CELL-DIVISION DOES NOT INCREASE TRANSEPITHELIAL PERMEABILITY OF LLC-PK1 CELL SHEETS

The transepithelial electrical resistance (TER) across LLC-PK1 cell sh eets is unstable for several days postseeding, even when the cells are trypsinized from a previously confluent culture and replated at confl uent density. We therefore followed the TER of LLC-PK1 cells plated at confluent density to elucidate characteristics of the TER fluctuation s after plating of the cells. Control cultures reached a maximum TER o f 1800 Omega . cm(2) approximately 24 h after plating. TER then declin ed sharply, reaching generally stable values (approximate to 175 Omega . cm(2)) only after 4 days. Cell cycle activity ([H-3]-thymidine inco rporation) peaked approximately 22 h after plating prior to the peak i n TER values, and then followed a decline similar to that of the TER. Treatment of cells with EGF at 24 h after plating caused the TER value s reached at 3-4 days postseeding to exceed time matched controls by a pproximately 100%. This EGF-treated group showed a concomitant increas e in [H-3]thymidine incorporation and cell density compared to control . Transepithelial fluxes of [C-14]D-mannitol across control vs EGF-tre ated cell sheets were not, however, significantly different at the 4-d ay time point, indicating that a change in tight junction sieving on t he basis of size had not occurred Diffusion and bi-ionic potential stu dies indicated that the change in TER in EGF-treated cells was instead due to altered charge selectivity at the tight junction and/or interc ellular space. We conclude: (1) TER across LLC-PK1 cell sheets does no t stabilize until 4 days after seeding at confluent density and (2) ce ll division and resultant increased cell density in LLC-PK1 cell sheet s can correlate with elevated TER values, due to altered ion permeabil ity of the paracellular pathway. Permeability to both Na+ and Cl- decr eased as a result of EGF treatment but the decline in chloride permeab ility was significantly greater, Not only was there a decrease in the permeabilities of all halide anions after exposure of cell sheets to E GF, but the permeability sequence changed after EGF exposure. (C) 1995 Academic Press, Inc.