Interleukin-6 Modulation of Intestinal Epithelial Tight Junction Permeability Is Mediated by JNK Pathway Activation of Claudin-2 Gene

Rana Al-Sadi,Shuhong Guo,Thomas Y Ma,Lisa Ereifej,Mariam Hashimi,Michel Boivin,Dongmei Ye,Shrikant Anant

doi:10.1371/journal.pone.0085345

Abstract

Defective intestinal epithelial tight junction (TJ) barrier has been shown to be a pathogenic factor in the development of intestinal inflammation. Interleukin-6 (IL-6) is a pleiotropic, pro-inflammatory cytokine which plays an important role in promoting inflammatory response in the gut and in the systemic circulation. Despite its key role in mediating variety inflammatory response, the effect of IL-6 on intestinal epithelial barrier remains unclear. The purpose of this study was to investigate the effect of IL-6 on intestinal epithelial TJ barrier and to delineate the intracellular mechanisms involved using in-vitro (filter-grown Caco-2 monolayers) and in-vivo model (mouse intestinal perfusion) systems. Our results indicated that IL-6 causes a site-selective increase in Caco-2 intestinal epithelia TJ permeability, causing an increase in flux of small-sized molecules having molecular radius <4 Å. The size-selective increase in Caco-2 TJ permeability was regulated by protein-specific increase in claudin-2 expression. The IL-6 increase in TJ permeability required activation of JNK signaling cascade. The JNK pathway activation of AP-1 resulted in AP-1 binding to its binding sequence on the claudin-2 promoter region, leading to promoter activation and subsequent increase in claudin-2 gene transcription and protein synthesis and TJ permeability. Our in-vivo mouse perfusion showed that IL-6 modulation of mouse intestinal permeability was also mediated by AP-1 dependent increase in claudin-2 expression. In conclusion, our studies show for the first time that the IL-6 modulation of intestinal TJ permeability was regulated by JNK activation of AP-1 and AP-1 activation of claudin-2 gene.

Highlights

Intestinal mucosal surface is covered by a single layer of columnar epithelial cells
The dose- and time-course of IL-6 effect on Caco-2 tight junction (TJ) barrier function was assessed by measuring Caco-2 trans-epithelial electrical resistance (TER)
IL-6 caused a dose-dependent drop in Caco-2 TER (Fig. 1A) during the 48-hr experimental period; IL-6 concentration of 10 ng/ml caused a maximal drop in TER (p,0.001) and increasing the IL-6 concentration above 10 ng/ml did not cause additional drop in TER. (In all subsequent studies, unless otherwise indicated, a dose of 10 ng/ml of IL-6 was used)

Summary

Introduction

Intestinal mucosal surface is covered by a single layer of columnar epithelial cells. The intestinal epithelial cells and the tight junctions form the intrinsic intestinal epithelial barrier, serving as a physical and functional barrier against trans-epithelial permeation of luminal substances [1,2]. The intercellular tight junctions act as gate or barrier against paracellular permeation of hydrophilic molecules in-between adjacent cells [1,3]. It is well-established that in intestinal permeability disorders, the defective intestinal TJ barrier allows paracellular permeation of luminal antigens which can initiate or propagate inflammatory response [1,4]. A persistent increase in intestinal permeability was predictive of early recurrence of Crohn’s disease and poor prognosis, while normalization of intestinal permeability following medical therapy predicted sustained clinical remission [14,15]

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