The development of colitogenic CD4(+) T cells is regulated by IL-7 in collaboration with NK cell function in a murine model of colitis.

We previously reported that IL-7(-/-)RAG(-/-) mice receiving naive T cells failed to induce colitis. Such abrogation of colitis may be associated with not only incomplete T cell maintenance due to the lack of IL-7, but also with the induction of colitogenic CD4(+) T cell apoptosis at an early stage of colitis development. Moreover, NK cells may be associated with the suppression of pathogenic T cells in vivo, and they may induce apoptosis of CD4(+) T cells. To further investigate these roles of NK cells, RAG(-/-) and IL-7(-/-)RAG(-/-) mice that had received naive T cells were depleted of NK cells using anti-asialo GM1 and anti-NK1.1 Abs. NK cell depletion at an early stage, but not at a later stage during colitogenic effector memory T cell (T(EM)) development, resulted in exacerbated colitis in recipient mice even in the absence of IL-7. Increased CD44(+)CD62L(-) T(EM) and unique CD44(-)CD62L(-) T cell subsets were observed in the T cell-reconstituted RAG(-/-) recipients when NK cells were depleted, although Fas, DR5, and IL-7R expressions in this subset differed from those in the CD44(+)CD62L(-) T(EM) subset. NK cell characteristics were the same in the presence or absence of IL-7 in vitro and in vivo. These results suggest that NK cells suppress colitis severity in T cell-reconstituted RAG(-/-) and IL-7(-/-)RAG(-/-) recipient mice through targeting of colitogenic CD4(+)CD44(+)CD62L(-) T(EM) and, possibly, of the newly observed CD4(+)CD44(-)CD62L(-) subset present at the early stage of T cell development.

Upregulated IL-7 receptor α expression on colitogenic memory CD4+ T cells may participate in the development and persistence of chronic colitis.

We have previously demonstrated that IL-7 is essential for the persistence of colitis as a survival factor of colitogenic IL-7Rα-expressing memory CD4(+) T cells. Because IL-7Rα is broadly expressed on various immune cells, it is possible that the persistence of colitogenic CD4(+) T cells is affected by other IL-7Rα-expressing non-T cells. To test this hypothesis, we conducted two adoptive transfer colitis experiments using IL-7Rα(-/-) CD4(+)CD25(-) donor cells and IL-7Rα(-/-) × RAG-2(-/-) recipient mice, respectively. First, IL-7Rα expression on colitic lamina propria (LP) CD4(+) T cells was significantly higher than on normal LP CD4(+) T cells, whereas expression on other colitic LP immune cells, (e.g., NK cells, macrophages, myeloid dendritic cells) was conversely lower than that of paired LP cells in normal mice, resulting in predominantly higher expression of IL-7Rα on colitogenic LP CD4(+) cells, which allows them to exclusively use IL-7. Furthermore, RAG-2(-/-) mice transferred with IL-7Rα(-/-) CD4(+)CD25(-) T cells did not develop colitis, although LP CD4(+) T cells from mice transferred with IL-7Rα(-/-) CD4(+)CD25(-) T cells were differentiated to CD4(+)CD44(high)CD62L(-) effector-memory T cells. Finally, IL-7Rα(-/-) × RAG-2(-/-) mice transferred with CD4(+)CD25(-) T cells developed colitis similar to RAG-2(-/-) mice transferred with CD4(+)CD25(-) T cells. These results suggest that IL-7Rα expression on colitogenic CD4(+) T cells, but not on other cells, is essential for the development of chronic colitis. Therefore, therapeutic approaches targeting the IL-7/IL-7R signaling pathway in colitogenic CD4(+) T cells may be feasible for the treatment of inflammatory bowel diseases.

IL-2 is positively involved in the development of colitogenic CD4+ IL-7R alpha high memory T cells in chronic colitis.

