Publications by authors named "Manon Blain"

22 Publications

  • Page 1 of 1

Size and ligand effects of gold nanoclusters in alteration of organellar state and translocation of transcription factors in human primary astrocytes.

Nanoscale 2021 Feb;13(5):3173-3183

Department of Pharmacology & Therapeutics, McGill University, Montréal, QC H3G 1Y6, Canada.

Ultra-small gold nanoclusters (AuNCs) with designed sizes and ligands are gaining popularity for biomedical purposes and ultimately for human imaging and therapeutic applications. Human non-tumor brain cells, astrocytes, are of particular interest because they are abundant and play a role in functional regulation of neurons under physiological and pathological conditions. Human primary astrocytes were treated with AuNCs of varying sizes (Au10, Au15, Au18, Au25) and ligand composition (glutathione, polyethylene glycol, N-acetyl cysteine). Concentration and time-dependent studies showed no significant cell loss with AuNC concentrations <10 μM. AuNC treatment caused marked differential astrocytic responses at the organellar and transcription factor level. The effects were exacerbated under severe oxidative stress induced by menadione. Size-dependent effects were most remarkable with the smallest and largest AuNCs (10, 15 Au atoms versus 25 Au atoms) and might be related to the accessibility of biological targets toward the AuNC core, as demonstrated by QM/MM simulations. In summary, these findings suggest that AuNCs are not inert in primary human astrocytes, and that their sizes play a critical role in modulation of organellar and redox-responsive transcription factor homeostasis.
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http://dx.doi.org/10.1039/d0nr06401gDOI Listing
February 2021

Vitamin D Regulates MerTK-Dependent Phagocytosis in Human Myeloid Cells.

J Immunol 2020 Jul 15;205(2):398-406. Epub 2020 Jun 15.

Neuroimmunology Unit, Department of Neurology and Neurosurgery, Montreal Neurological Institute, McGill University, Montreal, Quebec H3A 0G4, Canada;

Vitamin D deficiency is a major environmental risk factor for the development of multiple sclerosis. The major circulating metabolite of vitamin D (25-hydroxyvitamin D) is converted to the active form (calcitriol) by the hydroxylase enzyme In multiple sclerosis lesions, the tyrosine kinase MerTK expressed by myeloid cells regulates phagocytosis of myelin debris and apoptotic cells that can accumulate and inhibit tissue repair and remyelination. In this study, we explored the effect of calcitriol on homeostatic (M-CSF, TGF-β-treated) and proinflammatory (GM-CSF-treated) human monocyte-derived macrophages and microglia using RNA sequencing. Transcriptomic analysis revealed significant calcitriol-mediated effects on both Ag presentation and phagocytosis pathways. Calcitriol downregulated MerTK mRNA and protein expression in both myeloid populations, resulting in reduced capacity of these cells to phagocytose myelin and apoptotic T cells. Proinflammatory myeloid cells expressed high levels of compared with homeostatic myeloid cells. Only proinflammatory cells in the presence of TNF-α generated calcitriol from 25-hydroxyvitamin D, resulting in repression of MerTK expression and function. This selective production of calcitriol in proinflammatory myeloid cells has the potential to reduce the risk for autoantigen presentation while retaining the phagocytic ability of homeostatic myeloid cells.
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http://dx.doi.org/10.4049/jimmunol.2000129DOI Listing
July 2020

Metabolic Control of Astrocyte Pathogenic Activity via cPLA2-MAVS.

Cell 2019 12 5;179(7):1483-1498.e22. Epub 2019 Dec 5.

Ann Romney Center for Neurologic Diseases, Brigham and Women's Hospital, Harvard Medical School, Boston, MA 02115, USA; Broad Institute of MIT and Harvard, Cambridge, MA 02142, USA. Electronic address:

Metabolism has been shown to control peripheral immunity, but little is known about its role in central nervous system (CNS) inflammation. Through a combination of proteomic, metabolomic, transcriptomic, and perturbation studies, we found that sphingolipid metabolism in astrocytes triggers the interaction of the C2 domain in cytosolic phospholipase A2 (cPLA2) with the CARD domain in mitochondrial antiviral signaling protein (MAVS), boosting NF-κB-driven transcriptional programs that promote CNS inflammation in experimental autoimmune encephalomyelitis (EAE) and, potentially, multiple sclerosis. cPLA2 recruitment to MAVS also disrupts MAVS-hexokinase 2 (HK2) interactions, decreasing HK enzymatic activity and the production of lactate involved in the metabolic support of neurons. Miglustat, a drug used to treat Gaucher and Niemann-Pick disease, suppresses astrocyte pathogenic activities and ameliorates EAE. Collectively, these findings define a novel immunometabolic mechanism that drives pro-inflammatory astrocyte activities, outlines a new role for MAVS in CNS inflammation, and identifies candidate targets for therapeutic intervention.
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http://dx.doi.org/10.1016/j.cell.2019.11.016DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6936326PMC
December 2019

Astrocytes in the Pathogenesis of Multiple Sclerosis: An In Situ MicroRNA Study.

