Chemokine-mediated cell migration into the central nervous system in progressive multifocal leukoencephalopathy.

Progressive multifocal leukoencephalopathy (PML) has been associated with different forms of immune compromise. This study analyzes the chemokine signals and attracted immune cells in cerebrospinal fluid (CSF) during PML to define immune cell subpopulations relevant for the PML immune response. In addition to chemokines that indicate a general state of inflammation, like CCL5 and CXCL10, the CSF of PML patients specifically contains CCL2 and CCL4. Single-cell transcriptomics of CSF cells suggests an enrichment of distinct CD4+ and CD8+ T cells expressing chemokine receptors CCR2, CCR5, and CXCR3, in addition to ITGA4 and the genetic PML risk genes STXBP2 and LY9. This suggests that specific immune cell subpopulations migrate into the central nervous system to mitigate PML, and their absence might coincide with PML development. Monitoring them might hold clues for PML risk, and boosting their recruitment or function before therapeutic immune reconstitution might improve its risk-benefit ratio.

Multiple sclerosis endophenotypes identified by high-dimensional blood signatures are associated with distinct disease trajectories.

One of the biggest challenges in managing multiple sclerosis is the heterogeneity of clinical manifestations and progression trajectories. It still remains to be elucidated whether this heterogeneity is reflected by discrete immune signatures in the blood as a surrogate of disease pathophysiology. Accordingly, individualized treatment selection based on immunobiological principles is still not feasible. Using two independent multicentric longitudinal cohorts of patients with early multiple sclerosis (n = 309 discovery and n = 232 validation), we were able to identify three distinct peripheral blood immunological endophenotypes by a combination of high-dimensional flow cytometry and serum proteomics, followed by unsupervised clustering. Longitudinal clinical and paraclinical follow-up data collected for the cohorts revealed that these endophenotypes were associated with disease trajectories of inflammation versus early structural damage. Investigating the capacity of immunotherapies to normalize endophenotype-specific immune signatures revealed discrete effect sizes as illustrated by the limited effect of interferon-ß on endophenotype 3-related immune signatures. Accordingly, patients who fell into endophenotype 3 subsequently treated with interferon-ß exhibited higher disease progression and MRI activity over a 4-year follow-up compared with treatment with other therapies. We therefore propose that ascertaining a patient's blood immune signature before immunomodulatory treatment initiation may facilitate prediction of clinical disease trajectories and enable personalized treatment decisions based on pathobiological principles.

Single-cell profiling reveals preferential reduction of memory B cell subsets in cladribine patients that correlates with treatment response.

Background: Cladribine is a highly effective immunotherapy that is applied in two short-term courses over 2 years and reduces relapse rate and disease progression in patients with relapsing multiple sclerosis (MS). Despite the short treatment period, cladribine has a long-lasting effect on disease activity even after recovery of lymphocyte counts, suggesting a yet undefined long-term immune modulating effect. Objectives: Our aim was to provide a more profound understanding of the detailed effects of cladribine, also with regard to the patients' therapy response. Design: We performed an open-labeled, explorative, prospective, single-arm study, in which we examined the detailed lymphocyte subset development of MS patients who received cladribine treatment over 2 years. Methods: We performed in-depth profiling of the effects of cladribine on peripheral blood lymphocytes by flow cytometry, bulk RNA sequencing of sorted CD4+ T cells, CD8+ T cells, and CD19+ B cells as well as single-cell RNA sequencing of peripheral blood mononuclear cells in a total of 23 MS patients before and at different time points up to 24 months after cladribine treatment. Data were correlated with clinical and cranial magnetic resonance imaging (MRI) disease activity. Results: Flow cytometry revealed a predominant and sustained reduction of memory B cells compared to other B cell subsets after cladribine treatment, whereas T cell subsets were slightly reduced in a more uniform pattern. The overall transcriptional profile of total blood B cells exhibited reduced expression of proinflammatory and T cell activating genes, while single-cell transcriptomics revealed that gene expression within each B cell cluster did not change over time. Stable patients displayed stronger reductions of selected memory B cell clusters as compared to patients with clinical or cerebral MRI disease activity. Conclusion: We describe a pronounced and sustained effect of cladribine on the memory B cell compartment, and the resulting change in B cell subset composition causes a significant alteration of B cell transcriptional profiles resulting in reduced proinflammatory and T cell activating capacities. The extent of reduction in selected memory B cell clusters by cladribine may predict treatment response.