IL-2 and IL-7 share a common gamma-chain receptor and are critical for T-cell homeostasis. We aimed to clarify the reciprocal roles of IL-2 and IL-7 in the development and persistence of chronic colitis. We performed a series of adoptive transfers of IL-2(-/-) CD4(+)CD45RB(high) T cells into RAG-2(-/-) mice and assessed the role of IL-2 in the induction of IL-7R alpha on colitogenic CD4(+) T cells and the development of chronic colitis. RAG-2(-/-) mice transferred with WT but not with IL-2(-/-) CD4(+)CD45RB(high) T cells developed Th1/Th17-mediated colitis. Consistently, re-expression of IL-7R alpha was severely impaired on IL-2(-/-) but not on WT CD4(+) T cells from the transferred mice. To exclude a contribution of the preclinical autoimmunity of IL-2(-/-)mice, WT Ly5.1(+) or IL-2(-/-) Ly5.2(+) CD4(+)CD45RB(high) T cells from GFP mice previously transplanted with the same number of WT and IL-2(-/-) BM cells were transferred into RAG-2(-/-) mice. RAG-2(-/-) mice transferred with IL-2(-/-)-derived CD4(+)CD45RB(high) T cells did not develop colitis, but their splenic CD4(+) T cells changed from effector-memory to central-memory type. These results show that IL-2 is critically involved in the establishment and maintenance of IL-7-dependent colitogenic memory CD4(+)IL-7R alpha(high) T cells.

RANK-RANKL signaling pathway is critically involved in the function of CD4+CD25+ regulatory T cells in chronic colitis.

It is now clear that functional CD4(+)CD25(+) regulatory T (T(R)) cells exist as part of the normal immune population and prevent the development of intestinal inflammation. We have recently shown that CD4(+)CD25(+) T(R) cells reside in the intestine and control intestinal homeostasis in humans and mice. In this study, we demonstrate that the TNF family molecule RANKL and its receptor RANK are critically involved in controlling the function of CD4(+)CD25(+) T(R) cells in the intestine. We first found that RANKL was preferentially expressed on both CD4(+)CD25(+) T(R) cells and colitogenic CD4(+) T cells, whereas RANK was expressed on dendritic cells. Although neutralizing anti-RANKL mAb did not affect T(R) activity of CD4(+)CD25(+) T(R) cells to suppress the proliferation of CD4(+) responder cells in vitro, in vivo administration of anti-RANKL mAb abrogated CD4(+)CD25(+) T(R) cell-mediated suppression of colitis induced by adoptive transfer of CD4(+)CD45RB(high) T cells into SCID mice. Interestingly, an adoptive transfer experiment using Ly5.1(+)CD4(+)CD45RB(high) cells and Ly5.2(+)CD4(+)CD25(+) T(R) cells revealed that the ratio of CD4(+)CD25(+) T(R) cells in total CD4(+) T cells in inflamed mucosa was significantly decreased by anti-RANKL mAb treatment. Consistent with this, the expression of RANK on lamina propria CD11c(+) cells from colitic mice was significantly increased as compared with that from normal mice, and in vitro treatment with anti-RANKL mAb suppressed the expansion of CD4(+)Foxp3(+) T(R) cells in culture with colitic lamina propria CD11c(+) cells. Together, these results suggest that the RANK-RANKL signaling pathway is critically involved in regulating the function of CD4(+)CD25(+) T(R) cells in colitis.

Long-lived colitogenic CD4+ memory T cells residing outside the intestine participate in the perpetuation of chronic colitis.

To understand the perpetuation of inflammatory bowel disease (IBD), it is important to clarify whether the colitogenic CD4(+) T cells are self-limited effector or long-lived memory T cells. We here investigate the latency of colitogenic CD4(+) T cells in the remission stage of colitis under germfree (GF) conditions. We isolated splenic (SP) CD4(+) T cells from colitic CD4(+)CD45RB(high) T cell-injected SCID mice maintained under specific pathogen-free (SPF) conditions and transferred them into SPF or GF SCID mice. Donor colitic SP CD4(+) T cells have a characteristic CD44(high)CD62L(-)IL-7Ralpha(high) effector-memory T-type phenotype. Six weeks after transfer of cells to GF SCID mice, one group of mice was continued in GF conditions (GF-->GF), and the other was transferred into SPF conditions (GF-->SPF). GF-->SPF but not GF-->GF SCID mice developed colitis with elevated production of Th1 and Th17 cytokines at 4 wk after transfer. Surprisingly, a large number of CD4(+) effector-memory T cells and a small but substantial number of central-memory T cells remained resident in SP and bone marrow, but not in lamina propria, of the GF-->GF SCID recipients. Consistent with this, GF-->SPF but not GF-->GF SCID mice rapidly developed colitis. Taken together, these findings suggest that long-lived colitogenic memory CD4(+) cells can be established even in the presence of commensal Ags, reside outside the intestine in the absence of commensal bacteria, and participate in the perpetuation of colitis. Thus, blocking a stimulus of colitogenic memory CD4(+) cells such as IL-7 may have therapeutic benefit for treatment of inflammatory bowel disease.