J Neuropathol Exp Neurol 2019 12;78(12):1130-1146

Department of Pathology, The Ottawa Hospital, University of Ottawa.

Astrocytes are increasingly recognized as active contributors to the disease process in multiple sclerosis (MS), rather than being merely reactive. We investigated the expression of a selected microRNA (miRNA) panel that could contribute both to the injury and to the recovery phases of the disease. Individual astrocytes were laser microdissected from brain sections. We then compared the miRNAs' expressions in MS and control brain samples at different lesional stages in white versus grey matter regions. In active MS lesions, we found upregulation of ischemia-related miRNAs in white but not grey matter, often with reversion to the normal state in inactive lesions. In contrast to our previous findings on MS macrophages, expression of 2 classical inflammatory-related miRNAs, miRNA-155 and miRNA-146a, was reduced in astrocytes from active and chronic active MS lesions in white and grey matter, suggesting a lesser direct pathogenetic role for these miRNAs in astrocytes. miRNAs within the categories regulating aquaporin4 (-100, -145, -320) and glutamate transport/apoptosis/neuroprotection (-124a, -181a, and -29a) showed some contrasting responses. The regional and lesion-stage differences of expression of these miRNAs indicate the remarkable ability of astrocytes to show a wide range of selective responses in the face of differing insults and phases of resolution.
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http://dx.doi.org/10.1093/jnen/nlz098DOI Listing
December 2019

Environmental Control of Astrocyte Pathogenic Activities in CNS Inflammation.

Cell 2019 01 17;176(3):581-596.e18. Epub 2019 Jan 17.

Ann Romney Center for Neurologic Diseases, Brigham and Women's Hospital, Harvard Medical School, Boston, MA 02115, USA; Broad Institute of MIT and Harvard, Cambridge, MA 02142, USA. Electronic address:

Genome-wide studies have identified genetic variants linked to neurologic diseases. Environmental factors also play important roles, but no methods are available for their comprehensive investigation. We developed an approach that combines genomic data, screens in a novel zebrafish model, computational modeling, perturbation studies, and multiple sclerosis (MS) patient samples to evaluate the effects of environmental exposure on CNS inflammation. We found that the herbicide linuron amplifies astrocyte pro-inflammatory activities by activating signaling via sigma receptor 1, inositol-requiring enzyme-1α (IRE1α), and X-box binding protein 1 (XBP1). Indeed, astrocyte-specific shRNA- and CRISPR/Cas9-driven gene inactivation combined with RNA-seq, ATAC-seq, ChIP-seq, and study of patient samples suggest that IRE1α-XBP1 signaling promotes CNS inflammation in experimental autoimmune encephalomyelitis (EAE) and, potentially, MS. In summary, these studies define environmental mechanisms that control astrocyte pathogenic activities and establish a multidisciplinary approach for the systematic investigation of the effects of environmental exposure in neurologic disorders.
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http://dx.doi.org/10.1016/j.cell.2018.12.012DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6440749PMC
January 2019

Helper CD4 T cells expressing granzyme B cause glial fibrillary acidic protein fragmentation in astrocytes in an MHCII-independent manner.

Glia 2019 04 16;67(4):582-593. Epub 2018 Nov 16.

Department of Exercise Science, Department of Biology, PERFORM Centre, Concordia University, Montréal, Quebec, Canada.

