Obesity intensifies sex-specific interferon signaling to selectively worsen central nervous system autoimmunity in females.

Obesity has been implicated in the rise of autoimmunity in women. We report that obesity induces a serum protein signature that is associated with T helper 1 (Th1), interleukin (IL)-17, and multiple sclerosis (MS) signaling pathways selectively in human females. Females, but not male mice, subjected to diet-induced overweightness/obesity (DIO) exhibited upregulated Th1/IL-17 inflammation in the central nervous system during experimental autoimmune encephalomyelitis, a model of MS. This was associated with worsened disability and a heightened expansion of myelin-specific Th1 cells in the peripheral lymphoid organs. Moreover, at steady state, DIO increased serum levels of interferon (IFN)-α and potentiated STAT1 expression and IFN-γ production by naive CD4+ T cells uniquely in female mice. This T cell phenotype was driven by increased adiposity and was prevented by the removal of ovaries or knockdown of the type I IFN receptor in T cells. Our findings offer a mechanistic explanation of how obesity enhances autoimmunity.

Serum Proteomics Distinguish Subtypes of NMO Spectrum Disorder and MOG Antibody-Associated Disease and Highlight Effects of B-Cell Depletion.

BACKGROUND AND OBJECTIVES: AQP4 antibody-positive NMOSD (AQP4-NMOSD), MOG antibody-associated disease (MOGAD), and seronegative NMOSD (SN-NMOSD) are neuroautoimmune conditions that have overlapping clinical manifestations. Yet, important differences exist in these diseases, particularly in B-cell depletion (BCD) efficacy. Yet, the biology driving these differences remains unclear. Our study aims to clarify biological pathways distinguishing these diseases beyond autoantibodies and investigate variable BCD effects through proteomic comparisons. METHODS: In a retrospective study, 1,463 serum proteins were measured in 53 AQP4-NMOSD, 25 MOGAD, 18 SN-NMOSD, and 49 healthy individuals. To identify disease subtype-associated signatures, we examined serum proteins in patients without anti-CD20 B-cell depletion (NoBCD). We then assessed the effect of BCD treatment within each subtype by comparing proteins between BCD-treated and NoBCD-treated patients. RESULTS: In NoBCD-treated patients, serum profiles distinguished the 3 diseases. AQP4-NMOSD showed elevated type I interferon-induced chemokines (CXCL9 and CXCL10) and TFH chemokine (CXCL13). MOGAD exhibited increased cytotoxic T-cell proteases (granzyme B and granzyme H), while SN-NMOSD displayed elevated Wnt inhibitory factor 1, a marker for nerve injury. Across all subtypes, BCD-treated patients showed reduction of B-cell-associated proteins. In AQP4-NMOSD, BCD led to a decrease in several inflammatory pathways, including IL-17 signaling, cytokine storm, and macrophage activation. By contrast, BCD elevated these pathways in patients with MOGAD. BCD had no effect on these pathways in SN-NMOSD. DISCUSSION: Proteomic profiles show unique biological pathways that distinguish AQP4-NMOSD, MOGAD, or SN-NMOSD. Furthermore, BCD uniquely affects inflammatory pathways in each disease type, providing an explanation for the disparate therapeutic response in AQP4-NMOSD and MOGAD.

Autoantibodies identify primary Sjögren's syndrome in patients lacking serum IgG specific for Ro/SS-A and La/SS-B.

