Myosin light chain kinase inhibition: correction of increased intestinal epithelial permeability in vitro.

PURPOSE: To examine whether myosin light chain kinase (MLCK) inhibitors can reduce intestinal epithelial permeability increases in vitro. MATERIALS AND 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.

Effects of Lactobacillus salivarius 433118 on intestinal inflammation, immunity status and in vitro colon function in two mouse models of inflammatory bowel disease.

The probiotic, Lactobacillus salivarius subsp. salivarius 433118 (UCC118), was investigated for its potential to attenuate colitis, modulate immune responses and alter intestinal barrier dysfunction in two different mouse models of inflammatory bowel disease (IBD). Following oral treatment with UCC118, faecal microbial analysis indicated that viable intact bacteria reached the colons of interleukin (IL)-10(-/-) mice and dextran sodium sulphate (DSS)-treated mice. Neither prophylactic nor therapeutic UCC118 treatment significantly prevented or attenuated inflammation in either model. In all studies, the probiotic-treated mice had comparable cytokine responses as vehicle-treated animals. Mannitol permeability was increased across colonic mucosae mounted in Ussing chambers from DSS-treated mice, but not in IL-10(-/-) mice. However, colonic mucosae from UCC118-treated mice had unchanged transepithelial electrical resistance (TEER) values and mannitol fluxes compared to controls. In two different mouse colitis models examined under a range of histological and functional criteria, the data therefore suggest that this Lactobacillus subsp. has limited potential as a prophylactic or therapeutic treatment for inflammatory bowel disease. While several studies have shown therapeutic activity for this probiotic in mouse models of IBD, our data suggest that there are inter-study variables in formulation, study design, animal models and assessment criteria that may impact on interpretation of probiotic efficacy.