GPR109A expressed on medullary thymic epithelial cells affects thymic Treg development.

Regulatory T cells (Treg) maintain immune homeostasis due to their anti-inflammatory functions. They can be generated either centrally in the thymus or in peripheral organs. Metabolites such as short-chain fatty acids produced by intestinal microbiota can induce peripheral Treg differentiation, by activating G-protein-coupled-receptors like GPR109A. In this study, we identified a novel role for GPR109A in thymic Treg development. We found that Gpr109a-/- mice had increased Treg under basal conditions in multiple organs compared with WT mice. GPR109A was not expressed on T cells but on medullary thymic epithelial cells (mTECs), as revealed by single-cell RNA sequencing in both mice and humans and confirmed by flow cytometry in mice. mTECs isolated from Gpr109a-/- mice had higher expression of autoimmune regulator (AIRE), the key regulator of Treg development, while the subset of mTECs that did not express Gpr109a in the WT displayed increased Aire expression and also enhanced signaling related to mTEC functionality. Increased thymic Treg in Gpr109a-/- mice was associated with protection from experimental autoimmune encephalomyelitis, with ameliorated clinical signs and reduced inflammation. This work identifies a novel role for GPR109A and possibly the gut microbiota, on thymic Treg development via its regulation of mTECs.

NK cell profiling in West Nile virus encephalitis reveals potential metabolic basis for functional inhibition.

Natural killer (NK) cells are cytotoxic lymphocytes important for viral defense. West Nile virus (WNV) infection of the central nervous system (CNS) causes marked recruitment of bone marrow (BM)-derived monocytes, T cells and NK cells, resulting in severe neuroinflammation and brain damage. Despite substantial numbers of NK cells in the CNS, their function and phenotype remain largely unexplored. Here, we demonstrate that NK cells mature from the BM to the brain, upregulate inhibitory receptors and show reduced cytokine production and degranulation, likely due to the increased expression of the inhibitory NK cell molecule, MHC-I. Intriguingly, this correlated with a reduction in metabolism associated with cytotoxicity in brain-infiltrating NK cells. Importantly, the degranulation and killing capability were restored in NK cells isolated from WNV-infected tissue, suggesting that WNV-induced NK cell inhibition occurs in the CNS. Overall, this work identifies a potential link between MHC-I inhibition of NK cells and metabolic reduction of their cytotoxicity during infection.

Interleukin-6 and interferon-alpha differentially regulate microglia function.

Previous reports have shown that IL-6 and IFN-⍺ induce distinct transcriptomic and morphological changes in microglia. Here, we demonstrate that IL-6 increases tissue surveillance, migration and phagocytosis in primary murine microglia, whereas IFN-⍺ inhibits these functions. Our results provide a crucial link between transcriptome and function. It holds the potential to serve as the foundation for future studies aimed at identifying therapeutic targets for cytokine-mediated neuroinflammatory diseases.

Integrating transcriptomic datasets across neurological disease identifies unique myeloid subpopulations driving disease-specific signatures.

Microglia and bone marrow-derived monocytes are key elements of central nervous system (CNS) inflammation, both capable of enhancing and dampening immune-mediated pathology. However, the study-specific focus on individual cell types, disease models or experimental approaches has limited our ability to infer common and disease-specific responses. This meta-analysis integrates bulk and single-cell transcriptomic datasets of microglia and monocytes from disease models of autoimmunity, neurodegeneration, sterile injury, and infection to build a comprehensive resource connecting myeloid responses across CNS disease. We demonstrate that the bulk microglial and monocyte program is highly contingent on the disease environment, challenging the notion of a universal microglial disease signature. Integration of six single-cell RNA-sequencing datasets revealed that these disease-specific signatures are likely driven by differing proportions of unique myeloid subpopulations that were individually expanded in different disease settings. These subsets were functionally-defined as neurodegeneration-associated, inflammatory, interferon-responsive, phagocytic, antigen-presenting, and lipopolysaccharide-responsive cellular states, revealing a core set of myeloid responses at the single-cell level that are conserved across CNS pathology. Showcasing the predictive and practical value of this resource, we performed differential expression analysis on microglia and monocytes across disease and identified Cd81 as a new neuroinflammatory-stable gene that accurately identified microglia and distinguished them from monocyte-derived cells across all experimental models at both the bulk and single-cell level. Together, this resource dissects the influence of disease environment on shared immune response programmes to build a unified perspective of myeloid behavior across CNS pathology.

