scholarly journals Dysregulated copper transport in multiple sclerosis may cause demyelination via astrocytes

2021 ◽  
Vol 118 (27) ◽  
pp. e2025804118
Author(s):  
Emanuela Colombo ◽  
Daniela Triolo ◽  
Claudia Bassani ◽  
Francesco Bedogni ◽  
Marco Di Dario ◽  
...  
Keyword(s):  
Multiple Sclerosis ◽  
Ex Vivo ◽  
Calcium Flux ◽  
Neurotrophin Receptor ◽  
Copper Uptake ◽  
Copper Transporter ◽  
Glia Cells ◽  
Uptake And Release ◽  
Intracellular Calcium Flux

Demyelination is a key pathogenic feature of multiple sclerosis (MS). Here, we evaluated the astrocyte contribution to myelin loss and focused on the neurotrophin receptor TrkB, whose up-regulation on the astrocyte finely demarcated chronic demyelinated areas in MS and was paralleled by neurotrophin loss. Mice lacking astrocyte TrkB were resistant to demyelination induced by autoimmune or toxic insults, demonstrating that TrkB signaling in astrocytes fostered oligodendrocyte damage. In vitro and ex vivo approaches highlighted that astrocyte TrkB supported scar formation and glia proliferation even in the absence of neurotrophin binding, indicating TrkB transactivation in response to inflammatory or toxic mediators. Notably, our neuropathological studies demonstrated copper dysregulation in MS and model lesions and TrkB-dependent expression of copper transporter (CTR1) on glia cells during neuroinflammation. In vitro experiments evidenced that TrkB was critical for the generation of glial intracellular calcium flux and CTR1 up-regulation induced by stimuli distinct from neurotrophins. These events led to copper uptake and release by the astrocyte, and in turn resulted in oligodendrocyte loss. Collectively, these data demonstrate a pathogenic demyelination mechanism via the astrocyte release of copper and open up the possibility of restoring copper homeostasis in the white matter as a therapeutic target in MS.

scholarly journals Anti-CD20 therapy depletes activated myelin-specific CD8+T cells in multiple sclerosis

2019 ◽  
Vol 116 (51) ◽  
pp. 25800-25807 ◽  
Author(s):  
Joseph J. Sabatino ◽  
Michael R. Wilson ◽  
Peter A. Calabresi ◽  
Stephen L. Hauser ◽  
Jonathan P. Schneck ◽  
...  
Keyword(s):  
Multiple Sclerosis ◽  
T Cells ◽  
T Cell ◽  
Ex Vivo ◽  
Antibody Therapy ◽  
Myelin Proteins ◽  
T Cell Epitopes ◽  
Control Subjects ◽  
Anti Cd20

CD8+T cells are believed to play an important role in multiple sclerosis (MS), yet their role in MS pathogenesis remains poorly defined. Although myelin proteins are considered potential autoantigenic targets, prior studies of myelin-reactive CD8+T cells in MS have relied on in vitro stimulation, thereby limiting accurate measurement of their ex vivo precursor frequencies and phenotypes. Peptide:MHC I tetramers were used to identify and validate 5 myelin CD8+T cell epitopes, including 2 newly described determinants in humans. The validated tetramers were used to measure the ex vivo precursor frequencies and phenotypes of myelin-specific CD8+T cells in the peripheral blood of untreated MS patients and HLA allele-matched healthy controls. In parallel, CD8+T cell responses against immunodominant influenza epitopes were also measured. There were no differences in ex vivo frequencies of tetramer-positive myelin-specific CD8+T cells between MS patients and control subjects. An increased proportion of myelin-specific CD8+T cells in MS patients exhibited a memory phenotype and expressed CD20 compared to control subjects, while there were no phenotypic differences observed among influenza-specific CD8+T cells. Longitudinal assessments were also measured in a subset of MS patients subsequently treated with anti-CD20 monoclonal antibody therapy. The proportion of memory and CD20+CD8+T cells specific for certain myelin but not influenza epitopes was significantly reduced following anti-CD20 treatment. This study, representing a characterization of unmanipulated myelin-reactive CD8+T cells in MS, indicates these cells may be attractive targets in MS therapy.

scholarly journals Signatures of immune reprogramming in anti-CD52 therapy of MS: markers for risk stratification and treatment response

2019 ◽  
Vol 1 (1) ◽  
Author(s):  
Laura Bierhansl ◽  
Tobias Ruck ◽  
Steffen Pfeuffer ◽  
Catharina C. Gross ◽  
Heinz Wiendl ◽  
...  
Keyword(s):  
Multiple Sclerosis ◽  
Adverse Events ◽  
Treatment Response ◽  
De Novo ◽  
Ex Vivo ◽  
Phase Iii ◽  
Serum Samples ◽  
Two Phase ◽  
Relapsing Remitting

