Central Nervous System Myeloid Cells Research Articles

Dietary wheat amylase trypsin inhibitors exacerbate CNS inflammation in experimental multiple sclerosis

ObjectiveWheat has become a main staple globally. We studied the effect of defined pro-inflammatory dietary proteins, wheat amylase trypsin inhibitors (ATI), activating intestinal myeloid cells via toll-like receptor 4, in...

466 Robust Model of Microglia Replacement With Circulation-Derived Microglia-Like Cells (CDMCs) in CNS Malignancies

INTRODUCTION: Tumor-associated microglia and macrophages (TAMs) represent a main cell type of brain malignancies and demonstrate complex interactions with cancerous cells and the tumor microenvironment. These interactions have important implications for the progression and treatment of malignant central nervous system (CNS) tumors. TAMs have therefore emerged as therapeutic targets, though their potential to date has been unfulfilled. METHODS: C57BL/6 mice were preconditioned for BMT with retroorbital busulfan injections (25 mg/kg), and native microglia were depleted via PLX5622 (1200 mg/kg, chow). Following transplantation, animals underwent intracranial stereotactic injection of syngeneic cell lines of either high-grade glioma (GL261) or breast cancer metastasis (E0771). Following animal sacrifice, we used single cell RNA sequencing of CD45-positive cells to profile the transcriptomic identity of glioma tumor-associated CDMCs (TA-CDMCs). RESULTS: Lineage tracing proved high intra- and peri-tumoral chimerism of BMT-graft derived cells among CNS myeloid cells in both models two weeks after tumor cell injection, demonstrating a robust ability to modify the tumor-immune microenvironment in animal models. TA-CDMCs showed a similarly activated morphology as naÏve TAMs under immunohistochemistry. Pathway enrichment analysis revealed an upregulation of immune response pathways in TA-CDMCs compared to regular TAMs, including the overexpression of genes relevant for the regulation of T cell activation, suggesting possible modification of the glioma immune-environment following CDMC repopulation. CONCLUSIONS: The present study offered a robust proof-of-concept that highly efficient TAM replacement could be achieved in orthotopic murine brain tumor models. Additionally, it appears that replaced CDMCs retain unique transcriptional patterns in the tumor-immune microenvironment, with the upregulation of several key immune response pathways. These efforts may represent a novel route of tumor-directed immunotherapy and offer exciting opportunities for ex-vivo CDMC manipulation to specifically target neoplastic cells.

Endothelial cell‐derived oxysterol ablation attenuates experimental autoimmune encephalomyelitis

The vasculature is a key regulator of leukocyte trafficking into the central nervous system (CNS) during inflammatory diseases including multiple sclerosis (MS). However, the impact of endothelial‐derived factors on CNS immune responses remains unknown. Bioactive lipids, in particular oxysterols downstream of Cholesterol‐25‐hydroxylase (Ch25h), promote neuroinflammation but their functions in the CNS are not well‐understood. Using floxed‐reporter Ch25h knock‐in mice, we trace Ch25h expression to CNS endothelial cells (ECs) and myeloid cells and demonstrate that Ch25h ablation specifically from ECs attenuates experimental autoimmune encephalomyelitis (EAE). Mechanistically, inflamed Ch25h‐deficient CNS ECs display altered lipid metabolism favoring polymorphonuclear myeloid‐derived suppressor cell (PMN‐MDSC) expansion, which suppresses encephalitogenic T lymphocyte proliferation. Additionally, endothelial Ch25h‐deficiency combined with immature neutrophil mobilization into the blood circulation nearly completely protects mice from EAE. Our findings reveal a central role for CNS endothelial Ch25h in promoting neuroinflammation by inhibiting the expansion of immunosuppressive myeloid cell populations.