IL-7 is essential for lymphopenia-driven turnover of colitogenic CD4(+) memory T cells in chronic colitis.

We previously demonstrated that IL-7 is essential for the persistence of T-cell-mediated colitis, by showing that adoptive transfer of CD4(+)CD45RB(high) T cells into IL-7(-/-) x RAG-1(-/-) mice did not induce colitis; and that intestinal IL-7 is not essential for this colitis model, by showing that IL-7(-/-) x RAG-1(-/-) mice parabiosed with colitic CD4(+)CD45RB(high) T-cell-transferred RAG-1(-/-) mice developed colitis. Here, we investigated the role of IL-7 in the maintenance of colitogenic CD4(+) T cells by surgically separating these parabionts. Surprisingly, the separated IL-7(-/-) x RAG-1(-/-) mice were consistently diseased after separation, although no IL-7 mRNA was detected in the tissues of separated IL-7(-/-) x RAG-1(-/-) partners. CD4(+) T cells isolated from the separated RAG-1(-/-) or IL-7(-/-) x RAG-1(-/-) mice were then transferred into new RAG-1(-/-) or IL-7(-/-) x RAG-1(-/-) mice. Regardless of the source of donor cells, RAG-1(-/-) recipients developed colitis, whereas IL-7(-/-) x RAG-1(-/-) recipients did not. Collectively, these results demonstrate that IL-7 is essential for lymphopenia-driven turnover of colitogenic CD4(+) T cells rather than the maintenance of those cells in established colitic mice. They also provide a basis for the timing of IL-7/IL-7R blockade for the treatment of inflammatory bowel diseases.

FTY720 suppresses the development of colitis in lymphoid-null mice by modulating the trafficking of colitogenic CD4+ T cells in bone marrow.

2-amino-2-(2-[4-octylphenyl]ethyl)-1,3-propanediol hydrochloride (FTY720) suppresses T-cell egress from LN, thereby preventing pathogenic T cells from migrating toward disease sites. However, little is known about whether FTY720 could control the trafficking of T cells without the presence of lymphoid tissues. Here we demonstrate that FTY720 treatment suppresses the recirculation of CD4(+) T cells in splenectomized (SPX) lymphotoxin-alpha(-/-) (LT-alpha(-/-)) mice that lack LN and spleen, as shown by peripheral blood (PB) lymphopenia in FTY720-treated SPX LT-alpha(-/-) mice. In a short-term transfer experiment, the cell number of transferred Ly5.1(+)CD4(+) T cells recovered from host FTY720-treated SPX LT-alpha(-/-) mice (Ly5.2(+)) was markedly decreased in PB, but conversely increased in BM. Notably, FTY720 treatment prevented the development of colitis that is otherwise induced in untreated SPX LT-alpha(-/-) x RAG-2(-/-) mice upon transfer of colitic lamina propria CD4(+) T cells. In such mice, the number of CD4(+) T cells in PB or lamina propria of FTY720-treated SPX LT-alpha(-/-) x RAG-2(-/-) recipients was significantly reduced, but that in the BM was significantly increased as compared with untreated control mice. Altogether, the present results indicate that FTY720 treatment may offer an additional role to direct trafficking of CD4(+) T cells in BM, resulting in the prevention of colitis.

Homeostatic (IL-7) and effector (IL-17) cytokines as distinct but complementary target for an optimal therapeutic strategy in inflammatory bowel disease.