Contactin-associated protein 2 autoantibodies can be associated with multifocal motor-like neuropathy: a case report.

Autoantibodies against contactin-associated protein 2 (CASPR2) are usually associated with autoimmune encephalitis and neuromyotonia. Their association with inflammatory neuropathies has been described in case reports albeit all with distal symmetric manifestation. Here, we report a patient who developed distal arm paresis, dominantly of the right arm, over the course of 1 year. Electroneurography showed a conduction block of motor nerve conduction, nerve ultrasonography a swelling of the right median and ulnar nerve and flow cytometry an increase in natural killer (NK cells) in the blood and natural killer T (NKT) cells in the cerebrospinal fluid (CSF), therefore indicating a multifocal motor neuropathy-like (MMN-like) phenotype. CASPR2 autoantibodies were detected in serum and CSF. Through immunotherapy with intravenous immunoglobulins the patient showed clinical and neurographic improvement. We therefore describe the first association of CASPR2 autoantibodies with a MMN-like clinical manifestation, extending the spectrum of CASPR2-associated diseases.

Supporting the differential diagnosis of connective tissue diseases with neurological involvement by blood and cerebrospinal fluid flow cytometry.

OBJECTIVE: Neurological manifestations of autoimmune connective tissue diseases (CTD) are poorly understood and difficult to diagnose. We here aimed to address this shortcoming by studying immune cell compositions in CTD patients with and without neurological manifestation. METHODS: Using flow cytometry, we retrospectively investigated paired cerebrospinal fluid (CSF) and blood samples of 28 CTD patients without neurological manifestation, 38 CTD patients with neurological manifestation (N-CTD), 38 non-inflammatory controls, and 38 multiple sclerosis (MS) patients, a paradigmatic primary neuroinflammatory disease. RESULTS: We detected an expansion of plasma cells in the blood of both N-CTD and CTD compared to non-inflammatory controls and MS. Blood plasma cells alone distinguished the clinically similar entities N-CTD and MS with high discriminatory performance (AUC: 0.81). Classical blood monocytes indicated higher disease activity in systemic lupus erythematosus (SLE) patients. Surprisingly, immune cells in the CSF did not differ significantly between N-CTD and CTD, while CD4+ T cells and the CD4+/CD8+ ratio were elevated in the blood of N-CTD compared to CTD. Several B cell-associated parameters partially overlapped in the CSF in MS and N-CTD. We built a machine learning model that distinguished N-CTD from MS with high discriminatory power using either blood or CSF. CONCLUSION: We here find that blood flow cytometry alone surprisingly suffices to distinguish CTD with neurological manifestations from clinically similar entities, suggesting that a rapid blood test could support clinicians in the differential diagnosis of N-CTD.

Stroke in Patients with Bacterial Meningitis: A Cohort Study and Meta-Analysis.

OBJECTIVE: The purpose of this study was to characterize patients with ischemic stroke due to bacterial meningitis. METHODS: In a single-center retrospective study, we analyzed 102 patients with bacterial meningitis of which 19 had an ischemic stroke. Clinical characteristics, cerebrospinal fluid (CSF) analyses, and spatiotemporal distribution of infarcts were assessed. In addition, we searched PubMed from database inception to August 2021 for observational studies on ischemic stroke in patients with bacterial meningitis, and performed a meta-analysis to investigate the frequency and timing of stroke as well as its effect on mortality. RESULTS: In our cohort, 15 (78.9%) patients with stroke had an modified Rankin scale (mRS)  ≥  3 at discharge compared to 33 (39.8%) in patients without stroke (p < 0.01). Of 1,692 patients with bacterial meningitis from 15 cohort studies included in our meta-analysis, cerebral infarcts were found in 332 (16%, 95% confidence interval [CI] = 0.13-0.20) patients. The occurrence of stroke was strongly associated with a higher mortality (odds ratio [OR] = 2.38, 95% CI = 1.70-3.34, p < 0.0001). There was no association of any specific causative pathogen with the occurrence of stroke. Infarcts were mainly distributed in territories of arteries located in the vicinity to the infection focus and peaked at 3 to -7 days and at 2 weeks after onset of meningitis. In patients with ischemic stroke, vasculopathy was found in 63.2% and additional intracerebral hemorrhage in 15.8%. INTERPRETATION: This study found that ischemic stroke due to bacterial meningitis is caused by cerebral vasculopathy located in the vicinity of the infection focus, and that the time course of infarctions might enable a therapeutic intervention. ANN NEUROL 2023;93:1094-1105.

