IL-12, independently of IFN-gamma, plays a crucial role in the pathogenesis of a murine psoriasis-like skin disorder.

The onset of acute psoriasis and the exacerbation of chronic psoriasis are often associated with a history of bacterial infection. We demonstrate that while only few scid/scid mice develop disease when CD4+CD45Rbhigh T cells are transferred alone, coadministration of LPS plus IL-12 or staphylococcal enterotoxin B into scid/scid mice 1 day after CD4+CD45Rbhigh T cell transfer greatly enhances disease penetrance and severity. Most importantly, the skin lesions induced by this method exhibit many of the histologic hallmarks observed in human psoriasis. Skin infiltrating CD4+ T cells were predominantly memory/effector cells (CD45Rblow) and exhibited a highly polarized Th1 phenotype. To test whether the development of pathogenic T cells was dependent on their production of IFN-gamma, we transferred IFN-gamma-/- CD4+CD45Rbhigh T cells into scid/scid or into T, B and NK cell-deficient scid/beige mice. Surprisingly, the incidence of psoriasis was similar to scid/scid animals that received IFN-gamma+/+ T cells, although acanthosis of the skin was attenuated. In contrast, the development of psoriasis was abolished if anti-IL-12 mAb was administered on day 7 and 35 after T cell transfer. Skin-derived IFN-gamma-/- inflammatory cells, but not cells from anti-IL-12-treated animals, secreted substantial amounts of TNF-alpha, suggesting that the inflammatory effect of IFN-gamma-/- T cells may be partly exerted by TNF-alpha and that the therapeutic effect of anti-IL-12 may depend on its ability to down-regulate both TNF-alpha and IFN-gamma. Overall, these results suggest that IL-12, independently of IFN-gamma, is able to induce pathogenic, inflammatory T cells that are able to induce psoriasiform lesions in mice.     

Blockade of late stages of autoimmune diabetes by inhibition of the receptor for advanced glycation end products

Ligation of the receptor for advanced glycation end products (RAGE) occurs during inflammation. Engagement of RAGE results in enhanced expression of addressins and it is therefore, not surprising that previous studies have shown a role of RAGE/ligand interactions in immune responses including cell/cell contact but the role of RAGE in spontaneous autoimmunity has not been clearly defined. To study the role of RAGE/ligand interactions in autoimmune diabetes, we tested the ability of soluble RAGE, a scavenger of RAGE ligands, in late stages of diabetes development in the NOD mouse-disease transferred with diabetogenic T cells and recurrent disease in NOD/scid recipients of syngeneic islet transplants. RAGE expression was detected on CD4(+), CD8(+), and B cells from diabetic mice and transferred to NOD/scid recipients. RAGE and its ligand, S100B, were found in the islets of NOD/scid mice that developed diabetes. Treatment of recipient NOD/scid mice with soluble RAGE prevented transfer of diabetes and delayed recurrent disease in syngeneic islet transplants. RAGE blockade was associated with increased expression of IL-10 and TGF-beta in the islets from protected mice. RAGE blockade reduced the transfer of disease with enriched T cells, but had no effect when diabetes was transferred with the activated CD4(+) T cell clone, BDC2.5. We conclude that RAGE/ligand interactions are involved in the differentiation of T cells to a mature pathogenic phenotype during the late stages of the development of diabetes.     

Critical role for IL-4 in the development of transplant arteriosclerosis in the absence of CD40-CD154 costimulation.

