TNF-α Modulation of Intestinal Tight Junction Permeability is Mediated by NIK/IKK-α Axis Activation of the Canonical NF-κB Pathway

Tumor necrosis factor (TNF)-alpha, a key mediator of intestinal inflammation, causes an increase in intestinal epithelial tight junction (TJ) permeability by activating myosin light chain kinase (MLCK; official name MYLK3) gene. However, the precise signaling cascades that mediate the INF-alpha-induced activation of MLCK gene and increase in TJ permeability remain unclear. Our aims were to delineate the upstream signaling mechanisms that regulate the TNF-alpha modulation of intestinal TJ barrier function with the use of in vitro and in vivo intestinal epithelial model systems. TNF-alpha caused a rapid activation of both canonical and noncanonical NF-kappa B pathway. NF-kappa B-inducing kinase (NIK) and mitogen-activated protein kinase kinase-1 (MEKK-1) were activated in response to TNF-alpha. NIK mediated the TNF-alpha activation of inhibitory kappa B kinase (IKK)-alpha, and MEKK1 mediated the activation of IKK complex, including IKK-beta. NIK/IKK-alpha axis regulated the activation of both NF-kappa B p50/p65 and RelB/p52 pathways. Surprisingly, the siRNA induced knockdown of NIK, but not MEKK-1, prevented the TNF-alpha activation of both NF-kappa B p50/p65 and RelB/p52 and the increase in intestinal TJ permeability. Moreover, NIK/IKK-alpha/NF-kappa B p50/p65 axis mediated the TNF-alpha induced MLCK gene activation and the subsequent MLCK increase in intestinal TJ permeability. In conclusion, our data show that NIK/IKK-alpha/regulates the activation of NF-kappa B p50/p65 and plays an integral role in the TNF-alpha induced activation of MLCK gene and increase in intestinal TJ permeability.