In a recent study of MDCK II cells investigators demonstrate that these cells have endogenously low ERK1/2 activity that corresponds to high expression of claudin 2. ERK1/2 inhibition in all of these studies prevented elevation of TER in the MDCK CP-868596 II cell line. Recently investigators have determined that claudin 2 forms cation selective channels in the tight junction complex, alteration in claudin 2 expres sion results in perturbations in ionic permeability. Con sistent with these studies we find a dose dependent decrease in claudin 2 expression in MDCK cells treated with TNF IFN, this loss of claudin 2 correlates to a sub stantial reduction in ionic permeability. Elevation in TER was inhibited by treatment with the ERK1/2 inhibitor but not by inhibiting the p38 signaling pathway.

These find ings are consistent with the current literature demonstrat ing that claudin 2 levels are regulated following ERK1/2 activation in MDCK cells and its expression level will influence recorded TER from MDCK cultures. The cellular tight junction response to proinflammatory cytokines is variable based on cell type and numerous physiological variables. Measurable changes in tight junc tion protein expression or localization that are predicted to play a key role in maintaining barrier function are typ ically more unpredictable. We report a statistically signifi cant elevation in the protein expression of claudin 1, but not occludin or claudin 3, following exposure to TNF IFN. However, occludin protein levels are slightly ele vated in response to several doses of TNF IFN tested compared to control.

In this study, we report a dose dependent decrease in claudin 2 expression following exposure to TNF IFN. The heterogeneous response of tight junctional proteins to cytokine exposure may be due to junctional remodeling which may involve additional protein synthesis and altered turnover rates. In other stud ies, researchers have reported decreases, increases or no change in tight protein expression following challenge with proinflammatory mediator. For instance TNF increased permeability while decreasing ZO 1 expression through increased NFB signaling, in a study using Caco 2 cells. Investigators report increased paracellular flux with a decrease in TER following TNF IFN exposure using a mouse cholangiocyte model. interestingly major structural changes to the tight junction proteins were not detected.

Finally, using T84 cells investigators find that inhibition of MEK signaling impairs both basal and cytokine induced tight junction formation demonstrating an increased claudin 1 and claudin 2 protein expression in response to the cytokine IL 17. Although it might be tractable to pre dict that exposure to proinflammatory cytokines would be correlated to decreased expression of tight junction pro teins, our study is in agreement with other studies, finding moderate Batimastat effects on expression.