title: Permeability enhancement in Caco-2 cell monolayers by sodium salicylate and sodium taurodihydrofusidate: assessment of effect-reversibility and imaging of transepithelial transport routes by confocal laser scanning microscopy.
abstract: The effects of sodium salicylate and sodium tauro-24,25-dihydrofusidate (STDHF) on the aqueous permeability of confluent monolayers of Caco-2 cells were studied. Measurements of transepithelial electrical resistance (TEER) showed a concentration-dependent effect of both compounds after apical incubation for 1 hr. Reductions in TEER resulting from EC50 concentrations (2.8 mM for STDHF; 173 mM for salicylate) were reversible within 5.75 hr. The transpithelial fluxes of two hydrophilic model compounds, sodium fluorescein F (molecular weight 376) and a fluorescein isothiocyanate-labeled dextran (mean molecular weight 4000) was significantly increased by STDHF (2.8 mM). Sodium salicylate (173 mM) only enhanced the transport of sodium fluorescein significantly. At the EC50 concentrations, confocal laser scanning microscopy (CLSM) visualized both fluorescent tracers mainly in the paracellular route. With higher enhancer concentrations (373 mM sodium salicylate and 8 mM STDHF), both transport markers appeared intracellularly as a result of cell death. STDHF rapidly extracted an exogenous lipophilic membrane probe, 5-(N-hexadecanoyl)aminofluorescein (HEDAF), from the apical part of Caco-2 plasma membranes, indicating qualitatively that STDHF interacts with the lipid portion of cell membranes. These results suggest that both sodium salicylate and STDHF can be used to reversibly increase paracellular permeability of Caco-2 cell monolayers, whereby STDHF appears to be advantageous compared to sodium salicylate. By adapting the Costar cell culture system to CLSM, we have shown that this technique is suitable to study membrane interactions qualitatively and for visualizing transport routes of hydrophilic tracers through nonfixed, filter-grown monolayers.
authors.affiliation: Leiden/Amsterdam Center for Drug Research, Leiden University, The Netherlands.
authors.forename: M A
authors.initials: MA
authors.lastname: Hurni
authors.forename: A B
authors.initials: AB
authors.lastname: Noach
authors.forename: M C
authors.initials: MC
authors.lastname: Blom-Roosemalen
authors.forename: A G
authors.initials: AG
authors.lastname: de Boer
authors.forename: J F
authors.initials: JF
authors.lastname: Nagelkerke
authors.forename: D D
authors.initials: DD
authors.lastname: Breimer
mesh_terms: D000277:Adjuvants, Pharmaceutic / Q000494:pharmacology* / Q000633:toxicity; D001692:Biological Transport / Q000187:drug effects; D002462:Cell Membrane / Q000378:metabolism; D002463:Cell Membrane Permeability / Q000187:drug effects*; D003110:Colonic Neoplasms; D003911:Dextrans / Q000493:pharmacokinetics; D004594:Electrophysiology; D004848:Epithelium / Q000378:metabolism; D019793:Fluorescein; D016650:Fluorescein-5-isothiocyanate / Q000031:analogs & derivatives / Q000493:pharmacokinetics; D005452:Fluoresceins / Q000493:pharmacokinetics; D005672:Fusidic Acid / Q000031:analogs & derivatives* / Q000494:pharmacology / Q000633:toxicity; D006801:Humans; D007408:Intestinal Absorption / Q000187:drug effects*; D007834:Lasers; D050356:Lipid Metabolism; D008853:Microscopy / Q000379:methods; D012980:Sodium Salicylate / Q000494:pharmacology* / Q000633:toxicity; D014343:Trypan Blue / Q000493:pharmacokinetics; D014407:Tumor Cells, Cultured
chemical_list: D000277:Adjuvants, Pharmaceutic; D003911:Dextrans; D005452:Fluoresceins; C015219:fluorescein isothiocyanate dextran; D005672:Fusidic Acid; C053824:tauro-24,25-dihydrofusidate; C058719:(hexadecanoyl)aminofluorescein; D016650:Fluorescein-5-isothiocyanate; D014343:Trypan Blue; D019793:Fluorescein; D012980:Sodium Salicylate