Intracellular pH regulatory mechanism in a human renal proximal cell line (HKC-8): Evidence for Na⁺/H⁺ exchanger, Cl-/HCO₃^- exchanger and Na⁺-HCO₃^- cotransporter

In the present study we investigated whether an immortalized human renal proximal cell line, HKC-8, expresses a recently cloned Na⁺-HCO₃^- cotransporter (NBC-1) and, if so, which isoform (kNBC-1 from kidney or pNBC-1 from pancreas) is expressed in this cell line. Cell pH (pH(i)) measurements using a pH-sensitive fluorescence probe in the absence of HCO₃^-/CO₂ revealed the presence of a Na⁺/H⁺ exchanger that required high concentrations of amiloride for full inhibition. In the presence of HCO₃^-/CO₂ another pH(i) recovery process, dependent on Na⁺ but independent of Cl^-, was identified. This process was electrogenic and was inhibited by 4,4'-diisothiocyanatodihydrostilbene-2,2'-disulphonic acid (DIDS), being consistent with the Na⁺-HCO₃^- cotransporter. In addition, the pH(i) responses to Cl^- removal were compatible with the presence of a Na⁺-independent Cl^-/HCO₃^- exchanger that was also inhibited by DIDS. Reverse transcriptase polymerase chain reaction (RT-PCR) using primers designed for specific and common regions detected mRNAs of both kNBC-1 and pNBC-1 and Western blot analysis confirmed the expression of NBC-1 protein. These results indicate that HKC-8 has transport activities similar to intact proximal tubules and also suggest that both kNBC-1 and pNBC-1 may contribute to the Na⁺-HCO₃^- cotransport activity in this cell line.