Intracellular pH regulatory mechanism in a human renal proximal cell line (HKC-8): evidence for Na+/H+ exchanger, CI-/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 (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.
Hara
C
C
Department of Internal Medicine, Faculty of Medicine, University of Tokyo, Japan.
Satoh
H
H
Usui
T
T
Kunimi
M
M
Noiri
E
E
Tsukamoto
K
K
Taniguchi
S
S
Uwatoko
S
S
Goto
A
A
Racusen
L C
LC
Inatomi
J
J
Endou
H
H
Fujita
T
T
Seki
G
G
D000143:Acids / Q000494:pharmacology; D017920:Antiporters / Q000378:metabolism*; D001639:Bicarbonates / Q000494:pharmacology; D002245:Carbon Dioxide / Q000494:pharmacology; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism*; D002461:Cell Line, Transformed; D027963:Chloride-Bicarbonate Antiporters; D002712:Chlorides / Q000494:pharmacology; D006801:Humans; D006859:Hydrogen / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D007425:Intracellular Membranes / Q000378:metabolism*; D007687:Kidney Tubules, Proximal / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D020033:Protein Isoforms / Q000235:genetics; D012333:RNA, Messenger / Q000378:metabolism; D027982:Sodium-Bicarbonate Symporters; D017923:Sodium-Hydrogen Exchangers / Q000378:metabolism*
D000143:Acids; D017920:Antiporters; D001639:Bicarbonates; D002352:Carrier Proteins; D027963:Chloride-Bicarbonate Antiporters; D002712:Chlorides; D020033:Protein Isoforms; D012333:RNA, Messenger; D027982:Sodium-Bicarbonate Symporters; D017923:Sodium-Hydrogen Exchangers; D002245:Carbon Dioxide; D006859:Hydrogen