Myosin light chain kinase inhibition: correction of increased intestinal epithelial permeability in vitro.
OBJECTIVE
To examine whether myosin light chain kinase (MLCK) inhibitors can reduce intestinal epithelial permeability increases in vitro.


METHODS
Isolated rat, mouse and human colonic tissue mucosae and Caco-2 monolayers were exposed to cytochalasin D (cD) and sodium caprate (C10), in the absence and presence of the MLCK inhibitors, ML-9 and D PIK. Transepithelial electrical resistance (TEER) and Papp of [14C]-mannitol or FITC-dextran 4000 (FD-4) were measured. Western blots were used to measure MLC phosphorylation.


RESULTS
Increases in Papp of [14C]-mannitol and decreases in TEER were induced by tight junction openers. These changes were attenuated by ML-9. D-PIK offset the FD-4 Papp increase induced by C10 in Caco-2 only, while ML-9 and PIK inhibited MLC directly, cD induced constriction of peri-junctional actin in Caco-2 monolayers, but this was prevented by ML-9. Although mannitol fluxes across colonic mucosae from dextran-sulphate (DSS)-treated mice were higher than control, they were not ameliorated by either ML-9 or PIK in vitro.


CONCLUSIONS
ML-9 inhibits paracellular permeability increases in several intestinal epithelial models. D-PIK reduced stimulated paracellular fluxes in Caco-2 monolayers, but not in tissue. Pre-established increases were not modified by two MLCK inhibitors in a mouse model of IBD.
Feighery
Linda M
LM
School of Agriculture, Food Science and Veterinary Medicine, University College Dublin, Belfield, Dublin 4, Ireland.
Cochrane
Sean W
SW
Quinn
Teresa
T
Baird
Alan W
AW
O'Toole
Daniel
D
Owens
Sian-Eleri
SE
O'Donoghue
Diarmuid
D
Mrsny
Randall J
RJ
Brayden
David J
DJ
D000818:Animals; D015153:Blotting, Western; D017097:Electric Impedance; D006801:Humans; D015212:Inflammatory Bowel Diseases / Q000378:metabolism; D007413:Intestinal Mucosa / Q000378:metabolism*; D008297:Male; D008353:Mannitol / Q000493:pharmacokinetics; D051379:Mice; D009219:Myosin-Light-Chain Kinase / Q000037:antagonists & inhibitors*; D010539:Permeability; D010766:Phosphorylation; D047428:Protein Kinase Inhibitors / Q000494:pharmacology*; D051381:Rats; D017208:Rats, Wistar
D047428:Protein Kinase Inhibitors; D008353:Mannitol; D009219:Myosin-Light-Chain Kinase