Evaluation of viability of excised rat intestinal segments in the Ussing chamber: investigation of morphology, electrical parameters, and permeability characteristics.

PURPOSE: To clarify relations between alterations in electrical and permeability data with time and to elaborate accompanying structural changes of intestinal segments in Ussing chamber experiments. METHODS: Excised intestinal segments from the rat were studied in a modified Ussing chamber. Experiments were run up to 180 minutes during which the electrical parameters, PD, SCC, and R, were measured and the permeability coefficients (Papp) of mannitol and propranolol were determined. Each segment was observed under the light microscope for morphological evaluation. RESULTS: PD and SCC values showed a decrease for most segments while the R values remained steady throughout the experiment. The Papp for propranolol increased aborally to the small intestine. For mannitol, the reversed was observed. In some cases, there was a time-dependent change in permeability for these marker molecules. The main morphological changes observed were a decreased nucleo-apical distance, decreased villi amplification factor, initial edema, cell sloughing, and epithelial restitution. CONCLUSIONS: The time-dependent changes in permeability coefficients of mannitol and propranolol are suggested to be related to changes in electrical parameters and morphological alterations. Presented data illustrates the importance of information regarding time-dependent structural changes for correct interpretation of permeability data.

Sodium caprate elicits dilatations in human intestinal tight junctions and enhances drug absorption by the paracellular route.

The effects of the absorption enhancer sodium caprate on human intestinal epithelial cells were investigated using Caco-2 cell monolayers. The effects on epithelial integrity and drug transport are dependent on time and concentration and are decreased by Ca2+, most likely through the formation of Ca2+ soaps. Morphological data indicate that exposure to sodium caprate results in cytoskeletal changes and in structural alterations of the tight junctions in the form of dilatations, while the effects on the apical cell membranes are limited. We conclude that sodium caprate increases the absorption of drugs mainly by the paracellular route.

Epithelial transport of drugs in cell culture. VIII: Effects of sodium dodecyl sulfate on cell membrane and tight junction permeability in human intestinal epithelial (Caco-2) cells.

This study demonstrates how the common pharmaceutical wetting agent sodium dodecyl sulfate (SDS) increases the absorption of drugs and peptides across the human intestinal epithelium. First, an assay that could follow the reversible and irreversible time-dependent effects of SDS on the permeability of Caco-2 cell monolayers with high reproducibility was developed. SDS (0.40 mM) exposure for 20 min resulted in reversible absorption enhancement of mannitol (M(r), 182 g/mol), 1-deamino-8-D-arginine-vasopressin (M(r), 1071 g/mol), and polyethylene glycol (M(r), 4000 g/mol). A longer (2 h) exposure to SDS resulted in irreversible absorption enhancement. Second, transepithelial electrical resistance measurements (TEER) together with fluorescence and transmission electron microscopy were used to study the effects of SDS on epithelial integrity, cell membranes, intracellular calcium concentration, cytoskeleton, and tight junctions. The effect of SDS (0.40 mM) on epithelial integrity was immediate. A significant decrease in transepithelial electrical resistance measurements was obtained with 1 min after exposure to SDS that was concomitant with increases in the permeability of the apical cell membranes and intracellular calcium concentration. SDS shortened the microvilli of the cells and produced apical (but not basolateral) membrane wounds, actin disbandment, disorganization of the terminal web, and structural separation of the tight junctions. The absorption enhancement was not reduced after repair of the apical cell membranes, indicating that SDS enhances drug and peptide absorption across the intestinal epithelium by the paracellular pathway.

Epithelial transport of drugs in cell culture. VII: Effects of pharmaceutical surfactant excipients and bile acids on transepithelial permeability in monolayers of human intestinal epithelial (Caco-2) cells.

The effects of anionic (sodium dodecyl sulfate and sodium dioctyl sufosuccinate) and nonionic (polysorbate 80 and polyoxyl 40 hydrogenated castor oil) synthetic surfactants and bile acids (sodium taurocholate, sodium taurodeoxycholate, and sodium taurodihydrofusidate) on epithelial integrity were studied in monolayers of human intestinal epithelial (Caco-2) cells grown on microporous polycarbonate filters. The effects of the surfactants on intracellular enzyme activity, cell monolayer permeability, and morphology were studied. The effects on permeability were studied by two methods: measurements of transport of marker molecules (mannitol and polyethylene glycol) and measurements of transepithelial electrical resistance. All surfactants demonstrated concentration-dependent effects on intracellular enzyme activities, permeability, and morphology. The effects of the anionic surfactants were more pronounced than those of the nonionic surfactants. The effects on transepithelial electrical resistance correlated with intracellular dehydrogenase activity. Fluxes of marker molecules were the most sensitive measure of epithelial integrity. The results indicate that the hydrophilic marker molecules permeate the epithelial monolayers through different pathways at different concentrations of the surfactants. The effects of the surfactants were reversible at intermediate concentrations, even though the morphology of the monolayers had changed. The results agree with published data obtained with experimental animals and indicate that Caco-2 cells can be used to study the concentration-dependent effects of surfactants and other pharmaceutical additives on intestinal epithelial permeability.