BACTERIAL TRANSLOCATION ACROSS ENTEROCYTES - RESULTS OF A STUDY OF BACTERIAL-ENTEROCYTE INTERACTIONS UTILIZING CACO-2 CELLS

Due to the inherent limitations of in vivo studies, the cellular mecha nisms underlying the process of bacterial translocation (BT) across th e intestinal epithelial barrier are poorly understood. Thus, we have u tilized the Caco-2 intestinal cell line to study this process. Caco-2 cells were grown to confluence on semipermeable membranes contained in the upper compartment of a 2 compartment system. Cellular confluence and tight junction integrity was verified by measurements of the trans epithelial electrical resistance in ohms cm2. BT was measured by cultu ring the bacteria (nonpathogenic Escherichia coli) that were able to c ross the Caco-2 monolayer and were present in the bottom compartment, as well as by monitoring the passage of 1-mum fluorescent beads. Caco- 2 cells were pretreated with several metabolic inhibitors: 1.0 mM sodi um azide (oxidative phosphorylation), 10 mug/ml nocodazole (microtubul e), 10 muM phalloidine (microfilament), and 5.0 mug/ml cytochalasin D (microfilament). To investigate the mechanisms of BT. Both bacteria an d fluorescent beads crossed the Caco-2 monolayer. Azide had no effect on BT while both nocodazole (n = 17) and phalloidine (n = 14) signific antly decreased translocation of E. coli versus control monolayers (p < .05). Cytochalasin D increased BT versus control membranes, however this was associated with loss of tight junction integrity (transepithe lial electrical resistance decreased from 201 +/- 79 to 87 +/- 6.4). B T across Caco-2 cells appears to be a polar process which is to some e xtent microtubule- and microfilament-dependent.