PURPOSE OF REVIEW: This review focuses on CD4+ T cells involved in the mediation of inflammatory tissue damage in murine models of inflammatory bowel diseases (IBDs). In particular, we describe the distinct roles of the homeostatic cytokine IL-7, which is essential to the maintenance of colitogenic memory CD4+ cells, and the newly discovered effector cytokine IL-17. We also discuss the close correlation between colitogenic Th17-type CD4+ T cells and inducible CD4+CD25+Foxp3+ regulatory T cells. RECENT FINDINGS: IBDs are characterized by wasting and chronic intestinal inflammation induced by many different cytokine-mediated pathways. It is clearly recognized that medical and surgical interventions do not cure Crohn's disease because relapse is the rule after remission. Until a few years ago, IBD was classified into Th1-dependent, that is, Crohn's disease, and Th2-dependent, that is, ulcerative colitis, phenotypes. However, in recent years, it has been shown that new T-cell subclasses, that is, Th17 and regulatory T cells (T(R)), exist independently of Th1 and Th2 and that they play a central role in modulating IBD. SUMMARY: The persistence of IL-7-dependent colitogenic memory CD4+ T cells is critical to the maintenance of experimental colitis. On the other hand, though Th1 and Th2 colitogenic memory CD4+ cells exist, in recent years the central role of IL-17-producing Th17-type cells in IBD has attracted renewed interest. The development of molecularly targeted therapies aimed at a variety of different Th-dependent pathogenic mechanisms may represent a novel approach to IBD therapy.

Extracorporeal elimination of TNF-alpha-producing CD14(dull)CD16(+) monocytes in leukocytapheresis therapy for ulcerative colitis.

BACKGROUND: In recent years leukocytapheresis using a leukocyte removal filter (known as lymphocytapheresis, LCAP) has been applied to the treatment of various autoimmune diseases including ulcerative colitis (UC). In the present study we aimed to clarify how LCAP therapy modifies inflammatory responses by modulating circulating TNF-alpha-producing monocytes. METHODS: Mononuclear cells were obtained from blood before and after the first treatment, and the expression profiles of various immune cells (naive versus. memory, regulatory CD4(+)CD25(bright) versus non-regulatory CD4(+)CD25(-) T cells, and CD14(+)CD16(-) versus CD14(dull)CD16(+) monocytes) were assessed. To evaluate immunological differences between CD14(+)CD16(-) and CD14(dull)CD16(+) monocytes, the expression of TNF-alpha, IL-6, IL-12, IL-10, IL-18, surface toll-like receptor 2 (TLR2), TLR4, and other activation markers including HLA-DR, CD80 and CD86, as well as cytokine profiles, were analyzed. RESULTS: LCAP treatment selectively removed CD14(dull)CD16(+) monocytes, which preferentially produce TNF-alpha and IL-12 and express HLA-DR, CD80, CD86, and TLR2, compared with the major fraction of CD14(+)CD16(-) monocytes, which conversely produce a higher amount of IL-10. In addition, the CD4(+)CD45RO(+)CD62L(-)/CD4(+)CD45RO(+)CD62L(+) ratio was significantly lower after LCAP therapy. However, the CD4(+)CD25(bright)/total CD4(+) ratio did not change. CONCLUSIONS: The present findings revealed the real target of proinflammatory CD14(dull)CD16(+) monocytes removed during LCAP treatment of UC and that LCAP might be used as an extracorporeal anti-TNF-alpha therapy, expanding the clinical applications of this procedure to include the treatment of Crohn's disease.

TH1/TH2-mediated colitis induced by adoptive transfer of CD4+CD45RBhigh T lymphocytes into nude mice.

