EFFECTS OF EPIDERMAL GROWTH-FACTOR AND PHORBOL ESTER ON THYROID EPITHELIAL INTEGRITY

The effects of epidermal growth factor (EGF) and phorbol ester (tetra- O-decanoylphorbol-16-acetate; TPA) on thyroid epithelial integrity wer e studied in filter-cultured monolayers of porcine thyrocytes, which b efore experiments were growth-arrested and had a high transepithelial resistance (R(TE) > 6 . 10(3) Omega . cm(2)) and polarized, thyroid-sp ecific functions. Both EGF and TPA stimulated dose-dependently the cel lular incorporation of [H-3]thymidine, which maximally (at 10 ng/ml EG E for 48 h) corresponded to a 65% increase of the DNA content. The EGF -treated cells proliferated mainly within the original monolayer, whic h became folded due to the increased cell number; clusters of epitheli al cells also assembled between the monolayer and the filter. Although the transepithelial potential difference was reduced, from 15-30 mV i n controls to 2-10 mV, the epithelial barrier function was maintained (R(TE) 1-3 . 10(3) Omega . cm(2); impermeability to [H-3]inulin). EGF did not change the ultrastructural polarity of the plasma membrane or the distinct distribution of ZO-1 and cadherin immunoreactivities to j unctions, but cytoplasmic cadherin present in controls disappeared aft er EGF. In cultures acutely depleted of extracellular Ca2+ EGF pretrea tment for 48 h antagonized the preventive effect of thyrotropin on par acellular leakage and loss of cell-cell adhesion. TPA (0.1 mu M) induc ed a temporary barrier dysfunction (maximal after 24 h) accompanied by pronounced alterations of cell shape and actin-based cytoskeleton, di ssociation of junctional cadherin, and shedding of cells into the apic al medium. In long-term (2-5 days) TPA-treated cultures the epithelial morphotype and barrier function recovered. The combined stimulation w ith EGF and TPA caused a persistent derangement of the cell layer incl uding attenuation of ZO-1 at cell-cell contacts, paracellular leakage of [H-3]inulin, and cell detachment. We conclude that EGF is able to r elease porcine thyroid epithelial cells from contact inhibition of gro wth along with intact cell polarity and tight junctions. Yet, when act ing together with phorbol ester EGF provokes a lasting morphological t ransformation. Impaired positive control of Ca2+-dependent cell-cell a dhesion in EGF-treated cultures suggests a latent defect with possible transforming potential in the cadherin-based regulation of the juncti onal complex. (C) 1995 Academic Press, Inc.