During inflammatory processes of the central nervous system, helper T cells have the capacity to cross the blood-brain barrier and injure or kill neural cells through cytotoxic mechanisms. Glial fibrillary acidic protein (GFAP) is an intermediate filament protein that is part of the astrocyte cytoskeleton that can become fragmented in neuroinflammatory conditions. The mechanism of action by which helper T cells with cytotoxic properties injure astrocytes is not completely understood. Primary human astrocytes were obtained from fetal brain tissue. Human helper (CD4 ) T cells were isolated from peripheral blood mononuclear cells and activated with the superantigen staphylococcal enterotoxin E (SEE). Granzyme B was detected by enzyme linked immunosorbent assay and intracellular flow cytometry. GFAP fragmentation was monitored by western blotting. Cell death was monitored by lactic acid dehydrogenase release and terminal biotin-dUTP nick labeling (TUNEL). Astrocyte migration was monitored by scratch assay. Adult human oligodendrocytes were cultured with sublethally injured astrocytes to determine support function. Helper T cells activated with SEE expressed granzyme B but not perforin. Helper T cells released granzyme B upon contact with astrocytes and caused GFAP fragmentation in a caspase-dependent, MHCII-independent manner. Sublethally injured astrocytes were not apoptotic; however, their processes were thin and elongated, their migration was attenuated, and their ability to support oligodendrocytes was reduced in vitro. Helper T cells can release granzyme B causing sublethal injury to astrocytes, which compromises the supportive functions of astrocytes. Blocking these pathways may lead to improved resolution of neuroinflammatory lesions.
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http://dx.doi.org/10.1002/glia.23503DOI Listing
April 2019

Microglial control of astrocytes in response to microbial metabolites.

Nature 2018 05 16;557(7707):724-728. Epub 2018 May 16.

Ann Romney Center for Neurologic Diseases, Brigham and Women's Hospital, Harvard Medical School, Boston, MA, USA.

Microglia and astrocytes modulate inflammation and neurodegeneration in the central nervous system (CNS). Microglia modulate pro-inflammatory and neurotoxic activities in astrocytes, but the mechanisms involved are not completely understood. Here we report that TGFα and VEGF-B produced by microglia regulate the pathogenic activities of astrocytes in the experimental autoimmune encephalomyelitis (EAE) mouse model of multiple sclerosis. Microglia-derived TGFα acts via the ErbB1 receptor in astrocytes to limit their pathogenic activities and EAE development. Conversely, microglial VEGF-B triggers FLT-1 signalling in astrocytes and worsens EAE. VEGF-B and TGFα also participate in the microglial control of human astrocytes. Furthermore, expression of TGFα and VEGF-B in CD14 cells correlates with the multiple sclerosis lesion stage. Finally, metabolites of dietary tryptophan produced by the commensal flora control microglial activation and TGFα and VEGF-B production, modulating the transcriptional program of astrocytes and CNS inflammation through a mechanism mediated by the aryl hydrocarbon receptor. In summary, we identified positive and negative regulators that mediate the microglial control of astrocytes. Moreover, these findings define a pathway through which microbial metabolites limit pathogenic activities of microglia and astrocytes, and suppress CNS inflammation. This pathway may guide new therapies for multiple sclerosis and other neurological disorders.
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http://dx.doi.org/10.1038/s41586-018-0119-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6422159PMC
May 2018

Comparative morphology and phagocytic capacity of primary human adult microglia with time-lapse imaging.

J Neuroimmunol 2017 09 22;310:143-149. Epub 2017 May 22.

Department of Exercise Science, Concordia University, Montréal, QC H4B 1R6, Canada; Department of Biology, Concordia University, Canada; PERFORM Centre, Concordia University, Canada; Center for Structural and Functional Genomics, Concordia University, Canada. Electronic address:

Microglia provide immune surveillance within the brain and spinal cord. Various microglial morphologies include ramified, amoeboid, and pseudopodic. The link between form and function is not clear, especially for human adult microglia which are limited in availability for study. Here, we examined primary human microglia isolated from normal-appearing white matter. Pseudopodic and amoeboid microglia were effective phagocytes, taking up E. coli bioparticles using ruffled cell membrane sheets and retrograde transport. Pseudopodic and amoeboid microglia were more effective phagocytes as compared to ramified microglia or monocyte-derived dendritic cells. Thus, amoeboid and pseudopodic microglia may both be effective as brain scavengers.
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http://dx.doi.org/10.1016/j.jneuroim.2017.05.012DOI Listing
September 2017

Sphingosine 1-phosphate receptor modulation suppresses pathogenic astrocyte activation and chronic progressive CNS inflammation.

Proc Natl Acad Sci U S A 2017 02 6;114(8):2012-2017. Epub 2017 Feb 6.