OBJECTIVE: Identify autoantibodies in anti-Ro/SS-A negative primary Sjögren's syndrome (SS). METHODS: This is a proof-of-concept, case-control study of SS, healthy (HC) and other disease (OD) controls. A discovery dataset of plasma samples (n=30 SS, n=15 HC) was tested on human proteome arrays containing 19 500 proteins. A validation dataset of plasma and stimulated parotid saliva from additional SS cases (n=46 anti-Ro+, n=50 anti-Ro-), HC (n=42) and OD (n=54) was tested on custom arrays containing 74 proteins. For each protein, the mean+3 SD of the HC value defined the positivity threshold. Differences from HC were determined by Fisher's exact test and random forest machine learning using 2/3 of the validation dataset for training and 1/3 for testing. Applicability of the results was explored in an independent rheumatology practice cohort (n=38 Ro+, n=36 Ro-, n=10 HC). Relationships among antigens were explored using Search Tool for the Retrieval of Interacting Genes/Proteins (STRING) interactome analysis. RESULTS: Ro+ SS parotid saliva contained autoantibodies binding to Ro60, Ro52, La/SS-B and muscarinic receptor 5. SS plasma contained 12 novel autoantibody specificities, 11 of which were detected in both the discovery and validation datasets. Binding to ≥1 of the novel antigens identified 54% of Ro- SS and 37% of Ro+ SS cases, with 100% specificity in both groups. Machine learning identified 30 novel specificities showing receiver operating characteristic area under the curve of 0.79 (95% CI 0.64 to 0.93) for identifying Ro- SS. Sera from Ro- cases of an independent cohort bound 17 of the non-canonical antigens. Antigenic targets in both Ro+ and Ro- SS were part of leukaemia cell, ubiquitin conjugation and antiviral defence pathways. CONCLUSION: We identified antigenic targets of the autoantibody response in SS that may be useful for identifying up to half of Ro seronegative SS cases.

Dual Role of B Cells in Multiple Sclerosis.

B cells have emerged as an important immune cell type that can be targeted for therapy in multiple sclerosis (MS). Depleting B cells with anti-CD20 antibodies is effective in treating MS. Yet, atacicept treatment, which blocks B-cell Activating Factor (BAFF) and A Proliferation-Inducing Ligand (APRIL), two cytokines important for B cell development and function, paradoxically increases disease activity in MS patients. The reason behind the failure of atacicept is not well understood. The stark differences in clinical outcomes with these therapies demonstrate that B cells have both inflammatory and anti-inflammatory functions in MS. In this review, we summarize the importance of B cells in MS and discuss the different B cell subsets that perform inflammatory and anti-inflammatory functions and how therapies modulate B cell functions in MS patients. Additionally, we discuss the potential anti-inflammatory functions of BAFF and APRIL on MS disease.

Neuronal deletion of MnSOD in mice leads to demyelination, inflammation and progressive paralysis that mimics phenotypes associated with progressive multiple sclerosis.

Neuronal oxidative stress has been implicated in aging and neurodegenerative disease. Here we investigated the impact of elevated oxidative stress induced in mouse spinal cord by deletion of Mn-Superoxide dismutase (MnSOD) using a neuron specific Cre recombinase in Sod2 floxed mice (i-mn-Sod2 KO). Sod2 deletion in spinal cord neurons was associated with mitochondrial alterations and peroxide generation. Phenotypically, i-mn-Sod2 KO mice experienced hindlimb paralysis and clasping behavior associated with extensive demyelination and reduced nerve conduction velocity, axonal degeneration, enhanced blood brain barrier permeability, elevated inflammatory cytokines, microglia activation, infiltration of neutrophils and necroptosis in spinal cord. In contrast, spinal cord motor neuron number, innervation of neuromuscular junctions, muscle mass, and contractile function were not altered. Overall, our findings show that loss of MnSOD in spinal cord promotes a phenotype of demyelination, inflammation and progressive paralysis that mimics phenotypes associated with progressive multiple sclerosis.

Biomarker panel increases accuracy for identification of an MS relapse beyond sNfL.