Enhanced pathogenicity of Th17 cells due to natalizumab treatment: Implications for MS disease rebound.

After natalizumab (NAT) cessation, some multiple sclerosis (MS) patients experience a severe disease rebound. The rebound pathophysiology is still unclear; however, it has been linked to interleukin-17-producing T-helper (Th17) cells. We demonstrate that during NAT treatment, MCAM+CCR6+Th17 cells gradually acquire a pathogenic profile, including proinflammatory cytokine production, pathogenic transcriptional signatures, brain endothelial barrier impairment, and oligodendrocyte damage via induction of apoptotic pathways. This is accompanied by an increase in Th17 cell frequencies in the cerebrospinal fluid of NAT-treated patients. Notably, Th17 cells derived from NAT-treated patients, who later developed a disease rebound upon treatment cessation, displayed a distinct transcriptional pathogenicity profile associated with altered migratory properties. Accordingly, increased brain infiltration of patient Th17 cells was illustrated in a humanized mouse model and brain histology from a rebound patient. Therefore, peripheral blood-accumulated MCAM+CCR6+Th17 cells might be involved in rebound pathophysiology, and monitoring of changes in Th17 cell pathogenicity in patients before/during NAT treatment cessation might enable rebound risk assessment in the future.

A single-cell analysis framework allows for characterization of CSF leukocytes and their tissue of origin in multiple sclerosis.

Peripheral central nervous system (CNS)-infiltrating lymphocytes are a hallmark of relapsing-remitting multiple sclerosis. Tissue-resident memory T cells (TRM) not only populate the healthy CNS parenchyma but also are suspected to contribute to multiple sclerosis pathology. Because cerebrospinal fluid (CSF), unlike CNS parenchyma, is accessible for diagnostics, we evaluated whether human CSF, apart from infiltrating cells, also contains TRM cells and CNS-resident myeloid cells draining from the parenchyma or border tissues. Using deep generative models, we integrated 41 CSF and 14 CNS parenchyma single-cell RNA sequencing (scRNAseq) samples from eight independent studies, encompassing 120,629 cells. By comparing CSF immune cells collected during multiple sclerosis relapse with cells collected during therapeutic very late antigen-4 blockade, we could identify immune subsets with tissue provenance across multiple lineages, including CNS border-associated macrophages, CD8 and CD4 TRM cells, and tissue-resident natural killer cells. All lymphocytic CNS-resident cells shared expression of CXCR6 but showed differential ITGAE expression (encoding CD103). A common signature defined CD4 and CD8 TRM cells by expression of ZFP36L2, DUSP1, and ID2. We further developed a user interface-driven application based on this analysis framework for atlas-level cell identity transfer onto new CSF scRNAseq data. Together, these results define CNS-resident immune cells involved in multiple sclerosis pathology that can be detected and monitored in CSF. Targeting these cell populations might be promising to modulate immunopathology in progressive multiple sclerosis and other neuroinflammatory diseases.

Immunoadsorption versus double-dose methylprednisolone in refractory multiple sclerosis relapses.

OBJECTIVE: Intravenous methylprednisolone is the standard treatment for a multiple sclerosis relapse; however, this fails to improve symptoms in up to one quarter of patients. Immunoadsorption is an accepted treatment for refractory relapses, but prospective comparator-controlled studies are missing. METHODS: In this observational study, patients with steroid-refractory acute multiple sclerosis relapses receiving either six courses of tryptophan-immunoadsorption or double-dose methylprednisolone therapy were analysed. Outcomes were evaluated at discharge and three months later. Immune profiling of blood lymphocytes and proteomic analysis were performed by multi-parameter flow cytometry and Olink analysis, respectively (NCT04450030). RESULTS: 42 patients were enrolled (methylprednisolone: 26 patients; immunoadsorption: 16 patients). For determination of the primary outcome, treatment response was stratified according to relative function system score changes ("full/best" vs. "average" vs. "worse/none"). Upon discharge, the adjusted odds ratio for any treatment response ("full/best" + "average" vs. "worse/none") was 10.697 favouring immunoadsorption (p = 0.005 compared to methylprednisolone). At follow-up, the adjusted odds ratio for the best treatment response ("full/best" vs. "average" + "worse/none") was 103.236 favouring IA patients (p = 0.001 compared to methylprednisolone). Similar results were observed regarding evoked potentials and quality of life outcomes, as well as serum neurofilament light-chain levels. Flow cytometry revealed a profound reduction of B cell subsets following immunoadsorption, which was closely correlated to clinical outcomes, whereas methylprednisolone had a minimal effect on B cell populations. Immunoadsorption treatment skewed the blood cytokine network, reduced levels of B cell-related cytokines and reduced immunoglobulin levels as well as levels of certain coagulation factors. INTERPRETATION: Immunoadsorption demonstrated favourable outcomes compared to double-dose methylprednisolone. Outcome differences were significant at discharge and follow-up. Further analyses identified modulation of B cell function as a potential mechanism of action for immunoadsorption, as reduction of B cell subsets correlated with clinical improvement.

