Proinflammatory cytokines tumor necrosis factor-alpha and interferon-gamma modulate epithelial barrier function in Madin-Darby canine kidney cells through mitogen activated protein kinase signaling.
BACKGROUND
The tight junction is a dynamic structure that is regulated by a number of cellular signaling processes. Occludin, claudin-1, claudin-2 and claudin-3 are integral membrane proteins found in the tight junction of MDCK cells. These proteins are restricted to this region of the membrane by a complex array of intracellular proteins which are tethered to the cytoskeleton. Alteration of these tight junction protein complexes during pathological events leads to impaired epithelial barrier function that perturbs water and electrolyte homeostasis. We examined MDCK cell barrier function in response to challenge by the proinflammatory cytokines tumor necrosis factor-alpha (TNFalpha) and interferon-gamma (IFNgamma).


RESULTS
Exposure of MDCK cells to TNFalpha/IFNgamma resulted in a marked sustained elevation of transepithelial electrical resistance (TER) as well as elevated paracellular permeability. We demonstrate that the combination of TNFalpha/IFNgamma at doses used in this study do not significantly induce MDCK cell apoptosis. We observed significant alterations in occludin, claudin-1 and claudin-2 protein expression, junctional localization and substantial cytoskeletal reorganization. Pharmacological inhibition of ERK1/2 and p38 signaling blocked the deleterious effects of the proinflammatory cytokines on barrier function.


CONCLUSIONS
These data strongly suggest that downstream effectors of MAP kinase signaling pathways mediate the TNFalpha/IFNgamma-induced junctional reorganization that modulates MDCK cell barrier function.
Patrick
David M
DM
Southwestern Graduate School of Biomedical Sciences, UT Southwestern Medical Center, Dallas, TX 75390-9004, USA. David.Patrick@UTSouthwestern.edu
Leone
Amanda K
AK
Shellenberger
Jeffry J
JJ
Dudowicz
Kara A
KA
King
Jonathan M
JM
D000818:Animals; D015153:Blotting, Western; D002460:Cell Line; D002463:Cell Membrane Permeability / Q000187:drug effects; D062445:Claudin-1; D062465:Claudin-3; D004285:Dogs; D017097:Electric Impedance; D004847:Epithelial Cells / Q000187:drug effects / Q000201:enzymology / Q000378:metabolism*; D018836:Inflammation Mediators / Q000494:pharmacology; D007371:Interferon-gamma / Q000494:pharmacology* / Q000633:toxicity; D007668:Kidney / Q000166:cytology; D020935:MAP Kinase Signaling System / Q000187:drug effects; D008565:Membrane Proteins / Q000032:analysis / Q000378:metabolism; D020928:Mitogen-Activated Protein Kinases / Q000037:antagonists & inhibitors; D062793:Occludin; D022502:Stress Fibers / Q000648:ultrastructure; D019108:Tight Junctions / Q000737:chemistry / Q000187:drug effects / Q000378:metabolism; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology* / Q000633:toxicity
D062445:Claudin-1; D062465:Claudin-3; D018836:Inflammation Mediators; D008565:Membrane Proteins; D062793:Occludin; D014409:Tumor Necrosis Factor-alpha; D007371:Interferon-gamma; D020928:Mitogen-Activated Protein Kinases