Blockade of the CD40-CD154 pathway can inhibit CD4(+) T cell activation but is unable to prevent immune responses mediated by CD8(+) T cells. However, even in the absence of CD8(+) T cells, inhibition of the CD40-CD154 pathway is insufficient to prevent the development of transplant arteriosclerosis. This study investigated the mechanisms of transplant arteriosclerosis in the absence of the CD40 pathway. C57BL/6 CD40(-/-) (H2(b)) recipients were transplanted with MHC-mismatched BALB/c (H2(d)) aortas. Transplant arteriosclerosis was evident in both CD40(-/-) and CD40(+/-) mice (intimal proliferation was 59 +/- 5% for CD40(-/-) mice vs 58 +/- 4% for CD40(+/-) mice) in the presence or absence of CD8(+) T cells (intimal proliferation was 46 +/- 7% for CD40(-/-) anti-CD8-treated mice vs 50 +/- 10% for CD40(+/-) anti-CD8-treated mice), confirming that CD8(+) T cells are not essential effector cells for the development of this disease. In CD40(-/-) recipients depleted of CD8(+) T cells, the number of eosinophils infiltrating the graft was markedly increased (109 +/- 24 eosinophils/grid for CD40(-/-) anti-CD8-treated mice vs 28 +/- 7 for CD40(+/-) anti-CD8-treated mice). The increased presence of eosinophils correlated with augmented intragraft production of IL-4. To test the hypothesis that IL-4 was responsible for the intimal proliferation, CD8 T cell-depleted CD40(-/-) recipients were treated with anti-IL-4 mAb. This resulted in significantly reduced eosinophil infiltration into the graft (12 +/- 5 eosinophils/grid for CD40(-/-) anti-CD8(+), anti-IL-4-treated mice vs 109 +/- 24 for CD40(-/-) anti-CD8-treated mice), intragraft eotaxin, CCR3 mRNA production, and the level of intimal proliferation (18 +/- 5% for CD40(-/-) anti-CD8(+)-, anti-IL-4-treated mice vs 46 +/- 7% for CD40(-/-) anti-CD8-treated mice). In conclusion, elevated intragraft IL-4 production results in an eosinophil infiltrate and is an important mechanism for CD8(+) T cell-independent transplant arteriosclerosis in the absence of CD40-CD154 costimulation.     

CD40 ligand in pathogenesis of autoimmune ovarian disease of day 3-thymectomized mice: implication for CD40 ligand antibody therapy

The blockade of CD40 ligand (CD40L) is effective in autoimmune disease prevention. Recently, a brief period of CD40L mAb treatment was reported to induce tolerance and enhancement of CD4(+)CD25(+) regulatory T cell activity. We therefore determined the efficacy of CD40L mAb treatment in autoimmunity that resulted from CD4(+)CD25(+) regulatory T cell deficiency. Autoimmune ovarian disease (AOD) and oocyte autoantibody response of day 3-thymectomized (d3tx) mice were inhibited by continuous CD40L mAb treatment from day 3, or from days 10-14, whereas CD40L mAb treatment confined to the neonatal week was ineffective. The enhanced expression of memory markers (CD44 and CD62L(low)) on CD4(+) T cells of the d3tx mice was unaffected by CD40L mAb treatment. In contrast, their increased T cell activation markers (CD69 and CD25) were eliminated by CD40L mAb treatment. Moreover, ex vivo activated T cells of d3tx mice expressed elevated intracellular IFN-gamma, and this was also blocked by CD40L mAb. The memory T cells, although nonpathogenic in CD40L mAb-positive environment, transferred severe AOD to CD40L mAb(-) neonatal recipients. Most importantly, CD40L mAb treatment inhibited AOD in recipients of T cells from d3tx donors with severe AOD and led to regression of AOD in d3tx mice documented at 4 wk. Therefore, 1) the continuous presence of CD40L mAb both prevents and causes regression of AOD in the d3tx mice; and 2) the multiple steps of the d3tx autoimmune disease, including T cell activation, cytokine production, T cell-mediated inflammation, and tissue injury, are CD40L dependent.     

Anti-CD4 monoclonal antibody therapy suppresses autoimmune disease in MRL/Mp-lpr/lpr mice.

MRL/Mp-lpr/lpr (MRL/lpr) mice spontaneously develop systemic autoimmune disease, characterized by vasculitis, lymphadenopathy, glomerulonephritis, and autoantibody formation. The target organ inflammatory lesions are composed largely of CD4+ "helper" T cells, while the massively enlarged lymph nodes are composed primarily of CD3+ CD4- CD8- TCR alpha/beta + "double-negative" T cells. In this study we investigated the effect of treatment of MRL/lpr mice with anti-CD4 monoclonal antibody (mAb); control groups consisted of animals treated with normal saline or rat immunoglobulin (Ig). Anti-CD4 mAb treatment, which was started at 4 weeks and continued through 20 weeks of age, resulted in a dramatic reduction of both the frequency and severity of the autoimmune disease, as demonstrated histologically and serologically. Anti-CD4 mAb therapy markedly reduced the frequency of glomerulonephritis and eliminated vasculitis of the major renal arterial branches. Glomerulonephritis was detected in 9 of 9 saline-treated, 9 of 9 rat Ig-treated, but in only 1 of 9 anti-CD4 mAb-treated mice; vasculitis was detected in 6 of 9 saline-treated, 7 of 9 rat Ig-treated, but in none of 9 anti-CD4 mAb-treated mice. The frequency of antinuclear antibodies, titer of anti-dsDNA antibodies, and total Ig levels were all significantly reduced by anti-CD4 mAb therapy. These data support the hypothesis that CD4+ T cells play a central role in the disease process in this autoimmune strain.     