BACKGROUND: For relapsing-remitting multiple sclerosis (RRMS), there is a need for biomarker development beyond clinical manifestations and MRI. Soluble neurofilament light chain (sNfL) has emerged as a biomarker for inflammatory activity in RRMS. However, there are limitations to the accuracy of sNfL in identifying relapses. Here, we sought to identify a panel of biomarkers that would increase the precision of distinguishing patients in relapse compared to sNfL alone. METHODS: We used a multiplex approach to measure levels of 724 blood proteins in two distinct RRMS cohorts. Multiple t-tests with covariate correction determined biomarkers that were differentially regulated in relapse and remission. Logistic regression models determined the accuracy of biomarkers to distinguish relapses from remission. RESULTS: The discovery cohort identified 37 proteins differentially abundant in active RRMS relapse compared to remission. The verification cohort confirmed four proteins, including sNfL, were altered in active RRMS relapse compared to remission. Logistic regression showed that the 4-protein panel identified active relapse with higher accuracy (AUC = 0.87) than sNfL alone (AUC = 0.69). CONCLUSION: Our studies confirmed that sNfL is elevated during relapses in RRMS patients. Furthermore, we identified three other blood proteins, uPA, hK8 and DSG3 that were altered during relapse. Together, these four biomarkers could be used to monitor disease activity in RRMS patients.

Antigen presentation and interferon signatures in B cells driven by localized ablative cancer immunotherapy correlate with extended survival.

Rationale: B cells have emerged as key regulators in protective cancer immunity. However, the activation pathways induced in B cells during effective immunotherapy are not well understood. Methods: We used a novel localized ablative immunotherapy (LAIT), combining photothermal therapy (PTT) with intra-tumor delivery of the immunostimulant N-dihydrogalactochitosan (GC), to treat mice bearing mouse mammary tumor virus-polyoma middle tumor-antigen (MMTV-PyMT). We used single-cell RNA sequencing to compare the transcriptional changes induced by PTT, GC and PTT+GC in B cells within the tumor microenvironment (TME). Results: LAIT significantly increased survival in the tumor-bearing mice, compared to the treatment by PTT and GC alone. We found that PTT, GC and PTT+GC increased the proportion of tumor-infiltrating B cells and induced gene expression signatures associated with B cell activation. Both GC and PTT+GC elevated gene expression associated with antigen presentation, whereas GC elevated transcripts that regulate B cell activation and GTPase function and PTT+GC induced interferon response genes. Trajectory analysis, where B cells were organized according to pseudotime progression, revealed that both GC and PTT+GC induced the differentiation of B cells from a resting state towards an effector phenotype. The analyses confirmed upregulated interferon signatures in the differentiated tumor-infiltrating B cells following treatment by PTT+GC but not by GC. We also observed that breast cancer patients had significantly longer survival time if they had elevated expression of genes in B cells that were induced by PTT+GC therapy in the mouse tumors. Conclusion: Our findings show that the combination of local ablation and local application of immunostimulant initiates the activation of interferon signatures and antigen-presentation in B cells which is associated with positive clinical outcomes for breast cancer. These findings broaden our understanding of LAIT's regulatory roles in remodeling TME and shed light on the potentials of B cell activation in clinical applications.

Interferon-ß Intensifies Interleukin-23-Driven Pathogenicity of T Helper Cells in Neuroinflammatory Disease.

Interferon (IFN)-ß is a popular therapy for multiple sclerosis (MS). However, 25-40% of patients are nonresponsive to this therapy, and it worsens neuromyelitis optica (NMO), another neuroinflammatory disease. We previously identified, in both NMO patients and in mice, that IFN-ß treatment had inflammatory effects in T Helper (TH) 17-induced disease through the production of the inflammatory cytokine IL-6. However, other studies have shown that IFN-ß inhibits the differentiation and function of TH17 cells. In this manuscript, we identified that IFN-ß had differential effects on discrete stages of TH17 development. During early TH17 development, IFN-ß inhibits IL-17 production. Conversely, during late TH17 differentiation, IFN-ß synergizes with IL-23 to promote a pathogenic T cell that has both TH1 and TH17 characteristics and expresses elevated levels of the potent inflammatory cytokines IL-6 and GM-CSF and the transcription factor BLIMP. Together, these findings help resolve a paradox surrounding IFN-ß and TH17-induced disease and illuminate the pathways responsible for the pathophysiology of NMO and MS patients who are IFN-ß nonresponders.

The Role of B Cells in Primary Progressive Multiple Sclerosis.