Early microglial response, myelin deterioration and lethality in mice deficient for very long chain ceramide synthesis in oligodendrocytes.

The sphingolipids galactosylceramide (GalCer), sulfatide (ST) and sphingomyelin (SM) are essential for myelin stability and function. GalCer and ST are synthesized mostly from C22-C24 ceramides, generated by Ceramide Synthase 2 (CerS2). To clarify the requirement for C22-C24 sphingolipid synthesis in myelin biosynthesis and stability, we generated mice lacking CerS2 specifically in myelinating cells (CerS2ΔO/ΔO ). At 6 weeks of age, normal-appearing myelin had formed in CerS2ΔO/ΔO mice, however there was a reduction in myelin thickness and the percentage of myelinated axons. Pronounced loss of C22-C24 sphingolipids in myelin of CerS2ΔO/ΔO mice was compensated by greatly increased levels of C18 sphingolipids. A distinct microglial population expressing high levels of activation and phagocytic markers such as CD64, CD11c, MHC class II, and CD68 was apparent at 6 weeks of age in CerS2ΔO/ΔO mice, and had increased by 10 weeks. Increased staining for denatured myelin basic protein was also apparent in 6-week-old CerS2ΔO/ΔO mice. By 16 weeks, CerS2ΔO/ΔO mice showed pronounced myelin atrophy, motor deficits, and axon beading, a hallmark of axon stress. 90% of CerS2ΔO/ΔO mice died between 16 and 26 weeks of age. This study highlights the importance of sphingolipid acyl chain length for the structural integrity of myelin, demonstrating how a modest reduction in lipid chain length causes exposure of a denatured myelin protein epitope and expansion of phagocytic microglia, followed by axon pathology, myelin degeneration, and motor deficits. Understanding the molecular trigger for microglial activation should aid the development of therapeutics for demyelinating and neurodegenerative diseases.

Targeting inflammatory monocytes by immune-modifying nanoparticles prevents acute kidney allograft rejection.

Inflammatory monocytes are a major component of the cellular infiltrate in acutely rejecting human kidney allografts. Since immune-modifying nanoparticles (IMPs) bind to circulating inflammatory monocytes via the specific scavenger receptor MARCO, causing diversion to the spleen and subsequent apoptosis, we investigated the therapeutic potential of negatively charged, 500-nm diameter polystyrene IMPs to prevent kidney allograft rejection. Kidney transplants were performed from BALB/c (H2d) to C57BL/6 (H2b) mice in two groups: controls (allo) and allo mice infused with IMPs. Groups were studied for 14 (acute rejection) or 100 (chronic rejection) days. Allo mice receiving IMPs exhibited superior survival and markedly less acute rejection, with better kidney function, less tubulitis, and diminished inflammatory cell density, cytokine and cytotoxic molecule expression in the allograft and lower titers of donor-specific IgG2c antibody in serum at day 14, as compared to allo mice. Cells isolated from kidneys from allo mice receiving IMPs showed reduced Ly6Chi monocytes, CD11b+ cells and NKT+ cells compared to allo mice. IMPs predominantly bound CD11b+ cells in the bloodstream and CD11b+ and CD11c-B220+ marginal zone B cells in the spleen. In the spleen, IMPs were found predominantly in red pulp, colocalized with MARCO and expression of cleaved caspase-3. At day 100, allo mice receiving IMPs exhibited reduced macrophage M1 responses but were not protected from chronic rejection. IMPs afforded significant protection from acute rejection, inhibiting both innate and adaptive alloimmunity. Thus, our current experimental findings, coupled with our earlier demonstration of IMP-induced protection in kidney ischemia-reperfusion injury, identify IMPs as a potential induction agent in kidney transplantation.

Integration, exploration, and analysis of high-dimensional single-cell cytometry data using Spectre.

