Epithelial permeability alterations in an in vitro air–liquid interface model of allergic fungal rhinosinusitis

Abstract

Background Chronic rhinosinusitis (CRS) is an inflammatory upper-airway disease with numerous etiologies. Patients with a characteristic subtype of CRS, allergic fungal rhinosinusitis (AFRS), display increased expression of Th2 cytokines and antigen-specific IgE. Various sinonasal inflammatory conditions are associated with alterations in epithelial barrier function. The aim of this study was to compare epithelial permeability and intercellular junctional protein expression amongst cultured primary sinonasal cells from AFRS patients versus non-inflammatory controls. Methods Epithelial cells isolated from paranasal sinus mucosa of AFRS and non-inflammatory control patients were grown to confluence on permeable supports and transitioned to air-liquid interface (ALI). Trans-epithelial resistance (TER) was measured with a horizontal Ussing chamber to characterize the functional permeability of each cell type. After TER recordings were complete, a panel of intercellular junctional proteins was assessed by Western blot and immunofluorescence labeling followed by confocal microscopy. Results After 12 samples were measured from each group, we observed a 41% mean decrease in TER in AFRS cells (296±89 ohms × cm2) compared to control (503±134 ohms × cm2, P=0.006). TER deficits observed in AFRS were associated with decreased expression of the tight junction proteins occludin and Junctional Adhesion Molecule-A (JAM-A), and increased expression of a leaky tight junction protein claudin-2. Conclusions Cultured sinonasal epithelium from AFRS patients displayed increased epithelial permeability and altered expression of intercellular junctional proteins. Given that these cells were not incubated with inflammatory cytokines in vitro, the cultured AFRS epithelial alterations may represent a retained modification in protein expression from the in vivo phenotype.

International Consensus Statement on Allergy and Rhinology: Rhinosinusitis

“…497 The barrier is maintained via ongoing production of AJC proteins as well as protective antiprotease molecules. The polypoid form of CRS and a Th2 cytokine milieu have been associated with significantly decreased levels of AJC proteins [498][499][500][501] as well as diminished intrinsic protective antiprotease activity. 486,502 Functional studies have also documented increased barrier permeability in CRSwNP.…”

“…486,502 Functional studies have also documented increased barrier permeability in CRSwNP. 500,501,503 It has been further proposed that the protein oncostatin M may play a key role mediating a leaky barrier in polyps. 503 Furthermore, a global gene microarray meta-analysis indicated that polyp epithelia exhibited a less mature gene signature typically associated with an epithelial to mesenchymal transition, which would predict a loss of barrier integrity.…”

Tight junctions in pulmonary epithelia during lung inflammation

“…In Calu3 cells, IL-4 induces disassembly of TJ molecules [100], IL-4 and IL-13 increase paracellular permeability in human bronchial epithelial cells [107, 118], in sinosonal epithelial cells [142] and in air-liquid interface cultivated paranasal sinus mucosa cells [16]. Although IL-4 and IL-13 show similar effects on TJ in these airway epithelial cell models, depending on the investigated model, they act via different pathways.…”

The Etiology and Pathogenesis of Chronic Rhinosinusitis: a Review of Current Hypotheses

“…Subsequent studies have suggested a weakened mechanical barrier as well, with diminished tight junctional proteins and increased susceptibility to exogenous protease degradation [79–81]. Functional studies demonstrated that the barrier was porous in CRSwNP, permitting increased access of foreign material across the epithelium [82, 83]. Recently, elevated levels of Oncostatin M, which is a cytokine that can cause the disassembly of tight junctions, have been reported in affected tissues, and may be hypothesized to play a role in barrier defects in CRS [84].…”