TMIC-85. MICROGLIA REPLACEMENT CHANGES THE TRANSCRIPTIONAL PROFILE OF TUMOR ASSOCIATED MYELOID CELLS IN MURINE MODELS OF BRAIN MALIGNANCIES

Abstract Tumor associated microglia and macrophages (TAMs) represent a main cell type of brain malignancies and demonstrate complex interactions with cancerous cells and the tumor microenvironment. These interactions have important implications for the progression and treatment of malignant central nervous system (CNS) tumors. In a non-oncological context, it has been shown that endogenous microglia can be replaced by peripheral hematopoietic cells by genetic or pharmacological intervention. A highly efficient replacement protocol (Shibuya et al, Sci Transl Med 2022, PMID:35294256) utilizing myeloablative bone marrow transplantation (BMT) followed by microglial depletion via pharmacological inhibition of the Colony-stimulating factor-1 receptor (CSF-1R) leads to almost complete repopulation of the CNS myeloid niche by circulation-derived myeloid cells (CDMCs). We investigated if this approach could be utilized to integrate CDMCs into brain tumors and study associated changes in the TAM signature. Microglia replacement in immune-competent mice was followed by intracranial transplantation of syngeneic cell lines of either high-grade glioma (GL261) or breast cancer metastasis (E0771). Lineage tracing proved high intra- and peri-tumoral chimerism of BMT-graft derived cells among CNS myeloid cells in both models two weeks after tumor cell injection. Tumor associated CDMCs (TA-CDMCs) showed a similarly activated morphology as naïve TAMs. We used single cell RNA sequencing of CD45-positive cells to further profile the transcriptomic identity of glioma TA-CDMCs. Pathway enrichment analysis revealed an upregulation of immune response pathways in TA-CDMCs compared to regular TAMs, including the overexpression of genes relevant for the regulation of T cell activation, suggesting possible modification of the glioma immune-environment. Strategies to manipulate the myeloid niche have the potential to provide further understanding of the pathobiology of TAMs and to build the basis for novel cell therapeutic approaches in the neuro-oncological domain.

Dopamine‐driven Increase in IL‐1β in Myeloid Cells is Mediated by Differential Dopamine Receptor Expression and Exacerbated by HIV

The neurological complications of HIV infection, known as neuroHIV, remain prevalent even in individuals on antiretroviral therapy (ART). Although the mechanism(s) underlying neuroHIV remain undefined, current data suggest that neuroinflammation is central to the development of HIV neuropathogenesis. Neuroinflammation can be exacerbated by substance use disorders, which are highly comorbid with HIV infection and substantively worsen clinical outcomes. Despite distinct mechanisms of action, all substances of abuse increase central nervous system (CNS) dopamine, suggesting that dopamine is a common mechanism by which addictive drugs potentiate neuroHIV. Our published data show that dopamine increases the production of inflammatory cytokines such as IL‐1β in human microglia and macrophages, but the mechanisms by which dopamine drives inflammation are not clear. Studies indicate that D1‐like dopamine receptors (DRD1, DRD5) mediate inflammatory activity while D2‐like dopamine receptors (DRD2, DRD3, and DRD4) mediate anti‐inflammatory activity, and we have shown that human macrophages and microglia have higher expression of D1‐like receptors. Therefore, we hypothesize that dopamine‐mediated production of IL‐1β in CNS myeloid cells is regulated by the ratio of different dopamine receptor expression levels and can be exacerbated by HIV infection. Our data in primary macrophages indicate that DRD1 expression is necessary for dopamine‐mediated increases in IL‐1β, but that changes in DRD2 expression can alter the magnitude of the dopamine‐mediated increase in IL‐1β. We confirm this in human microglia cell lines, showing that dopamine only increases IL‐1β gene and protein expression in microglia with a high ratio of D1‐like receptors to D2‐like receptors. Further, antagonizing dopamine receptor expression with the pan‐dopamine receptor antagonist flupentixol or decreasing the D1‐like/D2‐like ratio by overexpressing DRD2 diminishes the dopamine‐mediated increase of IL‐1β. We also show that the effects of dopamine on IL‐1β are potentiated in the presence of HIV infection in both microglial cell lines and iPSC‐derived microglia. Ongoing studies are examining the role of epigenetic regulation in the effects of dopamine and HIV on inflammation in these myeloid systems.Due to the prevalence of both substance use disorders and the increasing use of dopamine‐modulating therapeutics, a detailed understanding of dopamine‐mediated changes in inflammation will be critical to effectively tailor ART regimens to HIV‐infected individuals using these drugs.

