University of Groningen Differential Effects of TNF (TNFSF2) and IFN-gamma on Intestinal Epithelial Cell Morphogenesis and Barrier Function in Three-Dimensional Culture Juuti-Uusitalo,

Background: The cytokines TNF (TNFSF2) and IFNc are important mediators of inflammatory bowel diseases and contribute to enhanced intestinal epithelial permeability by stimulating apoptosis and/or disrupting tight junctions. Apoptosis and tight junctions are also important for epithelial tissue morphogenesis, but the effect of TNF and IFNc on the process of intestinal epithelial morphogenesis is unknown. Methods/Principal Findings: We have employed a three-dimensional cell culture system, reproducing in vivo-like multicellular organization of intestinal epithelial cells, to study the effect of TNF on intestinal epithelial morphogenesis and permeability. We show that human intestinal epithelial cells in three-dimensional culture assembled into luminal spheres consisting of a single layer of cells with structural, internal, and planar cell polarity. Exposure of preformed luminal spheres to TNF or IFNc enhanced paracellular permeability, but via distinctive mechanisms. Thus, while both TNF and IFNc, albeit in a distinguishable manner, induced the displacement of selected tight junction proteins, only TNF increased paracellular permeability via caspase-driven apoptosis and cell shedding. Infliximab and adalumimab inhibited these effects of TNF. Moreover, we demonstrate that TNF via its stimulatory effect on apoptosis fundamentally alters the process of intestinal epithelial morphogenesis, which contributes to the de novo generation of intestinal epithelial monolayers with increased permeability. Also IFNc contributes to the de novo formation of monolayers with increased permeability, but in a manner that does not involve apoptosis. Conclusions: Our study provides an optimized 3D model system for the integrated analysis of (real-time) intestinal epithelial paracellular permeability and morphogenesis, and reveals apoptosis as a pivotal mechanism underlying the enhanced permeability and altered morphogenesis in response to TNF, but not IFNc. Citation: Juuti-Uusitalo K, Klunder LJ, Sjollema KA, Mackovicova K, Ohgaki R, et al. (2011) Differential Effects of TNF (TNFSF2) and IFN-c on Intestinal Epithelial Cell Morphogenesis and Barrier Function in Three-Dimensional Culture. PLoS ONE 6(8): e22967. doi:10.1371/journal.pone. 0022967 Editor: Hang Thi Thu Nguyen, Emory University, United States of America Received March 1, 2011; Accepted July 4, 2011; Published August 11, 2011 Copyright: 2011 Juuti-Uusitalo et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Funding: This study was supported by Top Institute Food and Nutrition (http://www.tifn.nl). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Competing Interests: The authors have declared that no competing interests exist. * E-mail: s.c. d.van. ijzendoorn@med.umcg. nl ¤ Current address: Department of Animal Sciences, Wageningen UR, The Netherlands