Binding of the VCHSKT peptide to ceramide‐rich and cholesterol‐rich domains of cell membranes in rat pancreatic glands

Abstract

A peptide with antisecretory effect has been defined and was denoted the antisecretory factor (AF‐16) with a sequence of 16 amino acids. A shorter sequence, the AF‐6 peptide (VCHSKT, Pepceuticals Ltd, Leicester, England), is readily detected by Time of Flight Secondary Ion Mass Spectrometry (TOF‐SIMS), using trifluoroacetic acid (TFA) as ionization catalyzer. Analysis of a reference sample revealed a pseudo‐molecular ion at m/z 657 and M + Na+ at m/z 679. These peaks were also detected in tissue samples of animals exposed to the peptide. The AF‐6 peptide was injected intravenously or sprayed into the nostrils of Sprague–Dawley rats, 250–300 g (n = 5). The animals were anesthetized with isoflurane. Pancreas tissue samples were taken after 2 h and were high‐pressure frozen in a Lecia EMPACT, sprayed with 0.1% TFA, and analyzed using a TOF‐SIMS V instrument (ION‐TOF, GmbH, Münster, Germany) equipped with a Bi3+‐liquid metal ion gun at the University of Gothenburg. The distribution of the AF‐6 peptide could be seen in the pancreas of exposed animals and control animals as high‐intensity dots that were heterogeneously distributed in regions with a high intensity of phosphatidylcholine head group m/z 184, indicating a localization to the membrane portion of the pancreatic cells. The peptide was colocalized with a dot‐like distribution of galactocylceramide c18:0 and cholesterol. Cholesterol and gal‐Cer are known lipid components of specialized membrane areas called lipid rafts. The results indicate that the AF‐6 peptide binds to cellular receptors present in such regions of rat pancreatic cells. Copyright © 2012 John Wiley & Sons, Ltd.