c-Met+ Cytotoxic T Lymphocytes Exhibit Enhanced Cytotoxicity in Mice and Humans In Vitro Tumor Models.

CD8+ cytotoxic T lymphocytes (CTLs) play a crucial role in anti-tumor immunity. In a previous study, we identified a subset of murine effector CTLs expressing the hepatocyte growth factor (HGF) receptor, c-Met (c-Met+ CTLs), that are endowed with enhanced cytolytic capacity. HGF directly inhibited the cytolytic function of c-Met+ CTLs, both in 2D in vitro assays and in vivo, leading to reduced T cell responses against metastatic melanoma. To further investigate the role of c-Met+ CTLs in a three-dimensional (3D) setting, we studied their function within B16 melanoma spheroids and examined the impact of cell-cell contact on the modulation of inhibitory checkpoint molecules' expression, such as KLRG1, PD-1, and CTLA-4. Additionally, we evaluated the cytolytic capacity of human CTL clones expressing c-Met (c-Met+) and compared it to c-Met- CTL clones. Our results indicated that, similar to their murine counterparts, c-Met+ human CTL clones exhibited increased cytolytic activity compared to c-Met- CTL clones, and this enhanced function was negatively regulated by the presence of HGF. Taken together, our findings highlight the potential of targeting the HGF/c-Met pathway to modulate CTL-mediated anti-tumor immunity. This research holds promise for developing strategies to enhance the effectiveness of CTL-based immunotherapies against cancer.

CD4+c-Met+Itgα4+ T cell subset promotes murine neuroinflammation.

OBJECTIVE: c-Met, a tyrosine kinase receptor, is the unique receptor for hepatocyte growth factor (HGF). The HGF/c-Met axis is reported to modulate cell migration, maturation, cytokine production, and antigen presentation. Here, we report that CD4+c-Met+ T cells are detected at increased levels in experimental autoimmune encephalomyelitis (EAE), a mouse model of multiple sclerosis (MS). METHODS: c-Met expression by CD4+ T cells was analyzed mostly by flow cytometry and by immunohistochemistry from mice and human PBMCs. The in vivo role of CD4+c-Met+ T cells was assessed in EAE. RESULTS: CD4+c-Met+ T cells found in the CNS during EAE peak disease are characterized by a pro-inflammatory phenotype skewed towards a Th1 and Th17 polarization, with enhanced adhesion and transmigration capacities correlating with increased expression of integrin α4 (Itgα4). The adoptive transfer of Itgα4-expressing CD4+Vα3.2+c-Met+ T cells induces increased disease severity compared to CD4+Vα3.2+c-Met- T cells. Finally, CD4+c-Met+ T cells are detected in the brain of MS patients, as well as in the blood with a higher level of Itgα4. These results highlight c-Met as an immune marker of highly pathogenic pro-inflammatory and pro-migratory CD4+ T lymphocytes associated with neuroinflammation.

c-Met is expressed by highly autoreactive encephalitogenic CD8+ cells.

BACKGROUND: CD8+ T lymphocytes are critical mediators of neuroinflammatory diseases. Understanding the mechanisms that govern the function of this T cell population is crucial to better understanding central nervous system autoimmune disease pathology. We recently identified a novel population of highly cytotoxic c-Met-expressing CD8+ T lymphocytes and found that hepatocyte growth factor (HGF) limits effective murine cytotoxic T cell responses in cancer models. Here, we examined the role of c-Met-expressing CD8+ T cells by using a MOG35-55 T cell-mediated EAE model. METHODS: Mice were subcutaneously immunized with myelin oligodendrocyte glycoprotein peptide (MOG)35-55 in complete Freund's adjuvant (CFA). Peripheral and CNS inflammation was evaluated at peak disease and chronic phase, and c-Met expression by CD8 was evaluated by flow cytometry and immunofluorescence. Molecular, cellular, and killing function analysis were performed by real-time PCR, ELISA, flow cytometry, and killing assay. RESULTS: In the present study, we observed that a fraction of murine effector CD8+ T cells expressed c-Met receptor (c-Met+CD8+) in an experimental autoimmune encephalitis (EAE) model. Phenotypic and functional analysis of c-Met+CD8+ T cells revealed that they recognize the encephalitogenic epitope myelin oligodendrocyte glycoprotein37-50. We demonstrated that this T cell population produces higher levels of interferon-γ and granzyme B ex vivo and that HGF directly restrains the cytolytic function of c-Met+CD8+ T cells in cell-mediated cytotoxicity reactions CONCLUSIONS: Altogether, our findings suggest that the HGF/c-Met pathway could be exploited to modulate CD8+ T cell-mediated neuroinflammation.

