Use of alveolar cell monolayers of varying electrical resistance to measure pulmonary peptide transport

The apparent permeability coefficient (P-app) of two fluorescently tagged m odel hydrophilic peptides, acXASNH(2) and acXAS(GAS)(7)NH2, and C-14-mannit ol across monolayers of cultured rat alveolar epithelial cel:Ls of varying transepithelial electrical resistance (TER) has been examined. In line with their design features, the peptides were not degraded under the conditions of the test. Furthermore, no concentration dependence of transport of the tripeptide acXASNH(2) was observed over the concentration range studied, no r was any directional transport seen for either of the model peptides, indi cating that under the conditions of the test they were not substrates for a ny transporters or efflux pumps. From the hydrophilic nature of the peptide s (as assessed by their log P), and their inverse dependence of transport w ith molecular weight and TER, it was assumed that the peptides were transpo rted across the cell monolayer passively via the paracellular route. The ob served P-app for the transport of C-14-mannitol and the peptides across rat alveolar epithelial cell monolayers were found to be inversely (though not linearly) related to the measured TER and could be well-modeled assuming t he presence of two populations of "pores" in the cell monolayer, namely, cy lindrical pores of diameter 1.5 nm and large pores of diameter 20 nm. The r elative populations of the two types of pores varied with the TER of the mo nolayer, with the number of large pores decreasing with an increase in TER (and the number of small pores taken as fixed). These results suggest that if the cell monolayer is well characterized with respect to the passage of a range of probe molecules across monolayers of varying electrical resistan ce, it should be possible to predict the P-app of any hydrophilic peptide o r drug crossing the membrane by the paracellular route at any desired TER u sing a monolayer of any electrical resistance, above a minimum value. (C) 2 000 Wiley-Liss, Inc.