In vivo accumulation of T cells in response to IL-2/anti-IL-2 mAb complexes is dependent in part on the TNF family ligand 4-1BBL

Immune complexes combining IL-2 with particular anti-IL-2 antibodies can be used to selectively expand regulatory T cells or memory T cells. Combining IL-2 with anti-IL-2 (Clone S4B6) greatly enhances the biological potency of IL-2 in vivo leading to selective expansion of CD8 memory T cells and NK cells compared with regulatory T cells. Here we show that in vivo administration of IL-2/anti-IL-2 mAb (IL-2/mAb) complexes induces 4-1BB expression on both adoptively transferred antigen-specific memory CD8 T cells as well as on endogenous memory phenotype cells. Remarkably, the accumulation of adoptively transferred memory CD8 T cells following in vivo IL-2/mAb-complex treatment was found to be dependent in part on the presence of 4-1BBL in the host. These effects were independent of IL-2-induced cell division, suggesting that 4-1BBL-induced survival signals contribute to IL-2/mAb-complex-induced T-cell accumulation in vivo.     

Amelioration of experimental autoimmune encephalomyelitis with anti-OX40 ligand monoclonal antibody: a critical role for OX40 ligand in migration, but not development, of pathogenic T cells

OX40 (CD134) and its ligand (OX40L) have been implicated in T cell activation and migration. In this study, we examined the contribution of these molecules to the pathogenesis of experimental autoimmune encephalomyelitis (EAE) by administering a neutralizing mAb against murine OX40L (RM134L) to proteolipid protein (139-151) peptide-induced EAE in SJL mice. Administration of RM134L effectively ameliorated the disease in both actively induced and adoptively transferred EAE models. Histological examination showed that the RM134L treatment greatly reduced mononuclear cell infiltration into the spinal cord. The RM134L treatment did not inhibit the development of pathogenic T cells, given that proliferative response and IFN-gamma production by draining lymph node cells were not reduced or rather enhanced upon restimulation with proteolipid protein (139-151) in vitro, and these cells effectively transferred EAE to naive SJL mice. Flow cytometric analyses showed that the RM134L treatment inhibited the accumulation of OX40-expressing CD4(+) T cells and the migration of adoptively transferred CD4(+) T cells in the spinal cord. Immunohistochemical staining showed that OX40L was most prominently expressed on endothelial cells in the inflamed spinal cord. These results suggest that the OX40/OX40L interaction plays a critical role for the migration of pathogenic T cells into the CNS in the pathogenesis of EAE.     

In vivo effects of anti-IL-4 monoclonal antibody on neonatal induction of tolerance and on an associated autoimmune syndrome

The role of IL-4 in the cellular interactions leading to the induction of CTL tolerance to H-2b alloantigens and to the development of a lupus-like autoimmune disease in BALB/c mice after neonatal injection with (C57BL/6 x BALB/c)F1 cells was investigated in vivo by using an anti-IL-4 mAb. Treatment of F1 cell-injected BALB/c mice with 15 mg of anti-IL-4 mAb was shown to interfere with tolerance induction, as assessed by the high percentages of H-2b target cell lysis and the very low or undetectable levels of B cell chimerism markers observed in these mice. Treatment with 4.5 mg of anti-IL-4 mAb interfered with tolerance induction only in one-third of F1 cell-injected BALB/c mice, but that dose induces specific modulations of the autoimmune manifestations in all mice, leading to the nearly complete prevention of the disease. In particular, the production of anti-ssDNA IgG1 and of total IgG1 and IgE antibodies was seriously affected by the treatment, as well as the proliferation and membrane Ia and K expression of F1 donor splenic cells and thymic APC. Treatment of F1 cell-injected BALB/c mice between 24 and 48 h of life with 0.5 mg of anti-IL-4 mAb did not interfere with tolerance induction, but had similar effects on the autoimmune syndrome as treatment with 4.5 mg. These results suggest that, after F1 cell injection of newborn mice, IL-4 plays an important role in the cellular interactions leading to the induction of tolerance to the corresponding alloantigens and to the development of the associated autoimmune syndrome.     