The success of ocrelizumab in reducing confirmed disability accumulation in primary progressive multiple sclerosis (PPMS) via CD20-targeted depletion implicates B cells as causal agents in the pathogenesis of PPMS. This review explores the possible mechanisms by which B cells contribute to disease progression in PPMS, specifically exploring cytokine production, antigen presentation, and antibody synthesis. B cells may contribute to disease progression in PPMS through cytokine production, specifically GM-CSF and IL-6, which can drive naïve T-cell differentiation into pro-inflammatory Th1/Th17 cells. B cell production of the cytokine LT-α may induce follicular dendritic cell production of CXCL13 and lead indirectly to T and B cell infiltration into the CNS. In contrast, production of IL-10 by B cells likely induces an anti-inflammatory effect that may play a role in reducing neuroinflammation in PPMS. Therefore, reduced production of IL-10 may contribute to disease worsening. B cells are also capable of potent antigen presentation and may induce pro-inflammatory T-cell differentiation via cognate interactions. B cells may also contribute to disease activity via antibody synthesis, although it's unlikely the benefit of ocrelizumab in PPMS occurs via antibody decrement. Finally, various B cell subsets likely promulgate pro- or anti-inflammatory effects in MS.

Pertussis Toxin Inhibits Encephalitogenic T-Cell Infiltration and Promotes a B-Cell-Driven Disease during Th17-EAE.

Pertussis toxin (PTX) is a required co-adjuvant for experimental autoimmune encephalomyelitis (EAE) induced by immunization with myelin antigen. However, PTX's effects on EAE induced by the transfer of myelin-specific T helper cells is not known. Therefore, we investigated how PTX affects the Th17 transfer EAE model (Th17-EAE). We found that PTX significantly reduced Th17-EAE by inhibiting chemokine-receptor-dependent trafficking of Th17 cells. Strikingly, PTX also promoted the accumulation of B cells in the CNS, suggesting that PTX alters the disease toward a B-cell-dependent pathology. To determine the role of B cells, we compared the effects of PTX on Th17-EAE in wild-type (WT) and B-cell-deficient (µMT) mice. Without PTX treatment, disease severity was equivalent between WT and µMT mice. In contrast, with PTX treatment, the µMT mice had significantly less disease and a reduction in pathogenic Th17 cells in the CNS compared to the WT mice. In conclusion, this study shows that PTX inhibits the migration of pathogenic Th17 cells, while promoting the accumulation of pathogenic B cells in the CNS during Th17-EAE. These data provide useful methodological information for adoptive-transfer Th17-EAE and, furthermore, describe another important experimental system to study the pathogenic mechanisms of B cells in multiple sclerosis.

CNS Autoimmune Responses in BCMA-Deficient Mice Provide Insight for the Failure of Atacicept in MS.

OBJECTIVE: B cells have emerged as a therapeutic target for MS. Anti-CD20 antibodies, which deplete B cells, are effective therapies for MS. However, atacicept (TACI-Fc), which blocks BAFF and APRIL and reduces B cells, unexpectedly exacerbates MS. We tested the hypothesis that B cell maturation antigen (BCMA), a receptor for BAFF and APRIL, plays a role in the paradoxical effects of anti-CD20 antibody and TACI-Fc using experimental autoimmune encephalomyelitis (EAE). METHODS: EAE was induced in wild-type (BCMA+/+) and BCMA-deficient (BCMA-/-) mice with an immunization of rodent myelin oligodendrocyte glycoprotein (MOG)35-55 peptide. Treatment with anti-CD20 antibody, TACI-Fc, and isotype controls was administered by intraperitoneal injections. CNS infiltration was evaluated by histology; immune cell phenotypes were evaluated by flow cytometry; MOG-specific antibodies were determined by ELISA. Mixed bone marrow chimeras and cell culture assays were used to identify the specific subsets of immune cells affected by BCMA deficiency. RESULTS: First, we found that BCMA-/- mice had more severe EAE compared with BCMA+/+ mice and the increased disease was associated with elevated anti-MOG B-cell responses. Second, we found that anti-CD20 therapy attenuated EAE in BCMA-/- mice but not in BCMA+/+ mice. Third, TACI-Fc attenuated EAE in BCMA+/+ mice but not in BCMA-/- mice. Mixed bone marrow chimeric and cell culture experiments demonstrated that BCMA deficiency elevates inflammatory B-cell responses but inhibits inflammatory responses in macrophages. CONCLUSIONS: BCMA has multifaceted roles during inflammation that affects therapeutic efficacies of anti-CD20 and TACI-Fc in EAE. Our results from BCMA-deficient mice provide insights into the failure of atacicept in MS.