The Joint-Brain Axis: Insights From Rheumatoid Arthritis on the Crosstalk Between Chronic Peripheral Inflammation and the Brain.

Rheumatoid arthritis (RA) is a chronic inflammatory disease characterized by erosive polyarthritis. Beyond joint pathology, RA is associated with neuropsychiatric comorbidity including depression, anxiety, and an increased risk to develop neurodegenerative diseases in later life. Studies investigating the central nervous system (CNS) in preclinical models of RA have leveraged the understanding of the intimate crosstalk between peripheral and central immune responses. This mini review summarizes the current knowledge of CNS comorbidity in RA patients and known underlying cellular mechanisms. We focus on the differential regulation of CNS myeloid and glial cells in different mouse models of RA reflecting different patterns of peripheral immune activation. Moreover, we address CNS responses to anti-inflammatory treatment in human RA patients and mice. Finally, to illustrate the bidirectional communication between the CNS and chronic peripheral inflammation, we present the current knowledge about the impact of the CNS on arthritis. A comprehensive understanding of the crosstalk between the CNS and chronic peripheral inflammation will help to identify RA patients at risk of developing CNS comorbidity, setting the path for future therapeutic approaches in both RA and neuropsychiatric diseases.

Angiopoietin-2 blockade ameliorates autoimmune neuroinflammation by inhibiting leukocyte recruitment into the CNS.

Angiopoietin-2 (Ang2), a ligand of the endothelial Tie2 tyrosine kinase, is involved in vascular inflammation and leakage in critically ill patients. However, the role of Ang2 in demyelinating central nervous system (CNS) autoimmune diseases is unknown. Here, we report that Ang2 is critically involved in the pathogenesis of experimental autoimmune encephalomyelitis (EAE), a rodent model of multiple sclerosis. Ang2 expression was induced in CNS autoimmunity, and transgenic mice overexpressing Ang2 specifically in endothelial cells (ECs) developed a significantly more severe EAE. In contrast, treatment with Ang2-blocking Abs ameliorated neuroinflammation and decreased spinal cord demyelination and leukocyte infiltration into the CNS. Similarly, Ang2-binding and Tie2-activating Ab attenuated the development of CNS autoimmune disease. Ang2 blockade inhibited expression of EC adhesion molecules, improved blood-brain barrier integrity, and decreased expression of genes involved in antigen presentation and proinflammatory responses of microglia and macrophages, which was accompanied by inhibition of α5β1 integrin activation in microglia. Taken together, our data suggest that Ang2 provides a target for increasing Tie2 activation in ECs and inhibiting proinflammatory polarization of CNS myeloid cells via α5β1 integrin in neuroinflammation. Thus, Ang2 targeting may serve as a therapeutic option for the treatment of CNS autoimmune disease.

Myeloid cell plasticity in the evolution of central nervous system autoimmunity.

