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.
Saladik
D T
DT
Lankenau Medical Research Center, Wynnewood, Pennsylvania 19096, USA.
Soler
A P
AP
Lewis
S A
SA
Mullin
J M
JM
D000818:Animals; D000838:Anions / Q000378:metabolism; D002455:Cell Division / Q000187:drug effects / Q000502:physiology*; D002462:Cell Membrane / Q000187:drug effects / Q000502:physiology; D002463:Cell Membrane Permeability; D002712:Chlorides / Q000378:metabolism; D004247:DNA / Q000096:biosynthesis; D004594:Electrophysiology / Q000379:methods; D004815:Epidermal Growth Factor / Q000494:pharmacology; D004847:Epithelial Cells; D004848:Epithelium / Q000187:drug effects / Q000502:physiology; D007700:Kinetics; D018374:LLC-PK1 Cells; D008353:Mannitol / Q000378:metabolism; D008564:Membrane Potentials; D012964:Sodium / Q000378:metabolism; D013552:Swine; D013936:Thymidine / Q000378:metabolism; D019108:Tight Junctions / Q000187:drug effects / Q000502:physiology; D013997:Time Factors
D000838:Anions; D002712:Chlorides; D008353:Mannitol; D004815:Epidermal Growth Factor; D004247:DNA; D012964:Sodium; D013936:Thymidine