Ann Romney Center for Neurologic Diseases, Brigham and Women's Hospital, Harvard Medical School, Boston, MA 02115;

Multiple sclerosis (MS) is an autoimmune inflammatory demyelinating disease of the CNS that causes disability in young adults as a result of the irreversible accumulation of neurological deficits. Although there are potent disease-modifying agents for its initial relapsing-remitting phase, these therapies show limited efficacy in secondary progressive MS (SPMS). Thus, there is an unmet clinical need for the identification of disease mechanisms and potential therapeutic approaches for SPMS. Here, we show that the sphingosine 1-phosphate receptor (S1PR) modulator fingolimod (FTY720) ameliorated chronic progressive experimental autoimmune encephalomyelitis in nonobese diabetic mice, an experimental model that resembles several aspects of SPMS, including neurodegeneration and disease progression driven by the innate immune response in the CNS. Indeed, S1PR modulation by FTY720 in murine and human astrocytes suppressed neurodegeneration-promoting mechanisms mediated by astrocytes, microglia, and CNS-infiltrating proinflammatory monocytes. Genome-wide studies showed that FTY720 suppresses transcriptional programs associated with the promotion of disease progression by astrocytes. The study of the molecular mechanisms controlling these transcriptional modules may open new avenues for the development of therapeutic strategies for progressive MS.
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http://dx.doi.org/10.1073/pnas.1615413114DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5338419PMC
February 2017

Type I interferons and microbial metabolites of tryptophan modulate astrocyte activity and central nervous system inflammation via the aryl hydrocarbon receptor.

Nat Med 2016 06 9;22(6):586-97. Epub 2016 May 9.

Ann Romney Center for Neurologic Diseases, Brigham and Women's Hospital, Harvard Medical School, Boston, Massachusetts, USA.

Astrocytes have important roles in the central nervous system (CNS) during health and disease. Through genome-wide analyses we detected a transcriptional response to type I interferons (IFN-Is) in astrocytes during experimental CNS autoimmunity and also in CNS lesions from patients with multiple sclerosis (MS). IFN-I signaling in astrocytes reduces inflammation and experimental autoimmune encephalomyelitis (EAE) disease scores via the ligand-activated transcription factor aryl hydrocarbon receptor (AHR) and the suppressor of cytokine signaling 2 (SOCS2). The anti-inflammatory effects of nasally administered interferon (IFN)-β are partly mediated by AHR. Dietary tryptophan is metabolized by the gut microbiota into AHR agonists that have an effect on astrocytes to limit CNS inflammation. EAE scores were increased following ampicillin treatment during the recovery phase, and CNS inflammation was reduced in antibiotic-treated mice by supplementation with the tryptophan metabolites indole, indoxyl-3-sulfate, indole-3-propionic acid and indole-3-aldehyde, or the bacterial enzyme tryptophanase. In individuals with MS, the circulating levels of AHR agonists were decreased. These findings suggest that IFN-Is produced in the CNS function in combination with metabolites derived from dietary tryptophan by the gut flora to activate AHR signaling in astrocytes and suppress CNS inflammation.
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http://dx.doi.org/10.1038/nm.4106DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4899206PMC
June 2016

Regulation of astrocyte activation by glycolipids drives chronic CNS inflammation.

Nat Med 2014 Oct 14;20(10):1147-56. Epub 2014 Sep 14.

Center for Neurologic Diseases, Brigham and Women's Hospital, Harvard Medical School, Boston, Massachusetts, USA.

Astrocytes have complex roles in health and disease, thus it is important to study the pathways that regulate their function. Here we report that lactosylceramide (LacCer) synthesized by β-1,4-galactosyltransferase 6 (B4GALT6) is upregulated in the central nervous system (CNS) of mice during chronic experimental autoimmune encephalomyelitis (EAE), a model of multiple sclerosis (MS). LacCer acts in an autocrine manner to control astrocyte transcriptional programs that promote neurodegeneration. In addition, LacCer in astrocytes controls the recruitment and activation of microglia and CNS-infiltrating monocytes in a non-cell autonomous manner by regulating production of the chemokine CCL2 and granulocyte-macrophage colony-stimulating factor (GM-CSF), respectively. We also detected high B4GALT6 gene expression and LacCer concentrations in CNS MS lesions. Inhibition of LacCer synthesis in mice suppressed local CNS innate immunity and neurodegeneration in EAE and interfered with the activation of human astrocytes in vitro. Thus, B4GALT6 regulates astrocyte activation and is a potential therapeutic target for MS and other neuroinflammatory disorders.
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http://dx.doi.org/10.1038/nm.3681DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4255949PMC
October 2014

Isolating, culturing, and polarizing primary human adult and fetal microglia.

Methods Mol Biol 2013 ;1041:199-211

McGill University, Montreal, Canada.

