Entamoeba histolytica interactions with polarized human intestinal Caco-2 epithelial cells

Abstract

To model the initial pathogenic eflects of Entamoeba histolytica trophozoites on intestinal epithelial cells, the interactions of E. histolytica HM1-IMSS trophozoites with polarized human intestinal Caco-2 cell monolayers grown on permeabilized filters were examined. Trophozoites, when incubated with the apical surface of the monolayers at 37°C, induced a rapid decrease in transepithelial resistance over 15 to 60 min. The transmonolayer resistance response was not associated with changes in short-circuit current but was associated with an increase in mannitol flux, suggesting that the drop in resistance reflected a nonselective

Entamoeba histolytica disturbs the tight junction complex in human enteric T84 cell layers

“…We can only speculate about the molecular pathways followed by signals from trophozoites to tight junctions. It is the opinion of Li et al (21) that the loss of the apical brush border in the region of contact between the trophozoites and epithelial cells, as also demonstrated in others (20) and our (unpublished results) transmission electron micrographs, reflects a more generalized disruption of the cytoskeleton. The latter might implicate disturbance of the tight junction complex as it is connected to the actin cytoskeleton (38).…”

“…Decrease of TER was ascribed to an increase in paracellular rather than in transcellular permeability. The arguments lie in the relationship between [ 3 H]mannitol flux and TER of enteric T84 cell layers in our and in the others' experiments and the lack of changes in short-circuit current (21). A similar interpretation was given to the effect of the actin-disturbing agent cytochalasin D and of low Ca 2ϩ concentration on tight junctions, respectively, in mucosal preparations (28) and in MDCK cell layers (33).…”

“…Moreover, cytotoxicity as evidenced by 51 Cr release was not associated with a decrease of TER. Differences between our kinetic analysis of TER decrease as compared to 51 Cr release and the results obtained by others (21,36) may be ascribed to differences in target cell type, technique, and time of coculturing or to decreased virulence of trophozoites maintained in axenic culture.…”

“…A dose-and time-dependent, trophozoite-induced decrease of TER of enteric cell layers established on filters in two-compartment chambers was also observed by others (20,21), but a molecular explanation had, so far, not been provided. Decrease of TER was ascribed to an increase in paracellular rather than in transcellular permeability.…”

“…Sonicates of trophozoites were prepared by intermittent treatment (Vibra Cell VC50; Sonics & Materials, Danbury, Conn.) of 1 ϫ 10 5 live trophozoites for a total of 45 s. To evaluate cell death, live cocultures were stained with fluorescein diacetate and propidium iodide for examination by fluorescence microscopy (27). To test amebic cytotoxicity, we also used the 51 Cr release assay (21). T84 cells grown on top of 24-well filters were incubated with 60 Ci of 51 Cr in 800 l DMEM/HAMF12 with serum in the lower chamber for 2 h. The monolayers were washed four times on the basal side and two times on the apical side with unlabeled DMEM/HAMF12 with serum.…”