Intratumor heterogeneity and T cell exhaustion in primary CNS lymphoma.

BACKGROUND: Primary central nervous system lymphoma (PCNSL) is a rare lymphoma of the central nervous system, usually of diffuse large B cell phenotype. Stereotactic biopsy followed by histopathology is the diagnostic standard. However, limited material is available from CNS biopsies, thus impeding an in-depth characterization of PCNSL. METHODS: We performed flow cytometry, single-cell RNA sequencing, and B cell receptor sequencing of PCNSL cells released from biopsy material, blood, and cerebrospinal fluid (CSF), and spatial transcriptomics of biopsy samples. RESULTS: PCNSL-released cells were predominantly activated CD19+CD20+CD38+CD27+ B cells. In single-cell RNA sequencing, PCNSL cells were transcriptionally heterogeneous, forming multiple malignant B cell clusters. Hyperexpanded B cell clones were shared between biopsy- and CSF- but not blood-derived cells. T cells in the tumor microenvironment upregulated immune checkpoint molecules, thereby recognizing immune evasion signals from PCNSL cells. Spatial transcriptomics revealed heterogeneous spatial organization of malignant B cell clusters, mirroring their transcriptional heterogeneity across patients, and pronounced expression of T cell exhaustion markers, co-localizing with a highly malignant B cell cluster. CONCLUSIONS: Malignant B cells in PCNSL show transcriptional and spatial intratumor heterogeneity. T cell exhaustion is frequent in the PCNSL microenvironment, co-localizes with malignant cells, and highlights the potential of personalized treatments.

Alemtuzumab-induced immune phenotype and repertoire changes: implications for secondary autoimmunity.

Alemtuzumab is a monoclonal antibody that causes rapid depletion of CD52-expressing immune cells. It has proven to be highly efficacious in active relapsing-remitting multiple sclerosis; however, the high risk of secondary autoimmune disorders has greatly complicated its use. Thus, deeper insight into the pathophysiology of secondary autoimmunity and potential biomarkers is urgently needed. The most critical time points in the decision-making process for alemtuzumab therapy are before or at Month 12, where the ability to identify secondary autoimmunity risk would be instrumental. Therefore, we investigated components of blood and CSF of up to 106 multiple sclerosis patients before and after alemtuzumab treatment focusing on those critical time points. Consistent with previous reports, deep flow cytometric immune-cell profiling (n = 30) demonstrated major effects on adaptive rather than innate immunity, which favoured regulatory immune cell subsets within the repopulation. The longitudinally studied CSF compartment (n = 18) mainly mirrored the immunological effects observed in the periphery. Alemtuzumab-induced changes including increased numbers of naïve CD4+ T cells and B cells as well as a clonal renewal of CD4+ T- and B-cell repertoires were partly reminiscent of haematopoietic stem cell transplantation; in contrast, thymopoiesis was reduced and clonal renewal of T-cell repertoires after alemtuzumab was incomplete. Stratification for secondary autoimmunity did not show clear immununological cellular or proteomic traits or signatures associated with secondary autoimmunity. However, a restricted T-cell repertoire with hyperexpanded T-cell clones at baseline, which persisted and demonstrated further expansion at Month 12 by homeostatic proliferation, identified patients developing secondary autoimmune disorders (n = 7 without secondary autoimmunity versus n = 5 with secondary autoimmunity). Those processes were followed by an expansion of memory B-cell clones irrespective of persistence, which we detected shortly after the diagnosis of secondary autoimmune disease. In conclusion, our data demonstrate that (i) peripheral immunological alterations following alemtuzumab are mirrored by longitudinal changes in the CSF; (ii) incomplete T-cell repertoire renewal and reduced thymopoiesis contribute to a proautoimmune state after alemtuzumab; (iii) proteomics and surface immunological phenotyping do not identify patients at risk for secondary autoimmune disorders; (iv) homeostatic proliferation with disparate dynamics of clonal T- and B-cell expansions are associated with secondary autoimmunity; and (v) hyperexpanded T-cell clones at baseline and Month 12 may be used as a biomarker for the risk of alemtuzumab-induced autoimmunity.

