Extracellular vesicles released by microglia and macrophages carry endocannabinoids which foster oligodendrocyte differentiation.

Introduction: Microglia and macrophages can influence the evolution of myelin lesions through the production of extracellular vesicles (EVs). While microglial EVs promote in vitro differentiation of oligodendrocyte precursor cells (OPCs), whether EVs derived from macrophages aid or limit OPC maturation is unknown. Methods: Immunofluorescence analysis for the myelin protein MBP was employed to evaluate the impact of EVs from primary rat macrophages on cultured OPC differentiation. Raman spectroscopy and liquid chromatography-mass spectrometry was used to define the promyelinating lipid components of myelin EVs obtained in vitro and isolated from human plasma. Results and discussion: Here we show that macrophage-derived EVs do not promote OPC differentiation, and those released from macrophages polarized towards an inflammatory state inhibit OPC maturation. However, their lipid cargo promotes OPC maturation in a similar manner to microglial EVs. We identify the promyelinating endocannabinoids anandamide and 2-arachidonoylglycerol in EVs released by both macrophages and microglia in vitro and circulating in human plasma. Analysis of OPC differentiation in the presence of the endocannabinoid receptor antagonists SR141716A and AM630 reveals a key role of vesicular endocannabinoids in OPC maturation. From this study, EV-associated endocannabinoids emerge as important mediators in microglia/macrophage-oligodendrocyte crosstalk, which may be exploited to enhance myelin repair.

Long-Term Follow-Up of Custom-Made Porous Hydroxyapatite Cranioplasties: Analysis of Infections in Adult and Pediatric Patients.

In neurosurgery, cranioplasty (CP) stands as a pivotal surgical intervention, particularly following head trauma or various neurosurgical interventions. This study scrutinizes the intricacies of CP, emphasizing its prevalence and associated complications, with a specific focus on custom-made porous hydroxyapatite (PHA) implants. The investigation spans 687 patients (with 80 patients of pediatric age, less than 14 years old) across 26 neurosurgical centers in five European countries. Methodologically, this study delves into patient characteristics, complications, and infection data through a comprehensive post-marketing on-site surveillance approach. Notably, infections emerged as the primary complication, affecting 41 patients (6% of implants) with a clear distinction in onset patterns between pediatric (with more infections, 10% versus 5.4% in adults and an earlier onset of complications) and adult populations. Out of these 41 cases, cranioplasty explantation was required in 30 patients, 4.4% of the total population. Furthermore, bifrontal decompression correlated with a significantly elevated infection risk as compared to unilateral decompression (12.5% versus 5.1%) which remains after the examination of possible confounding factors. These findings provide substantial insights into the complexities of CP, suggesting the necessity for tailored strategies in pediatric and adult cases and cautioning against bifrontal decompressions. Despite acknowledging limitations and calling for prospective studies with long term follow-up, this research advances our understanding of the use of PHA CP, guiding clinical decision-making and emphasizing the importance of customized approaches for diverse patient cohorts.

Microglial TNFR2 signaling regulates the inflammatory response after CNS injury in a sex-specific fashion.

Microglia, the resident immune cells of the central nervous system (CNS), play a major role in damage progression and tissue remodeling after acute CNS injury, including ischemic stroke (IS) and spinal cord injury (SCI). Understanding the molecular mechanisms regulating microglial responses to injury may thus reveal novel therapeutic targets to promote CNS repair. Here, we investigated the role of microglial tumor necrosis factor receptor 2 (TNFR2), a transmembrane receptor previously associated with pro-survival and neuroprotective responses, in shaping the neuroinflammatory environment after CNS injury. By inducing experimental IS and SCI in Cx3cr1CreER:Tnfrsf1bfl/fl mice, selectively lacking TNFR2 in microglia, and corresponding Tnfrsf1bfl/fl littermate controls, we found that ablation of microglial TNFR2 significantly reduces lesion size and pro-inflammatory cytokine levels, and favors infiltration of leukocytes after injury. Interestingly, these effects were paralleled by opposite sex-specific modifications of microglial reactivity, which was found to be limited in female TNFR2-ablated mice compared to controls, whereas it was enhanced in males. In addition, we show that TNFR2 protein levels in the cerebrospinal fluid (CSF) of human subjects affected by IS and SCI, as well as healthy donors, significantly correlate with disease stage and severity, representing a valuable tool to monitor the inflammatory response after acute CNS injury. Hence, these results advance our understanding of the mechanisms regulating microglia reactivity after acute CNS injury, aiding the development of sex- and microglia-specific, personalized neuroregenerative strategies.