Development of memory-like autoregulatory CD8+ T cells is CD4+ T cell dependent

Progression of spontaneous autoimmune diabetes is associated with development of a disease-countering negative-feedback regulatory loop that involves differentiation of low-avidity autoreactive CD8(+) cells into memory-like autoregulatory T cells. Such T cells blunt diabetes progression by suppressing the presentation of both cognate and noncognate Ags to pathogenic high-avidity autoreactive CD8(+) T cells in the pancreas-draining lymph nodes. In this study, we show that development of autoregulatory CD8(+) T cell memory is CD4(+) T cell dependent. Transgenic (TG) NOD mice expressing a low-affinity autoreactive TCR were completely resistant to autoimmune diabetes, even after systemic treatment of the mice with agonistic anti-CD40 or anti-4-1BB mAbs or autoantigen-pulsed dendritic cells, strategies that dramatically accelerate diabetes development in TG NOD mice expressing a higher affinity TCR for the same autoantigenic specificity. Furthermore, whereas abrogation of RAG-2 expression, hence endogenous CD4(+) T cell and B cell development, decelerated disease progression in high-affinity TCR-TG NOD mice, it converted the low-affinity TCR into a pathogenic one. In agreement with these data, polyclonal CD4(+) T cells from prediabetic NOD mice promoted disease in high-affinity TCR-TG NOD.Rag2(-/-) mice, but inhibited it in low-affinity TCR-TG NOD.Rag2(-/-) mice. Thus, in chronic autoimmune responses, CD4(+) Th cells contribute to both promoting and suppressing pathogenic autoimmunity.     

Sequential appearance of host-derived T cell subsets during differentiation in nude mice grafted with rat fetal thymus

To elucidate the abnormality of T cell differentiation in nude mice grafted with rat fetal thymus that develop multiple-organ-localized autoimmune diseases, we examined sequential appearance of T cell subsets and expression of TCR genes in BALB/c nude mice after grafting with fetal F344 rat thymus. We observed progressive expression of TCR gamma/delta-alpha/beta genes in the lymph node (LN) cells from 8 to 12 wk after grafting. An appreciable number of CD4+ T cells but few CD8+ T cells were detected in the LN at 8 wk after grafting. CD8+ T cells increased slowly in number by 12 wk after grafting but remained at a low level in comparison with those in nude mice 12 wk after grafting with BALB/c thymus. In correlation with an increase in the number of T cells expressing TCR alpha/beta genes, alloreactivity as assessed by MLR was increased to a normal level. However, CTL activity against alloantigens remained at a low level in the LN cells at 12 wk. At this stage, organ-specific autoimmune diseases and a high level of anti-DNA autoantibodies were detected. In these mice host-reactive T cells such as V beta 3- or V beta 11-bearing T cells were virtually eliminated in the peripheral mature T cell pool, whereas T cells maturing in the fetal rat thymus significantly proliferated in response to donor-rat stimulator cells. These results suggest that the development of the autoimmune diseases may be ascribed to an impaired maturation of CD8+ T cells but not to failure in clonal elimination of host-reactive T cells in nude mice grafted with rat thymus.     

Development of murine lupus in CD4-depleted NZB/NZW mice. Sustained inhibition of residual CD4+ T cells is required to suppress autoimmunity.

