Extrathymic Aire-expressing cells are a distinct bone marrow-derived population that induce functional inactivation of CD4⁺ T cells.

The autoimmune regulator (Aire) is essential for prevention of autoimmunity; its role is best understood in the thymus, where it promotes self-tolerance through tissue-specific antigen (TSA) expression. Recently, extrathymic Aire-expressing cells (eTACs) have been described in murine secondary lymphoid organs, but the identity of such cells and their role in immune tolerance remains unclear. Here we have shown that eTACs are a discrete major histocompatibility complex class II (MHC II)(hi), CD80(lo), CD86(lo), epithelial cell adhesion molecule (EpCAM)(hi), CD45(lo) bone marrow-derived peripheral antigen-presenting cell (APC) population. We also have demonstrated that eTACs can functionally inactivate CD4⁺ T cells through a mechanism that does not require regulatory T cells (Treg) and is resistant to innate inflammatory stimuli. Together, these findings further define eTACs as a distinct tolerogenic cell population in secondary lymphoid organs.

Abbreviated exposure to cuprizone is sufficient to induce demyelination and oligodendrocyte loss.

Cuprizone intoxication is one of several animal models used to study demyelination and remyelination. Early treatment protocols exposed mice to cuprizone for 6 weeks to induce demyelination; however, more recent reports have varied exposure times from 4 to 5 weeks. The goal of this study was to determine the minimal exposure of cuprizone in C57BL/6 mice that would induce a pathology of robust demyelination and gliosis similar to that described for a 5- or 6-week treatment. We found that an abbreviated insult of only 2 weeks of exposure to cuprizone induced significant demyelination 3 weeks later (5-week time point) but was somewhat variable. Three weeks of exposure to cuprizone produced extensive demyelination by week 5, equivalent to that observed with 5 weeks of exposure. The depletion of mature oligodendrocytes, as well as microglia and astrocyte accumulation, showed trends similar to those with 5-week exposure to cuprizone. Once mature oligodendrocytes are perturbed after a 3-week treatment, the progression to demyelination occurs without requiring further exposure. Furthermore, the early removal of cuprizone did not accelerate remyelination, suggesting that other sequences of events must follow before repair can occur. Thus, a short, "hit and run" CNS insult triggers a cascade of events leading to demyelination 2-3 weeks later.

Epitope spreading initiates in the CNS in two mouse models of multiple sclerosis.

Chronic progression of two T cell-mediated central nervous system (CNS) demyelinating models of multiple sclerosis, relapsing EAE (R-EAE) and Theiler's murine encephalomyelitis virus-induced demyelinating disease (TMEV-IDD) is dependent on the activation of T cells to endogenous myelin epitopes (epitope spreading). Using transfer of carboxyfluorescein succinyl ester (CFSE)-labeled T-cell receptor (TCR)-transgenic T cells and mixed bone marrow chimeras, we show that activation of naive proteolipid protein (PLP)139-151-specific T cells in SJL mice undergoing PLP178-191-induced R-EAE or TMEV-IDD occurs directly in the CNS and not in the cervical lymph nodes or other peripheral lymphoid organs. Examination of the antigen-presentation capacity of antigen-presenting cell (APC) populations purified from the CNS of mice with PLP178-191-induced R-EAE shows that only F4/80-CD11c+CD45hi dendritic cells (DCs) efficiently present endogenous antigen to activate naive PLP139-151-specific T cells in vitro. In contrast, DCs as well as F4/80+CD45hi macrophages and F4/80+CD45lo microglia activate a PLP139-151-specific helper T cell line. The data suggest that naive T cells enter the inflamed CNS and are activated by local APCs, possibly DCs, to initiate epitope spreading.

Antigen presentation in the CNS by myeloid dendritic cells drives progression of relapsing experimental autoimmune encephalomyelitis.

Chronic progression of relapsing experimental autoimmune encephalomyelitis (R-EAE), a mouse model of multiple sclerosis (MS), is dependent on the activation of T cells to endogenous myelin epitopes, that is, epitope spreading. This review focuses on the cellular and molecular mechanisms underlying the process of epitope spreading. Surprisingly, activation of naïve T cells to endogenous myelin epitopes in SJL mice undergoing R-EAE occurs directly in the central nervous system (CNS), a site generally perceived to be immunologically privileged. Determination of the antigen presentation capacity of antigen-presenting cell (APC) populations purified from the CNS of mice with established R-EAE shows that peripherally derived CD11b(+)CD11c(+)CD45(hi) myeloid dendritic cells (mDCs) most efficiently present endogenous myelin antigens to activate both preactivated effector myelin-specific T cells and naïve T cells. The mDCs, which drive epitope spreading, preferentially polarize pathogenic Th17 responses correlating with their enhanced expression of TGF-beta1, IL-6, and IL-23. Both B220(+)CD11c(+) plasmacytoid (pDCs) and CD8alpha(+)CD11c(+) (CD8 DCs) were superior to CD11b(+)CD11c(-)CD45(hi) macrophages, but less efficient than mDCs at presenting endogenous peptide to induce Th17 cells. In contrast, CNS-resident CD11b(+)CD11c(-)CD45(low) microglia purified from the inflamed CNS were found to be largely incapable of activating either naïve or effector T cells.