G protein-coupled receptor 17 is regulated by WNT pathway during oligodendrocyte precursor cell differentiation.

G protein-coupled receptor 17 (GPR17) and the WNT pathway are critical players of oligodendrocyte (OL) differentiation acting as essential timers in developing brain to achieve fully-myelinating cells. However, whether and how these two systems are related to each other is still unknown. Of interest, both factors are dysregulated in developing and adult brain diseases, including white matter injury and cancer, making the understanding of their reciprocal interactions of potential importance for identifying new targets and strategies for myelin repair. Here, by a combined pharmacological and biotechnological approach, we examined regulatory mechanisms linking WNT signaling to GPR17 expression in OLs. We first analyzed the relative expression of mRNAs encoding for GPR17 and the T cell factor/Lymphoid enhancer-binding factor-1 (TCF/LEF) transcription factors of the canonical WNT/ß-CATENIN pathway, in PDGFRα+ and O4+ OLs during mouse post-natal development. In O4+ cells, Gpr17 mRNA level peaked at post-natal day 14 and then decreased concomitantly to the physiological uprise of WNT tone, as shown by increased Lef1 mRNA level. The link between WNT signaling and GPR17 expression was further reinforced in vitro in primary PDGFRα+ cells and in Oli-neu cells. High WNT tone impaired OL differentiation and drastically reduced GPR17 mRNA and protein levels. In Oli-neu cells, WNT/ß-CATENIN activation repressed Gpr17 promoter activity through both putative WNT response elements (WRE) and upregulation of the inhibitor of DNA-binding protein 2 (Id2). We conclude that the WNT pathway influences OL maturation by repressing GPR17, which could have implications in pathologies characterized by dysregulations of the OL lineage including multiple sclerosis and oligodendroglioma.

Selective Inhibition of Soluble Tumor Necrosis Factor Alters the Neuroinflammatory Response following Moderate Spinal Cord Injury in Mice.

Clinical and animal model studies have implicated inflammation and glial and peripheral immune cell responses in the pathophysiology of spinal cord injury (SCI). A key player in the inflammatory response after SCI is the pleiotropic cytokine tumor necrosis factor (TNF), which exists both in both a transmembrane (tmTNF) and a soluble (solTNF) form. In the present study, we extend our previous findings of a therapeutic effect of topically blocking solTNF signaling after SCI for three consecutive days on lesion size and functional outcome to study the effect on spatio-temporal changes in the inflammatory response after SCI in mice treated with the selective solTNF inhibitor XPro1595 and compared to saline-treated mice. We found that despite comparable TNF and TNF receptor levels between XPro1595- and saline-treated mice, XPro1595 transiently decreased pro-inflammatory interleukin (IL)-1ß and IL-6 levels and increased pro-regenerative IL-10 levels in the acute phase after SCI. This was complemented by a decrease in the number of infiltrated leukocytes (macrophages and neutrophils) in the lesioned area of the spinal cord and an increase in the number of microglia in the peri-lesion area 14 days after SCI, followed by a decrease in microglial activation in the peri-lesion area 21 days after SCI. This translated into increased myelin preservation and improved functional outcomes in XPro1595-treated mice 35 days after SCI. Collectively, our data suggest that selective targeting of solTNF time-dependently modulates the neuroinflammatory response by favoring a pro-regenerative environment in the lesioned spinal cord, leading to improved functional outcomes.

Altered Purinergic Signaling in Neurodevelopmental Disorders: Focus on P2 Receptors.

