STRUCTURAL AND FUNCTIONAL ROLES OF CYTOSKELETAL PROTEINS DURING REPAIR OF NATIVE GUINEA-PIG INTESTINAL EPITHELIUM

The cytoskeletal events that assist restitution of the native intestin al epithelium are poorly understood. To enhance our understanding of r epair mechanisms in the native intestinal epithelium we assessed the f unctional role of actin and the temporal and spatial alterations in ac tin and villin that occur in native enterocytes migrating in response to injury. Using a well-characterized in vitro Ussing chamber model of native intestine epithelial restitution, the actin inhibitor cytochal asin D (CD) was applied to determine the functional importance of acti n to restitution as assessed by sensitive electrophysiological means a nd structural techniques. Additionally we used phalloidin and indirect immunohistochemistry to localize and semi-quantitate F-actin and vill in in migrating cells during restitution. We report new data that show s that when cytoskeletal changes were impaired with CD, the epithelial monolayer was re-established in fewer than 20% of CD-treated villi, c ells bordering the epithelial defect did not assume the characteristic phenotype associated with migrating cells, and transepithelial resist ance did not return to pre-injury levels. F-actin and villin were pres ent at the leading edge of the migrating cells, basolateral F-actin wa s decreased, and cytoplasmic villin was increased as determined by pha lloidin and immunohistochemical methods. We conclude that in vitro rep air of the native intestinal epithelium is functionally and structural ly dependent on major changes in the cytoskeleton of cells involved in re-establishing the epithelial monolayer over a complex extracellular matrix. (C) 1996 Academic Press Limited.