Damage to the gastric epithelium activates cellular bicarbonate secretion via SLC26A9 Cl⁻/HCO₃⁻exchange

Abstract

Gastric surface pH (pH(o)) transiently increases in response to focal epithelial damage. The sources of that increase, either from paracellular leakage of interstitial fluid or transcellular acid/base fluxes, have not been determined. Using in vivo microscopy approaches we measured pH(o) with Cl-NERF, tissue permeability with intravenous fluorescent-dextrans to label interstitial fluid (paracellular leakage), and gastric epithelial intracellular pH (pH(i)) with SNARF-5F (cellular acid/base fluxes). In response to two-photon photodamage, we found that cell-impermeant dyes entered damaged cells from luminal or tissue compartments, suggesting a possible slow transcellular, but not paracellular, route for increased permeability after damage. Regarding cytosolic acid/base status, we found that damaged cells acidified (6.63 +/- 0.03) after photodamage, compared with healthy surface cells both near (7.12 +/- 0.06) and far (7.07 +/- 0.04) from damage (P < 0.05). This damaged cell acidification was further attenuated with 20 muM intravenous EIPA (6.34 +/- 0.05, P < 0.05) but unchanged by addition of 0.5 mM luminal H(2)DIDS (6.64 +/- 0.08, P > 0.05). Raising luminal pH did not realkalinize damaged cells, suggesting that the mechanism of acidification is not attributable to leakiness to luminal protons. Inhibition of apical HCO(3)(-) secretion with 0.5 mM luminal H(2)DIDS or genetic deletion of the solute-like carrier 26A9 (SLC26A9) Cl(-)/HCO(3)(-) exchanger blocked the pH(o) increase normally observed in control animals but did not compromise repair of damaged tissue. Addition of exogenous PGE(2) significantly increased pH(o) in wild-type, but not SLC26A9 knockout, animals, suggesting that prostaglandin-stimulated HCO(3)(-) secretion is fully mediated by SLC26A9. We conclude that cellular HCO(3)(-) secretion, likely through SLC26A9, is the dominant mechanism whereby surface pH transiently increases in response to photodamage.

Trefoil Factor 2 Requires Na/H Exchanger 2 Activity to Enhance Mouse Gastric Epithelial Repair

“…The activation of NHE2 might logically be expected to be related to the surface pH change that occurs after damage, and so it was exciting to note that (a) TFF2 null mice have a diminished surface pH increase after damage that was restored by exogenous rTFF3 (25) and that (b) increases in extracellular pH will kinetically activate Na/H exchangers including NHE2 (41). However, we have previously shown that damage causes activation of the SLC26A9 Cl/HCO 3 exchanger to partially mediate an increase in surface pH (30), and we now show that an extracellular pH increase of similar magnitude occurs after damage even in the presence of NHE2 inhibition or NHE2 genetic deletion (Fig. 7).…”

“…Alternatively, NHE2 could be activated by the damage-induced cytosolic acidification, but an EIPA-sensitive acidification was only observed in cells that are fated to be exfoliated from the monolayer (25) and so would not be a viable mechanism in the healthy restituting cells that are the presumed target of TFF-mediated repair. It remains possible that NHE2 in restituting cells is activated in a manner that masks cytosolic or surface pH change via other compensatory mechanisms (18,24,30), but at present, we can only conclude that the apical NHE2 does not mediate its novel beneficial effect on gastric repair via changes in surface pH.…”

“…Live Tissue Imaging and Inducing Focal Lesions in Gastric Surface Epithelium-Our methods for confocal/two-photon imaging of the exposed gastric mucosa and creation of microscopic lesions in the surface epithelium by two-photon photodamage have been reported previously (25,30,31). Briefly, the mucosal surface of the gastric corpus was observed using a Zeiss C-Apo ϫ40 objective, and data were collected in a time series.…”

“…The damagerepair cycle was measured independently several times per animal in different locations of the corpus. As described previously (25,30,31), post-acquisition image analysis (Metamorph software, Molecular Devices, Downington, PA) measured the lesioned area as defined by cellular loss of NAD(P)H autofluorescence in direct comparison with simultaneously recorded confocal reflectance images that confirmed location of cellular structures. The changing size of the damaged area over time was used to estimate rates of epithelial restitution, by fitting data to a single exponential decay curve as described and reporting the rate constant of repair as a measure of the restitution rate (25).…”

Antioxidant-mediated preventative effect of Dragon-pearl tea crude polyphenol extract on reserpine-induced gastric ulcers

“…Samples of 4-ml DTCP solution (25,50 or 100 µg/ml) were added to 1.0 ml DPPH methanol solution (1.5x10 -4 M). Following storage at room temperature for 30 min, the absorbance of the solution was determined at 520 nm using a spectrophotometer (UV-5100; Shanghai Metash Instruments Co. Ltd., Shanghi, China), and the remaining DPPH was quantified [(OD DPPH -OD sample )/OD DPPH ]x100%, where OD is optical density.…”