With the umbrella term 'neurodevelopmental disorders' (NDDs) we refer to a plethora of congenital pathological conditions generally connected with cognitive, social behavior, and sensory/motor alterations. Among the possible causes, gestational and perinatal insults have been demonstrated to interfere with the physiological processes necessary for the proper development of fetal brain cytoarchitecture and functionality. In recent years, several genetic disorders caused by mutations in key enzymes involved in purine metabolism have been associated with autism-like behavioral outcomes. Further analysis revealed dysregulated purine and pyrimidine levels in the biofluids of subjects with other NDDs. Moreover, the pharmacological blockade of specific purinergic pathways reversed the cognitive and behavioral defects caused by maternal immune activation, a validated and now extensively used rodent model for NDDs. Furthermore, Fragile X and Rett syndrome transgenic animal models as well as models of premature birth, have been successfully utilized to investigate purinergic signaling as a potential pharmacological target for these diseases. In this review, we examine results on the role of the P2 receptor signaling in the etiopathogenesis of NDDs. On this basis, we discuss how this evidence could be exploited to develop more receptor-specific ligands for future therapeutic interventions and novel prognostic markers for the early detection of these conditions.

The multiple faces of extracellular vesicles released by microglia: Where are we 10 years after?

As resident component of the innate immunity in the central nervous system (CNS), microglia are key players in pathology. However, they also exert fundamental roles in brain development and homeostasis maintenance. They are extremely sensitive and plastic, as they assiduously monitor the environment, adapting their function in response to stimuli. On consequence, microglia may be defined a heterogeneous community of cells in a dynamic equilibrium. Extracellular vesicles (EVs) released by microglia mirror the dynamic nature of their donor cells, exerting important and versatile functions in the CNS as unbounded conveyors of bioactive signals. In this review, we summarize the current knowledge on EVs released by microglia, highlighting their heterogeneous properties and multifaceted effects.

Rewiring of Glucose and Lipid Metabolism Induced by G Protein-Coupled Receptor 17 Silencing Enables the Transition of Oligodendrocyte Progenitors to Myelinating Cells.

In the mature central nervous system (CNS), oligodendrocytes (OLs) provide support and insulation to axons thanks to the production of a myelin sheath. During their maturation to myelinating cells, OLs require energy and building blocks for lipids, which implies a great investment of energy fuels and molecular sources of carbon. The oligodendroglial G protein-coupled receptor 17 (GPR17) has emerged as a key player in OL maturation; it reaches maximal expression in pre-OLs, but then it has to be internalized to allow terminal maturation. In this study, we aim at elucidating the role of physiological GPR17 downregulation in OL metabolism by applying transcriptomics, metabolomics and lipidomics on differentiating OLs. After GPR17 silencing, we found a significant increase in mature OL markers and alteration of several genes involved in glucose metabolism and lipid biosynthesis. We also observed an increased release of lactate, which is partially responsible for the maturation boost induced by GPR17 downregulation. Concomitantly, GPR17 depletion also changed the kinetics of specific myelin lipid classes. Globally, this study unveils a functional link between GPR17 expression, lactate release and myelin composition, and suggests that innovative interventions targeting GPR17 may help to foster endogenous myelination in demyelinating diseases.

Dynamics of Microglia Activation in the Ischemic Brain: Implications for Myelin Repair and Functional Recovery.

Ischemic stroke is a neurological disorder representing a leading cause of death and permanent disability world-wide, for which effective regenerative treatments are missing. Oligodendrocyte degeneration and consequent myelin disruption are considered major contributing factors to stroke-associated neurological deficits. Therefore, fostering myelin reconstruction by oligodendrocyte precursor cells (OPCs) has emerged as a promising therapeutic approach to enhance functional recovery in stroke patients. A pivotal role in regulating remyelination is played by microglia, the resident immune cells of the brain. Early after stroke, microglial cells exert beneficial functions, promoting OPC recruitment toward the ischemic lesion and preserving myelin integrity. However, the protective features of microglia are lost during disease progression, contributing to remyelination failure. Unveiling the mechanisms driving the pro-remyelination properties of microglia may provide important opportunities for both reducing myelin damage and promoting its regeneration. Here, we summarize recent evidence describing microglia activation kinetics in experimental models of ischemic injury, focusing on the contribution of these innate immune cells to myelin damage and repair. Some molecular signals regulating the pro-regenerative functions of microglia after stroke have been highlighted to provide new possible therapeutic targets involved in the protective functions of these cells. Finally, we analyzed the impact of microglia-to-OPCs communication via extracellular vesicles on post-stroke remyelination and functional recovery. The results collected in this review underline the importance of supporting the pro-remyelination functions of microglial cells after stroke.

