Introduction:
Coeliac disease (CeD) is a gluten-triggered immune-mediated enteropathy characterized by villous atrophy and crypt hyperplasia. IFNγ has been implicated in crypt hyperplasia, while recent transcriptomic studies have an expansion of the transient amplifying zone as the putative cause for crypt hyperplasia in CeD. Whether IFNγ can directly drive such a reprogramming of the epithelial independently of immune and stromal cells remains unclear.
Methods：
Human duodenal organoid-derived 2D epithelial monolayers were established from 3 healthy donors (HD) and 3 CeD patients. Barrier function and epithelial organization were assessed by Transepithelial Electrical Resistance (TEER) and immunofluorescence following IFNγ and/or TNFα exposure. For transcriptomic profiling, monolayers were treated with IFNγ (0.25, 0.5, or 1 ng/mL) for 6 h followed by bulk RNA sequencing.
Results:
Neither IFNγ nor TNFα alone significantly reduced TEER, whereas IFNγ pre-exposure sensitized the epithelium to a subsequent TNFα-induced reduction in TEER. IFNγ-exposed monolayers retained continuous ZO-1 staining, but showed increased Claudin-2 staining and disrupted F-actin organization.
Transcriptomic profiling revealed dose-dependent responses to IFNγ in both groups, with increased IFN/JAK–STAT-associated, inflammatory, and cell-cycle/proliferation related passways (NES >0, FDR <0.001). PCA further demonstrated a significant dose-dependent transcriptional shift, with PC1 scores changing progressively with increasing IFNγ concentrations in both groups. Although E2F targets, G2M checkpoint, and mitotic spindle were induced in both groups, their enrichment was lower in treated CeD than in treated HD (NES <0, FDR <0.001). OXPHOS was significantly suppressed by IFNγ in CeD (NES <0, FDR <0.001) but not HD, yet remained higher in CeD than HD under treated conditions (NES >0, FDR <0.01).
Conclusion:
These findings support a direct epithelial role for IFNγ in driving dose-dependent inflammatory and proliferative transcriptional responses, with distinct cell-cycle and metabolic features in CeD-derived epithelium that may contribute to the epithelial alterations observed in active CeD.