Myeloid cells, including macrophages and dendritic cells, are a prominent component of central nervous system (CNS) infiltrates during multiple sclerosis (MS) and the animal model experimental autoimmune encephalomyelitis (EAE). Although myeloid cells are generally thought to be proinflammatory, alternatively polarized subsets can serve noninflammatory and/or reparative functions. Here we investigate the heterogeneity and biological properties of myeloid cells during central nervous system autoimmunity. Myeloid cell phenotypes in chronic active MS lesions were analyzed by immunohistochemistry. In addition, immune cells were isolated from the CNS during exacerbations and remissions of EAE and characterized by flow cytometric, genetic, and functional assays. Myeloid cells expressing inducible nitric oxide synthase (iNOS), indicative of a proinflammatory phenotype, were detected in the actively demyelinating rim of chronic active MS lesions, whereas macrophages expressing mannose receptor (CD206), a marker of alternatively polarized human myeloid cells, were enriched in the quiescent lesion core. During EAE, CNS-infiltrating myeloid cells, as well as microglia, shifted from expression of proinflammatory markers to expression of noninflammatory markers immediately prior to clinical remissions. Murine CNS myeloid cells expressing the alternative lineage marker arginase-1 (Arg1) were partially derived from iNOS+ precursors and were deficient in activating encephalitogenic T cells compared with their Arg1- counterparts. These observations demonstrate the heterogeneity of CNS myeloid cells, their evolution during the course of autoimmune demyelinating disease, and their plasticity on the single cell level. Future therapeutic strategies for disease modification in individuals with MS may be focused on accelerating the transition of CNS myeloid cells from a proinflammatory to a noninflammatory phenotype. Ann Neurol 2018;83:131-141.

Ontogeny and homeostasis of CNS myeloid cells.

Myeloid cells in the central nervous system (CNS) represent a heterogeneous class of innate immune cells that contribute to the maintenance of tissue homeostasis differentially during development and adulthood. The subsets of CNS myeloid cells identified so far, including parenchymal microglia and non-parenchymal meningeal, perivascular and choroid-plexus macrophages, as well as disease-associated monocytes, have classically been distinguished on the basis of their surface epitope expression, localization and morphology. However, studies using cell-specific targeting, in vivo imaging, single-cell expression analysis and other sophisticated tools have now increased the depth of knowledge of this immune-cell compartment and call for reevaluation of the traditional views on the origin, fate and function of distinct CNS myeloid subsets. The concepts of CNS macrophage biology that are emerging from these new insights have broad implications for the understanding and treatment of CNS diseases.

Enhanced disease reduction using clozapine, an atypical antipsychotic agent, and glatiramer acetate combination therapy in experimental autoimmune encephalomyelitis.

BackgroundAtypical antipsychotic agents (AAP) alleviate the symptoms of severe mental health disorders, such as schizophrenia, by antagonizing dopamine and serotonin receptors. Recently, AAP have also been shown to exhibit immunomodulatory properties in the central nervous system (CNS).ObjectiveBuilding on research which demonstrated the ability of the AAP risperidone and clozapine to modify the disease course of experimental autoimmune encephalomyelitis (EAE), an animal model of multiple sclerosis (MS), we aimed to more fully investigate the potential of clozapine as a possible treatment for MS.ResultsWe report that orally administered clozapine significantly reduced the disease severity of EAE in a dose-dependent manner and was effective when administered prophylactically and therapeutically. In comparison to risperidone, quetiapine, and olanzapine, clozapine was the best at reducing disease severity. While clozapine-treated mice had only modest changes to peripheral leukocytes and cytokine responses, these animals had significantly fewer CNS-infiltrating CD4 T cells and myeloid cells. Furthermore, the CNS myeloid cells displayed a less activated phenotype in mice treated with clozapine. Finally, we found that co-administration of clozapine with glatiramer acetate enhanced disease protection compared to either treatment alone.ConclusionThese studies indicate that clozapine is an effective immunomodulatory agent with the potential to treat immune-mediated diseases such as MS.

Type-1 angiotensin receptor signaling in central nervous system myeloid cells is pathogenic during fatal alphavirus encephalitis in mice.