ELTD1 as a biomarker for multiple sclerosis: Pre-clinical molecular-targeted studies in a mouse experimental autoimmune encephalomyelitis model.

Multiple sclerosis (MS) and glioblastoma (GBM) are two distinct diseases that affect the central nervous system (CNS). However, perturbation in CNS vasculature are hallmarks of both diseases. ELTD1 (epidermal growth factor, latrophilin, and 7 transmembrane domain containing protein 1 on chromosome 1) is associated with vascular development, and has been linked with tumor angiogenesis. In glioblastomas, we detected over-expression of ELTD1, and found that an antibody targeting ELTD1 could increase animal survival and decrease tumor volumes in a xenograft GBM model. RNA-seq analysis of the preclinical data in the model for GBM identified that some of the molecular pathways affected by the anti-ELTD1 antibody therapy are also found to be associated with MS. In this study, we used molecular-targeted (mt) MR imaging and immunohistochemistry to assess ELTD1 levels in experimental autoimmune encephalomyelitis (EAE), a mouse model of MS. Specifically, we found that ELTD1 is readily detected in the brains of mice with EAE and is predominantly found in the corpus callosum. In addition, we found that the blood-brain barrier (BBB) was compromised in the brains of EAE mice using contrast-enhanced MRI (CE-MRI), as well as altered relative cerebral blood flow (rCBF) in the brains and cervical spinal cords of these mice using perfusion imaging, compared to controls. These findings indicate that ELTD1 may be a promising biomarker for CNS-inflammation in MS.

E3 Ubiquitin Ligase Von Hippel-Lindau Protein Promotes Th17 Differentiation.

Von Hippel-Lindau (VHL) is an E3 ubiquitin ligase that targets proteins, including HIF-1α, for proteasomal degradation. VHL and HIF regulate the balance between glycolysis and oxidative phosphorylation, which is critical in highly dynamic T cells. HIF-1α positively regulates Th17 differentiation, a complex process in which quiescent naive CD4 T cells undergo transcriptional changes to effector cells, which are commonly dysregulated in autoimmune diseases. The role of VHL in Th17 cells is not known. In this study, we hypothesized VHL negatively regulates Th17 differentiation and deletion of VHL in CD4 T cells would elevate HIF-1α and increase Th17 differentiation. Unexpectedly, we found that VHL promotes Th17 differentiation. Mice deficient in VHL in their T cells were resistant to an autoimmune disease, experimental autoimmune encephalomyelitis, often mediated by Th17 cells. In vitro Th17 differentiation was impaired in VHL-deficient T cells. In the absence of VHL, Th17 cells had decreased activation of STAT3 and SMAD2, suggesting that VHL indirectly or directly regulates these critical signaling molecules. Gene expression analysis revealed that in Th17 cells, VHL regulates many cellular pathways, including genes encoding proteins involved indirectly or directly in the glycolysis pathway. Compared with wild-type, VHL-deficient Th17 cells had elevated glycolysis and glycolytic capacity. Our finding has implications on the design of therapeutics targeting the distinct metabolic needs of T cells to combat chronic inflammatory diseases.

Transcriptomics and proteomics reveal a cooperation between interferon and T-helper 17 cells in neuromyelitis optica.