Blockade of inhibitory killer cell immunoglobulin-like receptors and IL-2 triggering reverses the functional hypoactivity of tumor-derived NK-cells in glioblastomas.

Killer cell immunoglobulin-like receptors (KIRs) comprise a group of highly polymorphic inhibitory receptors which are specific for classical HLA class-I molecules. Peripheral blood and freshly prepared tumor cell suspensions (n = 60) as well as control samples (n = 32) were investigated for the distribution, phenotype, and functional relevance of CD158ab/KIR2DL1,-2/3 expressing NK-cells in glioblastoma (GBM) patients. We found that GBM were scarcely infiltrated by NK-cells that preferentially expressed CD158ab/KIR2DL1,-2/3 as inhibitory receptors, displayed reduced levels of the activating receptors CD335/NKp46, CD226/DNAM-1, CD159c/NKG2C, and showed diminished capacity to produce IFN-γ and perforin. Functional hypoactivity of GBM-derived NK-cells persisted despite IL-2 preactivation. Blockade with a specific KIR2DL-1,2/3 monoclonal antibody reversed NK-cell inhibition and significantly enhanced degranulation and IFN-γ production of IL-2 preactivated NK-cells in the presence of primary GBM cells and HLA-C expressing but not HLA class-I deficient K562 cells. Additional analysis revealed that significant amounts of IL-2 could be produced by tumor-derived CD4+ and CD8+CD45RA- memory T-cells after combined anti-CD3/anti-CD28 stimulation. Our data indicate that both blockade of inhibitory KIR and IL-2 triggering of tumor-derived NK-cells are necessary to enhance NK-cell responsiveness in GBM.

Impact of disease-modifying therapies on humoral and cellular immune-responses following SARS-CoV-2 vaccination in MS patients.

The impact of distinct disease-modifying therapies (DMTs) on severe acute respiratory syndrome-coronavirus 2 (SARS-CoV-2) vaccination efficacy in patients with multiple sclerosis (MS) is still enigmatic. In this prospective comparative study, we investigated humoral and cellular immune-responses in patients with MS receiving interferon beta, natalizumab, and ocrelizumab pre-vaccination and 6 weeks post second SARS-CoV-2 vaccination. Healthy individuals and interferon beta-treated patients generated robust humoral and cellular immune-responses. Although humoral immune responses were diminished in ocrelizumab-treated patients, cellular immune-responses were reduced in natalizumab-treated patients. Thus, both humoral and cellular immune responses should be closely monitored in patients on DMTs. Whereas patients with a poor cellular immune-response may benefit from additional vaccination cycles, patients with a diminished humoral immune-response may benefit from a treatment with SARS-CoV-2 antibodies in case of an infection.

Increased Intrathecal B and Plasma Cells in Patients With Anti-IgLON5 Disease: A Case Series.

BACKGROUND AND OBJECTIVES: Despite detection of autoantibodies, anti-IgLON5 disease was historically considered a tau-associated neurodegenerative disease, with limited treatment options and detrimental consequences for the patients. Observations in increasing case numbers hint toward underlying inflammatory mechanisms that, early detection provided, open a valuable window of opportunity for therapeutic intervention. We aimed to further substantiate this view by studying the CSF of patients with anti-IgLON5. METHODS: We identified 11 patients with anti-IgLON5 from our database and compared clinical, MRI, and CSF findings with a cohort of 20 patients with progressive supranuclear palsy (PSP) (as a noninflammatory tauopathy) and 22 patients with functional neurologic disorder. RESULTS: Patients with anti-IgLON5 show inflammatory changes in routine CSF analysis, an increase in B-lymphocyte frequency, and the presence of plasma cells in comparison to the PSP-control group and functional neurologic disease controls. Patients with intrathecal plasma cells showed a clinical response to rituximab. DISCUSSION: Our findings indicate the importance of inflammatory mechanisms, in particular in early and acute anti-IgLON5 cases, which may support the use of immune-suppressive treatments in these cases. The main limitation of the study is the small number of cases due to the rarity of the disease.

