Zonula occludens toxin modulates tight junctions through protein kinase C-dependent actin reorganization, in vitro.

Abstract

The intracellular signaling involved in the mechanism of action of zonula occludens toxin (ZOT) was studied using several in vitro and ex vivo models. ZOT showed a selective effect among various cell lines tested, suggesting that it may interact with a specific receptor, whose surface expression on various cells differs. When tested in IEC6 cell monolayers, ZOT-containing supernatants induced a redistribution of the F-actin cytoskeleton. Similar results were obtained with rabbit ileal mucosa, where the reorganization of F-actin paralleled the increase in tissue permeability. In endothelial cells, the cytoskeletal rearrangement involved a decrease of the soluble G-actin pool (-27%) and a reciprocal increase in the filamentous F-actin pool (+22%). This actin polymerization was time-and dose-dependent, and was reversible. Pretreatment with a specific protein kinase C inhibitor, CGP41251, completely abolished the ZOT effects on both tissue permeability and actin polymerization.In IEC6 cells ZOT induced a peak increment of the PKCct isoform after 3 min incubation. Taken together, these results suggest that ZOT activates a complex intracellular cascade of events that regulate tight junction permeability, probably mimicking the effect of physiologic modulator(s) of epithelial barrier function. (J. Clin. Invest. 1995. 96:710-720.)

ICAM-1-Coupled Cytoskeletal Rearrangements and Transendothelial Lymphocyte Migration Involve Intracellular Calcium Signaling in Brain Endothelial Cell Lines

“…We observed in this paper that endothelial PKC activation is clearly involved, although to a lesser extent than calcium, in lymphocyte transmigration. PKC activation is correlated with tight junction regulation, via a mechanism involving actin reorganization (39). The role of PKCs during transendothelial migration may be due to their ability to mediate ICAM-1-coupled cytoskeletal rearrangements.…”

Modulation of intestinal tight junctions by Zonula occludens toxin permits enteral administration of insulin and other macromolecules in an animal model.

“…We have recently demonstrated that Zot activates a complex intracellular cascade of events that regulate intestinal permeability (9). Zot induces a dose-and time-dependent PKC␣-related polymerization of actin filaments strategically localized to regulate the paracellular pathway (9).…”

“…Zot (a) is not cytotoxic and does not affect the viability of the intestinal epithelium ex vivo (7,9); (b) fails to completely abolish the intestinal transepithelial resistance (7,9, and present paper), (c) interacts with a specific intestinal receptor whose regional distribution within the intestine varies (10), (d) is not effective in the large intestine where the presence of the colonic microflora could be potentially harmful if the mucosal barrier was compromised (10 and present paper), (e) does not induce acute systemic side-effects (for at least 80-90 h) when orally administered (present paper), and (f) induces a reversible increase of tissue permeability (7,9, and present paper). Our results demonstrate that coadministration of Zot with biologically active ingredients enhances intestinal absorption of the active molecule, and that this enhancement is effective for both relatively small (insulin, 5733 D) and large molecules (IgG, 140-160 kD).…”

“…The utility of the paracellular route for oral drug delivery has remained unexplored due to a limited understanding of tight junction (tj) 1 physiology and the lack of substances capable of increasing the tj permeability without irreversibly compromising intestinal integrity and function (3)(4)(5)(6). We have recently demonstrated that Zonula occludens toxin (Zot), a protein elaborated by Vibrio cholerae (7,8), induced modifications of cytoskeletal organization that lead to the opening of tj related to the PKC ␣ -dependent polymerization of actin monomers into actin filaments strategically localized to regulate the paracellular pathway (9). We have also shown that this modulation is reversible, time-and dose-dependent, and is confined to the small intestine, since Zot does not affect the colon permeability (7,10).…”

Role of the intestinal tight junction modulator zonulin in the pathogenesis of type I diabetes in BB diabetic-prone rats

“…Although it is well accepted that tj are dynamic structures, surprisingly little is known about their regulation. The discovery of zonula occludens toxin (Zot), a protein elaborated by Vibrio cholerae (18) and of its receptor (19), has shed some light on the intricate mechanisms involved in the modulation of the intestinal paracellular pathway (20) and led us to the discovery of its eukaryotic counterpart zonulin (21). This protein is involved in the innate immunity of the gut (3) and, when inappropriately up-regulated, appears to play a key role in the increased intestinal permeability and pathogenesis of autoimmune diseases such as CD (22).…”