Microglia are an important component of the innate immune system within the central nervous system (CNS). Isolation and in vitro culturing of microglia can provide insight towards the basic biology of these cells as well as their interactions with neurons, astrocytes, and oligodendrocytes. While studies of rodent microglia and microglial cell lines have provided a basis for our understanding of these cells, human adult microglia exhibit distinct properties when compared to rodent cells. Furthermore, the study of human fetal microglia provides a window into the developing CNS. This chapter describes the protocols used to isolate, purify, and culture both human adult and fetal microglia. Under basal culture conditions, human microglia survive for extended periods in the absence of growth factors, thus allowing their properties to be investigated under resting conditions. In addition, both human adult and fetal microglia can be used to study how they respond to different polarization conditions. As is the case with macrophages, it is also possible to polarize microglia towards the pro-inflammatory "M1" and the anti-inflammatory "M2" phenotypes, as described in this chapter.
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http://dx.doi.org/10.1007/978-1-62703-520-0_19DOI Listing
January 2014

Reduction of the peripheral blood CD56(bright) NK lymphocyte subset in FTY720-treated multiple sclerosis patients.

J Immunol 2011 Jul 27;187(1):570-9. Epub 2011 May 27.

Neuroimmunology Unit, Montreal Neurological Institute, McGill University, Montreal, Quebec H3A 2B4, Canada.

FTY720 (fingolimod) treatment of multiple sclerosis (MS) results in lymphopenia due to increased recruitment into and decreased egress from secondary lymphoid organs of CCR7(+) lymphocytes. Although absolute numbers of NK lymphocytes were reported as being unaltered in FTY720-treated MS patients (MS-FTY), such analyses did not detect a change in a minor subset. Because expression of CCR7 has been described on CD56(bright) NK cells, a minority population of NK cells, we investigated the effect of FTY720 treatment on the phenotype and function of human NK cells in the peripheral circulation of MS patients. MS-FTY patients displayed a decreased proportion of peripheral CD56(bright)CD62L(+)CCR7(+) NK cells compared with untreated MS and healthy donors. In vitro treatment with FTY720-P increased migration of untreated donor NK cells to CXCL12 while reducing the response to CX3CL1 with similar migration responses seen in NK cells from MS-FTY patients. FTY720-P inhibited sphingosine 1-phosphate-directed migration of CD56(bright) and CD56(dim) NK cells subsets from untreated healthy donors. IL-12- and IL-15-stimulated NK cells from MS-FTY patients displayed similar capacity to produce IFN-γ, TNF, IL-10, and MIP-1α cytokines/chemokines compared with NK cells from untreated healthy donors and displayed comparable levels of degranulation in response to K562 tumor cells compared with untreated donors. Subset alterations and function of NK cell populations will need to be considered as part of assessing overall immunosurveillance capacity of patients with MS who will receive sustained FTY720 therapy.
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http://dx.doi.org/10.4049/jimmunol.1003823DOI Listing
July 2011

Differential responses of human microglia and blood-derived myeloid cells to FTY720.

J Neuroimmunol 2011 Jan 9;230(1-2):10-6. Epub 2010 Sep 9.

Neuroimmunology Unit, Montréal Neurological Institute, McGill University, Montréal, Québec, Canada.

Human microglia, monocyte-derived dendritic cells (DCs) and macrophages ex vivo express relatively higher levels of sphingosine-1-phosphate (S1P) receptor 1 (S1P1) mRNA as compared to other receptor subtypes. The S1P agonist FTY720 decreased ERK phosphorylation and induced myosin light chain (MLC) II phosphorylation only in macrophages and DCs. FTY720 inhibited IL-12p70 production (CD40L induced) by DCs and macrophages but not microglia (poly I:C induced). IL-10 production was increased in DCs and unaffected in other myeloid cells. Despite similar receptor expression patterns, the distinct myeloid cell populations present in the human CNS, under steady-state or inflammatory conditions, exhibit differential responses to FTY720.
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http://dx.doi.org/10.1016/j.jneuroim.2010.08.006DOI Listing
January 2011

Innate immune-mediated neuronal injury consequent to loss of astrocytes.

J Neuropathol Exp Neurol 2008 Jun;67(6):590-9

Neuroimmunology Unit, Montreal Neurological Institute, McGill University, Montreal, Quebec, Canada.