BACKGROUND: Transfer of CD4CD45RB T cells from normal donors to SCID/Rag-1, 2-deficient mice, which lack T and B cells, leads to the development of a TH1-mediated inflammatory bowel disease (IBD)-like syndrome characterized by extensive mononuclear cell infiltrates and epithelial cell hyperplasia. Because it is well known that B cells are also involved in a multitude of mechanistic pathways in human IBD, this study attempts to establish a new model of colitis in nude mice. METHODS: We transferred CD4CD45RB T cells into athymic nude mice, which lack thymus-dependent T cells but retain normal B cells, to establish and investigate a B cell-involving chronic colitis model. As a control, CD4CD25 T cells were also used. RESULTS: Mice reconstituted with CD4CD45RB but not CD4CD25 T cells developed a wasting disease, with severe infiltrates of B cell aggregates as well as T cells, macrophages, and dendritic cells into the colon and elevated levels of interferon-gamma, tumor necrosis factor-alpha, interleukin (IL)-4, IL-5, and IL-10, by 7 weeks after T cell transfer. Furthermore, the infiltrated lamina propria B cells in colitic nude mice consisted predominantly of massive aggregated immunoglobulin (Ig) M- and scattered IgG-positive cells, but not IgA-positive cells. In contrast, mice reconstituted with CD4CD45RB and CD4CD45RB did not develop wasting disease or colitis. CONCLUSIONS: Collectively, the power of the colitis model induced by the adoptive transfer of CD4CD45RB T cells into nude mice is that one can investigate the roles of TH2-type cells and B cells in a regulatory T cell-depleted condition.

MyD88-deficient mice develop severe intestinal inflammation in dextran sodium sulfate colitis.

BACKGROUND: Gut commensal microbes affect the development and activation of the mucosal and systemic immune systems. However, the exact molecular mechanism of these microbes that is involved in the development of colitis remains unclear. METHODS: The present study was conducted to determine the distinct role of the innate immune system in the development of a dextran sulfate sodium (DSS) colitis model in MyD88(-/-) mice, because myeloid differentiation protein (MyD88) is a major adaptor molecule essential for signaling via Toll-like receptors (TLRs). To this end, MyD88(-/-) and wild-type (WT) mice received sterile distilled water containing 1.2% DSS for 8 days. The survival rate, total clinical score (body weight loss, stool consistency, and rectal bleeding), colon length, and histological score were assessed. The expression of surface markers (F4/80 and CD4) on infiltrating lamina propria mononuclear cells was analyzed immunohistochemistrically. RESULTS: MyD88(-/-) mice exhibited increased susceptibility to DSS-induced colitis, as reflected by significantly higher lethality and higher clinical and histological scores, and more severe colonic shortening compared to WT mice. Immunohistochemical analysis revealed a significant increase of both F4/80+ macrophages and CD4+ T cells in the inflamed mucosa in DSS-fed MyD88(-/-) mice compared to DSS-fed WT mice. CONCLUSIONS: These findings suggest that, via MyD88 signaling, the innate immune system in the gut plays an important protective role in colitis.

Macrophage-derived IL-18 targeting for the treatment of Crohn's disease.

Crohn's disease is an inflammatory bowel disease associated with several changes in the immune system, including an increased number of infiltrating macrophages. These macrophages release a variety of proinflammatory cytokines, such as tumor necrosis factor-alpha (TNF-alpha, macrophage infiltrating factor (MIF), interleukin (IL)-6, IL-12 and IL-18, which are critically involved in the onset and the development of Crohn's disease. We here focus on the role of macrophages, especially macrophage-derived IL-18 in both patients with Crohn's disease and a murine model of Crohn's disease.

ICOS costimulation in inflammatory bowel disease.

For years, medical researchers have striven to develop selective immunotherapies that could specifically ameliorate pathogenic immune responses without immunocompromising the patient. Blockade of many known receptors on T cells can inhibit the initiation of immune responses. However, this approach is problematic in that it is not possible to predict the onset of disease in patients. Current immunotherapies are unsatisfactory for the sporadic exacerbating type of diseases such as multiple sclerosis and inflammatory bowel disease (IBD), because they require either long-term treatment or acute treatment with high-dose immunosuppressants. With regard to this issue, the inducible and inflammatory site-specific molecule, inducible costimulator (ICOS), may be particularly useful as an ideal targeting molecule for the strategy of treatment of human IBD patients.

Role of the innate immune system in the development of chronic colitis.

Based on Pasteur's work on the microbial nature of fermentation, it was widely believed that the presence of bacteria in the intestine was essential for the life of the host. It has also been known for decades that gut commensal microbes effect the activation and development of the systemic immune system through gut-associated lymphoid tissues (GALT). Recent extensive studies have shown that recognition of microbes is mediated by a set of germline-encoded receptors, Toll-like receptors (TLRs), in mammals. This article reviews the role of the innate immunity system in the development of GALT and the pathogenesis of inflammatory bowel diseases (IBD).