Oligodendrocyte Dysfunction in Amyotrophic Lateral Sclerosis: Mechanisms and Therapeutic Perspectives.

Myelin is the lipid-rich structure formed by oligodendrocytes (OLs) that wraps the axons in multilayered sheaths, assuring protection, efficient saltatory signal conduction and metabolic support to neurons. In the last few years, the impact of OL dysfunction and myelin damage has progressively received more attention and is now considered to be a major contributing factor to neurodegeneration in several neurological diseases, including amyotrophic lateral sclerosis (ALS). Upon OL injury, oligodendrocyte precursor cells (OPCs) of adult nervous tissue sustain the generation of new OLs for myelin reconstitution, but this spontaneous regeneration process fails to successfully counteract myelin damage. Of note, the functions of OPCs exceed the formation and repair of myelin, and also involve the trophic support to axons and the capability to exert an immunomodulatory role, which are particularly relevant in the context of neurodegeneration. In this review, we deeply analyze the impact of dysfunctional OLs in ALS pathogenesis. The possible mechanisms underlying OL degeneration, defective OPC maturation, and impairment in energy supply to motor neurons (MNs) have also been examined to provide insights on future therapeutic interventions. On this basis, we discuss the potential therapeutic utility in ALS of several molecules, based on their remyelinating potential or capability to enhance energy metabolism.

The immune-inflammatory response of oligodendrocytes in a murine model of preterm white matter injury: the role of TLR3 activation.

A leading cause of preterm birth is the exposure to systemic inflammation (maternal/fetal infection), which leads to neuroinflammation and white matter injury (WMI). A wide range of cytokines and chemokines are expressed and upregulated in oligodendrocytes (OLs) in response to inflammation and numerous reports show that OLs express several receptors for immune related molecules, which enable them to sense inflammation and to react. However, the role of OL immune response in WMI is unclear. Here, we focus our study on toll-like receptor-3 (TLR3) that is activated by double-strand RNA (dsRNA) and promotes neuroinflammation. Despite its importance, its expression and role in OLs remain unclear. We used an in vivo mouse model, which mimics inflammation-mediated WMI of preterm born infants consisting of intraperitoneal injection of IL-1ß from P1 to P5. In the IL-1ß-treated animals, we observed the upregulation of Tlr3, IL-1ß, IFN-ß, Ccl2, and Cxcl10 in both PDGFRα+ and O4+ sorted cells. This upregulation was higher in O4+ immature OLs (immOLs) as compared to PDGFRα+ OL precursor cells (OPCs), suggesting a different sensitivity to neuroinflammation. These observations were confirmed in OL primary cultures: cells treated with TLR3 agonist Poly(I:C) during differentiation showed a stronger upregulation of Ccl2 and Cxcl10 compared to cells treated during proliferation and led to decreased expression of myelin genes. Finally, OLs were able to modulate microglia phenotype and function depending on their maturation state as assessed by qPCR using validated markers for immunomodulatory, proinflammatory, and anti-inflammatory phenotypes and by phagocytosis and morphological analysis. These results show that during inflammation the response of OLs can play an autonomous role in blocking their own differentiation: in addition, the immune activation of OLs may play an important role in shaping the response of microglia during inflammation.

Microglial vesicles improve post-stroke recovery by preventing immune cell senescence and favoring oligodendrogenesis.