BackgroundAlphaviruses can cause fatal encephalitis in humans. Natural infections occur via the bite of infected mosquitos, but aerosol transmissibility makes some of these viruses potential bioterrorism agents. Central nervous system (CNS) host responses contribute to alphavirus pathogenesis in experimental models and are logical therapeutic targets. We investigated whether reactive oxygen species (ROS) generated by nicotinamide adenine dinucleotide phosphate (NADPH) oxidase (Nox) activity within the CNS contributes to fatal alphavirus encephalitis in mice.MethodsInfected animals were treated systemically with the angiotensin receptor-blocking drug, telmisartan, given its ability to cross the blood-brain barrier, selectively block type-1 angiotensin receptors (AT1R), and inhibit Nox-derived ROS production in vascular smooth muscle and other extraneural tissues. Clinical, virological, biochemical, and histopathological outcomes were followed over time.ResultsThe importance of the angiotensin II (Ang II)/AT1R axis in disease pathogenesis was confirmed by demonstrating increased Ang II levels in the CNS following infection, enhanced disease survival when CNS Ang II production was suppressed, increased AT1R expression on microglia and tissue-infiltrating myeloid cells, and enhanced disease survival in AT1R-deficient mice compared to wild-type (WT) controls. Systemic administration of telmisartan protected WT mice from lethal encephalitis caused by two different alphaviruses in a dose-dependent manner without altering virus replication or exerting any anti-inflammatory effects in the CNS. Infection triggered up-regulation of multiple Nox subunits in the CNS, while drug treatment inhibited local Nox activity, ROS production, and oxidative neuronal damage. Telmisartan proved ineffective in Nox-deficient mice, demonstrating that this enzyme is its main target in this experimental setting.ConclusionsNox-derived ROS, likely arising from CNS myeloid cells triggered by AT1R signaling, are pathogenic during fatal alphavirus encephalitis in mice. Systemically administered telmisartan at non-hypotensive doses targets Nox activity in the CNS to exert a neuroprotective effect. Disruption of this pathway may have broader implications for the treatment of related infections as well as for other CNS diseases driven by oxidative injury.Electronic supplementary materialThe online version of this article (doi:10.1186/s12974-016-0683-7) contains supplementary material, which is available to authorized users.

Infiltrating T lymphocytes reduce myeloid phagocytosis activity in synucleinopathy model.

BackgroundSynucleinopathies comprise a group of neurodegenerative diseases associated with abnormal accumulation of α-synuclein. One of the key factors that contribute to the progression of synucleinopathies is neuroinflammation. However, the role of lymphocytes in synucleinopathies like Parkinson’s disease (PD) remains largely unclear.MethodsTo investigate how lymphocytes impact synucleinopathies, human wild-type α-synuclein (WTS) transgenic mice were crossed with mice lacking mature lymphocytes (Rag2−/−). In this in vivo model, we quantified α-synuclein aggregation in the substantia nigra (SN) and striatum and determined the numbers of innate and adaptive immune cells in the central nervous system (CNS). The activation state of resident and infiltrated CNS myeloid cells (M1 vs. M2) was further classified by gene and protein expression analyses. The impact of T and B lymphocytes on the phagocytic activity of microglia in the presence of α-synuclein aggregates was addressed in BV2 microglia in vitro.ResultsCompared to WTS+ Rag2+/+ mice, where T but not B lymphocytes infiltrated the CNS, decreased amounts of α-synuclein aggregates were found in WTS+ Rag2−/− mice devoid of mature lymphocytes. The presence of T lymphocytes did not alter the number of Iba1+ microglia but increased the frequency of the CD11b+ CD45hi population in the CNS, indicative of an increased number of infiltrated macrophages. Moreover, the M1 phenotype was more prominent in WTS+ Rag2+/+ mice, whereas the M2 activation state was dominating in the absence of lymphocytes in WTS+ Rag2−/− mice. In vitro, in the presence of T but not B lymphocytes, significantly less α-synuclein was phagocytosed by BV2 microglia, further supporting the prevalence of the M1 phenotype in the presence of T lymphocytes.ConclusionsPeripheral T lymphocytes strongly contribute to increased α-synuclein pathology via modulation of CNS myeloid cell function. In the presence of T lymphocytes, microglia phagocytosis of aggregated α-synuclein is reduced, which increases the severity of synucleinopathy.Electronic supplementary materialThe online version of this article (doi:10.1186/s12974-016-0632-5) contains supplementary material, which is available to authorized users.