As the size and complexity of high-dimensional (HD) cytometry data continue to expand, comprehensive, scalable, and methodical computational analysis approaches are essential. Yet, contemporary clustering and dimensionality reduction tools alone are insufficient to analyze or reproduce analyses across large numbers of samples, batches, or experiments. Moreover, approaches that allow for the integration of data across batches or experiments are not well incorporated into computational toolkits to allow for streamlined workflows. Here we present Spectre, an R package that enables comprehensive end-to-end integration and analysis of HD cytometry data from different batches or experiments. Spectre streamlines the analytical stages of raw data pre-processing, batch alignment, data integration, clustering, dimensionality reduction, visualization, and population labelling, as well as quantitative and statistical analysis. Critically, the fundamental data structures used within Spectre, along with the implementation of machine learning classifiers, allow for the scalable analysis of very large HD datasets, generated by flow cytometry, mass cytometry, or spectral cytometry. Using open and flexible data structures, Spectre can also be used to analyze data generated by single-cell RNA sequencing or HD imaging technologies, such as Imaging Mass Cytometry. The simple, clear, and modular design of analysis workflows allow these tools to be used by bioinformaticians and laboratory scientists alike. Spectre is available as an R package or Docker container. R code is available on Github (https://github.com/immunedynamics/spectre).

Microglia and monocytes in inflammatory CNS disease: integrating phenotype and function.

In neurological diseases, the actions of microglia, the resident myeloid cells of the CNS parenchyma, may diverge from, or intersect with, those of recruited monocytes to drive immune-mediated pathology. However, defining the precise roles of each cell type has historically been impeded by the lack of discriminating markers and experimental systems capable of accurately identifying them. Our ability to distinguish microglia from monocytes in neuroinflammation has advanced with single-cell technologies, new markers and drugs that identify and deplete them, respectively. Nevertheless, the focus of individual studies on particular cell types, diseases or experimental approaches has limited our ability to connect phenotype and function more widely and across diverse CNS pathologies. Here, we critically review, tabulate and integrate the disease-specific functions and immune profiles of microglia and monocytes to provide a comprehensive atlas of myeloid responses in viral encephalitis, demyelination, neurodegeneration and ischemic injury. In emphasizing the differential roles of microglia and monocytes in the severe neuroinflammatory disease of viral encephalitis, we connect inflammatory pathways common to equally incapacitating diseases with less severe inflammation. We examine these findings in the context of human studies and highlight the benefits and inherent limitations of animal models that may impede or facilitate clinical translation. This enables us to highlight common and contrasting, non-redundant and often opposing roles of microglia and monocytes in disease that could be targeted therapeutically.

High-parameter cytometry unmasks microglial cell spatio-temporal response kinetics in severe neuroinflammatory disease.

BACKGROUND: Differentiating infiltrating myeloid cells from resident microglia in neuroinflammatory disease is challenging, because bone marrow-derived inflammatory monocytes infiltrating the inflamed brain adopt a 'microglia-like' phenotype. This precludes the accurate identification of either cell type without genetic manipulation, which is important to understand their temporal contribution to disease and inform effective intervention in its pathogenesis. During West Nile virus (WNV) encephalitis, widespread neuronal infection drives substantial CNS infiltration of inflammatory monocytes, causing severe immunopathology and/or death, but the role of microglia in this remains unclear. METHODS: Using high-parameter cytometry and dimensionality-reduction, we devised a simple, novel gating strategy to identify microglia and infiltrating myeloid cells during WNV-infection. Validating our strategy, we (1) blocked the entry of infiltrating myeloid populations from peripheral blood using monoclonal blocking antibodies, (2) adoptively transferred BM-derived monocytes and tracked their phenotypic changes after infiltration and (3) labelled peripheral leukocytes that infiltrate into the brain with an intravenous dye. We demonstrated that myeloid immigrants populated only the identified macrophage gates, while PLX5622 depletion reduced all 4 subsets defined by the microglial gates. RESULTS: Using this gating approach, we identified four consistent microglia subsets in the homeostatic and WNV-infected brain. These were P2RY12hi CD86-, P2RY12hi CD86+ and P2RY12lo CD86- P2RY12lo CD86+. During infection, 2 further populations were identified as 'inflammatory' and 'microglia-like' macrophages, recruited from the bone marrow. Detailed kinetic analysis showed significant increases in the proportions of both P2RY12lo microglia subsets in all anatomical areas, largely at the expense of the P2RY12hi CD86- subset, with the latter undergoing compensatory proliferation, suggesting replenishment of, and differentiation from this subset in response to infection. Microglia altered their morphology early in infection, with all cells adopting temporal and regional disease-specific phenotypes. Late in disease, microglia produced IL-12, downregulated CX3CR1, F4/80 and TMEM119 and underwent apoptosis. Infiltrating macrophages expressed both TMEM119 and P2RY12 de novo, with the microglia-like subset notably exhibiting the highest proportional myeloid population death. CONCLUSIONS: Our approach enables detailed kinetic analysis of resident vs infiltrating myeloid cells in a wide range of neuroinflammatory models without non-physiological manipulation. This will more clearly inform potential therapeutic approaches that specifically modulate these cells.

