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Autosomal Recessive Spastic Ataxia of the Charlevoix Saguenay (ARSACS) is caused by mutation in the SACS gene resulting in loss of function of the protein sacsin. A key feature is the formation of abnormal bundles of neurofilaments (NF) in neurons and vimentin intermediate filaments (IF) in cultured fibroblasts, suggesting a role of sacsin in IF homeostasis. Sacsin contains a J domain (SacsJ) homologous to Hsp40, that can interact with Hsp70 chaperones. The SacsJ domain resolved NF bundles in cultured Sacs−/− neurons. Having studied the mechanism using NF assembled in vitro from purified NF proteins, we report that the SacsJ domain interacts with NF proteins to disassemble NFL filaments, and to inhibit their initial assembly. A cell-penetrating peptide derived from this domain, SacsJ-myc-TAT was efficient in disassembling NF bundles in cultured Sacs−/− motor neurons, restoring the NF network; however, there was some loss of vimentin IF and NF in cultured Sacs+/+ fibroblasts and motor neurons, respectively. These results suggest that sacsin through its SacsJ domain is a key regulator of NF and vimentin IF networks in cells.

Le 18 mars 2019 a eu lieu le forum sur la vitalité culturelle des quartiers à Montréal, organisé par Culture Montréal et le Centre de recherche sur les innovations sociales (CRISES). Ce forum a permis une réflexion d’une diversité d’acteurs et actrices de la culture de proximité et de la revitalisation des quartiers par la culture sur la base de la présentation d’une grille d’indicateurs de la vitalité culturelle des quartiers et des résultats de l’application de cette grille à l’arrondissement Rosemont—La Petite-Patrie. On March 18th 2019, a Forum on the Cultural Vitality of Neighborhoods was held, organized by Culture Montreal and the Centre for Research on Social Innovations (CRISES). This forum allowed a conversation between a vast array of actors of the culture of proximity and the revitalization of neighborhoods in Montreal. The conference was centered on the presentation of a grid of indicators of the cultural vitality of neighborhoods and the results of its application to the case of Rosemont-La Petite-Patrie, a large district of Montreal.

Viruses, particularly the Epstein-Barr virus (EBV) has long been suspected to exacerbate acute arthritic symptoms. However, the cell populations that contribute to import viruses into the inflamed tissues remain to be identified. In the present study, we have investigated the role of monocytes in the transport of Murid herpesvirus 68 (MHV-68), a mouse virus closely related to EBV, using the serum transfer-induced arthritis (STIA) model. We found compelling evidence that MHV-68 infection markedly increased disease severity in NR4A1-/- mice, which are deficient for Ly6Clow monocytes. In contrast, the MHV-68-induced enhancement of joint inflammation was lessened in CCR2-/- mice, suggesting the involvement of inflammatory Ly6Chigh monocytes in viral transport. We also observed that following selective depletion of monocyte subsets by administration of clodronate, MHV-68 transport into the synovium occurs only in the presence of Ly6Chigh monocytes. Tracking of adoptively transferred Ly6Chigh GFP infected monocytes into arthritic CCR2-/- mice by two-photon intravital microscopy showed that this monocyte subset has the capacity to deliver viruses to inflamed AR joints, as confirmed by the detection of viral DNA in inflamed tissues of recipient mice. We thus conclude that Ly6Chigh monocytes import MHV-68 when they are mobilized to the inflamed arthritic joint.

Multiple sclerosis (MS) is an autoimmune disease that affects hundreds of thousands of people worldwide. Given the autoimmune nature of the disease, a large part of the research has focused on autoreactive T and B cells. However, research on the involvement of myeloid cells in the pathophysiology of MS has received a strong and renewed attention over the recent years. Despite the multitude of inflammatory mediators involved in innate immunity, only a select group of cytokines are absolutely critical to the development of CNS autoimmunity, among which is interleukin (IL)-1. While the importance of the IL-1 system in experimental autoimmune encephalomyelitis (EAE) and MS has been recognized for about 20years, it is only recently that we have begun to understand that IL-1 plays multifaceted roles in disease initiation, development, amplification and chronicity. Here, we review the recent findings showing an implication of the IL-1 system in EAE and MS, and introduce a model that highlights how IL-1β and granulocyte-macrophage colony-stimulating factor (GM-CSF) are interacting together to create a vicious feedback cycle of CNS inflammation that ultimately leads to myelin and neuronal damage.