Abstract Background Multiple sclerosis is one of the most prevalent neurological diseases in young adults affecting over 2 million people worldwide. Alemtuzumab is a highly effective therapy in relapsing remitting MS. Alemtuzumab is a monoclonal CD52 antibody that proved its efficacy against an active comparator (interferon [IFN]-β1a) in a phase II trial and two phase III trials regarding clinical and MRI outcomes. Nevertheless, the exact mode of action is still unknown. Alemtuzumab is commonly associated with secondary autoimmune disorders significantly affecting the risk-benefit ratio. Therefore, new biomarkers predicting treatment response and adverse events are urgently needed. This study aims to further elucidate the mechanism of action of the neuroprotective potential of alemtuzumab in relapsing-remitting multiple sclerosis (RRMS). Methods/Design This is a 3-year multicentre, explorative study including overall 150 patients comprising three different groups: (i) de novo patients prior and after alemtuzumab treatment initiation, (ii) patients under alemtuzumab treatment and (iii) patients requiring more than two alemtuzumab infusions. Peripheral blood and serum samples will be collected semi-annually for several in vitro/ex vivo assays to detect and characterize immune cells including their functional activity. Furthermore, data of MRI scans and disease-related impairment (using EDSS and MSFC), as well as the number and time of relapses, will be assessed. The clinical study is registered at clinicaltrials.gov (NCT04082260). Perspective Our study will provide deep insights into the underlying immunological changes in a longitudinal analysis of alemtuzumab treated RRMS patients. By combining clinical, radiological and functional immune-phenotype data, we will be able to identify biomarkers and/or immune signatures predicting treatment response and adverse events. Thereby, the understanding of the mechanisms of action of alemtuzumab will improve its efficacy and safety for present and future patients.

A GMCSF/Interleukin-15 Fusokine Leads to the Generation of a Novel Type of CD2/MHCII Immune Regulatory Cell with Potent Immune Suppressive Properties as Demonstrated in the EAE Mouse Model of Multiple Sclerosis.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 2295-2295
Author(s):  
Moutih Rafei ◽  
Jeremy Hsieh ◽  
Meng Yang Li ◽  
Simone Zehntner ◽  
Kathy Forner ◽  
...  
Keyword(s):  
Multiple Sclerosis ◽  
T Cells ◽  
Autoimmune Disease ◽  
Ex Vivo ◽  
Regulatory Cell ◽  
Autoreactive T Cell ◽  
Experimental Allergic Encephalitis ◽  
Over Time

Abstract Multiple sclerosis (MS) is an autoimmune disease characterised by the infiltration of autoreactive T-cell causing damages to the central nervous system. So far, interferon-β and glatiramer acetate are the only two immunomodulatory coumpounds that have been approved as non-curative disease managing strategies. Therefore, there is an urgent need for the development of novel efficient therapies that can be both safe and potent in inhibiting MS progression and promote reversal of disease state. We have recently published a report describing a novel synthetic GMCSF and IL15 Fusion Transgene (GIFT15) and have described its paradoxical and potent immune suppressive properties in vivo [Rafei et al., Blood (March 2007)]. Its mechanism of action relies on STAT3 hyperactivation arising from aberrant signalling taking place downstream of the IL15 receptor. We have now further studied the effect of GIFT15 on mouse spleen cells in vitro and here demonstrate that it leads to the conversion of murine T-cells to a novel suppressive regulatory cell type. Indeed, GIFT15-treated splenocytes (hereafter GIFT15 regs) shed their TCR and loose expression of CD3, CD4 and CD8, retain CD2 expression and acquire expression of MHC II. Distinct to classic T-regulatory cells, GIFT15 regs do not express CD25 or FOXP3. GIFT15 regs were able to suppress an in vitro two-way MLR by a contact-dependent mechanism as well as by the contemporaneous production of interleukin (IL)-10. Furthermore, GIFT15 regs were able to block antigen-specific activation of CD4-T-cells in response to autologous macrophage stimulation. As a proof-of-principle in vivo study, GIFT15 regs were injected intravenously in mice with pre-established experimental allergic encephalitis (EAE) and disease score was monitored over time. Interestingly, mice recovered significantly faster than controls following administration GIFT15 regs and a blockade in EAE progression was also noticed over time. In conclusion, our data suggests that GIFT15 can be used as a method to ex vivo generate suppressor cells of a new type which are distinct from classic Tregs or Tr1 cells. We propose that GIFT15 regs derived from autologous lymphocytes may be exploited for the treatment of autoimmune disease such as MS and may also be of use for other autoimmune ailments as well.