Type I interferon (IFN-I) and T helper 17 (TH17) drive pathology in neuromyelitis optica spectrum disorder (NMOSD) and in TH17-induced experimental autoimmune encephalomyelitis (TH17-EAE). This is paradoxical because the prevalent theory is that IFN-I inhibits TH17 function. Here we report that a cascade involving IFN-I, IL-6 and B cells promotes TH17-mediated neuro-autoimmunity. In NMOSD, elevated IFN-I signatures, IL-6 and IL-17 are associated with severe disability. Furthermore, IL-6 and IL-17 levels are lower in patients on anti-CD20 therapy. In mice, IFN-I elevates IL-6 and exacerbates TH17-EAE. Strikingly, IL-6 blockade attenuates disease only in mice treated with IFN-I. By contrast, B-cell-deficiency attenuates TH17-EAE in the presence or absence of IFN-I treatment. Finally, IFN-I stimulates B cells to produce IL-6 to drive pathogenic TH17 differentiation in vitro. Our data thus provide an explanation for the paradox surrounding IFN-I and TH17 in neuro-autoimmunity, and may have utility in predicting therapeutic response in NMOSD.

B cell function impacts the efficacy of IFN-ß therapy in EAE.

Recent studies identified that interferon beta (IFN-ß) treatment skews B-cells towards a regulatory phenotype in multiple sclerosis. To assess B cell involvement during IFN-ß therapy, we compared IFN-ß treatment in a B cell-independent model and a B cell-dependent model of experimental autoimmune encephalomyelitis (EAE). We show that in B cell-independent EAE, IFN-ß ameliorates neuroinflammation. Conversely, in B cell-dependent EAE, IFN-ß has no effect on disease. Effective IFN-ß therapy in B cell-independent EAE was associated with reduced inflammatory T cells in the CNS and skewed splenic B cells towards an immature population and away from a germinal center population. These immune cell populations were unchanged in B cell-dependent EAE. Finally, we found that IFN-ß increased marginal zone B cells in both EAE models. These findings indicate that B cell function impacts IFN-ß efficacy during neuroinflammation.

Epstein Barr virus nuclear antigen 1 (EBNA-1) peptides recognized by adult multiple sclerosis patient sera induce neurologic symptoms in a murine model.

Multiple sclerosis (MS) is an autoimmune demyelinating disease with progressive neurodegeneration and complex etiology likely involving genetic and environmental factors. MS has been associated with Epstein Barr virus (EBV) infection, with patients often showing enhanced responses to EBV antigens. To determine whether abnormal EBV nuclear antigen-1 (EBNA-1) humoral immunity can serve as an initiator of autoimmune responses in MS, we investigated the fine specificities of the humoral immune response against EBNA-1 in MS patients using solid phase epitope mapping. Antibodies from MS patients recognized an EBNA-1 epitope spanning amino acids 411-426, previously unknown to be recognized specifically by untreated MS patients. Antibodies against this epitope cross-reacted to myelin basic protein (MBP). Furthermore, animals immunized with this EBNA-1 polypeptide mounted a response against MBP and developed signs of experimental autoimmune encephalitis (EAE). These data support a link between MS and EBV through antibodies that cross-react between EBV proteins and the MBP autoantigen.

Immunization-Expanded NKT Follicular Helper Cells Drive IgG1 Isotype Switch against an Exogenous T-Independent Polysaccharide but Do Not Promote Recall Responses.

The CD1d-binding glycolipid α-galactosylceramide (α-GC) is a potent adjuvant that activates NKT cells and in turn enhances T-dependent humoral immunity. Very little is known about how NKT cells and the NKT follicular helper (NKTfh) subset influence the immune response to T-independent polysaccharides. In this study, we used a Cre-Lox approach to generate mice devoid of the Bcl6 master transcription factor in CD4 lineage cells and thus devoid of NKTfh cells but not total NKT cells. It was observed that α-GC-driven IgG1 class switch against a polysaccharide Ag was dependent on the NKTfh subset. However, α-GC was unable to stimulate a polysaccharide-specific Ab recall response. It was observed that NKT-derived IL-21 was able to exert limited influence on the IgG1 response and was therefore likely to work in concert with other factors. This work shows that α-GC-driven NKTfh cells can direct polysaccharide-specific B cell responses by promoting IgG1 class switch but do not provide signals needed for generation of polysaccharide-specific B cell memory.

