Abstract
Background/Aims: To functionally characterize intracellular pH (pH<sub>i</sub>) regulating mechanisms, such as Na<sup>+</sup>-H<sup>+</sup> exchanger (NHE) and Na<sup>+</sup>-HCO<sub>3</sub><sup>-</sup> co-transporter (NBC), and further examine effects of ethanol on the pH<sub>i</sub> regulating mechanism in human oral epidermoid carcinoma (OEC-M1) cells. Methods: OEC-M1 cells were a gift from Tri-Service General Hospital. Changes of pH<sub>i</sub> were detected by microspectrofluroimetry with a pH-sensitive fluorescent dye, BCECF. Isoforms of transporters were examined by Western blot technique. Results: i) the steady-state pH<sub>i</sub> value shifted from alkaline (7.35∼7.49) to acidic (7.0∼7.03) following acid/base impacts; ii) in HEPES-buffer system, pH<sub>i</sub> recovery following induced-acidification was totally blocked by either removing [Na]<sub>o</sub><sup>+</sup> or adding HOE 694 (a NHE1 specific inhibitor), which demonstrates existence of NHE1; iii) in HCO<sub>3</sub><sup>-</sup>/CO<sub>2</sub>-buffer system, the pH<sub>i</sub> recovery following induced-acidification was entirely blocked by either removing [Na]<sub>o</sub><sup>+</sup> or adding HOE 694 plus DIDS (a NBC specific inhibitor), which suggests existence of Na<sup>+</sup>- and HCO<sub>3</sub><sup>–</sup>dependent acid-extruder, i.e. NBC; iv) the isoforms of the two acid extruders were NHE1, NBCn1, NBCe1 and NDCBE; v) ethanol (10-1000 mM) showed a biphasic and concentration-dependent effect on resting pH<sub>i</sub> (i.e. increase then decrease) by changing the activity of NHE1 and NBC accordingly; vi) treatment with ethanol for 24 hr (<unterline>></unterline> 300 mM) significantly inhibited the expression of NHE1, NBCn1 and NDCBE, while up-regulated NBCe1. Conclusions: Ethanol affects pH<sub>i</sub> in a concentration-dependent manner by changing function and expression of NHE1 and NBC isoforms in OEC-M1 cells.
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