Interferon-γ selectively increases epithelial permeability to large molecules by activating different populations of paracellular pores

Abstract

Impairment of the gut epithelial barrier by agents such as IFNγ may play a key role in the pathogenesis of inflammatory disorders by increasing the paracellular penetration of luminal macromolecules, potentially including bacterial antigens. Owing to limitations of current paracellular probes, little is known about the precise functional changes induced by IFNγ and how these relate to the development of increased macromolecular permeability. Here we investigate how IFNγ modulates this pathway in T84 monolayers using a novel profiling technique that resolves different populations of paracellular pores by simultaneous analysis of 24 permeability probes of defined molecular size. Two types of functional pore present in control monolayers, an abundant restrictive pore with a radius of ∼4.5 Å and a much larger but infrequent, non-restrictive pore, were differentially regulated by IFNγ. Incubation with IFNγ dose-dependently and reversibly increased the frequency of the non-restrictive pores while having no significant effect on the restrictive component. Cytokine-induced increases in β, the descriptor of the non-restrictive pore, correlated closely with increased permeability to large molecules (10 kDa) including E. coli-derived lipopolysaccharide, but not small (0.182 kDa) molecules. This effect was associated with changes in expression of the tight junction proteins occludin and claudin-1. These data suggest that IFNγ selectively increases the transepithelial flux of large molecules by activating specific pathways within the junctional pore. One hypothesis is that this process may be activated in the early stages of the inflammatory response, facilitating the passage of large and potentially antigenic molecules across the gut without gross disruption of the barrier to small molecules.