Prevalence of metabolic syndrome is comparable between inflammatory bowel disease patients and the general population.

BACKGROUND: Metabolic syndrome (MS) is associated with an increased risk of cardiovascular disease. However, its prevalence in inflammatory bowel disease (IBD) patients remains largely unknown. This study was planned to determine the prevalence of MS in Japanese IBD patients. METHODS: The prevalence of MS among outpatients with IBD aged 18 or older was studied using the modified National Cholesterol Education Program Adult Treatment Panel III definition. RESULTS: A total of 107 quiescent IBD patients, including 76 ulcerative colitis (UC) patients and 31 Crohn's disease (CD) patients, were studied. Sufficient data were collected from a total of 102 patients. Prevalence of MS was significantly higher in UC (23.0%) patients compared to CD patients (7.1%). MS prevalence was substantially higher among male IBD patients (21.1%) compared to female IBD patients (12.9%), particularly in patients over 30 years of age. No difference was observed in the prevalence of MS between our IBD cohort and the general population in both males and females aged 40 years and older (P = 0.707 in males, P = 0.328 in females). IBD patients with MS were also older than those without (50.2 ± 15.0 vs. 38.0 ± 11.9 years, P = 0.013). In a logistic regression analysis, age was the statistically significant predictor of MS among IBD patients. The odds ratio (95% confidence interval) was 1.064 (1.017-1.114). CONCLUSIONS: Prevalence of metabolic syndrome in our IBD patients was comparable to that of the general population. Because age was the independent risk factor for developing MS, evaluation for MS is needed for elderly IBD patients.

Persistent retention of colitogenic CD4+ memory T cells causes inflammatory bowel diseases to become intractable.

Despite the advent of an age when "malignant" leukemia is cured by bone marrow transplantation, "benign" inflammatory bowel diseases (IBDs) are still intractable lifelong diseases. Why is it that once an IBD develops it lasts a long time? We propose that, the same as in the response to vaccination, immune memory T cells that remember the disease are formed in IBDs and, perceiving them as "benign T-cell leukemia"-like lifelong pathology that hematogenously spreads throughout the body, we here propose that the bone marrow itself, which produces large amounts of the survival factor IL-7, is the reservoir for colitogenic CD4(+) memory T cells responsible for the intractability of IBDs.

Signaling pathway via TNF-alpha/NF-kappaB in intestinal epithelial cells may be directly involved in colitis-associated carcinogenesis.

Treatment with anti-TNF-alpha MAb has been accepted as a successful maintenance therapy for patients with inflammatory bowel diseases (IBD). Moreover, it has been recently reported that blockade of TNF receptor (TNFR) 1 signaling in infiltrating hematopoietic cells may prevent the development of colitis-associated cancer (CAC). However, it remains unclear whether the TNF-alpha signaling in epithelial cells is involved in the development of CAC. To investigate this, we studied the effects of anti-TNF-alpha MAb in an animal model of CAC by administration of azoxymethane (AOM) followed by sequential dextran sodium sulfate (DSS) ingestion. We observed that the NF-kappaB pathway is activated in colonic epithelia from DSS-administered mice in association with upregulation of TNFR2 rather than TNFR1. Immunoblot analysis also revealed that the TNFR2 upregulation accompanied by the NF-kappaB activation is further complicated in CAC tissues induced in AOM/DSS-administered mice compared with the nontumor area. Such NF-kappaB activity in the epithelial cells is significantly suppressed by the treatment of MP6-XT22, an anti-TNF-alpha MAb. Despite inability to reduce the severity of colitis, sequential administration of MP6-XT22 reduced the numbers and size of tumors in association with the NF-kappaB inactivation. Taken together, present studies suggest that the TNFR2 signaling in intestinal epithelial cells may be directly involved in the development of CAC with persistent colitis and imply that the maintenance therapy with anti-TNF-alpha MAb may prevent the development of CAC in patients with long-standing IBD.

Continuous generation of colitogenic CD4(+) T cells in persistent colitis.