Emerging Role of Follicular T Helper Cells in Multiple Sclerosis and Experimental Autoimmune Encephalomyelitis.

Multiple sclerosis (MS) is an autoimmune disorder where both T cells and B cells are implicated in pathology. However, it remains unclear how these two distinct populations cooperate to drive disease. There is ample evidence from studies in both MS patients and mouse models that Th17, B cells, and follicular T helper (TFH) cells contribute to disease. This review article describes the literature that identifies mechanisms by which Th17, TFH, and B cells cooperatively drive disease activity in MS and experimental autoimmune encephalomyelitis (EAE). The curation of this literature has identified that central nervous system (CNS) infiltrating TFH cells act with TH17 cell to contribute to an inflammatory B cell response in neuroinflammation. This demonstrates that TFH cells and their products are promising targets for therapies in MS.

Role of TFH Cells in Promoting T Helper 17-Induced Neuroinflammation.

Both T cells and B cells are implicated in the pathology of multiple sclerosis (MS), but how these cells cooperate to drive disease remains unclear. Recent studies using experimental autoimmune encephalomyelitis (EAE) demonstrated that the TH17 pathway is correlated with increased numbers of ectopic B-cell follicles in the central nervous system (CNS). As follicular T helper (TFH) cells are regulators of B cell responses, we sought to examine the role of TFH cells in EAE induced by the transfer of myelin-specific TH17 cells (TH17-EAE). In this study, we first confirmed previous reports that B-cells are a major cell type infiltrating the CNS during TH17-EAE. In addition, we found that B cells contribute to the severity of TH17-EAE. Class-switched B-cells in the CNS were positively correlated with disease and, strikingly, the severity TH17-EAE was diminished in B cell deficient mice. We next focused on the role TFH cells play in TH17-EAE. We found substantial numbers of CXCR5+PD1+CD4+ TFH cells in the CNS tissue of TH17-EAE mice and that at the peak of disease, the number of infiltrating TFHs was correlated with the number of infiltrating B-cells. Using congenic CD45.1+ donor mice and CD45.2+ recipient mice, we determined that the TFH cells were recipient-derived, whereas IL-17+ cells were donor-derived. We assessed whether myelin-specific TFH cells are capable of inducing EAE in recipient mice and found that transferring TFH cells failed to induce EAE. Finally, we tested the effects of blocking TFH trafficking in TH17-EAE using an antagonistic antibody against CXCL13, the chemokine ligand for CXCR5 on TFH cells. We found anti-CXCL13 treatment significantly reduced TH17-EAE disease. This treatment blocked CD4+ T cells from entering the CNS, but had no effect on infiltration of B cells. Strikingly, this antibody treatment had no measurable effect on TH17 disease in B cell-deficient mice. These data demonstrate that infiltrating TFH cells are a key cell type that contributes to an inflammatory B cell response in TH17-EAE and provide evidence for targeting TFH cells as a treatment for neuro-autoimmune diseases like MS.

Amelioration of ongoing experimental autoimmune encephalomyelitis with fluoxetine.

In patients with multiple sclerosis, the selective serotonin reuptake inhibitor, fluoxetine, resulted in less acute disease activity. We tested the immune modulating effects of fluoxetine in a mouse model of multiple sclerosis, i.e. experimental autoimmune encephalomyelitis (EAE). We show that fluoxetine delayed the onset of disease and reduced clinical paralysis in mice with established disease. Fluoxetine had abrogating effects on proliferation of immune cells and inflammatory cytokine production by both antigen-presenting cells and T cells. Specifically, in CD4 T cells, fluoxetine increased Fas-induced apoptosis. We conclude that fluoxetine possesses immune-modulating effects resulting in the amelioration of symptoms in EAE.