Distal renal tubular acidosis (RTA) with sensorineural deafness is a rare entity, inherited in an autosomal recessive manner. It is caused by mutations in the ATP6V1B1 gene, leading to defective function of H⁺-ATPase pump in the distal nephron, cochlea and endolymphatic sac. We report two siblings with distal RTA and sensorineural deafness having mutation C>T in the first coding exon of the gene, resulting in a non functional protein. The parents were found to be carriers for the mutation.

Key words: ATP6V1B1 gene, Renal tubular acidosis, Sensorineural deafness.

Distal renal tubular acidosis (RTA) is characterized by impaired urine acidification leading to severe hyperchloremic metabolic acidosis, hypokalemia, hypercalciuria, hypocitraturia, nephrocalcinosis and nephrolithiasis. The disease is characterized by failure to thrive, nephrocalcinosis, polyuria and urolithiasis. In untreated cases, the progression of nephrocalcinosis may lead to chronic renal failure(1-3). Mutations in the ATP6V1B1 gene, encoding the B1 subunit of vacuolar H⁺-ATPase, result in autosomal recessive distal RTA associated with nerve deafness (OMIM #267300). Genetic screening in multiple kindreds show different mutations in the gene and almost all affected individuals have sensorineural hearing loss. The majority of these mutations are likely to disrupt the structure or abrogate the production, of the normal B1 subunit protein. This leads to loss of expression of the gene in the human cochlea and in endolymphatic sac epithelium, in addition to the renal tubular defect(1,4).

Patient 1. This 3-yr-old girl, born of non-consanguineous marriage to a north-Indian Hindu family, presented with complaints of polyuria, polydipsia, failure to thrive and bony deformities. Similar history was present in an elder sibling who passed away at 1-yr of age, but was not investigated. The parents were apparently normal. Investigations showed metabolic acidosis (pH 7.08, bicarbonate 7.8 mEq/L) with plasma anion gap of 10 mEq/L; serum potassium levels ranged between 2.7-3.2 mEq/L (Table I). Urine pH was 6.5, urine anion gap was positive and there was evidence of hypercalciuria (urine calcium to creatinine ratio between 0.7-0.9 on multiple occasions). Following bicarbonate loading, the fractional excretion of bicarbonate was 7.2% and difference between urinary to plasma CO₂ (U-P CO₂) was 1.7 mm Hg. Ultrasonography of abdomen showed bilateral medullary nephrocalcinosis. Pure tone audiometry showed severe sensorineural nerve deafness.

TABLE I



Biochemical and Genetic features*

Patient 2. This 1-year old younger sibling of the first patient also presented with similar history of polyuria, polydipsia, failure to thrive and bony deformities. Investigations showed metabolic acidosis with plasma anion gap of 11.4 mEq/L; serum potassium levels ranged between 3.0-3.2 mEq/L (Table I). Urine pH was 6.12, urine anion gap was positive and there was evidence of hypercalciuria (calcium to creatinine ratio ranging between 1.8-2.2 on multiple occasions). Following bicarbonate loading, the fractional excretion of bicarbonate was 6.8% and U-P CO₂ was -4.6 mm Hg. Ultrasonography of abdomen also showed bilateral medullary nephrocalcinosis and audiometry showed severe sensorineural deafness.

A diagnosis of familial distal RTA with sensorineural deafness was made. Both patients were treated with Polycitra-K to provide 8 mEq/kg/day bicarbonate, and were provided aural rehabilitation with hearing aids and speech therapy.

ATP6VIBI gene sequencing: After informed consent, DNA was extracted from the peripheral blood from the patients and their parents. The 14 coding exons of ATP6V1B1 gene were amplified by polymerase chain reaction using the primers described previously(1). The amplified fragments were purified and sequenced on an automated ABI310 system, using BigDye chemistry (Applied Biosystems-Applera Corporaton, Drive Foster City, CA). Sequencing of the coding exons and the intronic flanking region showed DNA mutation at exon 1: C>T (R31X). Both siblings were homozygous, while parents were heterozygous for the mutation (Fig.1).

Fig.1 The electrophoregrams show the exon 1 sequences of the ATP6V1B1 gene of a normal control and the family. Both siblings were homozygous for the mutation exon 1: 91 C>T, and the both parents were carriers for the same mutation.

Discussion

Both patients in the present report had features of distal RTA with failure to thrive, polyuria, refractory rickets, hypokalemia and nephrocalcinosis. They also showed severe sensorineural deafness requiring rehabilitation. Investigations confirmed the diag-nosis of secretory distal RTA and homozygous mutations were found in the ATP6V1B1 gene in both cases.

Most cases of primary distal RTA in children result from defective function of the proton pump vacuolar H⁺-ATPase, located at the apical surface of the a-intercalated cells. The vacuolar H⁺-ATPase is formed by several subunits. Mutations in the ATP6V0A4 gene, which encodes for the a4 subunit, cause autosomal recessive dRTA (OMIM # 602722)(3-5). ATP6V1B1, a gene on chromosome 2, encodes for the B1-subunit of the vacuolar H⁺-ATPase. ATP6V1B1 is also expressed in the human cochlea and in endolymphatic sac epithelium(4). Endolymph is a unique extracellular fluid having low sodium and high potassium concentrations, which maximizes the sensitivity of hair cells. To preserve its pH at 7.4, there is presumably a requirement for proton pumping into endolymph. It is assumed that H⁺-ATPase contributes to maintenance of endolymph pH and defects in its B1 subunit cause irreversible hair cell damage because of abnormalities in electrolyte and pH(6).

Apart from the presence or absence of hearing loss, there do not appear to be major phenotypic differences at diagnosis between patients with homozygous ATP6V1B1 and ATP6V0A4 mutations. Long term follow-up of a cohort of affected individuals with ATP6V0A4 mutations shows mild and/or older-onset hearing impairment in some patients who were initially considered to have normal hearing by audiometry(3).

4. Karet FE, Finberg KE, Nelson RD, Nayir A, Mocan H, Sanjad SA, et al. Mutations in the gene encoding B1 subunit of H⁺-ATPase cause renal tubular acidosis with sensorineural deafness. Nat Genet 1999; 21: 84-90.