ABSTRACT
Tight junctions (TJs) are intercellular structures that control paracellular permeability and epithelial polarity. It is now accepted that TJs are highly dynamic structures that are regulated in response to exogenous and endogenous stimuli. Here, we provide details on the mechanism of action of AT-1002, the active domain of Vibrio cholerae's second toxin, zonula occludens toxin (ZOT). AT-1002, a hexamer peptide, caused the redistribution of ZO-1 away from cell junctions as seen by fluorescence microscopy. AT-1002 also activated src and mitogen activated protein (MAP) kinase pathways, increased ZO-1 tyrosine phosphorylation, and rearrangement of actin filaments. Functionally, AT-1002 caused a reversible reduction in transepithelial electrical resistance (TEER) and an increase in lucifer yellow permeability in Caco-2 cell monolayers. In vivo, co-administration of salmon calcitonin with 1 mg of AT-1002 resulted in a 5.2-fold increase in AUC over the control group. Our findings provide a mechanistic explanation for AT-1002-induced tight junction disassembly, and demonstrate that AT-1002 can be used for delivery of other agents in vivo.
Subject(s)
Cholera Toxin/chemistry , Oligopeptides/pharmacology , Tight Junctions/drug effects , Actin Cytoskeleton/metabolism , Actins/drug effects , Actins/metabolism , Animals , Area Under Curve , Caco-2 Cells , Calcitonin/pharmacokinetics , Drug Interactions , Electric Impedance , Endotoxins , Humans , Isoquinolines/metabolism , Male , Microscopy, Fluorescence , Mitogen-Activated Protein Kinases/drug effects , Mitogen-Activated Protein Kinases/metabolism , Phosphorylation/drug effects , Rats , Rats, Sprague-Dawley , Tight Junctions/metabolism , Tyrosine/metabolism , src-Family Kinases/drug effects , src-Family Kinases/metabolismABSTRACT
AT-1002 a 6-mer synthetic peptide belongs to an emerging novel class of compounds that reversibly increase paracellular transport of molecules across the epithelial barrier. The aim of this project was to elaborate on the structure-activity relationship of this peptide with the specific goal to replace the P2 cysteine amino acid. Herein, we report the discovery of peptides that exhibit reversible permeability enhancement properties with an increased stability profile.
