Renal assimilation of short chain peptides: Visualization of tubular peptide uptake

Purpose. Renal assimilation of short chain peptides plays an important role in systemic protein metabolism and amino acid homeostasis. The transepithelial peptide transport across the apical membrane of tubular cells is mediated almost exclusively by pH-dependent H⁺-peptide symport pathways. The current study was designed to identify by visualization functional peptide transport activity along the nephron structures. Methods. Visualization of peptide uptake was achieved by using the fluorescent dipeptide derivative D-Ala-Lys-AMCA and unlabelled cefadroxil and glycylglutamine as transport competitors to demonstrate specificity. To confirm these assays, rat specific cRNA probes were synthesized and non-isotopic high-resolution in-situ-hybridization and northern blot analysis were carried out to demonstrate the expression of the high-affinity peptide transporter PEPT2. Results. The reporter molecule was accumulated by cells of the proximal tubulus but not in glomerular or endothelial cells. Inhibition studies revealed competitive inhibition of D-Ala-Lys-AMCA uptake by the betalactam cefadroxil and the dipeptide glycylglutamine. The control organs intestine and spleen did not show uptake of the systemically administered molecule. Non-isotopic mRNA in-situ-hybridization, using an antisense probe for rat PEPT2 confirmed uptake assays by identifying PEPT2 expression throughout segments of the straight proximal tubule at the inner cortex and outer stripe. Conclusions. We demonstrate for the first time renal in vivo transport activity of a dipeptide that allows cells that participate in peptide reabsorption to be visualized. This functional assay may be used to investigate renal peptide transport mechanisms and test new compounds that are transported via proton-driven peptide transporters.