HLA-B27 transgenic rats exhibit striking Th17 expansion and activation in the colon that is temporally related to the development of colitis. At the cellular level, macrophages undergoing a UPR induced either by HLA-B27 misfolding or exposure to pharmacologic inducers of ER stress, are polarized to produce more IL-23 in response to LPS. Taken together, these data suggest that the IL-23/IL-17 axis may play a role in the pathogenesis of spondyloarthritis-like disease in HLA-B27 transgenic rats, and demonstrate a potential link between HLA-B27 misfolding and immune dysregulation.
In recent years, the IL-23/IL-17 axis and CD4+ Th17 cells have gained widespread attention for their role in immune-mediated inflammatory diseases in rodent models (17
) and several human diseases including inflammatory bowel disease (IBD) (21
). IL-17 is also overexpressed in subjects with HLA-B27-associated spondyloarthropathies (22
), and polymorphisms in the IL23R gene are associated with susceptibility to ankylosing spondylitis (25
). These findings provide strong support for involvement of the IL-23/IL-17 axis in human spondyloarthritis as well as in the rat model.
Th17 cells are important regulators of intestinal homeostasis, and are present in healthy lamina propria at much higher frequency than in peripheral tissues (27
). They also have the capacity to become pathogenic under the influence of increased local expression of IL-23 (28
). The increase in IL-23 subunit expression found in the colon of HLA-B27 transgenic rats occurs at least as early as the increase in IL-17 (6 weeks; ). We also found increased expression of IFN-γ in the colon (), and evidence of Th1 expansion and activation (). These early changes either precede or are coincident with, the development of diarrhea, which typically begins between 6 and 9 weeks of age. IFN-γ overexpression has been demonstrated previously along with increases in IL-1α, IL-1β, TNF-α, IL-6, MIP-2 and iNOS (1
). These data have been interpreted in support of colitis being a Th1-mediated process. However, this type of cytokine profile (i.e. IL-17 in addition to IFN-γ, TNF-α, IL-6, and IL-1β) is also seen in mouse models of inflammatory bowel disease shown to be driven by IL-23 (16
). Our studies demonstrate that colitis in HLA-B27 transgenic rats is associated with prominent Th17 expansion and activation, and are consistent with the idea that it could be driven by excess local production of IL-23 in the lamina propria. In support of this, the striking inflammatory phenotype of IL-23p19 transgenic mice includes gastrointestinal inflammation (17
The extent to which IL-17 mediates intestinal inflammation in animal models remains unclear. In IL-10-deficient mice IL-17 blockade is only effective in suppressing intestinal inflammation if IL-6 is also neutralized (20
), and in other T cell-dependent inflammatory bowel disease models inhibition of Th1 responses attenuates disease (16
). In uveitis and encephalomyelitis both Th1 and Th17 cells mediate pathology (34
). In HLA-B27 transgenic rats, IL-10 administration reduced IFN-γ, IL-1β and TNF-α expression in the colon without affecting the severity of colitis (32
), consistent with the idea that other cytokines such as IL-17 may be important. In addition, Th17 cells can secrete IL-22 and IL-21, which may contribute to their pathogenicity (12
). Thus, while IL-17 is pro-inflammatory and clearly responsible for pathology in some animal models of inflammatory bowel disease (16
), we do not at this time know its relative importance in the context of other pro-inflammatory cytokines in HLA-B27 transgenic rats.
Interestingly, IFN-γ is an antagonist of Th17 development in mice (37
), and genetic ablation of IFNG
results in Th17 expansion and exacerbates several types of Th17-mediated immunopathology (39
). However, in humans with psoriasis, IFN-γ has been shown to induce Th17 T cells, in part through enhancing IL-23 expression, and promote their trafficking and function (40
). In HLA-B27 transgenic but not WT rats, our data suggest that IFN-γ promotes IL-23 expression via HLA-B27-induced ER stress and UPR activation. In addition, preliminary studies suggest that a subset of Th17 CD4+ T cells in the colon of HLA-B27 transgenic rats also expresses IFN-γ (unpublished observations), consistent with evidence for IL-17/IFN-γ double positive T cells in humans (41
). It will be important to further explore the balance and interplay between Th17 and Th1 T cells and cytokines in mediating inflammation in HLA-B27 transgenic rats.