A proliferation-inducing ligand-mediated anti-inflammatory response of astrocytes in multiple sclerosis.

OBJECTIVE: The two related tumor necrosis factor members a proliferation-inducing ligand (APRIL) and B-cell activation factor (BAFF) are currently targeted in autoimmune diseases as B-cell regulators. In multiple sclerosis (MS), combined APRIL/BAFF blockade led to unexpected exacerbated inflammation in the central nervous system (CNS) of patients. Here, we investigate the role of the APRIL/BAFF axis in the CNS. METHODS: APRIL expression was analyzed in MS lesions by immunohistochemistry. The in vivo role of APRIL was assessed in the murine MS model, experimental autoimmune encephalitis (EAE). Functional in vitro studies were performed with human and mouse astrocytes. RESULTS: APRIL was expressed in lesions from EAE. In its absence, the disease was worst. Lesions from MS patients also showed APRIL expression upon infiltration of macrophages. Notably, all the APRIL secreted by these macrophages specifically targeted astrocytes. The upregulation of chondroitin sulfate proteoglycan, sometimes bearing chondroitin sulfate of type E sugar moieties, binding APRIL, in reactive astrocytes explained the latter selectivity. Astrocytes responded to APRIL by producing a sufficient amount of IL-10 to dampen antigen-specific T-cell proliferation and pathogenic cytokine secretion. Finally, an intraspinal delivery of recombinant APRIL before disease onset, shortly reduced EAE symptoms. Repeated intravenous injections of recombinant APRIL before and even at disease onset also had an effect. INTERPRETATION: Our data show that APRIL mediates an anti-inflammatory response from astrocytes in MS lesions. This protective activity is not shared with BAFF. ANN NEUROL 2019;85:406-420.

Transmembrane TNF and Partially TNFR1 Regulate TNFR2 Expression and Control Inflammation in Mycobacterial-Induced Pleurisy.

Pleural tuberculosis is one of the most frequent forms of extra-pulmonary tuberculosis observed in patients infected with Mycobacterium tuberculosis. Tumor Necrosis Factor (TNF) is a crucial cytokine needed to control tuberculosis infection that remains a leading cause of morbidity and mortality worldwide. TNF blockade compromises host immunity and may increase the risk of reactivation of latent infection resulting in overt pulmonary, pleural and extra-pulmonary tuberculosis. While TNF signaling is mainly considered pro-inflammatory, its requirement for the anti-inflammation process involved in the resolution of infection and tissue repair is less explored. Our study analyzes the role of TNF and TNF receptors in the control of the inflammatory process associated with Bacillus Calmette-Guérin (BCG)-induced pleurisy. This study shows that the absence of TNF causes exacerbated inflammation in the pleural cavity of BCG-infected mice which is controlled by the transmembrane TNF (tmTNF) expression. The lack of TNF is associated with an impaired cellular expression and shedding of TNFR2 in the pleural cavity. The presence of tmTNF restores the normal expression of TNFR2 on myeloid cells during BCG-induced pleurisy. We also show that absence of TNFR1 affects the expression of TNFR2 on pleural cells and inflammation in the pleural cavity of BCG-infected mice. In conclusion, tmTNF but not soluble TNF prevents pleural cavity inflammation leading to attenuation and the resolution of the inflammatory process caused by mycobacterial pleurisy in association with the expression of TNFR2 on myeloid cells.

In-vivo gp100-specific Cytotoxic CD8+ T Cell Killing Assay.