Innate and adaptive immune responses of the central nervous system.

The central nervous system (CNS) is an immunologically specialized organ. The blood-brain barrier regulates the passage of molecules and cells into the CNS. Robust immune responses occur in the CNS even though there is normally an absence of MHC molecules, lack of normal lymphatic drainage, and reduced immune surveillance. This review discusses the immunological elements of the healthy CNS and the pattern of responses that evolve during innate and adaptive immunity in this organ. We also discuss the contribution of astrocytes, cerebrovascular endothelial cells, microglia, macrophages, and dendritic cells to the integrity and pathology of the CNS during CD4+ T-cell autoimmune responses directed against neuroantigens.

CNS dendritic cells: critical participants in CNS inflammation?

Dendritic cells (DCs) are a heterogeneous population of migratory cells specialized for the uptake, processing, and presentation of antigen to T cells. They consist of a variety of mature subpopulations, classically divided into "lymphoid" and "myeloid" subsets. Although there likely exists significant plasticity and redundancy between DC subpopulations, unique differences have been noted in their abilities for T cell stimulation, tolerance induction, T helper cell polarization, cytokine secretion, and anatomic localization. Although DCs are conspicuously absent from the healthy CNS parenchyma, their presence in the vascular-rich regions of the healthy CNS has been well established and suggests they may have a role in immune surveillance. DCs do accumulate in the CNS parenchyma in a wide range of inflammatory responses including parasite, viral, or bacterial infection and CNS autoimmune disease. They also are present in CNS immune responses without overt T cell involvement, such as the inflammation accompanying CNS injury or neurodegeneration. Controversy remains on the role of CNS DCs during inflammation and whether they differentiate from CNS-resident microglia or infiltrate from a blood-borne population. This review will summarize DC subsets and function, overview the current research on DCs in the healthy and inflamed CNS, and address discrepancies between experimental studies.

Peripheral macrophage recruitment in cuprizone-induced CNS demyelination despite an intact blood-brain barrier.

The contribution of peripheral macrophage was assessed in cuprizone intoxication, a model of demyelination and remyelination in which the blood-brain barrier remains intact. Flow cytometry of brain cells isolated from cuprizone-treated mice revealed an increase in the percentage of Mac-1(+)/CD45(hi) peripheral macrophage. To confirm these results in situ, C57BL/6 mice were lethally irradiated, transplanted with bone marrow from GFP-transgenic mice, and exposed to cuprizone. GFP(+) peripheral macrophages were seen in the CNS after 2 weeks of treatment, and infiltration continued through 6 weeks. While the peripheral macrophages were far outnumbered by the resident microglia, their recruitment across the blood-brain barrier alludes to a potentially important role.

Characterization of a novel and spontaneous mouse model of inflammatory arthritis.

INTRODUCTION: Mouse models of rheumatoid arthritis (RA) have proven critical for identifying genetic and cellular mechanisms of the disease. Upon discovering mice in our breeding colony that had spontaneously developed inflamed joints reminiscent of RA, we established the novel IIJ (inherited inflamed joints) strain. The purpose of this study was to characterize the histopathological, clinical, genetic and immunological properties of the disease. METHODS: To begin the IIJ strain, an arthritic male mouse was crossed with SJL/J females. Inheritance of the phenotype was then tracked by intercrossing, backcrossing and outcrossing to other inbred strains. The histopathology of the joints and extraarticular organ systems was examined. Serum cytokines and immunoglobulins (Igs) were measured by ELISA and cytometric bead array. Transfer experiments tested whether disease could be mediated by serum alone. Finally, the cellular joint infiltrate and the composition of secondary lymphoid organs were examined by immunohistochemistry and flow cytometry. RESULTS: After nine generations of intercrossing, the total incidence of arthritis was 33% (304 of 932 mice), with females being affected more than males (38% vs. 28%; P < 0.001). Swelling, most notably in the large distal joints, typically became evident at an early age (mean age of 52 days). In addition to the joint pathology, which included bone and cartilage erosion, synovial hyperproliferation and a robust cellular infiltration of mostly Gr-1(+) neutrophils, there was also evidence of systemic inflammation. IL-6 was elevated in the sera of recently arthritic mice, and extraarticular inflammation was observed histologically in multiple organs. Total serum Ig and IgG1 levels were significantly elevated in arthritic mice, and autoantibodies such as rheumatoid factor and Ig reactive to joint components (collagen type II and joint homogenate) were also detected. Nevertheless, serum failed to transfer disease. A high percentage of double-negative (CD4(-)CD8(-)) CD3(+) TCRα/ß(+) T cells in the lymphoid organs of arthritic IIJ mice suggested significant disruption in the T-cell compartment. CONCLUSIONS: Overall, these data identify the IIJ strain as a new murine model of inflammatory, possibly autoimmune, arthritis. The IIJ strain is similar, both histologically and serologically, to RA and other murine models of autoimmune arthritis. It may prove particularly useful for understanding the female bias in autoimmune diseases.