Dimethyl fumarate treatment restrains the antioxidative capacity of T cells to control autoimmunity.

Dimethyl fumarate, an approved treatment for relapsing-remitting multiple sclerosis, exerts pleiotropic effects on immune cells as well as CNS resident cells. Here, we show that dimethyl fumarate exerts a profound alteration of the metabolic profile of human CD4+ as well as CD8+ T cells and restricts their antioxidative capacities by decreasing intracellular levels of the reactive oxygen species scavenger glutathione. This causes an increase in mitochondrial reactive oxygen species levels accompanied by an enhanced mitochondrial stress response, ultimately leading to impaired mitochondrial function. Enhanced mitochondrial reactive oxygen species levels not only result in enhanced T-cell apoptosis in vitro as well as in dimethyl fumarate-treated patients, but are key for the well-known immunomodulatory effects of dimethyl fumarate both in vitro and in an animal model of multiple sclerosis, i.e. experimental autoimmune encephalomyelitis. Indeed, dimethyl fumarate immune-modulatory effects on T cells were completely abrogated by pharmacological interference of mitochondrial reactive oxygen species production. These data shed new light on dimethyl fumarate as bona fide immune-metabolic drug that targets the intracellular stress response in activated T cells, thereby restricting mitochondrial function and energetic capacity, providing novel insight into the role of oxidative stress in modulating cellular immune responses and T cell-mediated autoimmunity.

Impact of T cells on neurodegeneration in anti-GAD65 limbic encephalitis.

OBJECTIVE: Direct pathogenic effects of autoantibodies to the 65 kDa isoform of glutamic acid decarboxylase (GAD65) in autoimmune limbic encephalitis (LE) have been questioned due to its intracellular localization. We therefore hypothesized a pathogenic role for T cells. METHODS: We assessed magnet resonance imaging, neuropsychological and peripheral blood, and CSF flow cytometry data of 10 patients with long-standing GAD65-LE compared to controls in a cross-sectional manner. These data were related to each other within the GAD65-LE group and linked to neuropathological findings in selective hippocampectomy specimen from another two patients. In addition, full-resolution human leukocyte antigen (HLA) genotyping of all patients was performed. RESULTS: Compared to controls, no alteration in hippocampal volume but impaired memory function and elevated fractions of activated HLADR+ CD4+ and CD8+ T cells in peripheral blood and cerebrospinal fluid were found. Intrathecal fractions of CD8+ T cells negatively correlated with hippocampal volume and memory function, whereas the opposite was true for CD4+ T cells. Consistently, antigen-experienced CD8+ T cells expressed increased levels of the cytotoxic effector molecule perforin in peripheral blood, and perforin-expressing CD8+ T cells were found attached mainly to small interneurons but also to large principal neurons together with wide-spread hippocampal neurodegeneration. 6/10 LE patients harbored the HLA-A*02:01 allele known to present the immunodominant GAD65114-123 peptide in humans. INTERPRETATION: Our data suggest a pathogenic effect of CD8+ T cells and a regulatory effect of CD4+ T cells in patients with long-standing GAD65-LE.

Evaluation of Age-Dependent Immune Signatures in Patients With Multiple Sclerosis.

