Basolateral glycylsarcosine (Gly-Sar) transport in Caco-2 cell monolayers is pH dependent

Objectives Transepithelial di/tripeptide transport in enterocytes occurs via the apical proton-coupled peptide transporter, hPEPT1 (SLC15A1) and a basolateral peptide transporter, which has only been characterized functionally. In this study we examined the pH dependency, substrate uptake kinetics and substrate specificity of the transporter. Methods We studied the uptake of [14C]Gly-Sar from basolateral solution into Caco-2 cell monolayers grown for 17-22 days on permeable supports, at a range of basolateral pH values. Key findings Basolateral Gly-Sar uptake was pH dependent, with a maximal uptake rate at a basolateral pH of 5.5. Uptake of Gly-Sar decreased in the presence of the protonophore nigericin, indicating that the uptake was proton-coupled. The uptake was saturable, with a maximal flux (Vmax) of 408 +/- 71, 307 +/- 25 and 188 +/- 19pmol/cm2/min (mean +/- S.E., n=3) at basolateral pH5.0, 6.0 and 7.4, respectively. The compounds Gly-Asp, Glu-Phe-Tyr, Gly-Glu-Gly, Gly-Phe-Gly, lidocaine and, to a smaller degree, para-aminohippuric acid were all shown to inhibit the basolateral uptake of Gly-Sar. Conclusions The study showed that basolateral Gly-Sar transport in the intestinal cell line Caco-2 is proton-coupled. The inhibitor profile indicated that the transporter has broad substrate specificity.