Cytochalasin B modulation of Caco-2 tight junction barrier: role of myosin light chain kinase.
The intracellular mechanisms that mediate cytochalasin-induced increase in intestinal epithelial tight junction (TJ) permeability are unclear. In this study, we examined the involvement of myosin light chain kinase (MLCK) in this process, using the filter-grown Caco-2 intestinal epithelial monolayers. Cytochalasin B (Cyto B) (5 microg/ml) produced an increase in Caco-2 MLCK activity, which correlated with the increase in Caco-2 TJ permeability. The inhibition of Cyto B-induced MLCK activation prevented the increase in Caco-2 TJ permeability. Additionally, myosin-Mg(2+)-ATPase inhibitor and metabolic inhibitors (which inhibit MLCK induced actin-myosin contraction) also prevented the Cyto B-induced increase in Caco-2 TJ permeability. Cyto B caused a late-phase (15-30 min) aggregation of actin fragments into large actin clumps, which was also inhibited by MLCK inhibitors. Cyto B produced a morphological disturbance of the ZO-1 TJ proteins, visually correlating with the functional increase in Caco-2 TJ permeability. The MLCK and myosin-Mg(2+)-ATPase inhibitors prevented both the functional increase in TJ permeability and disruption of ZO-1 proteins. These findings suggested that Cyto B-induced increase in Caco-2 TJ permeability is regulated by MLCK activation.
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Division of Gastroenterology, Department of Medicine, Department of Veterans Affairs Medical Center, California State University, Long Beach 90822, California. tyma@uci.edu
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Bui
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Pedram
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Mills
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Merryfield
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D008841:Actin Cytoskeleton / Q000187:drug effects / Q000378:metabolism; D000199:Actins / Q000378:metabolism; D001692:Biological Transport / Q000187:drug effects / Q000502:physiology; D018938:Caco-2 Cells; D003571:Cytochalasin B / Q000494:pharmacology*; D015638:Cytochalasin D / Q000494:pharmacology; D003931:Diacetyl / Q000031:analogs & derivatives* / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004847:Epithelial Cells / Q000166:cytology / Q000378:metabolism; D005947:Glucose / Q000378:metabolism; D006801:Humans; D007413:Intestinal Mucosa / Q000166:cytology / Q000378:metabolism; D008565:Membrane Proteins / Q000378:metabolism; D009219:Myosin-Light-Chain Kinase / Q000378:metabolism*; D009218:Myosins / Q000378:metabolism; D019384:Nucleic Acid Synthesis Inhibitors / Q000494:pharmacology; D010750:Phosphoproteins / Q000378:metabolism; D019108:Tight Junctions / Q000187:drug effects* / Q000201:enzymology*; D062826:Zonula Occludens-1 Protein
D000199:Actins; D004791:Enzyme Inhibitors; D008565:Membrane Proteins; D019384:Nucleic Acid Synthesis Inhibitors; D010750:Phosphoproteins; C569042:TJP1 protein, human; D062826:Zonula Occludens-1 Protein; C004717:diacetylmonoxime; D015638:Cytochalasin D; D003571:Cytochalasin B; D009219:Myosin-Light-Chain Kinase; D009218:Myosins; D005947:Glucose; D003931:Diacetyl