Cytotoxic CD8+ T lymphocytes (CTLs) represent a crucial component of the adaptive immune system and play a prominent role in the anti-tumor immune responses of both mice and humans. Cytotoxic CD8+ T cells are responsible for the lysis of cells expressing peptides associated with MHC class I molecules and derived from infection with a pathogen or from mutated antigens. In order to quantify in vivo this antigen-specific CD8+ T cell killing activity, we use the in vivo killing assay (IVKA). Here, we describe the protocol for the lysis of cells expressing a CD8+ T cell melanoma epitope of the hgp10025-33 protein (KVPRNQDWL). C57BL/6 recipient mice, receive first target cells, prepared from naive congenic (CD45.1) C57BL/6 spleen cells pulsed with the hgp10025-33 peptide and labeled with CFSE and of non-pulsed control cells labeled with Brilliant violet. One day later, the spleen cells of recipient mice are isolated and analyzed by FACS to measure the amount of CFSE cells and Brillant Violet (BV) cells. The percentage of lysis is calculated by the difference between CFSE versus BV. Measuring the ability of antigen-specific CD8+ T cells to lyse their antigen in vivo is very important to evaluate the adaptive cytotoxic response induced against a pathogen or a tumor antigen.

Transmembrane Tumor Necrosis Factor Controls Myeloid-Derived Suppressor Cell Activity via TNF Receptor 2 and Protects from Excessive Inflammation during BCG-Induced Pleurisy.

Pleural tuberculosis (TB) is a form of extra-pulmonary TB observed in patients infected with Mycobacterium tuberculosis. Accumulation of myeloid-derived suppressor cells (MDSC) has been observed in animal models of TB and in human patients but their role remains to be fully elucidated. In this study, we analyzed the role of transmembrane TNF (tmTNF) in the accumulation and function of MDSC in the pleural cavity during an acute mycobacterial infection. Mycobacterium bovis BCG-induced pleurisy was resolved in mice expressing tmTNF, but lethal in the absence of tumor necrosis factor. Pleural infection induced MDSC accumulation in the pleural cavity and functional MDSC required tmTNF to suppress T cells as did pleural wild-type MDSC. Interaction of MDSC expressing tmTNF with CD4 T cells bearing TNF receptor 2 (TNFR2), but not TNFR1, was required for MDSC suppressive activity on CD4 T cells. Expression of tmTNF attenuated Th1 cell-mediated inflammatory responses generated by the acute pleural mycobacterial infection in association with effective MDSC expressing tmTNF and interacting with CD4 T cells expressing TNFR2. In conclusion, this study provides new insights into the crucial role played by the tmTNF/TNFR2 pathway in MDSC suppressive activity required during acute pleural infection to attenuate excessive inflammation generated by the infection.

Identification of a novel population of highly cytotoxic c-Met-expressing CD8+ T lymphocytes.

CD8+ cytotoxic T lymphocytes (CTLs) are critical mediators of anti-tumor immunity, and controlling the mechanisms that govern CTL functions could be crucial for enhancing patient outcome. Previously, we reported that hepatocyte growth factor (HGF) limits effective murine CTL responses via antigen-presenting cells. Here, we show that a fraction of murine effector CTLs expresses the HGF receptor c-Met (c-Met+ CTLs). Phenotypic and functional analysis of c-Met+ CTLs reveals that they display enhanced cytolytic capacities compared to their c-Met- CTL counterparts. Furthermore, HGF directly restrains the cytolytic function of c-Met+ CTLs in cell-mediated cytotoxicity reactions in vitro and in vivo and abrogates T-cell responses against metastatic melanoma in vivo Finally, we establish in three murine tumor settings and in human melanoma tissues that c-Met+ CTLs are a naturally occurring CD8+ T-cell population. Together, our findings suggest that the HGF/c-Met pathway could be exploited to control CD8+ T-cell-mediated anti-tumor immunity.

Activation of human B cells negatively regulates TGF-ß1 production.

BACKGROUND: Accumulating evidence indicate that B cells can exhibit pro- or anti-inflammatory activities. Similar to interleukin (IL)-10-competent B cells, we recently showed that transforming growth factor (TGF)-ß1-producing regulatory B cells limit the induction of autoimmune neuroinflammation in mice, making them potentially important in maintaining peripheral immune tolerance in central nervous system inflammatory demyelinating disorders such as multiple sclerosis. METHODS: In this study, we compared B cell production of TGF-ß1 and IL-10, the two most studied regulatory cytokines, and the pro-inflammatory B cell-derived IL-6 and tumor necrosis factor cytokines under basal conditions and following polyclonal stimulation with dual B cell receptor (BCR) cross-linking and Toll-like receptor (TLR)9 engagement. RESULTS: We showed that resting TGF-ß1-producing B cells fall within both the naïve (CD27-) and memory (CD27+) B cell compartments. We found no spontaneous B cell-derived IL-10, IL-6 or tumor necrosis factor (TNF) production. Human B cell activation with anti-Ig antibodies plus CPG-B leads to only modest IL-10 production by memory CD19+CD27+ B cells while expression levels of IL-6 and TNF by both naive and memory B cells were strongly induced. Remarkably, stimulated B cells showed significantly reduced capacity to produce TGF-ß1. CONCLUSIONS: These findings indicate that B cell activation may facilitate the development of excessive immune responses and autoimmunity by restricting B cell-derived TGF-ß1 production by resting B cells and favoring in turns the proinflammatory actions of activated cytokine-producing B cells.

