Cell division does not increase transepithelial permeability of LLC-PK1 cell sheets.

The transepithelial electrical resistance (TER) across LLC-PK1 cell sheets is unstable for several days postseeding, even when the cells are trypsinized from a previously confluent culture and replated at confluent density. We therefore followed the TER of LLC-PK1 cells plated at confluent density to elucidate characteristics of the TER fluctuations after plating of the cells. Control cultures reached a maximum TER of 1800 omega.cm2 approximately 24 h after plating. TER then declined sharply, reaching generally stable values (approximately 175 omega.cm2) only after 4 days. Cell cycle activity ([3H]-thymidine incorporation) peaked approximately 22 h after plating, prior to the peak in 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 values reached at 3-4 days postseeding to exceed time matched controls by approximately 100%. This EGF-treated group showed a concomitant increase in [3H]thymidine incorporation and cell density compared to control. Transepithelial fluxes of [14C]D-mannitol across control vs EGF-treated cell sheets were not, however, significantly different at the 4-day time point, indicating that a change in tight junction sieving on the basis of size had not occurred. Diffusion and bi-ionic potential studies indicated that the change in TER in EGF-treated cells was instead due to altered charge selectivity at the tight junction and/or intercellular space. We conclude: (1) TER across LLC-PK1 cell sheets does not stabilize until 4 days after seeding at confluent density and (2) cell division and resultant increased cell density in LLC-PK1 cell sheets can correlate with elevated TER values, due to altered ion permeability of the paracellular pathway. Permeability to both Na+ and Cl- decreased as a result of EGF treatment but the decline in chloride permeability was significantly greater. Not only was there a decrease in the permeabilities of all halide anions after exposure of cell sheets to EGF, but the permeability sequence changed after EGF exposure.