Regulation of paracellular permeability in Caco-2 cell monolayers by protein kinase C

W. F. Stenson, R. A. Easom, T. E. Riehl, J. Turk

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Abstract

Caco-2 cells are an enterocyte-like cell line derived from a human colonic adenocarcinoma. Paracellular permeability was assessed in monolayers of these cells by transmonolayer resistance and by the permeation of [3H]mannitol across the monolayer. Paracellular permeability was increased by the protein kinase C (PKC) activator phorbol 12-myristate 13-acetate (50 nM), carbachol (500 μM), and the combination of carbachol (50 μM) and monolein (100 μM), an inhibitor of diacylglycerol kinase, as manifested by a decrease in transmonolayer resistance and an increase in mannitol permeation. The effects of all of these stimuli on transmonolayer resistance were inhibited by staurosporine (3 nM), an inhibitor of PKC. The effects of carbachol plus monolein were also inhibited by atropine (0.1 μM), a muscarinic antagonist. Treatment of the monolayers with each of the stimuli was associated with translocation of PKC activity from cytosol to a membrane-associated state. Stimulation of Caco-2 cell monolayers with phorbol myristate acetate or with the combination of carbachol and monolein was also associated with phosphorylation of the MARCKS protein, an endogenous substrate of PKC. These data support the hypothesis that intestinal paracellular permeability is regulated by the activity of enterocyte PKC and demonstrate that the increase in paracellular permeability induced by binding of carbachol to the muscarinic receptor is mediated by activation of PKC.

Original languageEnglish
Pages (from-to)G955-G962
JournalAmerican Journal of Physiology - Gastrointestinal and Liver Physiology
Volume265
Issue number5 28-5
DOIs
StatePublished - 1993

Keywords

  • atropine
  • carbachol
  • intestine
  • monolein
  • phorbol myristate acetate
  • staurosporine
  • tight junction

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