The findings presented here are consistent with a model where HLA-B27 misfolding might promote a chronic inflammatory process such as colitis. Colonization of the gastrointestinal tract with commensal organisms results in a low level immune response including IFN-γ production (42
), which is normally controlled (43
). However, in HLA-B27 transgenic rats, IFN-γ could have a paradoxical effect by increasing HLA-B27 expression and generating ER stress, thus superimposing the UPR on macrophage activation. Macrophages may then become sensitized to pathogen-associated molecular patterns such as LPS that signal through pattern recognition receptors including the TLRs (). Increased expression of IL-23 in response to microbial products would activate CD4+ Th17 cells to produce IL-17 leading to tissue specific inflammation and damage. Production of IFN-γ by Th1 and Th17 T cells could perpetuate HLA-B27 misfolding and UPR activation (), particularly in the presence of other cytokines such as TNF-α that synergize with IFNs to increase class I expression. IFN-γ also primes antigen-presenting cells to produce more IL-12 (14
). Consistent with this possibility, we find upregulation of IL-12 subunits (p35 and p40) in antigen-presenting cells isolated from the lamina propria (). While there appears to be a shift in the Th17/Th1 balance associated with UPR activation and increased IL-23 expression, there is still considerable Th1 activation, which may play an important role in the inflammatory disease.
Figure 6 Proposed mechanism linking UPR activation as a consequence of HLA-B27 misfolding to activation of the IL-23/Il-17 axis. HLA-B27 upregulation may occur initially as a result of antigen presenting cell stimulation with TLR agonists from commensal microorganisms (more ...)
There are several other hypotheses to explain the role of HLA-B27 in disease (reviewed in (44
)). Evidence that CD8αβ T cells are not required for spondyloarthritis-like disease in rats argues against antigen presentation as an initiating event (3
). Dendritic cell dysfunction that could reduce tolerance to microbial flora has been reported (45
), and cell surface dimers of HLA-B27 heavy chains have been hypothesized to modulate the immune response and lead to inflammation (46
). Our studies do not rule out the involvement of alternative mechanisms, and it is conceivable that more than one mechanism is responsible.
The rats used for these studies did not develop arthritis, consistent with previous observations that this component of the inflammatory phenotype is rare in younger animals, particularly on the F344 background ((1
) and unpublished observations). In future studies it will be important to examine the IL-23/IL-17 axis in arthritis, including the spondylitis phenotype that occurs when additional hβ2
m is expressed in HLA-B27 transgenic rats (47
). Overexpression of additional hβ2
m was reported to curb HLA-B27 misfolding and caused a small reduction in BiP mRNA levels in splenocytes (47
). However, these studies did not examine macrophages or the response to HLA-B27 upregulation, which is important for maximal UPR activation (7
Our results suggest a novel mechanism linking HLA-B27 misfolding and the generation of ER stress to augmented TLR4-mediated induction of IL-23p19 via activation of the UPR. This may sustain CD4+ Th17 cells and drive the production of IL-17 and IFN-γ from double positive T cells. The IL-23/IL-17 axis has been implicated in the pathogenesis of several immune-mediated inflammatory diseases in humans, including psoriasis and Crohn's disease, as well as animal models. Our results strongly support a role for this axis in the pathogenesis of colitis in HLA-B27 transgenic rats. Considering genetic studies implicating IL23R polymorphisms in susceptibility to ankylosing spondylitis, this work suggests a mechanism that might link HLA-B27 misfolding to the IL-23/IL-17 axis in humans, and should prompt further inquiry into the role of these cytokines in the pathogenesis of human spondyloarthritis.