Interferon-gamma suppresses Na+ -H+ exchanger in cultured human endolymphatic sac epithelial cells.
Adequate regulation of endolymphatic pH is essential for maintaining inner ear function. The Na(+)-H(+) exchanger (NHE) is a major determinant of intracellular pH (pH(i)), and facilitates Na(+) and fluid absorption in various epithelia. We determined the functional and molecular expression of NHEs in cultured human endolymphatic sac (ES) epithelial cells and examined the effect of IFN-gamma on NHE function. Serial cultures of human ES epithelial cells were generated from tissue samples. The molecular expression of NHE1, -2, and -3 isoforms was determined by real-time RT-PCR. The functional activity of NHE isoforms was measured microfluorometrically using a pH-sensitive fluorescent dye, 2',7'-bis(carbonylethyl)-5(6)-carboxyfluorescein (BCECF), and a NHE-inhibitor, 3-methylsulfonyl-4-piperidinobenzoyl guanidine methanesulfonate (HOE694). NHE1, -2, and -3 mRNAs were expressed in human ES epithelial cells. Functional activity of NHE1 and -2 was confirmed in the luminal membrane of ES epithelial cells by sequentially suppressing Na(+)-dependent pH(i) recovery from intracellular acidification using different concentrations of HOE694. Treatment with IFN-gamma (50 nM for 24 h) suppressed mRNA expression of NHE1 and -2. IFN-gamma also suppressed functional activity of both NHE1 and -2 in the luminal membrane of ES epithelial cells. This study shows that NHEs are expressed in cultured human ES epithelial cells and that treatment with IFN-gamma suppresses the expression and functional activity of NHE1 and -2.
Son
Eun Jin
EJ
Department of Otorhinolaryngology, Yonsei University College of Medicine, Seoul, South Korea.
Moon
In Seok
IS
Kim
Sung Huhn
SH
Kim
Su Jin
SJ
Choi
Jae Young
JY
D000818:Animals; D002021:Buffers; D027682:Cation Transport Proteins / Q000235:genetics / Q000378:metabolism; D002417:Cattle; D048430:Cell Shape / Q000187:drug effects; D002478:Cells, Cultured; D004712:Endolymphatic Sac / Q000166:cytology*; D004847:Epithelial Cells / Q000166:cytology* / Q000187:drug effects* / Q000378:metabolism / Q000648:ultrastructure; D005786:Gene Expression Regulation / Q000187:drug effects; D006531:HEPES; D006801:Humans; D006863:Hydrogen-Ion Concentration / Q000187:drug effects; D007371:Interferon-gamma / Q000494:pharmacology*; D020033:Protein Isoforms / Q000235:genetics / Q000378:metabolism; D012964:Sodium / Q000494:pharmacology; D000074082:Sodium-Hydrogen Exchanger 1; D000074083:Sodium-Hydrogen Exchanger 3; D017923:Sodium-Hydrogen Exchangers / Q000235:genetics / Q000378:metabolism*; D012996:Solutions
D002021:Buffers; D027682:Cation Transport Proteins; D020033:Protein Isoforms; C490998:SLC9A1 protein, human; C081517:SLC9A2 protein, human; D000074082:Sodium-Hydrogen Exchanger 1; D000074083:Sodium-Hydrogen Exchanger 3; D017923:Sodium-Hydrogen Exchangers; D012996:Solutions; D007371:Interferon-gamma; D012964:Sodium; D006531:HEPES