IMPORTANCE OF THE PARACELLULAR PATHWAY FOR THE TRANSPORT OF A NEW BISPHOSPHONATE USING THE HUMAN CACO-2 MONOLAYERS MODEL

The transport of a new bisphosphonate, Tiludronate, was investigated o n the human adenocarcinoma cell line, CACO-2. Experiments were perform ed 7-16 days after cells achieved confluence, conditions under which t hey form well-differentiated monolayers joined by tight junctions. Til udronate transport rate across CACO-2 monolayers was independent of th e temperature (4-degrees versus 37-degrees), of the polarity of the ce ll membrane (apical-to-basolateral versus basolateral-to-apical), and of the presence of metabolic poisons (sodium azide). Its transport was enhanced by either the presence of EGTA in the incubation buffer, i.e . when extracellular Ca2+ concentration was reduced, or by the pretrea tment of monolayers with EGTA, i.e. when the intercellular spaces and the tight junctions were widened. Based on these different observation s, we could suggest that Tiludronate mainly used the paracellular path way to cross the intestinal epithelium. An increase in the Tiludronate permeability coefficient was also observed following treatment of cel ls with high Tiludronate concentrations, as a consequence of the direc t effect of this compound on the extracellular Ca2+ ions. Hence, for h igh drug concentrations, i.e. 20 mM, we observed a decrease in free ex tracellular Ca2+ concentration, an increase in the transepithelial ele ctrical resistance and an increase in the transport of [C-14]polyethyl eneglycol ([C-14]PEG400), a probe for the paracellular pathway. The re sults indicate that Tiludronate is transported across CACO-2 monolayer s by the paracellular route. Moreover, it can affect its own transport by its concentration-dependent effect on tight junction widening.