CYTOSKELETAL REGULATION OF CACO-2 INTESTINAL MONOLAYER PARACELLULAR PERMEABILITY

An abnormal increase in intestinal paracellular permeability may be an important pathogenic factor in various intestinal diseases. The intra cellular factors and processes that regulate and cause alteration of i ntestinal paracellular permeability are not well understood. The purpo se of this study was to examine some of the intracellular processes in volved in cytoskeletal regulation of intestinal epithelial paracellula r permeability using the filter-grown Caco-2 intestinal epithelial mon olayers. Cytochalasin-b and colchicine were used to disrupt the cytosk eletal elements, actin microfilaments, and microtubules. Cytochalasin- b (5 mu g/ml) and colchicine (2 x 10(-5)M) at the doses used caused ma rked depolymerization and disruption of actin microfilaments and micro tubules, respectively. Cytochalasin-b-induced disruption of actin micr ofilaments resulted in perturbation of tight junctions and desmosomes and an increase in Caco-2 monolayer paracellular permeability. The cyt ochalasin-b-induced disruption of actin microfilaments and subsequent changes in intercellular junctional complexes and paracellular permeab ility were not affected by inhibitors of protein synthesis (actinomyci n-D or cycloheximide) or microtubule function (colchicine), but were i nhibited by metabolic energy inhibitors (2,4-dinitrophenol or sodium a zide). The cytochalasin-b-induced disturbance in Caco-2 actin microfil aments and intercellular junctional complexes and increase in paracell ular permeability were rapidly reversed. The paracellular pathway ''re -tightening'' following cytochalasin-b removal was not affected by act inomycin-D, cycloheximide, or colchicine, but was inhibited by 2,4-din itrophenol and sodium azide. The colchicine-induced disruption of micr otubules did not have significant effect on actin microfilaments, inte rcellular junctions, or paracellular permeability. These findings sugg est that cytochalasin-b-induced increase in Caco-2 monolayer paracellu lar permeability was due to actin microfilament mediated perturbation of intercellular junctional complexes. The re-tightening of paracellul ar pathways (following removal of cytochalasin-b) resulted from energy -mediated re-assembly of pre-existing actin microfilaments and interce llular junctional complexes. This re-closure process did not require p rotein synthesis or microtubule-mediated shuttling process. (C) 1995 W iley-Liss, Inc.