Are ex vivo neutralising antibodies against IFN-β always detrimental to therapeutic efficacy in multiple sclerosis?

2007 ◽  
Vol 13 (5) ◽  
pp. 616-621 ◽  
Author(s):  
PS Sorensen ◽  
N. Koch-Henriksen ◽  
K. Bendtzen
Keyword(s):  
Multiple Sclerosis ◽  
Treatment Effect ◽  
Cytopathic Effect ◽  
Ex Vivo ◽  
Affinity Maturation ◽  
Plasma Samples ◽  
Neutralising Antibodies ◽  
Relapse Rates

Neutralising antibodies (NAbs) against interferon (IFN)-β reduce the treatment effect in multiple sclerosis (MS). However, data from pivotal trials of IFN-β in MS suggest that NAb-positive patients may have a reduced relapse rate during the first six to 12 months of therapy. We collected clinical data and plasma samples for NAb measurements prospectively, every six months, in 468 patients treated with the same IFN-β preparation for at least 24 months. NAbs were measured blindly with a cytopathic effect (CPE) assay. During treatment months 0-6, patients who became NAb-positive had significantly fewer relapses compared to patients who maintained the NAb-negative status, whereas the opposite was observed after month 6. This is in accordance with observations in randomised studies of the three different IFN-β preparations, showing that patients who become NAb-positive have lower relapse rates during the first six or 12 months of therapy. We hypothesise that low affinity NAbs, present early after the start of IFN-β therapy, though neutralising in vitro in sensitive assays increase the half-life of IFN-β in vivo and, thereby, enhance the therapeutic effect. With affinity maturation, NAbs effectively prevent IFN-β binding to its receptors also in vivo and, hence, abolish the treatment effect. Multiple Sclerosis 2007; 13: 616-621. http://msj.sagepub.com

scholarly journals Hydroxycarboxylic Acid Receptor 2, a Pleiotropically Linked Receptor for the Multiple Sclerosis Drug, Monomethyl Fumarate. Possible Implications for the Inflammatory Response

2021 ◽  
Vol 12 ◽  
Author(s):  
Benedetta Parodi ◽  
Alessia Sanna ◽  
Alessia Cedola ◽  
Antonio Uccelli ◽  
Nicole Kerlero de Rosbo
Keyword(s):  
Multiple Sclerosis ◽  
Side Effects ◽  
Transforming Growth Factor Beta ◽  
Cell Activation ◽  
Transforming Growth Factor ◽  
Ex Vivo ◽  
Acid Receptor ◽  
Hydroxycarboxylic Acid ◽  
Monomethyl Fumarate

Monomethyl fumarate (MMF), metabolite of dimethyl fumarate (DMF), an immunosuppressive drug approved for the treatment of multiple sclerosis (MS), is a potent agonist for hydroxycarboxylic acid receptor 2 (HCAR2), eliciting signals that dampen cell activation or lead to inflammation such as the skin flushing reaction that is one of the main side effects of the treatment, together with gastrointestinal inflammation. Our aim is to further understand the molecular basis underlying these differential effects of the drug. We have used wild-type and HCAR2 knock-out mice to investigate, in vitro and ex vivo under steady-state and pathological conditions, the HCAR2-mediated signaling pathways activated by MMF in dendritic cells (DC), which promote differentiation of T cells, and in intestinal epithelial cells (IEC) where activation of a pro-inflammatory pathway, such as the cyclooxygenase-2 pathway involved in skin flushing, could underlie gastrointestinal side effects of the drug. To understand how DMF treatment might impact on gut inflammation induced by experimental autoimmune encephalomyelitis (EAE), the animal model for MS, we have used 3D X-ray phase contrast tomography and flow cytometry to monitor possible intestinal alterations at morphological and immunological levels, respectively. We show that HCAR2 is a pleiotropically linked receptor for MMF, mediating activation of different pathways leading to different outcomes in different cell types, depending on experimental in-vitro and in-vivo conditions. In the small intestine of EAE-affected mice, DMF treatment affected migration of tolerogenic DC from lamina propria to mesenteric lymph nodes, and/or reverted their profile to pro-inflammatory, probably as a result of reduced expression of aldehyde dehydrogenase and transforming growth factor beta as well as the inflammatory environment. Nevertheless, DMF treatment did not amplify the morphological alterations induced by EAE. On the basis of our further understanding of MMF signaling through HCAR2, we suggest that the pleiotropic signaling of fumarate via HCAR2 should be addressed for its pharmaceutical relevance in devising new lead compounds with reduced inflammatory side effects.