Central nervous system myeloid cells as drug targets: current status and translational challenges.

Myeloid cells of the central nervous system (CNS), which include parenchymal microglia, macrophages at CNS interfaces and monocytes recruited from the circulation during disease, are increasingly being recognized as targets for therapeutic intervention in neurological and psychiatric diseases. The origin of these cells in the immune system distinguishes them from ectodermal neurons and other glia and endows them with potential drug targets distinct from classical CNS target groups. However, despite the identification of several promising therapeutic approaches and molecular targets, no agents directly targeting these cells are currently available. Here, we assess strategies for targeting CNS myeloid cells and address key issues associated with their translation into the clinic.

Effects of fumarates on circulating and CNS myeloid cells in multiple sclerosis.

Dimethyl fumarate (DMF), a therapy for relapsing-remitting multiple sclerosis (RRMS), is implicated as acting on inflammatory and antioxidant responses within both systemic immune and/or central nervous system (CNS) compartments. Orally administered DMF is rapidly metabolized to monomethyl fumarate (MMF). Our aim was to analyze the impact of fumarates on antiinflammatory and antioxidant profiles of human myeloid cells found in the systemic compartment (monocytes) and in the inflamed CNS (blood-derived macrophages and brain-derived microglia). We analyzed cytokine and antioxidant expression in monocytes from untreated or DMF-treated RRMS patients and controls, and in monocyte-derived macrophages (MDMs) and microglia isolated from adult and fetal human brain tissue. Monocytes from multiple sclerosis (MS) patients receiving DMF had reduced expression of the proinflammatory micro-RNA miR-155 and of antioxidant genes HMOX1 and OSGIN1 compared to untreated MS patients; similar changes were observed in patients receiving FTY720 and/or natalizumab. In vitro addition of DMF but not MMF to MDMs and microglia inhibited lipopolysaccharide-induced production of inflammatory cytokines and increased expression of the antioxidant gene HMOX1 in the absence of significant cytotoxicity. Our in vivo-based observations that effects of DMF therapy on systemic myeloid cell gene expression are also observed with FTY720 and natalizumab therapy suggests that the effect may be indirect, reflecting reduced overall disease activity. Our in vitro results demonstrate significant effects of DMF but not MMF on inflammation and antioxidant responses by MDMs and microglia, questioning the mechanisms whereby DMF therapy would modulate myeloid cell properties within the CNS.

Systemic inflammation in early neonatal mice induces transient and lasting neurodegenerative effects.

BackgroundThe inflammatory mediator lipopolysaccharide (LPS) has been shown to induce acute gliosis in neonatal mice. However, the progressive effects on the murine neurodevelopmental program over the week that follows systemic inflammation are not known. Thus, we investigated the effects of repeated LPS administration in the first postnatal week in mice, a condition mimicking sepsis in late preterm infants, on the developing central nervous system (CNS).MethodsSystemic inflammation was induced by daily intraperitoneal administration (i.p.) of LPS (6 mg/kg) in newborn mice from postnatal day (PND) 4 to PND6. The effects on neurodevelopment were examined by staining the white matter and neurons with Luxol Fast Blue and Cresyl Violet, respectively. The inflammatory response was assessed by quantifying the expression/activity of matrix metalloproteinases (MMP), toll-like receptor (TLR)-4, high mobility group box (HMGB)-1, and autotaxin (ATX). In addition, B6 CX3CR1gfp/+ mice combined with cryo-immunofluorescence were used to determine the acute, delayed, and lasting effects on myelination, microglia, and astrocytes.ResultsLPS administration led to acute body and brain weight loss as well as overt structural changes in the brain such as cerebellar hypoplasia, neuronal loss/shrinkage, and delayed myelination. The impaired myelination was associated with alterations in the proliferation and differentiation of NG2 progenitor cells early after LPS administration, rather than with excessive phagocytosis by CNS myeloid cells. In addition to disruptions in brain architecture, a robust inflammatory response to LPS was observed. Quantification of inflammatory biomarkers revealed decreased expression of ATX with concurrent increases in HMGB1, TLR-4, and MMP-9 expression levels. Acute astrogliosis (GFAP+ cells) in the brain parenchyma and at the microvasculature interface together with parenchymal microgliosis (CX3CR1+ cells) were also observed. These changes preceded the migration/proliferation of CX3CR1+ cells around the vessels at later time points and the subsequent loss of GFAP+ astrocytes.ConclusionCollectively, our study has uncovered a complex innate inflammatory reaction and associated structural changes in the brains of neonatal mice challenged peripherally with LPS. These findings may explain some of the neurobehavioral abnormalities that develop following neonatal sepsis.

