Cytochalasin-Induced Actin Disruption of Polarized Enterocytes Can Augment Internalization of Bacteria

Cytochalasin-induced actin disruption has often been associated with decreased bacterial internalization by cultured epithelial cells, although polarized enterocytes have not been systematically studied. In assays using confluent polarized HT-29 enterocytes, cytochalasin D appeared to increase internalization of wild-type Salmonella typhimurium, Proteus mirabilis, and Escherichia coli. HeLa and HEp-2 epithelial cells, as well as HT-29 and Caco-2 enterocytes, were used to clarify this unexpected observation. Resulting data showed that cytochalasin D was associated with increased internalization of S. typhimurium and P. mirabilis by both HT-29 and Caco-2 enterocytes and with increased internalization of E. coli by HT-29 enterocytes; with either HeLa or HEp-2 cells, cytochalasin was associated with no change or a decrease in internalization of these same bacterial strains. Cytochalasin caused decreased internalization of Listeria monocytogenes by HT-29, Caco-2, HeLa, and HEp-2 cells, indicating that cytochalasin did not consistently augment bacterial internalization by polarized enterocytes. Fluorescein-labeled phalloidin confirmed marked disruption of filamentous actin in cytochalasin-treated HT-29, Caco-2, HeLa, and HEp-2 cells. Cytochalasin had no noticeable effect on epithelial viability but caused distorted apical microvilli, cell rounding, and separation of adjacent enterocytes in confluent cultures (with a corresponding decrease in transepithelial electrical resistance). Scanning electron microscopy showed that cytochalasin-induced enhanced bacterial internalization was associated with preferential bacterial adherence on the exposed enterocyte lateral surface. Colchicine, used to disrupt microtubules, had no noticeable effect on bacterial internalization by HT-29 or Caco-2 enterocytes. These data indicated that for HT-29 and Caco-2 enterocytes, cytochalasin-induced disruption of filamentous actin might augment internalization of some bacterial species by a mechanism that appeared to involve exposure of the enterocyte lateral surface.