Abstract
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 (pHi) 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 pHi 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 pHi 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.
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