Inflammatory bowel diseases take chronic courses due to the expansion of colitogenic CD4(+) cells. However, it is unclear whether the persistent disease is driven by continuous reactivation of colitogenic memory CD4(+) cells to generate effector CD4(+) cells or by continuous generation of effector CD4(+) cells from naïve cells. To clarify this issue, we performed a series of sequential adoptive transfers of Ly5.2(+) and Ly5.1(+) CD4(+)CD45RB(high) cells into RAG-2(-/-) mice at different time points. We show here that the secondarily transferred CD4(+)CD45RB(high) cells can be converted to CD4(+)CD44(high)CD62L(-)IL-7Ralpha(high) effector-memory T cells even in the presence of pre-existing effector-memory CD4(+) cells. Although the total cell numbers of CD4(+) cells in established colitic mice were consistently equivalent irrespective of the number of primarily transferred cells, the ratio of primarily and secondarily transferred cells was dependent on the ratio of the transferred cell numbers, but not on the order of the transfer. Of note, we found that primarily transferred CD4(+) cells produced significantly lower amounts of IFN-gamma and IL-17 than CD4(+) cells arising from secondary transfer. In conclusion, the continuous generation of colitogenic CD4(+) cells that compensate for exhausted CD4(+) cells may be one of the mechanisms involved in the persistence of colitis.

Immunosenescent colitogenic CD4(+) T cells convert to regulatory cells and suppress colitis.

Inflammatory bowel diseases progress steadily by the expansion of colitogenic CD4(+) cells. However, it remains unknown whether colitogenic CD4(+) cells are long-living like memory cells or exhausted like effector cells. To assess the longevity of colitogenic lamina propria (LP) CD4(+) cells, we performed sequential transfers of LP CD4(+) cells from colitic CD4(+)CD45RB(high) cell-transferred SCID mice into new SCID mice. Although SCID mice transferred with colitic LP CD4(+) cells stably developed colitis until at least the sixth transfer, the interval to the development of colitis gradually lengthened as the number of transfers increased. The incidence of colitis gradually decreased after the seventh transfer. Furthermore, non-colitic LP CD4(+) cells from mice transferred over seven times expressed significantly higher levels of PD-1 and produced significantly lower amounts of IFN-gamma, TNF-alpha, and IL-17 than colitic LP CD4(+) cells recovered after the first transfer. Most notably, we found that re-transfer of non-colitic LP CD4(+) cells recovered after multiple transfers prevented the development of colitis in SCID mice co-transferred with CD4(+)CD45RB(high) cells. Thus, colitogenic LP CD4(+) cells may be exhausted over time, become non-functional, convert to regulatory cells, and finally suppress colitis in the process of immunosenescence.

Systemic, but not intestinal, IL-7 is essential for the persistence of chronic colitis.

We previously demonstrated that IL-7 is produced by intestinal goblet cells and is essential for the persistence of colitis. It is well known, however, that goblet cells are decreased or depleted in the chronically inflamed mucosa of animal colitis models or human inflammatory bowel diseases. Thus, in this study, we assess whether intestinal IL-7 is surely required for the persistence of colitis using a RAG-1/2-/- colitis model induced by the adoptive transfer of CD4+CD45RBhigh T cells in combination with parabiosis system. Surprisingly, both IL-7-/-xRAG-1-/- and IL-7+/+xRAG-1-/- host mice developed colitis 4 wk after parabiosis to a similar extent of colitic IL-7+/+xRAG-1-/- donor mice that were previously transferred with CD4+CD45RBhigh T cells. Of note, although the number of CD4+ T cells recovered from the spleen or the bone marrow of IL-7-/-xRAG-1-/- host mice was significantly decreased compared with that of IL-7+/+xRAG-1-/- host mice, an equivalent number of CD4+ T cells was recovered from the lamina propria of both mice, indicating that the expansion of CD4+ T cells in the spleen or in the bone marrow is dependent on IL-7, but not in the lamina propria. Development of colitis was never observed in parabionts between IL-7+/+xRAG-1-/- host and noncolitic IL-7-/-xRAG-1-/- donor mice that were transferred with CD4+CD45RBhigh T cells. Collectively, systemic, but not intestinal, IL-7 is essential for the persistence of colitis, suggesting that therapeutic approaches targeting the systemic IL-7/IL-7R signaling pathway may be feasible in the treatment of inflammatory bowel diseases.