BACKGROUND AND OBJECTIVES: In MS, an age-related decline in disease activity and a decreased efficacy of disease-modifying treatment have been linked to immunosenescence, a state of cellular dysfunction associated with chronic inflammation. METHODS: To evaluate age-related immunologic alterations in MS, we compared immune signatures in peripheral blood (PB) and CSF by flow cytometry in patients with relapsing-remitting (RR) (PB n = 38; CSF n = 51) and primary progressive (PP) MS (PB n = 40; CSF n = 36) and respective controls (PB n = 40; CSF n = 85). RESULTS: Analysis revealed significant age-related changes in blood immune cell composition, especially in the CD8 T-cell compartment of healthy donors (HDs) and patients with MS. However, HDs displayed a strong age-dependent decline in the expression of the immunoregulatory molecules KLRG1, LAG3, and CTLA-4 on memory CD8 T cells, whereas this age-dependent reduction was completely abrogated in patients with MS. An age-dependent increase in the expression of the costimulatory molecule CD226 on memory CD8 T cells was absent in patients with MS. CD226 expression correlated with disability in younger (≤50 years) patients with MS. CSF analysis revealed a significant age-dependent decline in various immune cell populations in PPMS but not RRMS, suggesting a differential effect of aging on the intrathecal compartment in PPMS. DISCUSSION: Our data illustrate that aging in MS is associated with a dysbalance between costimulatory and immunoregulatory signals provided by CD8 T cells favoring a proinflammatory phenotype and, more importantly, a pattern of premature immune aging in the CD8 T-cell compartment of young patients with MS with potential implications for disease severity.

Resident human dermal γδT-cells operate as stress-sentinels: Lessons from the hair follicle.

Murine γδT-cells have stress-surveillance functions and are implicated in autoimmunity. Yet, whether human γδT-cells are also stress sentinels and directly promote autoimmune responses in the skin is unknown. Using a novel (mini-)organ assay, we tested if human dermis resident γδT-cells can recognize stressed human scalp hair follicles (HFs) to promote an alopecia areata (AA)-like autoimmune response. Accordingly, we show that γδT-cells from healthy human scalp skin are activated (CD69+), up-regulate the expression of NKG2D and IFN-γ, and become cytotoxic when co-cultured with autologous stressed HFs ex vivo. These autologous γδT-cells induce HF immune privilege collapse, dystrophy, and premature catagen, i.e. three hallmarks of the human autoimmune HF disorder, AA. This is mediated by CXCL12, MICA, and in part by IFN-γ and CD1d. In conclusion, human dermal γδT-cells exert physiological stress-sentinel functions in human skin, where their excessive activity can promote autoimmunity towards stressed HFs that overexpress CD1d, CXCL12, and/or MICA.

Amyotrophic lateral sclerosis patients show increased peripheral and intrathecal T-cell activation.

Several studies suggest a role for the peripheral immune system in the pathophysiology of amyotrophic lateral sclerosis. However, comprehensive studies investigating the intrathecal immune system in amyotrophic lateral sclerosis are rare. To elucidate whether compartment-specific inflammation contributes to amyotrophic lateral sclerosis pathophysiology, we here investigated intrathecal and peripheral immune profiles in amyotrophic lateral sclerosis patients and compared them with controls free of neurological disorders (controls) and patients with dementia or primary progressive multiple sclerosis. Routine CSF parameters were examined in 308 patients, including 132 amyotrophic lateral sclerosis patients. In a subgroup of 41 amyotrophic lateral sclerosis patients, extensive flow-cytometric immune cell profiling in peripheral blood and CSF was performed and compared with data from 26 controls, 25 dementia and 21 multiple sclerosis patients. Amyotrophic lateral sclerosis patients presented with significantly altered proportions of monocyte subsets in peripheral blood and increased frequencies of CD4+ and CD8+ T cells expressing the activation marker HLA-DR in peripheral blood (CD8+) and CSF (CD4+ and CD8+) compared with controls. While dementia and multiple sclerosis patients exhibited a comparable increase in intrathecal CD8+ T-cell activation, CD8+ T-cell activation in the peripheral blood in amyotrophic lateral sclerosis was higher than in multiple sclerosis patients. Furthermore, intrathecal CD4+ T-cell activation in amyotrophic lateral sclerosis surpassed levels in dementia patients. Intrathecal T-cell activation resulted from in situ activation rather than transmigration of activated T cells from the blood. While T-cell activation did not correlate with amyotrophic lateral sclerosis progression, patients with rapid disease progression showed reduced intrathecal levels of immune-regulatory CD56bright natural killer cells. The integration of these parameters into a composite score facilitated the differentiation of amyotrophic lateral sclerosis patients from patients of all other cohorts. To conclude, alterations in peripheral monocyte subsets, as well as increased peripheral and intrathecal activation of CD4+ and CD8+ T cells concomitant with diminished immune regulation by CD56bright natural killer cells, suggest an involvement of these immune cells in amyotrophic lateral sclerosis pathophysiology.