Functional and Molecular Expression of the Proton-Coupled Oligopeptide Transporters in Spleen and Macrophages from Mouse and Human

The aim of this study was to determine the expression and function of proton-coupled oligopeptide transporters (POTs) in spleen and macrophages and their contribution to innate immune response induced by bacterial peptidomimetics gamma-iE-DAP and MDP. Quantitative real-time PCR (qRT-PCR) and Western blot results revealed the mRNA and protein expression of PepT2, PhT1, and PhT2, but not PepT1, in the spleen of mice and humans. In comparison to lymphocytes of the spleen, macrophages had higher transcript levels of PepT2 and PhT2. The cellular uptake of Ala-Lys-AMCA in mouse splenic macrophages was pH-dependent with maximum uptake at pH 6.0, and the kinetic parameters were K-m = 75.5 +/- 14.3 mu M and V-max = 25.4 +/- 2.1 pmol/min per mg protein. The uptake of Ala-Lys-AMCA by mouse splenic macrophages was not inhibited by histidine but was significantly inhibited by glycyl-sarcosine (GlySar) and carnosine (P < 0.01), and by bacterial peptidomimetics gamma-iE-DAP and MDP, ligands of nucleotide-binding oligomerization domain (NOD)-containing proteins. Carnosine and GlySar, but not histidine, attenuated the inflammatory response induced by gamma-iE-DAP and MDP in mouse splenic macrophages. Functional expression of POTs was also demonstrated in THP-1 cells, and dipeptides reduced the immune response induced by gamma-iE-DAP. In conclusion, our findings are novel by providing important information on the molecular and functional expression of POTs in the spleen. Moreover, it appears that the PepT2-mediated uptake of gamma-iE-DAP and MDP in macrophages further contributes to the innate immune response.