Chronic administration of anti-CD4 mAb prevents autoimmune disease in NZB/NZW F1 (B/W) mice. This may be due either to CD4 cell depletion or to inhibition of CD4 cell function. To evaluate the relative importance of these mechanisms, we devised a system in which the consequences of cell depletion could be analyzed independent of the inhibitory effects of chronic mAb therapy. This was accomplished by performing adult thymectomy before mAb administration. Specifically, female B/W mice underwent thymectomy or sham thymectomy at age 6 wk, followed at age 3 mo by a short course of either anti-CD4 (2 mg/wk for 3 wk) or saline. Treatment with anti-CD4 depleted 90% of circulating CD4 cells, but a small subpopulation (10%) of CD4 cells was refractory to depletion. In non-thymectomized mice, the CD4 population gradually reconstituted after cessation of therapy. In contrast, in thymectomized mice, recovery of CD4 cells was prevented by the absence of the thymus. Despite the striking reduction in CD4 cells in thymectomized mice, severe autoimmune disease developed, with autoantibody levels, proteinuria, and mortality comparable with non-thymectomized, nondepleted controls. The unexpected development of lupus nephritis in thymectomized, CD4-depleted B/W mice suggested that the thymus might be required to achieve the benefits of therapy with anti-CD4. To exclude this possibility, we demonstrated that chronic therapy with anti-CD4 prevents autoimmunity in thymectomized B/W mice. These findings imply that: 1) substantial depletion of CD4 T cells is not sufficient to suppress autoimmunity; 2) suppression of autoimmunity requires sustained functional inhibition of CD4 T cells; and 3) a small subpopulation of CD4 cells that is refractory to depletion by anti-CD4 is sufficient to promote the full expression of murine lupus in B/W mice.     

Anti-IL-2 treatment impairs the expansion of T(reg) cell population during acute malaria and enhances the Th1 cell response at the chronic disease

Plasmodium chabaudi infection induces a rapid and intense splenic CD4(+) T cell response that contributes to both disease pathogenesis and the control of acute parasitemia. The subsequent development of clinical immunity to disease occurs concomitantly with the persistence of low levels of chronic parasitemia. The suppressive activity of regulatory T (T(reg)) cells has been implicated in both development of clinical immunity and parasite persistence. To evaluate whether IL-2 is required to induce and to sustain the suppressive activity of T(reg) cells in malaria, we examined in detail the effects of anti-IL-2 treatment with JES6-1 monoclonal antibody (mAb) on the splenic CD4(+) T cell response during acute and chronic P. chabaudi AS infection in C57BL/6 mice. JES6-1 treatment on days 0, 2 and 4 of infection partially inhibits the expansion of the CD4(+)CD25(+)Foxp3(+) cell population during acute malaria. Despite the concomitant secretion of IL-2 and expression of high affinity IL-2 receptor by large CD4(+) T cells, JES6-1 treatment does not impair effector CD4(+) T cell activation and IFN-γ production. However, at the chronic phase of the disease, an enhancement of cellular and humoral responses occurs in JES6-1-treated mice, with increased production of TNF-α and parasite-specific IgG2a antibodies. Furthermore, JES6-1 mAb completely blocked the in vitro proliferation of CD4(+) T cells from non-treated chronic mice, while it further increased the response of CD4(+) T cells from JES6-1-treated chronic mice. We conclude that JES6-1 treatment impairs the expansion of T(reg) cell population during early P. chabaudi malaria and enhances the Th1 cell response in the late phase of the disease.     

In vitro analysis of interferon gamma (IFN-gamma) and interleukin-12 (IL-12) production and their effects in ileal Crohn's disease

