Abstracts
1998 Digestive Disease Week

#1066

LOSS OF MATRIX-DEPENDENT CYTOSKELETAL TYROSINE KINASE SIGNALS MAY REGULATE INTESTINAL EPITHELIAL DIFFERENTIA-TION DURING MUCOSAL HEALING. Y.-W. Liu, M.A. Sanders, M.D. Basson. Depts. of Surgery, Yale University, New Haven, CT and CT VA Health Care System, West Haven, CT.

Intestinal epithelial restitution and the migratory phenotype appear regulated by the extracellular matrix. Since integrin-associated adhesion to matrix triggers tyrosine kinase (TK) activity, we hypothesized that the intestinal epithelial migratory phenotype might be modulated by matrix-specific TK signals, particularly via Focal Adhesion Kinase (FAK). Caco-2 cells were seeded at two densities on collagen I, laminin, fibronectin, and tissue culture plastic so that at four days the first cells were confluent while the second were not contact-inhibited and expressed migratory lamellipodia. After fractionation into Triton X-100 soluble (membrane/cytoskeletal) and insoluble (cytosolic) fractions, TK activity (n_6) was assayed by ELISA and FAK activity (tyrosine autophosphorylation) by immunoprecipitating with anti-FAK and Western blotting for phosphotyrosine (n_4). Basal cytoskeletal TK activity in static cells was itself matrix-dependent and, unlike cytosolic TK, correlated with adhesion, highest on collagen (0.74 ± 0.06 pmol/mg/min) and lowest on plastic (0.49 ± 0.07 pmol/mg/min). Migrating cells generally exhibited an increase in cytosolic TK of approximately 0.28 pmol/mg/min independently of matrix. However, an additional increment in cytosolic TK correlated inversely with changes in cytoskeletal TK, suggesting TK translocation during cell motility from the cytoskeletal to the cytosolic compartment. Indeed, cytoskeletal FAK activity was decreased by 53.1 ± 11.1% during migration on collagen. (n=4, p<0.02) Furthermore, TK inhibition by genistein both substantially inhibited migration and stimulated expression of brush border enzymes downregulated during motility. Although enterocyte-matrix interactions alter both cytosolic and cytoskeletal TK activity, matrix-dependent cytoskeletal events are likely to regulate adhesion and differentiation in static cells. Loss of matrix-dependent cytoskeletal TK signals such as FAK during cell motility may trigger a phenotypic switch to the "dedifferentiated" migrating intestinal epithelial phenotype.

Copyright 1996 - 1998, SSAT, Inc. Revised 29 June 1998.