High-Dimensional Methods of Single-Cell Microglial Profiling to Enhance Understanding of Neuropathological Disease.

Microglia are the innate myeloid cells of the central nervous system (CNS) parenchyma, functionally implicated in almost every defined neuroinflammatory and neurodegenerative disorder. Current understanding of disease pathogenesis for many neuropathologies is limited and/or lacks reliable diagnostic markers, vaccines, and treatments. With the increasing aging of society and rise in neurogenerative diseases, improving our understanding of their pathogenesis is essential. Analysis of microglia from murine disease models provides an investigative tool to unravel disease processes. In many neuropathologies, bone-marrow-derived monocytes are recruited to the CNS, adopting a phenotype similar to that of microglia. This significantly confounds the accurate identification of cell-type-specific functions and downstream therapeutic targeting. The increased capacity to analyze more phenotypic markers using spectral-cytometry-based technologies allows improved separation of microglia from monocyte-derived cells. Full-spectrum profiling enables enhanced marker resolution, time-efficient analysis of >40 fluorescence parameters, and extraction of cellular autofluorescence parameters. Coupling this system with additional cytometric technologies, including cell sorting and high-parameter imaging, can improve the understanding of microglial phenotypes in disease. To this end, we provide detailed, step-by-step protocols for the analysis of murine brain tissue by high-parameter ex vivo cytometric analysis using the Aurora spectral cytometer (Cytek), including best practices for unmixing and autofluorescence extraction, cell sorting for single-cell RNA analysis, and imaging mass cytometry. Together, this provides a toolkit for researchers to comprehensively investigate microglial disease processes at protein, RNA, and spatial levels for the identification of therapeutic targets in neuropathology. © 2024 The Authors. Current Protocols published by Wiley Periodicals LLC. Basic Protocol 1: Processing the mouse brain into a single-cell suspension for microglia isolation Basic Protocol 2: Staining single-cell mouse brain suspensions for microglial phenotyping by spectral cytometry Basic Protocol 3: Flow cytometric sorting of mouse microglia for ex vivo analysis Basic Protocol 4: Processing the mouse brain for imaging mass cytometry for spatial microglia analysis.

CD8+ T Cells Mediate Lethal Lung Pathology in the Absence of PD-L1 and Type I Interferon Signalling following LCMV Infection.

CD8+ T cells are critical to the adaptive immune response against viral pathogens. However, overwhelming antigen exposure can result in their exhaustion, characterised by reduced effector function, failure to clear virus, and the upregulation of inhibitory receptors, including programmed cell death 1 (PD-1). However, exhausted T cell responses can be "re-invigorated" by inhibiting PD-1 or the primary ligand of PD-1: PD-L1. Further, the absence of the type I interferon receptor IFNAR1 also results in T cell exhaustion and virus persistence in lymphocytic choriomeningitis virus Armstrong (LCMV-Arm)-infected mice. In this study, utilizing single- and double-knockout mice, we aimed to determine whether ablation of PD-1 could restore T cell functionality in the absence of IFNAR1 signalling in LCMV-Arm-infected mice. Surprisingly, this did not re-invigorate the T cell response and instead, it converted chronic LCMV-Arm infection into a lethal disease characterized by severe lung inflammation with an infiltration of neutrophils and T cells. Depletion of CD8+ T cells, but not neutrophils, rescued mice from lethal disease, demonstrating that IFNAR1 is required to prevent T cell exhaustion and virus persistence in LCMV-Arm infection, and in the absence of IFNAR1, PD-L1 is required for survival. This reveals an important interplay between IFNAR1 and PD-L1 with implications for therapeutics targeting these pathways.