Neuronal injury and loss are recognized features of neuroinflammatory disorders, including acute and chronic encephalitides and multiple sclerosis; destruction of astrocytes has been demonstrated in cases of Rasmussen encephalitis. Here, we show that innate immune cells (i.e. natural killer [NK] and gammadelta T cells) cause loss of neurons from primary human neuron-enriched cultures by destroying the supporting astrocytes. Interleukin 2-activated NK cells caused loss of astrocytes within 1 hour, whereas neurons were lost at 4 hours. Time-lapse imaging indicated that delayed neuron loss was due to early destruction of supporting astrocytes. Selective blocking of astrocyte death with anti-NKG2D antibodies reduced neuron loss, as did blocking of CD54 on astrocytes. gammadelta T cells also induced astrocyte cytotoxicity, leading to subsequent neuronal displacement. In astrocytes, NK cells caused caspase-dependent fragmentation of the intermediate filament proteins glial fibrillary acidic protein and vimentin, whereas anti-CD3-activated T cells produced fragmentation to a lesser extent and without measurable cytotoxicity. Glial fibrillary acidic protein fragmentation was also demonstrated in lysates from chronic multiple sclerosis plaques but not from normal control white matter. These data suggest that non-major histocompatibility complex-restricted immune effector cells may contribute to neuron loss in neuroinflammatory disorders indirectly through injury of glia.
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http://dx.doi.org/10.1097/NEN.0b013e3181772cf6DOI Listing
June 2008

Th1 polarization of CD4+ T cells by Toll-like receptor 3-activated human microglia.

J Neuropathol Exp Neurol 2007 Sep;66(9):848-59

Neuroimmunology Unit, Montreal Neurological Institute, McGill University, Montreal, Quebec, Canada.

Toll-like receptors (TLRs) are expressed by human microglia and translate environmental cues into distinct activation programs. We addressed the impact of TLR ligation on the capacity of human microglia to activate and polarize CD4 T cell responses. As microglia exist under distinct states of activation, we examined both ramified and ameboid microglia isolated from adult and fetal CNS, respectively. In vitro, ligation of TLR3 significantly increased major histocompatibility complex and costimulatory molecule expression on adult microglia and induced high levels of interferon-alpha, interleukin-12p40, and interleukin-23. TLR4 and, in particular, TLR2 had a more limited capacity to induce such responses. Coculturing allogeneic CD4 T cells with microglia preactivated with TLR3 did not increase T cell proliferation above basal levels but consistently led to elevated levels of interferon-gamma secretion and Th1 polarization. Fetal microglial TLR3 responses were comparable; in contrast, TLR2 and TLR4 decreased major histocompatibility complex class II expression on fetal cells and reduced CD4 T cell proliferation to levels below those found in untreated cocultures. All 3 TLRs induced comparable interleukin-6 secretion by microglia. Our findings illustrate how activation of human microglia via TLRs, particularly TLR3, can change the profile of local CNS immune responses by translating Th1 polarizing signals to CD4 T cells.
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http://dx.doi.org/10.1097/nen.0b013e3181492a7DOI Listing
September 2007

NKG2D-mediated cytotoxicity toward oligodendrocytes suggests a mechanism for tissue injury in multiple sclerosis.

J Neurosci 2007 Jan;27(5):1220-8

Neuroimmunology Unit, Montreal Neurological Institute, Montreal, Quebec, Canada H3A 2B4.

NKG2D is an activating or coactivating receptor expressed on human natural killer (NK) cells, CD8+ T cells, and gamma/delta T cells. NKG2D ligands have been detected on many tumor cell types and can be induced on nontransformed cells by environmental signals including DNA damage and inflammation. We investigated the contribution of NKG2D-NKG2D ligand interaction on CNS-directed immune responses. We observed that primary cultures of human adult oligodendrocytes and fetal astrocytes expressed ligands for NKG2D in vitro whereas neurons, microglia, and adult astrocytes did not. Disruption of the NKG2D-NKG2D ligand interaction using blocking antibodies significantly inhibited killing of primary human oligodendrocytes mediated by activated human NK cells, gamma/delta T cells, and allo-reactive CD8+ T cells. NKG2D ligands [major histocompatibility complex class I chain-related molecules A and B (MICA/B)] were detected in groups of cells and colocalized with an oligodendrocyte marker (adenomatous polyposis coli) in white matter sections obtained from multiple sclerosis lesions but not in normal control samples. CD8+ T cells could be detected in close proximity to MICA/B+ cells within multiple sclerosis lesions, supporting an in vivo interaction between these immune effectors and stressed MICA/B-expressing oligodendrocytes. These results imply that NKG2D-NKG2D ligand interaction can potentially contribute to cytotoxic responses mediated by activated immune effector cells in the inflamed CNS, as observed in multiple sclerosis.
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http://dx.doi.org/10.1523/JNEUROSCI.4402-06.2007DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6673175PMC
January 2007

TLR signaling tailors innate immune responses in human microglia and astrocytes.

