Acidosis induces hyperpermeability in Caco-2BBe cultured intestinal epithelial monolayers

We previously demonstrated that ileal mucosal acidosis in pigs reversibly increases intestinal permeability to hydrophilic macromolecules, even in the absence of tissue hypoxia [A. L. Salzman, H. Wang, P. S. Wollert, T. J. Vandermeer, C. C. Compton, A. G. Denenberg, and M. P. Fink. Am. J. Physiol. 266 (Gastrointest. Liver Physiol. 29): G633-G646, 1994]. In an effort to further explore the mechanism(s) underlying this phenomenon, we examined the effect of acidic pH on the permeability characteristics of cultured Caco-2BBe (human intestinal epithelial) cells grown as monolayers on permeable supports. Permeability was determined by measuring the mucosal-to-basolateral flux of fluorescein disulfonic acid (FS; mol wt 478 Da), fluorescein isothiocyanate-labeled dextran (FD4; average mol wt 4 kDa), or [H-3]mannitol. Incubation of monolayers under hypercarbic conditions or with acidified bicarbonate-free medium significantly increased permeability to FS, FD4, and mannitol in a manner dependent on both time and pH. Incubation in medium at pH 5.43 for 24 h increased the release of lactate dehydrogenase and decreased the intensity of staining with calcein-acetoxymethyl ester, findings that are indicative of plasma membrane injury; nevertheless, the percentage of nonviable cells did not increase. Ultrastructural analyses revealed evidence of increased paracellular trafficking of horseradish peroxidase after incubation of monolayers under acidic conditions. Fluorescence confocal microscopy and temperature studies demonstrated that incubation at pH 5.43 induced an increase in both the intracellular uptake of FD4 and the activation energy for FS permeation across Caco-2BBe monolayers, respectively, suggesting increased transcellular permeation. Exposure to acidic conditions also decreased cellular levels of ATP. We conclude that acidosis increases both paracellular and transcellular permeability to hydrophilic macromolecules and leads to depletion of ATP.