B cell-derived transforming growth factor-ß1 expression limits the induction phase of autoimmune neuroinflammation.

Studies in experimental autoimmune encephalomyelitis (EAE), a murine model of multiple sclerosis (MS), have shown that regulatory B cells modulate the course of the disease via the production of suppressive cytokines. While data indicate a role for transforming growth factor (TGF)-ß1 expression in regulatory B cell functions, this mechanism has not yet been tested in autoimmune neuroinflammation. Transgenic mice deficient for TGF-ß1 expression in B cells (B-TGF-ß1-/-) were tested in EAE induced by recombinant mouse myelin oligodendrocyte glycoprotein (rmMOG). In this model, B-TGF-ß1-/- mice showed an earlier onset of neurologic impairment compared to their littermate controls. Exacerbated EAE susceptibility in B-TGF-ß1-/- mice was associated with augmented CNS T helper (Th)1/17 responses. Moreover, selective B cell TGF-ß1-deficiency increased the frequencies and activation of myeloid dendritic cells, potent professional antigen-presenting cells (APCs), suggesting that B cell-derived TGF-ß1 can constrain Th1/17 responses through inhibition of APC activity. Collectively our data suggest that B cells can down-regulate the function of APCs, and in turn encephalitogenic Th1/17 responses, via TGF-ß1, findings that may be relevant to B cell-targeted therapies.

Novel Cell-Penetrating Peptide-Based Vaccine Induces Robust CD4+ and CD8+ T Cell-Mediated Antitumor Immunity.

Vaccines that can coordinately induce multi-epitope T cell-mediated immunity, T helper functions, and immunologic memory may offer effective tools for cancer immunotherapy. Here, we report the development of a new class of recombinant protein cancer vaccines that deliver different CD8(+) and CD4(+) T-cell epitopes presented by MHC class I and class II alleles, respectively. In these vaccines, the recombinant protein is fused with Z12, a novel cell-penetrating peptide that promotes efficient protein loading into the antigen-processing machinery of dendritic cells. Z12 elicited an integrated and multi-epitopic immune response with persistent effector T cells. Therapy with Z12-formulated vaccines prolonged survival in three robust tumor models, with the longest survival in an orthotopic model of aggressive brain cancer. Analysis of the tumor sites showed antigen-specific T-cell accumulation with favorable modulation of the balance of the immune infiltrate. Taken together, the results offered a preclinical proof of concept for the use of Z12-formulated vaccines as a versatile platform for the development of effective cancer vaccines.

Hepatocyte growth factor: A regulator of inflammation and autoimmunity.

Hepatocyte growth factor (HGF) is a pleiotropic cytokine that has been extensively studied over several decades, but was only recently recognized as a key player in mediating protection of many types of inflammatory and autoimmune diseases. HGF was reported to prevent and attenuate disease progression by influencing multiple pathophysiological processes involved in inflammatory and immune response, including cell migration, maturation, cytokine production, antigen presentation, and T cell effector function. In this review, we discuss the actions and mechanisms of HGF in inflammation and immunity and the therapeutic potential of this factor for the treatment of inflammatory and autoimmune diseases.

Hepatocyte growth factor limits autoimmune neuroinflammation via glucocorticoid-induced leucine zipper expression in dendritic cells.

