As mentioned at the outset of this review, the cytokines driving ulcerative colitis were identified as having Th2-like characteristics in the initial studies attempting to place it within the Th1/Th2 spectrum. These consisted of lamina propria cells producing increased amounts of IL-5 in the conspicuous absence of increased amounts of IL-4, the more defining Th2 cytokine; hence, the descriptive phrase, Th2-like3
. Also absent was any hint of an increased IFN-γ response, thus ruling out the presence of a Th1-driven inflammation. More recently, several studies have appeared showing that IL-17 levels were increased in ulcerative colitis compared to controls, but in the most reliable studies in which protein rather than mRNA was measured this increase was found to be far less than that found in Crohn’s disease61
In an attempt to gain insight into the origin of this kind of inflammation investigators studied several colitis models driven by a Th2 response, such as TCRα chain deficiency and late-phase IL-10 deficiency64
. However, these models differed from ulcerative colitis by the fact that the dominant cytokine abnormality was an elevated IL-4 response, which is not observed in UC. A breakthrough came when it was discovered that intra-rectal administration of oxazolone, an agent that like TNBS binds to self-proteins and renders them immunogenic, causes an intense but short-lived colitis that exhibits characteristics of ulcerative colitis65
. The latter consisted of a superficial inflammation associated with micro-ulcerations of the epithelium, edema of the bowel wall and an inflammatory infiltrate containing granulocytes. In addition, the inflammation was most intense in the distal half of the colon, as frequently found in ulcerative colitis.
Initial cytokine analysis of oxazolone-colitis (oxa-colitis) revealed the presence of cells producing greatly increased amounts of IL-4 but not IFN-γ and increased amounts of TGF-β, a cytokine whose production is favored by Th2 conditions. Furthermore, treatment of oxa-colitis with anti-IL-4 prevented disease whereas as treatment with anti-IL-12p40 exacerbated disease. Thus, there could be no question that oxa-colitis, in contrast to TNBS-colitis, fit into the Th2 T cell spectrum. It should be noted that these studies of oxa-colitis was conducted with SJL/J mice, a strain which proved to be more susceptible than C57BL/6 to oxa-colitis, as was the case for TNBS-colitis.
In a second study of oxa-colitis, mice were pre-sensitized to oxazolone by skin painting and could thus be induced with a lower intra-rectal dose of oxazolone that caused a less intense and more persistent inflammation66
. This modified oxa-colitis model revealed that the cytokine response, given more time, evolved from an initial IL-4 response lasting only several days into a more prolonged IL-13 response. In addition, the latter was shown to emanate from CD4+
NKT cells and not from conventional CD4+
T cells. Indeed, these studies proved that IL-13 and NKT cells are the driving force of oxa-colitis in that treatment of the latter with an IL-13 blocker such as IL-13Rα2-Fc or with antibodies that delete NKT cells, leads to amelioration of the inflammation. In addition, mice with CD1-deficiency that lack the ability to present antigens to NKT cells as well as mice with Jα281-deficiency that lack the invariant T cell receptors (TCRs) that NKT cells usually use to recognize antigen were also unable to support oxa-colitis. Finally, stimulation of lamina propria T cells with CD1-expressing antigen-presenting cells loaded with a-galactosyl-ceramide, the canonical antigen that has been shown to stimulate NKT cells expressing the “invariant” TCR associated with NKT cells, led to high level production of Th2 cytokines, including IL-13. Taken together, these data offered definitive proof that oxa-colitis was due to NKT cells producing IL-13.
The above studies of oxa-colitis set the stage for studies of ulcerative colitis, which would determine if an NKT cell/IL-13 abnormality also was present in this disease. Initial studies showed that stimulated lamina propria T cells from UC patients did indeed produce high levels of IL-13 but not IL-4 (or IFN-γ) whereas cells from CD patients produced only slightly increased amounts of IL-13 (compared to controls)11
. Similarly, flow cytometric studies revealed that UC patients had high levels of lamina propria CD4+
T cells producing IL-13 whereas CD patients had only minor increases in such cells. In further studies, lamina propria T cells from UC patients were shown to produce IL-13 when stimulated with B cells stably transfected with CD1d, i.e., cells which preferentially activate NKT cells. It was noted, however, that stimulation of UC lamina propria cells with a-galactosyl-ceramide did not lead to activation of NKT cells as determined by up-regulation of Vα and Vβ chain expression. It was thus evident that UC lamina propria harbored increased numbers of IL-13-producing NKT cells but that these cells were not typical NKT cells bearing invariant T cell receptors (capable of responding to a-galactosyl-ceramide).
In a final series of studies, the potential of either NKT cells and/or IL-13 to mediate tissue damage was evaluated. It was found that purified lamina propria CD4+
T cells were cytotoxic to HT-29 epithelial cells pre-stimulated with LPS to up-regulate CD1d and that such cytotoxicity was augmented by IL-13. Furthermore, it was observed that IL-13 (but not IL-4) lowers the electrical resistance of HT-29 epithelial cell monolayers by inducing epithelial cell apoptosis and increasing expression of pore-forming tight-junction protein claudin-267
. Interestingly, the latter was also observed in patients with ulcerative colitis. These studies suggest that the presence of NKT cells in the lamina propria of ulcerative colitis can lead to ulceration and inflammation by NKT cell-mediated cytotoxicity; in addition, such tissue injury is abetted by IL-13, which augments NKT cell cytotoxicity and has effects on epithelial cell integrity (See ).
The Pathogenesis of Ulcerative Colitis
The above studies of ulcerative colitis provide a basic framework with which to understand the immunopathogenesis of UC; nevertheless, they leave certain basic questions unanswered including the identification of the glycolipid antigen or antigens that stimulate UC NKT cells and the mechanism by which IL-13 stimulates NKT cell cytotoxic activity. In addition, in that they focus on IL-13 as a major factor in UC, they fail to explain why a polymorphism of the gene encoding the IL-23 receptor would result in protection from the development of UC (as well as CD), although one possibility here is that since IL-23R is found on NKT cells the polymorphism may be affecting NKT cell activity68
. Finally, it is important to mention that the above studies offer no formal proof that either IL-13 or NKT cells (or both) are pathologic factors in UC as they manifestly are in oxa-colitis. Nothing short of clinical studies in which the efficacy of IL-13 or NKT cell inhibitors is tested under blinded conditions will suffice to establish this possibility. This said, one recently published clinical study does offer promising correlative evidence that IL-13 is a pathologic factor in UC. This consisted of a study of a group of patients with UC given IFN-β therapy in which it was found that significant response to this therapy was clearly correlated with decreased IL-13 production by lamina propria T cells69