In vitro and ex vivo models of multiple sclerosis

2016 ◽  
Vol 21 (9) ◽  
pp. 1504-1511 ◽  
Author(s):  
Martin Madill ◽  
Denise Fitzgerald ◽  
Kara E. O’Connell ◽  
Kumlesh K. Dev ◽  
Sanbing Shen ◽  
...  
Keyword(s):  
Multiple Sclerosis ◽  
Ex Vivo

scholarly journals Growth and Differentiation of Circulating Stem Cells After Extensive Ex Vivo Expansion

2021 ◽  
Author(s):  
Silvia Barbon ◽  
Senthilkumar Rajendran ◽  
Thomas Bertalot ◽  
Monica Piccione ◽  
Marco Gasparella ◽  
...  
Keyword(s):  
Stem Cells ◽  
Cell Therapy ◽  
Muscle Damage ◽  
Adult Stem Cells ◽  
Ex Vivo ◽  
Myogenic Differentiation ◽  
Calcium Flux ◽  
Morphological Studies

Abstract Background: Stem cell therapy is gaining momentum as an effective treatment strategy for degenerative diseases. Adult stem cells isolated from various sources (i.e., cord blood, bone marrow, adipose tissue) are being considered as a realistic option due to their well-documented therapeutic potentials. Our previous studies standardized a method to isolate circulating multipotent cells (CMCs) that are able to sustain long term in vitro culture and differentiate towards mesodermal lineages. Methods: In this work, long-term cultures of CMCs were stimulated to study in vitro neuronal and myogenic differentiation. After induction, cells were analysed at different time points. Morphological studies were performed by scanning electron microscopy and specific neuronal and myogenic marker expression were evaluated using RT-PCR, flow cytometry and western blot. For myogenic plasticity study, CMCs were transplanted into in vivo model of chemically-induced muscle damage. Results: After neurogenic induction, CMCs showed characteristic dendrite-like morphology and expressed specific neuronal markers both at mRNA and protein level. The calcium flux activity of CMCs under stimulation with potassium chloride and the secretion of noradrenalin confirmed their ability to acquire a functional phenotype. In parallel, the myogenic potential of CMCs was confirmed by their ability to form syncytium-like structures in vitro and express myogenic markers both at early and late phases of differentiation. Interestingly, in a rat model of bupivacaine-induced muscle damage, CMCs integrated within the host tissue taking part in tissue repair. Conclusion: Overall, collected data demonstrated long-term cultured CMCs retain proliferative and differentiative potentials suggesting to be a good candidate for cell therapy.

Doxorubicin impairs cardiomyocyte viability by suppressing transcription factor EB expression and disrupting autophagy

2016 ◽  
Vol 473 (21) ◽  
pp. 3769-3789 ◽  
Author(s):  
Jordan J. Bartlett ◽  
Purvi C. Trivedi ◽  
Pollen Yeung ◽  
Petra C. Kienesberger ◽  
Thomas Pulinilkunnil
Keyword(s):  
Cell Death ◽  
Transcription Factor ◽  
Cathepsin B ◽  
Ex Vivo ◽  
Calcium Flux ◽  
Autophagic Flux ◽  
Mitochondrial Energy Metabolism ◽  
Transcription Factor Eb

Doxorubicin (DOX) is an effective anti-cancer agent. However, DOX treatment increases patient susceptibility to dilated cardiomyopathy. DOX predisposes cardiomyocytes to insult by suppressing mitochondrial energy metabolism, altering calcium flux, and disrupting proteolysis and proteostasis. Prior studies have assessed the role of macroautophagy in DOX cardiotoxicity; however, limited studies have examined whether DOX mediates cardiac injury through dysfunctions in inter- and/or intra-lysosomal signaling events. Lysosomal signaling and function is governed by transcription factor EB (TFEB). In the present study, we hypothesized that DOX caused myocyte injury by impairing lysosomal function and signaling through negative regulation of TFEB. Indeed, we found that DOX repressed cellular TFEB expression, which was associated with impaired cathepsin proteolytic activity across in vivo, ex vivo, and in vitro models of DOX cardiotoxicity. Furthermore, we observed that loss of TFEB was associated with reduction in macroautophagy protein expression, inhibition of autophagic flux, impairments in lysosomal cathepsin B activity, and activation of cell death. Restoration and/or activation of TFEB in DOX-treated cardiomyocytes prevented DOX-induced suppression of cathepsin B activity, reduced DOX-mediated reactive oxygen species (ROS) overproduction, attenuated activation of caspase-3, and improved cellular viability. Collectively, loss of TFEB inhibits lysosomal autophagy, rendering cardiomyocytes susceptible to DOX-induced proteotoxicity and injury. Our data reveal a novel mechanism wherein DOX primes cardiomyocytes for cell death by depleting cellular TFEB.