J Immunol 2005 Oct;175(7):4320-30

Neuroimmunology Unit, Montreal Neurological Institute, McGill University, Montreal, Quebec, Canada.

The specific signals mediating the activation of microglia and astrocytes as a prelude to, or consequence of, CNS inflammation continue to be defined. We investigated TLRs as novel receptors mediating innate immune responses in human glial cells. We find that microglia express mRNA for TLRs 1-9, whereas astrocytes express robust TLR3, low-level TLR 1, 4, 5, and 9, and rare-to-undetectable TLR 2, 6, 7, 8, and 10 mRNA (quantitative real-time PCR). We focused on TLRs 3 and 4, which can signal through both the MyD88-dependent and -independent pathways, and on the MyD88-restricted TLR2. By flow cytometry, we established that microglia strongly express cell surface TLR2; TLR3 is expressed at higher levels intracellularly. Astrocytes express both cell surface and intracellular TLR3. All three TLRs trigger microglial activation upon ligation. TLR3 signaling induces the strongest proinflammatory polarizing response, characterized by secretion of high levels of IL-12, TNF-alpha, IL-6, CXCL-10, and IL-10, and the expression of IFN-beta. CXCL-10 and IL-10 secretion following TLR4 ligation are comparable to that of TLR3; however, other responses were lower or absent. TLR2-mediated responses are dominated by IL-6 and IL-10 secretion. Astrocytes respond to TLR3 ligation, producing IL-6, CXCL-10, and IFN-beta, implicating these cells as contributors to proinflammatory responses. Initial TLR-mediated glial activation also regulates consequent TLR expression; while TLR2 and TLR3 are subject to positive feedback, TLR4 is down-regulated in microglia. Astrocytes up-regulate all three TLRs following TLR3 ligation. Our data indicate that activation of innate immune responses in the CNS is not homogeneous but rather tailored according to cell type and environmental signal.
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http://dx.doi.org/10.4049/jimmunol.175.7.4320DOI Listing
October 2005

Interferon beta promotes nerve growth factor secretion early in the course of multiple sclerosis.

Arch Neurol 2005 Apr;62(4):563-8

Neuroimmunology Unit and McConnell Brain Imaging Center, Montreal Neurological Institute, McGill University.

Background: Interferon beta therapy has been shown to reduce the rate of clinical relapse and the frequency of magnetic resonance imaging-defined T2- weighted lesions in patients with multiple sclerosis (MS). When given early, interferon beta also reduces the rate of development of brain atrophy and improves axonal integrity. Nerve growth factor (NGF) can retard the severity and course of experimental allergic encephalomyelitis.

Objective: To determine whether interferon beta effects on patients with MS could be related to modulation of neurotrophin production within the central nervous system.

Design: We studied neurotrophin production by human glial and brain endothelial cells in response to coculture with MS patient-derived lymphocytes, and correlated levels of NGF secretion with clinical and magnetic resonance imaging-defined markers of disease.

Results: We demonstrate that production of NGF by human brain microvascular endothelial cells is triggered by interaction with T lymphocytes derived from MS patients. No such response was observed using human adult microglia or human fetal astrocytes. Nerve growth factor production by endothelial cells was potentiated by pretreating lymphocytes with interferon beta in vitro, and by using lymphocytes derived from MS patients treated with interferon beta in vivo. By using this assay, we show that levels of NGF induced by lymphocytes from MS patients inversely correlate with magnetic resonance imaging measures of brain atrophy and axonal injury.

Conclusion: These findings suggest that interferon beta-mediated production of NGF at the level of the blood-brain barrier, whether acting as an immunomodulator or directly on neural cells, is another potential mechanism contributing to the magnetic resonance imaging-defined effect of interferon beta on brain atrophy when given early in the course of MS.
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http://dx.doi.org/10.1001/archneur.62.4.563DOI Listing
April 2005

Distinctive properties of human adult brain-derived myelin progenitor cells.

Am J Pathol 2004 Dec;165(6):2167-75

Neuroimmunology Unit, Montreal Neurological Institute, McGill University, Montreal, Quebec H3A 2B4, Canada.