Crohn's disease is an inflammatory disease of the gut in which tumour necrosis factor (TNF) and T helper 1 (Th1) cytokines (interleukin (IL)-12, interferon (IFN)-gamma) are thought to play a major role. After the successes obtained with neutralisation of TNF, interest is now growing for therapy aiming at neutralisation of Th1-associated cytokines. Since cytokines are linked in a delicate network, in vitro cultures of ileal lamina propria mononuclear cells (LPMC) were set up for evaluation of a) IFN-gamma and IL-12 production, b) effects of rhIFN-gamma and rhIL-12 and c) effects of anti-IFN-gamma and anti-IL-12 on pro-inflammatory cytokines and IL-10 production. LPMC were isolated from surgical specimens of a total of 27 Crohn's disease and 17 caecum carcinoma (control) patients. Cells were stimulated with CD40L (which triggers myeloid CD40-expressing cells) or anti-CD3 +CD80 (which triggers T cells). LPMC from involved ileal, Crohn's disease produced, in both non-stimulated and stimulated conditions, more IFN-gamma and IL-12p70 than LPMC from non-involved tissue or from control patients. rhIFN-gamma significantly enhanced TNF production in both controls and in ileal Crohn's disease patients, while rhIL-12 enhanced IFN-gamma but not TNF production. LPMC from involved tissue were more sensitive to IL-12 than control LPMC. LP-T cell-dependent activation of monocytes was then studied by co-culture of anti-CD3/CD80-stimulated LPMC with fresh monocytes, which resulted in high IL-12, IFN-gamma, TNF and IL-10 production. The data show that neutralisation of either IL-12 or IFN-gamma with mAb in these cultures also affects secretion of the reciprocal cytokine and (in the case of anti-IL-12) also that of the anti-inflammatory cytokine IL-10. However, no effect of anti-IL-12 or anti-IFN-gamma on production of TNF, a cytokine with an important pathogenic role in Crohn's disease, could be found. Therapies aiming at neutralisation of IFN-gamma or IL-12 are therefore unlikely to replace anti-TNF, but they might provide an additive or synergistic effect.     

Suppression of murine allergic airway disease by IL-2:anti-IL-2 monoclonal antibody-induced regulatory T cells

Regulatory T cells (Treg) play a decisive role in many diseases including asthma and allergen-induced lung inflammation. However, little progress has been made developing new therapeutic strategies for pulmonary disorders. In the current study we demonstrate that cytokine:antibody complexes of IL-2 and anti-IL-2 mAb reduce the severity of allergen-induced inflammation in the lung by expanding Tregs in vivo. Unlike rIL-2 or anti-IL-2 mAb treatment alone, IL-2:anti-IL-2 complexes dampened airway inflammation and eosinophilia while suppressing IL-5 and eotaxin-1 production. Mucus production, airway hyperresponsiveness to methacholine, and parenchymal tissue inflammation were also dramatically reduced following IL-2:anti-IL-2 treatment. The suppression in allergic airway disease was associated with a marked expansion of Tregs (IL-10(+)CD4(+)CD25(+) and Foxp3(+)CD4(+)CD25(+)) in the tissues, with a corresponding decrease in effector T cell responses. The ability of IL-2:anti-IL-2 complexes to suppress airway inflammation was dependent on Treg-derived IL-10, as IL-10(+/+), but not IL-10(-/-) Tregs, were capable of mediating the suppression. Furthermore, a therapeutic protocol using a model of established airway allergy highlighted the ability of IL-2:anti-IL-2 complexes to expand Tregs and prevent successive airway inflammation and airway hyperresponsiveness. This study suggests that endogenous Treg therapy may be a useful tool to combat the rising incidence of allergic airway disease.     

Virus and autoimmunity: induction of autoimmune disease in mice by mouse T lymphotropic virus (MTLV) destroying CD4+ T cells

Neonatal infection of the mouse T lymphotropic virus (MTLV), a member of herpes viridae, causes various organ-specific autoimmune diseases, such as autoimmune gastritis, in selected strains of normal mice. The infection selectively depletes CD4+ T cells in the thymus and periphery for 2-3 wk from 1 wk after infection. Thymectomy 3 wk after neonatal MTLV infection enhances the autoimmune responses and produces autoimmune diseases at higher incidences and in a wider spectrum of organs than MTLV infection alone. On the other hand, inoculation of peripheral CD4+ cells from syngeneic noninfected adult mice prevents the autoimmune development. These autoimmune diseases can be adoptively transferred to syngeneic athymic nude mice by CD4+ T cells. The virus is not detected by bioassay in the organs/tissues damaged by the autoimmune responses. Furthermore, similar autoimmune diseases can be induced in normal mice by manipulating the neonatal thymus/T cells (e.g., by neonatal thymectomy) without virus infection. These results taken together indicate that neonatal MTLV infection elicits autoimmune disease by primarily affecting thymocytes/T cells, not self Ags. It may provoke or enhance thymic production of CD4+ pathogenic self-reactive T cells by altering the thymic clonal deletion mechanism, or reduce the production of CD4+ regulatory T cells controlling self-reactive T cells, or both. The possibility is discussed that other T cell-tropic viruses may cause autoimmunity in humans and animals by affecting the T cell immune system, not the self Ags to be targeted by the autoimmunity.     