The p53 Transcriptional Network Influences Microglia Behavior and Neuroinflammation.

The tumor-suppressor protein p53 belongs to a family of proteins that play pivotal roles in multiple cellular functions including cell proliferation, cell death, genome stability, and regulation of inflammation. Neuroinflammation is a common feature of central nervous system (CNS) pathology, and microglia are the specialized resident population of CNS myeloid cells that initiate innate immune responses. Microglia maintain CNS homeostasis through pathogen containment, phagocytosis of debris, and initiation of tissue-repair cascades. However, an unregulated pro-inflammatory response can lead to tissue injury and dysfunction in both acute and chronic inflammatory states. Therefore, regulation of the molecular signals that control the induction, magnitude, and resolution of inflammation are necessary for optimal CNS health. We and others have described a novel mechanism by which p53 transcriptional activity modulates microglia behaviors in vitro and in vivo. Activation of p53 induces expression of microRNAs (miRNAs) that support microglia pro-inflammatory functions and suppress anti-inflammatory and tissue repair behaviors. In this review, we introduce the previously described roles of the p53 signaling network and discuss novel functions of p53 in the microglia-mediated inflammatory response in CNS health and disease. Ultimately, improved understanding of the molecular regulators modulated by p53 transcriptional activity in microglia will enhance the development of rational therapeutic strategies to harness the homeostatic and tissue repair functions of microglia.

Differential modulation of EAE by α9*‐ and β2*‐nicotinic acetylcholine receptors

Nicotine is a potent inhibitor of the immune response and is protective against experimental autoimmune encephalomyelitis (EAE). Initial studies suggested that the cholinergic system modulates inflammation via the α7-nicotinic acetylcholine receptor (nAChR) subtype. We recently have shown that effector T cells and myeloid cells constitutively express mRNAs encoding nAChR α9 and β2 subunits and found evidence for immune system roles for non-α7-nAChRs. In the present study, we assessed the effects of nAChR α9 or β2 subunit gene deletion on EAE onset and severity, with or without nicotine treatment. We report again that disease onset is delayed and severity is attenuated in nicotine-treated, wild-type mice, an effect that also is observed in α9 subunit knock-out (KO) mice irrespective of nicotine treatment. On the other hand, β2 KO mice fail to recover from peak measures of disease severity regardless of nicotine treatment, despite retaining sensitivity to nicotine’s attenuation of disease severity. Prior to disease onset, we found significantly less reactive oxygen species production in the CNS of β2 KO mice, elevated proportions of CNS myeloid cells but decreased ratios of CNS macrophages/microglia in α9 or β2 KO mice, and some changes in iNOS, TNF-α and IL-1β mRNA levels in α9 KO and/or β2 KO mice. Our data thus suggest that β2*- and α9*-nAChRs, in addition to α7-nAChRs, play different roles in endogenous and nicotine-dependent modulation of immune functions and could be exploited as therapeutic targets to modulate inflammation and autoimmunity.