Contrasting myelin damage through the generation of new myelinating oligodendrocytes represents a promising approach to promote functional recovery after stroke. Here, we asked whether activation of microglia and monocyte-derived macrophages affects the regenerative process sustained by G protein-coupled receptor 17 (GPR17)-expressing oligodendrocyte precursor cells (OPCs), a subpopulation of OPCs specifically reacting to ischemic injury. GPR17-iCreERT2:CAG-eGFP reporter mice were employed to trace the fate of GPR17-expressing OPCs, labeled by the green fluorescent protein (GFP), after permanent middle cerebral artery occlusion. By microglia/macrophages pharmacological depletion studies, we show that innate immune cells favor GFP+ OPC reaction and limit myelin damage early after injury, whereas they lose their pro-resolving capacity and acquire a dystrophic "senescent-like" phenotype at later stages. Intracerebral infusion of regenerative microglia-derived extracellular vesicles (EVs) restores protective microglia/macrophages functions, limiting their senescence during the post-stroke phase, and enhances the maturation of GFP+ OPCs at lesion borders, resulting in ameliorated neurological functionality. In vitro experiments show that EV-carried transmembrane tumor necrosis factor (tmTNF) mediates the pro-differentiating effects on OPCs, with future implications for regenerative therapies.

TNF Production and Release from Microglia via Extracellular Vesicles: Impact on Brain Functions.

Tumor necrosis factor (TNF) is a pleiotropic cytokine powerfully influencing diverse processes of the central nervous system (CNS) under both physiological and pathological conditions. Here, we analyze current literature describing the molecular processes involved in TNF synthesis and release from microglia, the resident immune cells of the CNS and the main source of this cytokine both in brain development and neurodegenerative diseases. A special attention has been given to the unconventional vesicular pathway of TNF, based on the emerging role of microglia-derived extracellular vesicles (EVs) in the propagation of inflammatory signals and in mediating cell-to-cell communication. Moreover, we describe the contribution of microglial TNF in regulating important CNS functions, including the neuroinflammatory response following brain injury, the neuronal circuit formation and synaptic plasticity, and the processes of myelin damage and repair. Specifically, the available data on the functions mediated by microglial EVs carrying TNF have been scrutinized to gain insights on possible novel therapeutic strategies targeting TNF to foster CNS repair.

Under-Age Children Returning From Jihadist Group Operation Areas: How Can We Make a Diagnosis and Construct a Narrative With a Fragmentary Anamnesis?

INTRODUCTION: Since 2011, the French government estimates that about 500 French children have been born in or taken by their parents to areas where terrorist operations prevail. Since May 2017, 75 children who returned to France have benefited from a dedicated health care system. METHOD: This article is the result of clinical interviews conducted with 53 patients evaluated and taken care of at Avicenne Hospital in Bobigny. To our knowledge, no studies have been published on this subject. RESULTS: A total of 32 evaluations have been completed, all of which indicated the need for care for these children. Of these children, 64% are under 5 years old, and 59% were born in France. Their clinical profiles are heterogeneous and fluctuate with time. DISCUSSION: The multiple adverse events experienced by these children and the uniqueness of children born to families suspected by authorities of having participated in activities related to terrorism make this situation unprecedented. How can we make a diagnosis of PTSD without the help of a precise anamnesis? How can we help these children form a structuring narrative that avoids the pitfalls inherent to generalized fascination?

Development of the first in vivo GPR17 ligand through an iterative drug discovery pipeline: A novel disease-modifying strategy for multiple sclerosis.

The GPR17 receptor, expressed on oligodendroglial precursors (OPCs, the myelin producing cells), has emerged as an attractive target for a pro-myelinating strategy in multiple sclerosis (MS). However, the proof-of-concept that selective GPR17 ligands actually exert protective activity in vivo is still missing. Here, we exploited an iterative drug discovery pipeline to prioritize novel and selective GPR17 pro-myelinating agents out of more than 1,000,000 compounds. We first performed an in silico high-throughput screening on GPR17 structural model to identify three chemically-diverse ligand families that were then combinatorially exploded and refined. Top-scoring compounds were sequentially tested on reference pharmacological in vitro assays with increasing complexity, ending with myelinating OPC-neuron co-cultures. Successful ligands were filtered through in silico simulations of metabolism and pharmacokinetics, to select the most promising hits, whose dose and ability to target the central nervous system were then determined in vivo. Finally, we show that, when administered according to a preventive protocol, one of them (named by us as galinex) is able to significantly delay the onset of experimental autoimmune encephalomyelitis (EAE), a mouse model of MS. This outcome validates the predictivity of our pipeline to identify novel MS-modifying agents.