T cell antigen receptors in autoimmunity

Three mAb to variable region determinants of the alpha/beta-chain TCR were used to detect discrete populations of peripheral blood T cells. T cells sharing a TCR determinant defined by such an antibody presumably use the same or similar TCR V or J genes for their alpha- or beta-chains. Thus analysis with these mAb provides a tool to investigate TCR gene usage and expression. Since autoantigen specific T cells may play an important role in initiating autoimmune diseases, TCR were analyzed in different autoimmune diseases and control groups including rheumatoid arthritis, Graves disease, idiopathic thrombocytopenic purpura, psoriasis, SLE, insulin-dependent diabetes mellitus, and in nonautoimmune control diseases and normals. Purified T cells were stained by indirect immunofluorescence with three mAb to TCR variable regions: mAb S511 stains 1.8 +/- 0.9% (mean +/- 2 SD), mAb C37 stains 3.4 +/- 1.5% and mAb OT145 stains from 0 to 6% of T cells from normal donors. Several individuals were identified with expanded subsets of positive T cells. One patient with adult ITP followed during a 12-mo period consistently had elevated percentages of T cells staining with the mAb OT145 (15.9 to 24.5%). These cells were found to be exclusively CD8+. By Southern blotting DNA prepared from these OT145+, CD8+ cells, but not DNA from the patient's OT145- T cells, revealed a clonal rearrangement using a beta-chain C region probe. Thus this patient had a monoclonal expansion of CD8+, OT145+ cells. Hyperexpression of a TCR variable region, as defined by the available mAb, could not be associated with any of the diseases studied. Examination of T cells at the site of autoimmunity, such as T cells from rheumatoid arthritis synovial fluid, revealed normal percentages of cells staining with these mAb. Immunoperoxidase staining of psoriatic lesional skin showed no striking enrichment of T cells bearing one or the other TCR type.     

Activation of regulatory cells suppresses experimental allergic encephalomyelitis via secretion of IL-10

Suppression of CD4+ Th1 cell-mediated autoimmune disease via immune deviation is an attractive potential therapeutic approach. CD4+ Th2 T cells specific for myelin basic protein, induced by immunization of young adult male SJL mice, suppress or modify the progression of CNS autoimmune disease. This report demonstrates that activation of non-neuroantigen-specific Th2 cells is sufficient to suppress both clinical and histological experimental allergic encephalomyelitis (EAE). Th2 cells were obtained following immunization of male SJL mice with keyhole limpet hemocyanin. Transfer of these cells did not modify EAE, a model of human multiple sclerosis, in the absence of cognate Ag. Disease suppression was obtained following adoptive transfer and subcutaneous immunization. Suppression was not due to the deletion of myelin basic protein-specific T cells, but resulted from the presence of IL-10 as demonstrated by the inhibition of Th2-mediated EAE suppression via passive transfer with either anti-IL-10 or anti-IL-10R mAb. These data demonstrate that peripheral activation of a CD4+ Th2 population specific for an Ag not expressed in the CNS modifies CNS autoimmune disease via IL-10. These data suggest that either peripheral activation or direct administration of IL-10 may be of benefit in treating Th1-mediated autoimmune diseases.     

Early rise in inflammation and microcirculatory disorder determine the development of autoimmune pancreatitis in the MRL/Mp-mouse

Autoimmune pancreatitis (AIP) is a rare cause of chronic pancreatitis and mimics pancreatic cancer. Although there is strong interest in research, etiology and pathophysiology of AIP are still unknown. Therefore, we analyzed a total of 92 MRL/Mp-mice of either sex, which are prone to develop AIP, in four different age groups (8-12, 16-20, 24-28, and 32-40 wk). Using intravital fluorescence microscopy, histology, laboratory analysis, and Western blot, onset, severity, and pathophysiological mechanisms of AIP were evaluated. Female animals showed in vivo an age-dependent increase of intrapancreatic leukocyte accumulation, as well as a loss in functional capillary perfusion. In contrast, intrapancreatic inflammation in male mice was less pronounced and not age dependent. Furthermore, pancreatic tissue specimen of female animals exhibited major organ destruction with significantly higher values of mean pathological scores (1.5 +/- 0.3 vs. < or =0.2; P < 0.05), as well as significantly increased CD4-, CD8-, CD11b-, and CD138-positive cells compared with male animals of the same age. Interestingly, there was a significant positive correlation between intravascular leukocyte adherence and the histopathological score of the pancreas, indicating a determining role of the innate immune system for the late onset of AIP. The present study shows that the onset of AIP is characterized by an inflammatory response and microcirculatory failure, most probably constituting initiators and propagators of this autoimmune disease.     