We used expression of the ganglioside A2B5 to isolate putative myelin progenitor cells from adult human central nervous system parenchyma and compared their phenotypic (expression of myelin lineage molecules) and functional (survival, proliferation) properties with mature oligodendrocytes (OLGs) derived from the same adult material and with A2B5(+) cells isolated from human fetal brain. A2B5(+) cells represented 3 to 5% of the total cell suspension derived from adult specimens. Results of protein (immunostaining) and RNA (polymerase chain reaction) analyses indicated that the adult A2B5(+) cells were more committed to the OLG lineage than their fetal counterparts while continuing to retain properties of progenitor cells compared to the postmitotic mature OLGs. Although the adult A2B5(+) cells retained the capacity to divide, albeit at a reduced rate compared to fetal A2B5(+) cells, they showed reduced survival and process outgrowth compared not only to fetal cells but also to mature OLGs. Our results confirm the presence of progenitor cells committed to the OLG lineage in the adult human central nervous system but raise the issues regarding the intrinsic capacity of these cells to contribute to the process of remyelination that may be necessary during demyelinating diseases.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC1618716PMC
http://dx.doi.org/10.1016/S0002-9440(10)63266-XDOI Listing
December 2004

Regulation of cellular and molecular trafficking across human brain endothelial cells by Th1- and Th2-polarized lymphocytes.

J Neuropathol Exp Neurol 2004 Mar;63(3):223-32

Neuroimmunology Unit, Montreal Neurological Institute, McGill University, Montreal, Quebec, Canada.

We used adult human brain-derived endothelial cells (HBECs) to model migration of peripheral blood lymphocytes across the blood brain barrier (BBB) as occurs in MS. We demonstrate that enhanced expression of adhesion molecule ICAM-1 and production of chemokines CXCL10/IP-10, CCL2/MCP-1, and CXCL8/IL-8 by HBECs induced by supernatants derived from allogeneic or myelin basic protein-reactive Th1 cells is only partially reversed with anti-IFNgamma antibody. This effect is not reproduced with IFNgamma or TNFalpha alone, implicating the interaction of multiple factors in the overall functional response. Supernatants from Th2 cells neither suppressed nor amplified Th1-induced effects. Although both Th1 and Th2 supernatants modulated the expression and localization of tight junction molecules zonula occludens (ZO)-1 and ZO-2, neither supernatant altered the permeability of HBEC monolayers to albumin or increased subsequent T cell migration rates. Prior migration of Th1 or Th2 cells across HBECs did enhance subsequent passage of cells and soluble molecules. Our results suggest that initial infiltration of either Th1 or Th2 polarized lymphocytes across the BBB contributes to the continuation of an inflammatory response in the central nervous system.
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http://dx.doi.org/10.1093/jnen/63.3.223DOI Listing
March 2004

Differential effects of Th1 and Th2 lymphocyte supernatants on human microglia.

Glia 2003 Apr;42(1):36-45

Neuroimmunology Unit, Montreal Neurological Institute, McGill University, Quebec, Canada.

We assessed the effects of soluble molecules (supernatants) produced by pro- (Th1) and anti- (Th2) inflammatory T-cell lines on the capacity of adult human CNS-derived microglia to express or produce selected cell surface and soluble molecules that regulate immune reactivity or impact on tissue protection/repair within the CNS. Treatment of microglia with supernatants from allo-antigen and myelin basic protein-specific Th1 cell lines augmented expression of cell surface molecules MHC class II, CD80, CD86, CD40, and CD54, enhanced the functional antigen-presenting cell capacity of microglia in a mixed lymphocyte reaction, and increased cytokine/chemokine secretion (TNFalpha, IL-6, and CXCL10/IP-10). These Th1-induced effects were not reproduced by interferon-gamma (IFNgamma) alone and were only incompletely blocked by anti-IFNgamma antibody. Th2 cell supernatant treatments did not alter costimulatory/adhesion molecule expression or induce cytokine/chemokine production by microglia. Th2 treatment, furthermore, failed to reduce the induction observed in response to Th1 supernatants. Neither Th1 nor Th2 supernatants induced production of the neurotrophin molecules, nerve growth factor, or brain-derived neurotrophic factor. Our results suggest that soluble molecules released by Th1 and not Th2 cells that infiltrate the CNS can stimulate resident microglia to acquire enhanced effector and accessory cell functions; the Th1-induced effects were not downregulated by Th2 supernatant-mediated bystander suppression.
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http://dx.doi.org/10.1002/glia.10201DOI Listing
April 2003