Lentiviral-mediated administration of IL-25 in the CNS induces alternative activation of microglia

Interleukin-25 (IL-25) is the only anti-inflammatory cytokine of the IL-17 family, and it has been shown to be efficacious in inhibiting neuroinflammation. Known for its effects on cells of the adaptive immune system, it has been more recently described to be effective also on cells of the innate immune system, namely macrophages. We used a lentiviral-mediated gene therapy approach to deliver IL-25 to the central nervous system (CNS) in two mouse models of neuroinflammation, entorhinal cortex lesion and experimental autoimmune encephalomyelitis. In both, we found that IL-25 gene therapy was able to modulate CNS myeloid cells, either infiltrating macrophages or resident microglia, towards an anti-inflammatory, tissue-protective phenotype, as testified by the increase in markers such as Arginase-1 (Arg1), Mannose receptor 1 (CD206) and Chitinase 3-like 3 (Ym1). As a consequence, neuroinflammation was partly inhibited and the CNS protected from immune-mediated damage. To our knowledge, this is the first example of M2 shift (alternative activation) induced in vivo on CNS-resident myeloid cells by gene therapy, and may constitute a promising strategy to investigate the potential role of protective microglia in neurological disorders.

Licensing of myeloid cells promotes central nervous system autoimmunity and is controlled by peroxisome proliferator-activated receptor γ

During central nervous system autoimmunity, interactions between infiltrating immune cells and brain-resident cells are critical for disease progression and ultimately organ damage. Here, we demonstrate that local cross-talk between invading autoreactive T cells and auto-antigen-presenting myeloid cells within the central nervous system results in myeloid cell activation, which is crucial for disease progression during experimental autoimmune encephalomyelitis, the animal model of multiple sclerosis. This T cell-mediated licensing of central nervous system myeloid cells triggered astrocytic CCL2-release and promoted recruitment of inflammatory CCR2(+)-monocytes, which are the main effectors of disease progression. By employing a cell-specific knockout model, we identify the nuclear receptor peroxisome proliferator-activated receptor γ (PPARγ) in myeloid cells as key regulator of their disease-determining interactions with autoreactive T cells and brain-resident cells, respectively. LysM-PPARγ(KO) mice exhibited disease exacerbation during the effector phase of experimental autoimmune encephalomyelitis characterized by enhanced activation of central nervous system myeloid cells accompanied by pronounced local CCL2 production and inflammatory monocyte invasion, which finally resulted in increased demyelination and neuronal damage. Pharmacological PPARγ activation decreased antigen-specific T cell-mediated licensing of central nervous system myeloid cells, reduced myeloid cell-mediated neurotoxicity and hence dampened central nervous system autoimmunity. Importantly, human monocytes derived from patients with multiple sclerosis clearly responded to PPARγ-mediated control of proinflammatory activation and production of neurotoxic mediators. Furthermore, PPARγ in human monocytes restricted their capacity to activate human astrocytes leading to dampened astrocytic CCL2 production. Together, interference with the disease-promoting cross-talk between central nervous system myeloid cells, autoreactive T cells and brain-resident cells represents a novel therapeutic approach that limits disease progression and lesion development during ongoing central nervous system autoimmunity.

Selective Chemokine Receptor Usage by Central Nervous System Myeloid Cells in CCR2-Red Fluorescent Protein Knock-In Mice

BackgroundMonocyte subpopulations distinguished by differential expression of chemokine receptors CCR2 and CX3CR1 are difficult to track in vivo, partly due to lack of CCR2 reagents.Methodology/Principal FindingsWe created CCR2-red fluorescent protein (RFP) knock-in mice and crossed them with CX3CR1-GFP mice to investigate monocyte subset trafficking. In mice with experimental autoimmune encephalomyelitis, CCR2 was critical for efficient intrathecal accumulation and localization of Ly6Chi/CCR2hi monocytes. Surprisingly, neutrophils, not Ly6Clo monocytes, largely replaced Ly6Chi cells in the central nervous system of these mice. CCR2-RFP expression allowed the first unequivocal distinction between infiltrating monocytes/macrophages from resident microglia.Conclusion/SignificanceThese results refine the concept of monocyte subsets, provide mechanistic insight about monocyte entry into the central nervous system, and present a novel model for imaging and quantifying inflammatory myeloid populations.