Cryptosporidium infection in an adult mouse model. Independent roles for IFN-gamma and CD4+ T lymphocytes in protective immunity.

Cryptosporidium is a protozoan parasite that can cause chronic life-threatening diarrhea in immunocompromised persons. Host immune responses are poorly understood, an impediment to development of effective therapy. In mice, normal adult BALB/c animals resist infection whereas chronic symptomatic cryptosporidiosis develops in adult nude mice and in neonatally infected BALB/c mice treated with anti-CD4 mAb. To define further the immune defects that allow mice to be infected with Cryptosporidium, adult BALB/c mice were treated with cytolytic anti-CD4 or anti-CD8 or with neutralizing anti-IFN-gamma or anti-IL-2 mAb. Chronic infection, manifested by continuous shedding of sparse but statistically significant numbers of oocysts, occurred with anti-CD4 +/- anti-CD8 mAb treatment although anti-CD8 mAb treatment alone did not allow infection. Treatment with anti-IFN-gamma mAb greatly enhanced oocyst shedding but infection was self-limited. Treatment with a combination of anti-CD4 and anti-IFN-gamma mAb permitted both chronic infection and shedding of large numbers of oocysts. Furthermore mice treated initially with anti-CD4 mAb showed a substantial increase in oocyst shedding when later treated with anti-IFN-gamma mAb; and mice treated initially with both mAbs showed a decline in oocyst shedding when anti-IFN-gamma mAb was stopped. Anti-IFN-gamma mAb treatment of congenitally athymic adult BALB/c mice led to an approximately a 75-fold increase in oocyst shedding. Treatment of adult BALB/c mice with anti-IL-2 mAb did not permit Cryptosporidium infection. These results suggest that redundant immunologic mechanisms limit Cryptosporidium infection such that both CD4+ cells and IFN-gamma are required to prevent initiation of infection whereas either alone can limit the extent (IFN-gamma) or duration (CD4+ T cells) of infection. They also suggest that production of IFN-gamma by a non-T cell contributes to host immunity.     

Tissue specificity of E- and P-selectin ligands in Th1-mediated chronic inflammation.

The demonstrated role of E- and P-selectin ligands in the recruitment of Th1 cells raises the question of tissue specificity determination by pathogenic T cells. We took advantage of the fact that chronic Th1-mediated inflammation in the scid/scid CD4+CD45RBhigh T cell transfer model can occur at multiple tissue sites, resembling inflammatory bowel disease in the colon and psoriasis in the skin. We show that the majority of infiltrating effector T cells from psoriatic skin expresses high levels of functional P-selectin ligand (87 +/- 3%), detected by P-selectin-Ig (PIg), while a significantly smaller subset of T cells from colitic lesions expresses this ligand (24 +/- 2%). Similarly, E-selectin ligand is preferentially expressed on CD4+ T cells infiltrating the skin (24 +/- 2%), but only on very few CD4+ T cells infiltrating the colon (CIT; 1.3 +/- 0.8%). In contrast, CD4+ T cells infiltrating the skin express alpha4beta7 at a significantly lower level than CIT (mean fluorescence intensity, 28 vs 61, respectively), although, interestingly, alphaEbeta7 was expressed at high levels on both populations. Analysis of the disease-inducing potential of PIg+ and PIg- CD4+ CIT cells revealed that both populations not only express similar levels of the gut-homing molecule alpha4beta7 (mean fluorescence intensity, 50 vs 56, respectively), but do not differ in their capacity to express IFN-gamma. Furthermore, CIT depleted of cells expressing functional P-selectin ligand were able to induce colitis upon transfer, suggesting that induction of colitis in this model may be independent of E- and P-selectin. These results indicate that adhesion molecule expression and the homing pattern of inflammatory T cells are regulated by the local environment independently of their inflammatory capacity.