H+/solute-induced intracellular acidification leads to selective activation of apical Na+/H+ exchange in human intestinal epithelial cells
David T. Thwaites, … , Michael Glanville, Nicholas L. Simmons
David T. Thwaites, … , Michael Glanville, Nicholas L. Simmons
Published September 1, 1999
Citation Information: J Clin Invest. 1999;104(5):629-635. https://doi.org/10.1172/JCI7192.
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Article

H+/solute-induced intracellular acidification leads to selective activation of apical Na+/H+ exchange in human intestinal epithelial cells

Abstract

The intestinal absorption of many nutrients and drug molecules is mediated by ion-driven transport mechanisms in the intestinal enterocyte plasma membrane. Clearly, the establishment and maintenance of the driving forces — transepithelial ion gradients — are vital for maximum nutrient absorption. The purpose of this study was to determine the nature of intracellular pH (pHi) regulation in response to H+-coupled transport at the apical membrane of human intestinal epithelial Caco-2 cells. Using isoform-specific primers, mRNA transcripts of the Na+/H+ exchangers NHE1, NHE2, and NHE3 were detected by RT-PCR, and identities were confirmed by sequencing. The functional profile of Na+/H+ exchange was determined by a combination of pHi, 22Na+ influx, and EIPA inhibition experiments. Functional NHE1 and NHE3 activities were identified at the basolateral and apical membranes, respectively. H+/solute-induced acidification (using glycylsarcosine or β-alanine) led to Na+-dependent, EIPA-inhibitable pHi recovery or EIPA-inhibitable 22Na+ influx at the apical membrane only. Selective activation of apical (but not basolateral) Na+/H+ exchange by H+/solute cotransport demonstrates that coordinated activity of H+/solute symport with apical Na+/H+ exchange optimizes the efficient absorption of nutrients and Na+, while maintaining pHi and the ion gradients involved in driving transport.

Authors

David T. Thwaites, Dianne Ford, Michael Glanville, Nicholas L. Simmons

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Identification of Na+/H+ exchange activity in Caco-2 cell monolayers. NH...
Identification of Na+/H+ exchange activity in Caco-2 cell monolayers. NH4Cl-induced intracellular acidification in monolayers of Caco-2 cells. pHi was measured in monolayers of Caco-2 cells loaded with the pH-sensitive fluorochrome BCECF. The solutions (all pH 7.4) bathing both the apical (a) and basolateral (b) membranes were standard Na+-containing Krebs-Ringer (hatched bars), Na+-free Krebs-Ringer (open bars), and Na+-free Krebs-Ringer containing NH4Cl (cross-hatched bars). Cells were exposed to 30 mM NH4Cl for 15 minutes; NH4Cl was removed in Na+-free conditions followed by sequential addition of Na+ to the basolateral and then apical solutions.