Antibody ligation of the murine neutrophil surface protein Ly6G disrupts neutrophil migration in some contexts but not others. We tested whether this variability reflected divergent dependence of neutrophil migration on β2 integrins, adhesion molecules that interact with Ly6G at the neutrophil surface. In integrin-dependent murine arthritis, Ly6G ligation attenuated joint inflammation, even though mice lacking Ly6G altogether developed arthritis normally. By contrast, Ly6G ligation had no impact on integrin- independent neutrophil migration into inflamed lung. In peritoneum, the role of β2 integrins varied with stimulus, proving dispensable for neutrophil entry in E. coli peritonitis but contributory in IL-1-mediated sterile peritonitis. Correspondingly, Ly6G ligation attenuated only IL-1 peritonitis, disrupting the molecular association between integrins and Ly6G and inducing cell-intrinsic blockade restricted to integrin-dependent migration. Consistent with this observation, Ly6G ligation impaired integrin-mediated postadhesion strengthening for neutrophils arresting on activated cremaster endothelium in vivo. Together, these findings identify selective inhibition of integrin- mediated neutrophil emigration through Ly6G ligation, highlighting the marked site and stimulus specificity of β2 integrin dependence in neutrophil migration.KEY POINTSThe contribution of β2 integrins to neutrophil migration into inflamed tissues varies with site and stimulus.Ligation of Ly6G, a GPI-linked neutrophil surface protein, selectively attenuates β2 integrin-dependent neutrophil migration in vivo.Blockade correlates with disrupted interaction between Ly6G and β2 integrins and impaired integrin-mediated postadhesion strengthening.

Work in a mouse model suggests that central inhibition of IL-1β/IL-1R1 signaling during the early acute phase of neuroinflammation may be an effective means for preventing loss of neurological function in multiple sclerosis., Growing evidence supports a role for IL-1 in multiple sclerosis and experimental autoimmune encephalomyelitis (EAE), but how it impacts neuroinflammation is poorly understood. We show that susceptibility to EAE requires activation of IL-1R1 on radiation-resistant cells via IL-1β secreted by bone marrow–derived cells. Neutrophils and monocyte-derived macrophages (MDMs) are the main source of IL-1β and produce this cytokine as a result of their transmigration across the inflamed blood–spinal cord barrier. IL-1R1 expression in the spinal cord is found in endothelial cells (ECs) of the pial venous plexus. Accordingly, leukocyte infiltration at EAE onset is restricted to IL-1R1+ subpial and subarachnoid vessels. In response to IL-1β, primary cultures of central nervous system ECs produce GM-CSF, G-CSF, IL-6, Cxcl1, and Cxcl2. Initiation of EAE or subdural injection of IL-1β induces a similar cytokine/chemokine signature in spinal cord vessels. Furthermore, the transfer of Gr1+ cells on the spinal cord is sufficient to induce illness in EAE-resistant IL-1β knockout (KO) mice. Notably, transfer of Gr1+ cells isolated from C57BL/6 mice induce massive recruitment of recipient myeloid cells compared with cells from IL-1β KO donors, and this recruitment translates into more severe paralysis. These findings suggest that an IL-1β–dependent paracrine loop between infiltrated neutrophils/MDMs and ECs drives neuroinflammation.

Multiple sclerosis (MS) is a neuroinflammatory disease characterized by demyelinating plaques in the brain and spinal cord, causing progressive loss of functions. While the causes of MS remain undefined, treatments that target immune cells or their functions greatly improve the clinical outcome. Our laboratory has previously shown that production of the inflammatory cytokine interleukin (IL)-1β by myeloid cells is required for the development of experimental autoimmune encephalomyelitis (EAE), an MS model. Here, we show that activation of central nervous system (CNS) endothelial cells by IL-1β enables inflammatory monocytes to enter the CNS and differentiate into antigen-presenting cells. Additionally, we demonstrate that factors released from the interaction between IL-1β–producing myeloid cells and autoreactive CD4+ T cells are toxic to neurons.

Molecular interventions that limit pathogenic CNS inflammation are used to treat autoimmune conditions such as multiple sclerosis (MS). Remarkably, IL-1β–knockout mice are highly resistant to experimental autoimmune encephalomyelitis (EAE), an animal model of MS. Here, we show that interfering with the IL-1β/IL-1R1 axis severely impairs the transmigration of myeloid cells across central nervous system (CNS) endothelial cells (ECs). Notably, we report that IL-1β expression by inflammatory CCR2hi monocytes favors their entry into the spinal cord before EAE onset. Following activation with IL-1β, CNS ECs release GM-CSF, which in turn converts monocytes into antigen-presenting cells (APCs). Accordingly, spinal cord-infiltrated monocyte-derived APCs are associated with dividing CD4+ T cells. Factors released from the interaction between IL-1β–competent myeloid cells and CD4+ T cells are highly toxic to neurons. Together, our results suggest that IL-1β signaling is an entry point for targeting both the initiation and exacerbation of neuroinflammation.

