SPECIFIC MODULATION OF INTESTINAL EPITHELIAL BRUSH-BORDER ENZYME EXPRESSION BY A PHORBOL ESTER

Much is known about intestinal epithelial regulation by growth factors and nutrients but the intracellular signals governing cell phenotype are less well understood. In an initial attempt to evaluate the role o f protein kinase C in these events, we studied the effects of protein kinase C modulation by the phorbol ester TPA upon the differentiation, motility, and doubling time of the human intestinal epithelial Caco-2 cell line, a common model for enterocytic brush border enzyme express ion. We also compared the effects of TPA to those of 4 alpha-phorbol 1 2,13-didecanoate, which does not modulate protein kinase C activity. D ifferentiation was studied by quantitating brush order dipeptidyl pept idase (DPDD)-specific activity in protein-matched Caco-2 lysates via s ynthetic substrate digestion. Alkaline phosphatase (AP) was studied fo r comparison, Doubling time was assessed by log transformation of seri al cell counts and motility by monolayer expansion across type I colla gen. TPA (0.03-0.7 mu g/ml) dose-dependently stimulated DPDD, with a m aximal 455 +/- 26% increase at 0.7 mu g/ml (P < 0.01, n = 5). However, TPA dose-dependently inhibited AP to a maximal 91.6 +/- 0.3% decrease (P < 0.01, n = 5). TPA also dose-dependently prolonged the cell doubl ing time from 26.5 +/- 0.4 to 64.5 +/- 8.8 hr (n = 20, P < 0.01) with a maximal effect at 1.0 mu g/ml and inhibited migration with essential ly complete ablation of cell motility at 0.1 mu g/ml (n = 10, P < 0.00 1). By contrast, the control phorbol ester exerted equivalent effects on cell proliferation but promoted both AP and DP activity only slight ly (18.1 +/- 7.5 and 10.4 +/- 2.1%, respectively, n > 9, P < 0.05 for each). The control phorbol ester did not affect migration. Manipulatio n of protein kinase C may specifically alter the phenotype of a human intestinal epithelial cell line. (C) 1995 Academic Press, Inc.