Autoimmune neuroinflammation, including multiple sclerosis and its animal model, experimental autoimmune encephalomyelitis (EAE), a prototype for T cell-mediated autoimmunity, is believed to result from immune tolerance dysfunction leading to demyelination and substantial neurodegeneration. We previously showed that CNS-restricted expression of hepatocyte growth factor (HGF), a potent neuroprotective factor, reduced CNS inflammation and clinical deficits associated with EAE. In this study, we demonstrate that systemic HGF treatment ameliorates EAE through the development of tolerogenic dendritic cells (DCs) with high expression levels of glucocorticoid-induced leucine zipper (GILZ), a transcriptional repressor of gene expression and a key endogenous regulator of the inflammatory response. RNA interference-directed neutralization of GILZ expression by DCs suppressed the induction of tolerance caused by HGF. Finally, adoptive transfer of HGF-treated DCs from wild-type but not GILZ gene-deficient mice potently mediated functional recovery in recipient mice with established EAE through effective modulation of autoaggressive T cell responses. Altogether, these results show that by inducing GILZ in DCs, HGF reproduces the mechanism of immune regulation induced by potent immunomodulatory factors such as IL-10, TGF-ß1, and glucocorticoids and therefore that HGF therapy may have potential in the treatment of autoimmune dysfunctions.

TLR7 signaling exacerbates CNS autoimmunity through downregulation of Foxp3+ Treg cells.

The innate Toll-like receptor 7 (TLR7) detects infections by recognizing viral and bacterial single-stranded RNA. In addition to pathogen-derived RNA, immune cells expressing high levels of TLR7, such as B cells and dendritic cells (DCs), can be activated by self-RNA. During myelin-induced experimental autoimmune encephalomyelitis (EAE), an animal model of multiple sclerosis, TLR7 expression is increased within the central nervous system (CNS). To define the contribution of TLR7 to the development of EAE, we evaluated the course of the disease in C57BL/6-Tlr7-deficient mice compared with that in WT mice and found that TLR7-deficient mice had decreased disease severity. This protection was associated with decreased myelin oligodendrocyte glycoprotein-specific T-cell activation by primed DCs, decreased circulating autoantibodies, attenuated inflammation within the CNS, and increased Foxp3(+) regulatory T cells in the periphery and in the CNS. In conclusion, we show that TLR7 is involved in the maintenance of autoimmunity in the pathogenesis of EAE.

The neurotrophic hepatocyte growth factor induces protolerogenic human dendritic cells.

Hepatocyte growth factor (HGF) limits mouse autoimmune neuroinflammation by promoting the development of tolerogenic dendritic cells (DCs). Given the role played by DCs in the establishment of immunological tolerance, agents that coerce DCs to adopt a protolerogenic function are currently under investigation for multiple sclerosis (MS) therapy. Here, we studied the immunomodulatory effects of HGF on DCs derived from human monocytes. DCs differentiated in the presence of HGF adopt a protolerogenic phenotype with increased ability to generate regulatory T cells, a property that might be exploited therapeutically in T cell-mediated immune disorders such as MS.

The neurotrophic hepatocyte growth factor attenuates CD8+ cytotoxic T-lymphocyte activity.

BACKGROUND: Accumulating evidence suggests a deleterious role for CD8+ T cells in multiple sclerosis (MS) pathogenesis. We have recently reported that hepatocyte growth factor (HGF), a potent neuroprotective factor, limits CD4+ T cell-mediated autoimmune neuroinflammation by promoting tolerogenic dendritic cells (DCs) and subsequently regulatory T cells. Whether HGF modulates cell-mediated immunity driven by MHC class I-restricted CD8+ T cells remains to be determined. METHODS: Here we examined whether HGF regulates antigen-specific CD8+ T cell responses using an established model of murine cytotoxic T lymphocyte (CTL)-mediated killing. RESULTS: We found that HGF treatment of gp100-pulsed DCs reduced the activation of gp100-specific T cell receptor (Pmel-1) CD8+ T cells and subsequent MHC class I-restricted CTL-mediated cytolysis of gp100-pulsed target cells. The levels of perforin, granzyme B, IFN-γ, and the degranulation marker CD107a as well as Fas ligand were decreased among CD8+ T cells, suggestive of a dual inhibitory effect of HGF on the perforin/granzyme B- and Fas-based lytic pathways in cell-mediated cytotoxicity. Treatment of CD8+ T cells with concanamycin A, a potent inhibitor of the perforin-mediated cytotoxic pathway, abrogated CTL cytotoxicity indicating that blockade of the perforin-dependent killing is a major mechanism by which HGF diminished cytolysis of gp100-pulsed target cells. Moreover, HGF suppressed the generation of effector memory CTLs. CONCLUSIONS: Our findings indicate that HGF treatment limits both the generation and activity of effector CTL from naïve CD8+ T cells. Complementary to its impact on CD4+ T-cell CNS autoimmunity and myelin repair, our findings further suggest that HGF treatment could be exploited to control CD8+ T-cell-mediated, MHC I-restricted autoimmune dysfunctions such as MS.

