Fluorescein uptake by a monocarboxylic acid transporter in human intestinal Caco-2 cells

The fluorescein transport characteristics of the human intestinal epithelial Caco-2 cell line were examined in monolayer cultures. The initial uptake rate was concentration-dependent and saturable; the Michealis constant and the maximum velocity were 0.40 mM and 1.32 nmol/min/mg protein, respectively. A protonophore, carbonyl cyanide m-chlorophenyl-hydrazone, reduced uptake significantly. The replacement of extracellular sodium ions by lithium ions did not alter the initial uptake rate. These facts imply that the transport is driven by a proton gradient. The initial uptake rate was strongly dependent upon extracellular pH, and the uptake was optimal at approximately pH 5.5. Based on the protolytic constants, the main species of fluorescein in the pH range of 5.5 to 6.0 was calculated to be a monoanion, suggesting that fluorescein was taken up by Caco-2 cells as a monocarboxylate. The following findings support this conclusion: the uptake was inhibited significantly by monocarboxylate compounds such as salicylate and pravastatin, but not by di- or tricarboxylic acids or by acidic amino acids. Furthermore, salicylate-preloaded cells showed remarkably enhanced uptake of fluorescein, indicating that monocarboxylates and fluorescein share a common transport carrier. The transporter has a wide spectrum of substrate recognition and seems likely to be different from MCT1.