Adequate regulation of endolymphatic pH is essential for maintaining inner ear function. The Na+-H+ exchanger (NHE) is a major determinant of intracellular pH (pHi), 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-γ 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 pHi recovery from intracellular acidification using different concentrations of HOE694. Treatment with IFN-γ (50 nM for 24 h) suppressed mRNA expression of NHE1 and -2. IFN-γ 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-γ suppresses the expression and functional activity of NHE1 and -2.
All Science Journal Classification (ASJC) codes
- Molecular Biology
- Cell Biology