DIFFERENT SIZE LIMITATIONS FOR INCREASED TRANSEPITHELIAL PARACELLULARSOLUTE FLUX ACROSS PHORBOL ESTER AND TUMOR NECROSIS FACTOR-TREATED EPITHELIAL-CELL SHEETS

By observing increases in the transepithelial paracellular permeabilit y of a range of radiolabeled solutes and electron dense dyes, changes in molecular sieving caused by the cytokine, TNF (tumor necrosis facto r), and the phorbol ester, TPA (12-0-tetra-decanoylphorbol-13 acetate) , were characterized. Using C-14-labeled mannitol (mw 182), raffinose (mw 504), PEG (polyethylene glycol; mw 4000), and dextran (mw 70,000, 70,000 and 2,000,000), the transepithelial flux rates of these compoun ds were determined at the peak oi the transepithelial electrical resis tance (TER) changes caused by these two agents. TNF treatment resulted in increased permeability across LLC-PK1 epithelial cell sheets only to relatively small solutes, with an upper limit of approximately 4,00 0 mM,. The low molecular weight ''ceiling'' for the TNF-treated epithe lium is further evidence against TNF increasing transepithelial permea bility by means of inducing nonspecific, microscopic ''holes'' in the epitheiium, For which a ''ceiling'' would not exist. TPA treatment inc reases transepithelial paracellular permeability to a much broader ran ge of solutes, extending well beyond 2 million mw. Transmission electr on micrographs provide evidence that even the electron-dense dye compl ex, ruthenium red, can cross tight junctions of TPA-treated cell sheet s. However, cationic Ferritin cannot cross tight junctions of TPA-trea ted cell sheets. This shows that there is an upper limit to solutes ab le to cross TPA-treated cell sheets, but that this upper limit will in clude most proteins, which would then be able to cross tumor promoter- exposed (protein kinase C-activated) epithelial layers at accelerated rates. The biomedical implications for a high molecular weight cutoff in tumor promoter action in epithelial carcinogenesis, and for a low m olecular weight cutoff in cytokine-induced epithelial apoptosis in inf lammation, are discussed. (C) 1997 Wiley-Liss, Inc.