Abstract
Inflammatory bowel diseases (IBD), such as Crohn’s disease (CD) and ulcerative colitis, are chronic inflammatory conditions that target the intestinal epithelial cells (IEC), driving barrier dysfunction, impaired epithelial turnover events, and altered mucous secretion. Patients with IBD that are non-responsive (NR) to the first-line anti-TNF therapy have been shown to have persistent type I interferon (IFN-I) levels and depleted butyrate levels that may drive alterations in the IEC barrier. I hypothesised that IFN-Is and butyrate do not directly alter IEC lineage commitment, butyrate receptor expression, or epithelial cytokine signalling, but may drive altered downstream JAK-STAT signalling pathways in NR patients. I also hypothesised that butyrate supplementation, and IFN-I blockade in combination with anti-TNF therapy could restore this altered downstream JAK-STAT signalling pathway. To test this, human derived 3D organoids were exposed to TNF, IL-1β, and IL-6 cytokines, 0.1 mM butyrate, anti-IFN-I, and anti-TNF antibodies. These organoids were collected for qRT-PCR to measure IEC lineage markers, and butyrate receptor markers, immunofluorescence staining of IEC phosphorylated pSTAT3, and organoid supernatants were collected for cytokine bead array analysis. Organoids derived from CD patients showed reduced stem cell marker expression and heightened proinflammatory cytokine expression compared to those from non-IBD donors. The addition of the butyrate, anti-IFN-I, and anti-TNF antibodies did not alter epithelial lineage commitment or butyrate receptor expression in CD and non-IBD organoids. Butyrate supplementation and anti-IFN-I antibody acted synergistically to reduce epithelial pSTAT3 levels. This effect was further enhanced by anti-TNF treatment in non-responder CD organoids, while selectively reducing pSTAT3 in responder CD organoids. These findings demonstrate that NR CD patients may benefit from anti-IFN-I and butyrate in combination with anti-TNF to restore IEC barrier integrity by targeting downstream JAK-STAT signalling pathways. These findings also highlighted heterogeneity in patient responses and the importance of tailoring therapies for individuals. Future work is needed to determine whether these observations are consistent in a larger cohort, and whether complex immune and microbial interactions could alter IEC responses to butyrate and IFN-I signalling.