Viruses, particularly the Epstein-Barr virus (EBV) has long been suspected to exacerbate acute arthritic symptoms. However, the cell populations that contribute to import viruses into the inflamed tissues remain to be identified. In the present study, we have investigated the role of monocytes in the transport of Murid herpesvirus 68 (MHV-68), a mouse virus closely related to EBV, using the serum transfer-induced arthritis (STIA) model. We found compelling evidence that MHV-68 infection markedly increased disease severity in NR4A1

Mitochondrial DNA (mtDNA) is a highly potent inflammatory trigger and is reportedly found outside the cells in blood in various pathologies. Platelets are abundant in blood where they promote hemostasis. Although lacking a nucleus, platelets contain functional mitochondria. On activation, platelets produce extracellular vesicles known as microparticles. We hypothesized that activated platelets could also release their mitochondria. We show that activated platelets release respiratory-competent mitochondria, both within membrane-encapsulated microparticles and as free organelles. Extracellular mitochondria are found in platelet concentrates used for transfusion and are present at higher levels in those that induced acute reactions (febrile nonhemolytic reactions, skin manifestations, and cardiovascular events) in transfused patients. We establish that the mitochondrion is an endogenous substrate of secreted phospholipase A2 IIA (sPLA2-IIA), a phospholipase otherwise specific for bacteria, likely reflecting the ancestral proteobacteria origin of mitochondria. The hydrolysis of the mitochondrial membrane by sPLA2-IIA yields inflammatory mediators (ie, lysophospholipids, fatty acids, and mtDNA) that promote leukocyte activation. Two-photon microscopy in live transfused animals revealed that extracellular mitochondria interact with neutrophils in vivo, triggering neutrophil adhesion to the endothelial wall. Our findings identify extracellular mitochondria, produced by platelets, at the midpoint of a potent mechanism leading to inflammatory responses.

Disruption of the blood–brain and blood–spinal cord barriers (BBB and BSCB, respectively) and immune cell infiltration are early pathophysiological hallmarks of multiple sclerosis (MS), its animal model experimental autoimmune encephalomyelitis (EAE), and neuromyelitis optica (NMO). However, their contribution to disease initiation and development remains unclear. In this study, we induced EAE in lys-eGFP-ki mice and performed single, nonterminal intravital imaging to investigate BSCB permeability simultaneously with the kinetics of GFP+ myeloid cell infiltration. We observed a loss in BSCB integrity within a day of disease onset, which paralleled the infiltration of GFP+ cells into the CNS and lasted for ∼4 d. Neutrophils accounted for a significant proportion of the circulating and CNS-infiltrating myeloid cells during the preclinical phase of EAE, and their depletion delayed the onset and reduced the severity of EAE while maintaining BSCB integrity. We also show that neutrophils collected from the blood or bone marrow of EAE mice transmigrate more efficiently than do neutrophils of naive animals in a BBB cell culture model. Moreover, using intravital videomicroscopy, we demonstrate that the IL-1R type 1 governs the firm adhesion of neutrophils to the inflamed spinal cord vasculature. Finally, immunostaining of postmortem CNS material obtained from an acutely ill multiple sclerosis patient and two neuromyelitis optica patients revealed instances of infiltrated neutrophils associated with regions of BBB or BSCB leakage. Taken together, our data provide evidence that neutrophils are involved in the initial events that take place during EAE and that they are intimately linked with the status of the BBB/BSCB.

In vivo imaging using two-photon microscopy is an essential tool to explore the dynamic of physiological events deep within biological tissues for short or extended periods of time. The new capabilities offered by this technology (e.g. high tissue penetrance, low toxicity) have opened a whole new era of investigations in modern biomedical research. However, the potential of using this promising technique in tissues of living animals is greatly limited by the intrinsic irregular movements that are caused by cardiac and respiratory cycles and muscular and vascular tone. Here, we show real-time imaging of the brain, spinal cord, sciatic nerve and myenteric plexus of living mice using a new automated program, named Intravital_Microscopy_Toolbox, that removes frames corrupted with motion artifacts from time-lapse videos. Our approach involves generating a dissimilarity score against precalculated reference frames in a specific reference channel, thus allowing the gating of distorted, out-of-focus or translated frames. Since the algorithm detects the uneven peaks of image distortion caused by irregular animal movements, the macro allows a fast and efficient filtering of the image sequence. In addition, extra features have been implemented in the macro, such as XY registration, channel subtraction, extended field of view with maximum intensity projection, noise reduction with average intensity projections, and automated timestamp and scale bar overlay. Thus, the Intravital_Microscopy_Toolbox macro for ImageJ provides convenient tools for biologists who are performing in vivo two-photon imaging in tissues prone to motion artifacts.

Platelets survey blood vessels, searching for endothelial damage and preventing loss of vascular integrity. However, there are circumstances where vascular permeability increases, suggesting that platelets sometimes fail to fulfill their expected function. Human inflammatory arthritis is associated with tissue edema attributed to enhanced permeability of the synovial microvasculature. Murine studies have suggested that such vascular leak facilitates entry of autoantibodies and may thereby promote joint inflammation. Whereas platelets typically help to promote microvascular integrity, we examined the role of platelets in synovial vascular permeability in murine experimental arthritis. Using an in vivo model of autoimmune arthritis, we confirmed the presence of endothelial gaps in inflamed synovium. Surprisingly, permeability in the inflamed joints was abrogated if the platelets were absent. This effect was mediated by platelet serotonin accumulated via the serotonin transporter and could be antagonized using serotonin-specific reuptake inhibitor antidepressants. As opposed to the conventional role of platelets to microvascular leakage, this demonstration that platelets are capable of amplifying and maintaining permeability adds to the rapidly growing list of unexpected functions for platelets.