Abnormal Upregulation of GPR17 Receptor Contributes to Oligodendrocyte Dysfunction in SOD1 G93A Mice.

Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disease characterized by progressive loss of motor neurons (MN). Importantly, MN degeneration is intimately linked to oligodendrocyte dysfunction and impaired capacity of oligodendrocyte precursor cells (OPCs) to regenerate the myelin sheath enwrapping and protecting neuronal axons. Thus, improving OPC reparative abilities represents an innovative approach to counteract MN loss. A pivotal regulator of OPC maturation is the P2Y-like G protein-coupled receptor 17 (GPR17), whose role in ALS has never been investigated. In other models of neurodegeneration, an abnormal increase of GPR17 has been invariably associated to myelin defects and its pharmacological manipulation succeeded in restoring endogenous remyelination. Here, we analyzed GPR17 alterations in the SOD1G93A ALS mouse model and assessed in vitro whether this receptor could be targeted to correct oligodendrocyte alterations. Western-blot and immunohistochemical analyses showed that GPR17 protein levels are significantly increased in spinal cord of ALS mice at pre-symptomatic stage; this alteration is exacerbated at late symptomatic phases. Concomitantly, mature oligodendrocytes degenerate and are not successfully replaced. Moreover, OPCs isolated from spinal cord of SOD1G93A mice display defective differentiation compared to control cells, which is rescued by treatment with the GPR17 antagonist montelukast. These data open novel therapeutic perspectives for ALS management.

Regulation of Oligodendrocyte Functions: Targeting Lipid Metabolism and Extracellular Matrix for Myelin Repair.

: Myelin is an essential structure that protects axons, provides metabolic support to neurons and allows fast nerve transmission. Several neurological diseases, such as multiple sclerosis, are characterized by myelin damage, which is responsible of severe functional impairment. Myelin repair requires the timely recruitment of adult oligodendrocyte precursor cells (OPCs) at the lesion sites, their differentiation and maturation into myelinating oligodendrocytes. As a consequence, OPCs undergo profound changes in their morphology, functions, and interactions with other cells and extracellular environment, thus requiring the reorganization of both their lipid metabolism and their membrane composition, which is substantially different compared to other plasma membranes. Despite the growing knowledge in oligodendroglia biology and in the mechanisms involved in OPC-mediated regeneration, the identification of strategies to promote remyelination still remains a challenge. Here, we describe how altered lipid metabolism in oligodendrocytes influences the pathogenesis of demyelination, and we show that several FDA-approved drugs with a previously unknown remyelination potential do act on cholesterol and lipid biosynthetic pathways. Since the interplay between myelin lipids and axons is strictly coordinated by the extracellular matrix (ECM), we also discuss the role of different ECM components, and report the last findings on new ECM-modifiers able to foster endogenous remyelination.

Regulation and signaling of the GPR17 receptor in oligodendroglial cells.

Remyelination, namely, the formation of new myelin sheaths around denuded axons, counteracts axonal degeneration and restores neuronal function. Considerable advances have been made in understanding this regenerative process that often fails in diseases like multiple sclerosis, leaving axons demyelinated and vulnerable to damage, thus contributing to disease progression. The identification of the membrane receptor GPR17 on a subset of oligodendrocyte precursor cells (OPCs), which mediate remyelination in the adult central nervous system (CNS), has led to a huge amount of evidence that validated this receptor as a new attractive target for remyelinating therapies. Here, we summarize the role of GPR17 in OPC function, myelination and remyelination, describing its atypical pharmacology, its downstream signaling, and the genetic and epigenetic factors modulating its activity. We also highlight crucial insights into GPR17 pathophysiology coming from the demonstration that oligodendrocyte injury, associated with inflammation in chronic neurodegenerative conditions, is invariably characterized by abnormal and persistent GPR17 upregulation, which, in turn, is accompanied by a block of OPCs at immature premyelinating stages. Finally, we discuss the current literature in light of the potential exploitment of GPR17 as a therapeutic target to promote remyelination.