Nogo-A downregulation improves insulin secretion in mice.

Type 2 diabetes (T2D) is characterized by ß-cell dysfunction and the subsequent depletion of insulin production, usually in a context of increased peripheral insulin resistance. T2D patients are routinely treated with oral antidiabetic agents such as sulfonylureas or dipeptidyl peptidase-4 antagonists, which promote glucose- and incretin-dependent insulin secretion, respectively. Interestingly, insulin secretion may also be induced by neural stimulation. Here we report the expression of Nogo-A in ß-cells. Nogo-A is a membrane protein that inhibits neurite outgrowth and cell migration in the central nervous system. We observed that Nogo-A-deficient mice display improved insulin secretion and glucose clearance. This was associated with a stronger parasympathetic input and higher sensitivity of ß-cells to the cholinergic analog carbachol. Insulin secretion was also improved in diabetic db/db mice treated with neutralizing antibody against Nogo-A. Together, these findings suggest that promoting the vagal stimulation of insulin secretion through the selective inhibition of Nogo-A could be a novel therapeutic approach in T2D.

Interferon-ß induces hepatocyte growth factor in monocytes of multiple sclerosis patients.

Interferon-ß is a first-line therapy used to prevent relapses in relapsing-remitting multiple sclerosis. The clinical benefit of interferon-ß in relapsing-remitting multiple sclerosis is attributed to its immunomodulatory effects on inflammatory mediators and T cell reactivity. Here, we evaluated the production of hepatocyte growth factor, a neuroprotective and neuroinflammation-suppressive mediator, by peripheral blood mononuclear cells collected from interferon-ß--treated relapsing-remitting multiple sclerosis patients, relapsing remitting multiple sclerosis patients not treated with interferon-ß, and healthy volunteers. Using intracellular flow cytometry analysis, increased production of hepatocyte growth factor was observed in circulating CD14(+) monocytes from patients undergoing long-term treatment with interferon-ß versus untreated patients. Complementary in vitro studies confirmed that treatment with interferon-ß induced rapid and transient transcription of the hepatocyte growth factor gene in CD14(+) monocytes and that intracellular and secreted monocytic hepatocyte growth factor protein levels were markedly stimulated by interferon-ß treatment. Additional exploration revealed that "pro-inflammatory" (CD14(+)CD16(+)) monocytes produced similar levels of hepatocyte growth factor in response to interferon-ß as "classical" (CD14(+)CD16(-)) monocytes, and that CD14(+) monocytes but not CD4(+) T cells express the hepatocyte growth factor receptor c-Met. Our findings suggest that interferon-ß may mediate some of its therapeutic effects in relapsing-remitting multiple sclerosis through the induction of hepatocyte growth factor by blood monocytes by coupling immune regulation and neuroprotection.

IgG glycan hydrolysis by EndoS inhibits experimental autoimmune encephalomyelitis.

Studies in experimental autoimmune encephalomyelitis (EAE), a mouse model of multiple sclerosis, have shown that B cells markedly influence the course of the disease, although whether their effects are protective or pathological is a matter of debate. EndoS hydrolysis of the IgG glycan has profound effects on IgG effector functions, such as complement activation and Fc receptor binding, suggesting that the enzyme could be used as an immunomodulatory therapeutic agent against IgG-mediated diseases. We demonstrate here that EndoS has a protective effect in myelin oligodendrocyte glycoprotein peptide amino acid 35-55 (MOG(35-55))-induced EAE, a chronic neuroinflammatory demyelinating disorder of the central nervous system (CNS) in which humoral immune responses are thought to play only a minor role. EndoS treatment in chronic MOG(35-55)-EAE did not impair encephalitogenic T cell priming and recruitment into the CNS of mice, consistent with a primary role of EndoS in controlling IgG effector functions. In contrast, reduced EAE severity coincided with poor serum complement activation and deposition within the spinal cord, suggesting that EndoS treatment impairs B cell effector function. These results identify EndoS as a potential therapeutic agent against antibody-mediated CNS autoimmune disorders.