The Effects of Epstein-Barr Virus in a Mouse Model of Inflammatory Bowel Disease

Abstract

Background: The Epstein-Barr virus (EBV) has been shown to play a role in amplifying the pathogenesis of inflammatory bowel disease (IBD), a chronic inflammatory malady with a poorly defined etiology. In previous research conducted by our laboratory, EBV DNA was demonstrated to exacerbate IBD in mouse models, which leads to increased production of the pro-autoimmune cytokine IL-17A. We have also reported that the usage of a Toll-like receptor 9 (TLR9) antagonist managed to alleviate EBV DNA-exacerbated intestinal inflammation in a mouse model of IBD. Hence, the primary aim of this research is to assess the influence of EBV directly rather than DNA, on the exacerbation of IBD, and explore the potential mitigation of this exacerbation through TLR9 inhibition.

Methods: A C57BL/6J mouse model of dextran sodium sulfate (DSS)-induced acute colitis was employed in this study. A total of fifty-eight mice were obtained and distributed amongst five groups of ten and one group of eight. Four groups out of six were administered 1.5% DSS in drinking water from day 0 until day 7. The groups were then rectally administered sterile water or 432 x 103 EBV particles in sterile water on day 3. On day 4, two groups of mice were intra-peritoneally injected with 56µg of ODN 2088 (TLR9 inhibitor) in 100µl of sterile water. The effect of EBV and TLR9 inhibition on EBV-exacerbated colitis was assessed based on trends in the disease activity index (DAI), macroscopic assessments of the colon, histological damage grading of H&E-stained colon cross sections, and immunofluorescence staining of pro-inflammatory markers in colon cross sections.

Results: The group that was intra-rectally administered EBV in addition to DSS had a statistically significant increase in the DAI, and histological scores in comparison to the group that only received DSS. The group that received an intra-peritoneal injection of the TLR9 inhibitor in addition to EBV and DSS administration had a statistically significant decrease in the DAI, and histological scores compared to the group that received intra-rectal EBV in addition to the DSS administration. In alignment with the DAI and histology scores, the group that received EBV in addition to DSS had a statistically significant decrease in the colon length in comparison to the group that received DSS alone. The group that received an intra-peritoneal injection of the TLR9 inhibitor in addition to EBV and DSS exhibited a significant increase in colon length compared to the group that received EBV and DSS. Consistent with these findings, the group that received intra-rectal EBV alongside DSS administration had a significant increase in the levels of foci positive for IL-17A, FOXP3, double positive IL-17A+ IFNγ+, and triple positive IL-17A+IFNγ+FOXP3+ foci in comparison to the group that only received DSS. The group that received an intra-peritoneal injection of the TLR9 inhibitor in addition to EBV and DSS had a significant decrease in the number of IL-17A, IFNγ, double positive IL-17A+ IFNγ+ and triple positive IL-17A+IFNγ+FOXP3+ foci found in the gut compared to the group that received EBV and DSS.

Conclusions: Our study demonstrates that EBV exacerbates intestinal inflammation in a mouse model of IBD and that TLR9 inhibition alleviates some of the severity of EBV-exacerbated intestinal inflammation in this mouse model. This indicates that TLR9 inhibition can provide a prospective therapeutic avenue for managing IBD in subjects with EBV infection. Further investigations could explore different dosages of this inhibitor, along with alternative methods of delivery.