TIRC7 and HLA-DR axis contributes to inflammation in multiple sclerosis

2014 ◽  
Vol 20 (9) ◽  
pp. 1171-1181 ◽  
Author(s):  
JM Frischer ◽  
M Reindl ◽  
B Künz ◽  
T Berger ◽  
S Schmidt ◽  
...  
Keyword(s):  
Multiple Sclerosis ◽  
T Cells ◽  
T Cell ◽  
Cell Clone ◽  
Ex Vivo ◽  
Peripheral Blood Lymphocytes ◽  
Cytokine Expression ◽  
Hla Dr ◽  
T Cell Clone

Background and objective: Interactions between TIRC7 (a novel seven-transmembrane receptor on activated lymphocytes) and its ligand HLA-DR might be involved in the inflammatory process in multiple sclerosis (MS). Methods: Methods comprised immunohistochemistry and microscopy on archival MS autopsies, proliferation-, cytokine-, and surface-staining assays using peripheral blood lymphocytes (PBLs) from MS patients and an in vitro model. Results: TIRC7 was expressed in brain-infiltrating lymphocytes and strongly correlated with disease activity in MS. TIRC7 expression was reduced in T cells and induced in B cells in PBLs obtained from MS patients. After ex vivo activation, T cell expression of TIRC7 was restored in patients with active MS disease. The interaction of TIRC7+ T lymphocytes with cells expressing HLA-DR on their surface led to T cell proliferation and activation whereas an anti-TIRC7 mAb preventing interactions with its ligand inhibited proliferation and Th1 and Th17 cytokine expression in T cells obtained from MS patients and in myelin basic protein-specific T cell clone. Conclusion: Our findings suggest that TIRC7 is involved in inflammation in MS and anti-TIRC7 mAb can prevent immune activation via selective inhibition of Th1- and Th17-associated cytokine expression. This targeting approach may become a novel treatment option for MS.

scholarly journals A dual effect of ursolic acid to the treatment of multiple sclerosis through both immunomodulation and direct remyelination

2020 ◽  
Vol 117 (16) ◽  
pp. 9082-9093 ◽  
Author(s):  
Yuan Zhang ◽  
Xing Li ◽  
Bogoljub Ciric ◽  
Mark T. Curtis ◽  
Wan-Jun Chen ◽  
...  
Keyword(s):  
Multiple Sclerosis ◽  
Ursolic Acid ◽  
Ex Vivo ◽  
Oral Treatment ◽  
Acute Stage ◽  
Peroxisome Proliferator ◽  
Cns Inflammation ◽  
Neural Repair

Current multiple sclerosis (MS) medications are mainly immunomodulatory, having little or no effect on neuroregeneration of damaged central nervous system (CNS) tissue; they are thus primarily effective at the acute stage of disease, but much less so at the chronic stage. An MS therapy that has both immunomodulatory and neuroregenerative effects would be highly beneficial. Using multiple in vivo and in vitro strategies, in the present study we demonstrate that ursolic acid (UA), an antiinflammatory natural triterpenoid, also directly promotes oligodendrocyte maturation and CNS myelin repair. Oral treatment with UA significantly decreased disease severity and CNS inflammation and demyelination in experimental autoimmune encephalomyelitis (EAE), an animal model of MS. Importantly, remyelination and neural repair in the CNS were observed even after UA treatment was started on day 60 post immunization when EAE mice had full-blown demyelination and axonal damage. UA treatment also enhanced remyelination in a cuprizone-induced demyelination model in vivo and brain organotypic slice cultures ex vivo and promoted oligodendrocyte maturation in vitro, indicating a direct myelinating capacity. Mechanistically, UA induced promyelinating neurotrophic factor CNTF in astrocytes by peroxisome proliferator-activated receptor γ(PPARγ)/CREB signaling, as well as by up-regulation of myelin-related gene expression during oligodendrocyte maturation via PPARγ activation. Together, our findings demonstrate that UA has significant potential as an oral antiinflammatory and neural repair agent for MS, especially at the chronic-progressive stage.

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