120 pg/ml).\nThe trans-tubular potassium gradient (TTKG) is an index reflecting the conservation of potassium in the cortical collecting ducts of the kidneys. The following is the formula for calculating the TTKG: TTKG =[(urine potassium) × (plasma osmolality)] divided by [(plasma potassium) × (urine osmolality)]. Her TTKG (9.6) was higher, indicative of renal K+ wasting. Serum magnesium (Mg2+) was tested twice and the value was 1.82, 2.0 mg/dl respectively. Serum calcium (Ca2+) levels were normal, but urinary Ca2+ excretion was very low (urine Ca2+/Cr 0.01 mg/mg) ( and ). EKG disclosed a sinus rhythm. Abdominal sonography revealed bilateral normal size kidneys and adrenal glands without nephrocalcinosis or renal stones. With the above-mentioned findings, she was clinically diagnosed as BS type III with the presentation of inactivity, hypokalemia and metabolic alkalosis along with normal renal function.\nAdministration of intravenous KCl 60 mmol/ day followed by oral KCl 1.5 mEq/kg/day restored and maintained her serum K+ concentration at about 3.0 mmol/l. Fluids were also supplemented. Indomethacin (2 mg/kg/day) and spironolactone (2 mg/kg/day) were also regularly given to control the hypokalemia until now. She did not experience any episodes of Indomethacin overdose or received other potential nephrotoxic medications during therapy periods. The most prominent laboratory data was persistent hypokalemia during one year of follow-up till now, especially after the episodes of acute upper respiratory infection and acute gastroenteritis. Her renal function still persisted in good condition.\nMolecular analysis: The human-investigation committee of the Zhejiang University School of Medicine approved the study. The parents were informed about the purpose of this study and gave consent. The polymerase chain reaction (PCR) for amplifying DNA sequences and direct sequencing of all the exons of CLCKB gene was performed using peripheral blood genomic DNA of the patient and her non-consanguineous parent. All the primers were designed according to the sequence of NG_013079.1 (). Several reported SNPs and some new nucleotide variations were also found in the CLCKB gene of the patient, her mother and father (). Three of the nucleotide variations are synonymous mutations or silent SNPs, the other nucleotide variations can make the amino acids change, however, these new nucleotide variations were found in the patient and her healthy parents, we concluded that these new nucleotide variations may be SNPs.\nFamily tree of new mutation or SNPs in exon 20 of CLCNKB gene was also investigated in this family (). A novel heterozygous C to G transition at nucleotide 2471 in exon 20 were found in the patient, which has not been detected in her mother or her father (, ). Sequencing for reverse DNA strands confirmed the above result.