Received for publication October 24, 2005.
Revised December 16, 2005.
Accepted for publication December 20, 2005.

Inhibition of E-Cadherin-Mediated Homotypic Adhesion of Caco-2 Cells: A Novel Evaluation Assay for Peptide Activities in Modulating Cell-Cell Adhesion

Abstract

Transient modulation of E-cadherin-mediated cell-cell adhesion may improve paracellular drug delivery through biological barriers. Therefore, there is a need to develop an efficient method to evaluate cadherin peptides that can modulate the intercellular junctions. The objective of this study is to establish a novel assay to evaluate peptide activity in modulating E-cadherin-mediated homophilic interactions, based on the homotypic adhesion of Caco-2 cells. Fluorescence-labeled Caco-2 single cells were incubated with Caco-2 monolayers that were treated beforehand with Ca²⁺-free medium. The homotypic adhesion in the presence or absence of peptide and antibody was determined fluorometrically. The Ca²⁺-deficient pretreatment dramatically increased the number of single cells bound to the monolayers. Immunofluorescence staining showed that some of E-cadherins became accessible without surfactant-induced permeabilization of Caco-2 cell monolayers after the Ca²⁺-deficient pretreatment. The homotypic adhesion was largely dependent on extracellular Ca²⁺ concentrations and significantly inhibited by the presence of anti-E-cadherin monoclonal antibody DECMA-1. In contrast, DECMA-1 did not inhibit E-cadherin-independent adhesion, such as the homotypic adhesion of Caco-2 cells in the absence of Ca²⁺ or the heterotypic adhesion of Molt-3 T cells to Caco-2 monolayers. These results indicate the predominant involvement of E-cadherin-mediated cell-cell adhesion in this assay. E-cadherin-derived peptides, which had been shown in our previous studies to inhibit E-cadherin-mediated cell-cell adhesion, significantly inhibited homotypic adhesion in a dose-dependent manner. These results, taken together, suggest that the present assay can be used for evaluation of peptide, protein, or antibody activity in modulating the E-cadherin-mediated homophilic interactions in the context of whole live cells.

Key words: Adhesion, Caco-2 cell, Drug delivery, E-cadherin, Paracellular pathway, Peptide

This article has been cited by other articles:

				D. Thapa, J. S. Lee, S.-Y. Park, Y.-H. Bae, S.-K. Bae, J. B. Kwon, K.-J. Kim, M.-K. Kwak, Y.-J. Park, H. G. Choi, et al. Clotrimazole Ameliorates Intestinal Inflammation and Abnormal Angiogenesis by Inhibiting Interleukin-8 Expression through a Nuclear Factor-{kappa}B-Dependent Manner J. Pharmacol. Exp. Ther., November 1, 2008; 327(2): 353 - 364. [Abstract] [Full Text] [PDF]