CNS myeloid DCs presenting endogenous myelin peptides 'preferentially' polarize CD4+ T(H)-17 cells in relapsing EAE.

Peripherally derived CD11b(+) myeloid dendritic cells (mDCs), plasmacytoid DCs, CD8alpha(+) DCs and macrophages accumulate in the central nervous system during relapsing experimental autoimmune encephalomyelitis (EAE). During acute relapsing EAE induced by a proteolipid protein peptide of amino acids 178-191, transgenic T cells (139TCR cells) specific for the relapse epitope consisting of proteolipid protein peptide amino acids 139-151 clustered with mDCs in the central nervous system, were activated and differentiated into T helper cells producing interleukin 17 (T(H)-17 cells). CNS mDCs presented endogenously acquired peptide, driving the proliferation of and production of interleukin 17 by naive 139TCR cells in vitro and in vivo. The mDCs uniquely biased T(H)-17 and not T(H)1 differentiation, correlating with their enhanced expression of transforming growth factor-beta1 and interleukins 6 and 23. Plasmacytoid DCs and CD8alpha(+) DCs were superior to macrophages but were much less efficient than mDCs in presenting endogenous peptide to induce T(H)-17 cells. Our findings indicate a critical function for CNS mDCs in driving relapses in relapsing EAE.

Upregulation of the stress-associated gene p8 in mouse models of demyelination and in multiple sclerosis tissues.

Cuprizone-induced demyelination is a mouse model of multiple sclerosis (MS) as cuprizone-fed mice exhibit neuroinflammation and demyelination in the brain. Upon removal of cuprizone from the diet, inflammation is resolved and reparative remyelination occurs. In an Affymetrix GeneChip analysis, the stress-associated gene p8 was strongly upregulated (>10x) during cuprizone-induced demyelination but not remyelination. We verified this upregulation (>15x) of p8 in the CNS during demyelination by real-time polymerase chain reaction (PCR). This upregulation is brain-specific, as p8 is not elevated in the liver, lung, kidney, spleen, and heart of cuprizone-treated mice. We also localized the cellular source of p8 during cuprizone treatment, and further found elevated expression during embryogenesis but not in normal adult brain. Compared with wild-type controls, the death of oligodendrocytes in p8-/- mice is delayed, as is microglial recruitment to areas of demyelination. The corpus callosum of p8-/- mice demyelinates at a slower rate than wild-type mice, suggesting that p8 exacerbates CNS inflammation and demyelination. Enhanced expression of p8 is also observed in the spinal cords of mice with acute experimental autoimmune encephalomyelitis (EAE) induced by PLP139-151 peptide (10x). Increased expression is detected during disease onset and expression wanes during the remission phase. Finally, p8 is found upregulated (8x) in post-mortem tissue from MS patients and is higher in the plaque tissue compared with adjacent normal-appearing white and gray matter. Thus, p8 is an excellent candidate as a novel biomarker of demyelination.

Fate of donor hematopoietic cells in demyelinating mutant mouse, twitcher, following transplantation of GFP+ bone marrow cells.

The twitcher mouse is an authentic murine model of a genetic demyelinating disease globoid cell leukodystrophy. Allogeneic bone marrow transplantation (BMT) in twitcher mice resulted in the clinicopathological improvement. Thus, using green fluorescent protein (GFP) transgenic mice as the donor, we investigated the behavior and fate of the donor cells and the possibility of transdifferentiation of the donor cells into neuroglial cells in the chimeric twitcher mice. GFP(+) cells were found throughout the brain, most conspicuous in the areas of demyelination. The donor GFP(+) cells expressed RCA-1, a marker for microglia/macrophages but were never exceed 70% of the entire population of the RCA-1(+) microglia/macrophages. There was no convincing evidence that GFP(+) donor cells expressed markers for neurons, astrocytes, or oligodendrocytes. We concluded BMT is therapeutic for this model. However, this effect is not mediated by donor cells transdifferentiation in the brain.