Abstract
Inflammatory Bowel Disease (IBD), comprising Crohn’s Disease (CD) and Ulcerative Colitis (UC), is a generic terminology used to describe conditions involving chronic idiopathic inflammation in the gastrointestinal tract. It affects 1 in 1,000 individuals with a total economic cost of $2.7 billion per annum. Despite decades of research, the aetiology of IBD remains unclear. However, current hypothesis suggests that IBD stems from a complex interaction between various environmental, genetic and immunologic factors.
In both UC and CD, leukocyte infiltration into the intestine is a fundamental event in the disease development and progression, which is largely believed to be orchestrated by integrins, and chemokines. Researchers have already demonstrated that patients with active IBD have increased levels of chemokine receptors and corresponding ligands, namely CXCR3 and its IFN-γ-inducible ligands, CXCL9, CXCL10 and CXCL11. Despite this, the role of CXCR3 in the pathogenesis of IBD remains poorly understood.
Following 2.5% DSS-induced colitis, CXCR3-/- mice revealed attenuated colitis as shown by reduced weight loss, clinical, faecal and hispathological scores when compared to Wt mice. Furthermore, change in colon length following DSS challenge did not reflect those of Wt mice, suggesting reduced inflammation and colitis. Surprisingly, B cell migration increased in CXCR3-/- during DSS-induced colitis, as shown by increase IgG+ and significantly increased IgA+ plasma cells in the colon. This suggests that CXCR3 may not be important for B cell migration to the colon during DSS-induced colitis and/or other recruitment factors may compensate. Upon further analysis of cellular infiltrates, neutrophils, monocytes and CD4+ T cells were significantly reduced in the colon of CXCR3-/- mice following DSS challenge. TNF, IFN-γ and IL-6 were significantly reduced, while IL-4 was increased in DSS-challenged CXCR-/- mice compared to Wt animals, revealing a sway in the Th profile from Th1 to Th2.
To help illuminate the role of CXCR3 during bacterial-induced colitis, CXCR3-/- mice were orally challenged with Salmonella and harvested at various time-points to assess disease progression. Our findings showed that CXCR3 is important in mounting an effective immune response towards Salmonella. CXCR3-/- mice challenged with S.typhimurium develop augmented Salmonellosis, marked by increased bacterial loading in the caecum and bacterial dissemination to the spleen and liver. Interestingly, neutrophil and macrophage migration was not affected during Salmonellosis in CXCR3-/- mice; however, their efficiency and/or functionality may be significantly reduced as IFN-γ and GM-CSF secretion was attenuated during the early phase and late phase of Salmonellosis CXCR3-/- mice, respectively. Contrarily, CD4+ T cell recruitment to the caecum was limited in CXCR3-/- during Salmonellosis, suggesting CXCR3 is important for CD4+ T cell migration during bacterial-induced colitis. Analysis of the cytokine and chemokine profile showed significantly reduced levels of the pro-Th1 IFN‐γ, IL-2, IL-4, IL-5 and IL-6 and IL-17 in the caecum of CXCR3-/- mice following Salmonella challenge. Collectively, our results show that during a LPS-driven immune response, CXCR3 may not be required for migration of CXCR3+ leukocytes, as previously suggested during DSS-induced colitis. However, it appears that the activity and functionality of migrating leukocytes, namely neutrophils and monocytes/macrophages are more important in containing and clearing S.typhimurium in the caecum.
Microspheres (beads) have previously been demonstrated to be up-taken by neutrophils and monocytes, consequently limiting the ability of these cells to migrate to the inflamed site. In an effort to understand the effect of neutrophil and monocyte chemotaxis, mice were challenged with 2.5% DSS while treated intravenously with beads. Our results revealed that beads-treated mice challenged with DSS develop attenuated colitis, reflected by reduced body weight loss, clinical, faecal and histopathological scores as well as significantly increased colon lengths when compared to mock-treated DSS-challenged mice. As expected, neutrophils and monocytes recruitment to the inflamed colon was significantly mitigated, while CD4+ T cell migration did not change, suggesting that CD4+ T cells do not contribute significantly to developing colitis. IFN-γ and IL-12, important cytokines for Th1 differentiation, were significantly reduced in beads-treated, DSS-challenged mice, suggesting that neutrophils and monocytes may play an important role in sourcing early cytokines for the development of a Th1 response.
Our findings also suggests that early neutrophil and monocyte recruitment to the inflamed sit may source important cytokines for the development of a Th1 response as IFN-γ preliminary findings suggest an effective immune response can be mounted as beads administration did not reduce IFN-γ, a strong marker for Th1 responses and an important cytokine implicated in clearance and resolution
Collectively, the presented data further expands the current understanding of CXCR3 in intestinal immunity, with important implications in IBD and other inflammatory disorders. In addition, our findings have revealed an important aspect of the CXCR3 axis responsible for its pro-inflammatory actions and the subsequent host-tissue damage caused as a result. Utilizing beads to reduce the migration of neutrophils and monocytes to the inflamed site may be an attractive therapeutic tool in treating patients with IBD and possibly other inflammatory disorders.