Understanding machine learning applications in dementia research and clinical practice: a review for biomedical scientists and clinicians.

Several (inter)national longitudinal dementia observational datasets encompassing demographic information, neuroimaging, biomarkers, neuropsychological evaluations, and muti-omics data, have ushered in a new era of potential for integrating machine learning (ML) into dementia research and clinical practice. ML, with its proficiency in handling multi-modal and high-dimensional data, has emerged as an innovative technique to facilitate early diagnosis, differential diagnosis, and to predict onset and progression of mild cognitive impairment and dementia. In this review, we evaluate current and potential applications of ML, including its history in dementia research, how it compares to traditional statistics, the types of datasets it uses and the general workflow. Moreover, we identify the technical barriers and challenges of ML implementations in clinical practice. Overall, this review provides a comprehensive understanding of ML with non-technical explanations for broader accessibility to biomedical scientists and clinicians.

Clodronate is not protective in lethal viral encephalitis despite substantially reducing inflammatory monocyte infiltration in the CNS.

Bone marrow (BM)-derived monocytes induce inflammation and tissue damage in a range of pathologies. In particular, in a mouse model of West Nile virus (WNV) encephalitis (WNE), nitric oxide-producing, Ly6Chi inflammatory monocytes from the BM are recruited to the central nervous system (CNS) and contribute to lethal immune pathology. Reducing the migration of these cells into the CNS using monoclonal antibody blockade, immune-modifying particles or CSF-1R inhibitors reduces neuroinflammation, improving survival and/or clinical outcomes. Macrophages can also be targeted more broadly by administration of clodronate-encapsulated liposomes, which induce apoptosis in phagocytes. In this study, clodronate reduced the inflammatory infiltrate by 70% in WNE, however, surprisingly, this had no effect on disease outcome. More detailed analysis demonstrated a compensatory increase in neutrophils and enhanced activation status of microglia in the brain. In addition, we observed increased numbers of Ly6Chi BM monocytes with an increased proliferative capacity and expression of SCA-1 and CD16/32, potentially indicating output of immature cells from the BM. Once in the brain, these cells were more phagocytic and had a reduced expression of antigen-presenting molecules. Lastly, we show that clodronate also reduces non-myeloid cells in the spleen and BM, as well as ablating red blood cells and their proliferation. These factors likely impeded the therapeutic potential of clodronate in WNE. Thus, while clodronate provides an excellent system to deplete macrophages in the body, it has larger and broader effects on the phagocytic and non-phagocytic system, which must be considered in the interpretation of data.

Temporal tracking of microglial and monocyte single-cell transcriptomics in lethal flavivirus infection.

As the resident parenchymal myeloid population in the central nervous system (CNS), microglia are strategically positioned to respond to neurotropic virus invasion and have been implicated in promoting both disease resolution and progression in the acute and post-infectious phase of virus encephalitis. In a mouse model of West Nile virus encephalitis (WNE), infection of the CNS results in recruitment of large numbers of peripheral immune cells into the brain, the majority being nitric oxide (NO)-producing Ly6Chi inflammatory monocyte-derived cells (MCs). In this model, these cells enhance immunopathology and mortality. However, the contribution of microglia to this response is currently undefined. Here we used a combination of experimental tools, including single-cell RNA sequencing (scRNA-seq), microglia and MC depletion reagents, high-dimensional spectral cytometry and computational algorithms to dissect the differential contribution of microglia and MCs to the anti-viral immune response in severe neuroinflammation seen in WNE. Intriguingly, analysis of scRNA-seq data revealed 6 unique microglia and 3 unique MC clusters that were predominantly timepoint-specific, demonstrating substantial transcriptional adaptation with disease progression over the course of WNE. While microglia and MC adopted unique gene expression profiles, gene ontology enrichment analysis, coupled with microglia and MC depletion studies, demonstrated a role for both of these cells in the trafficking of peripheral immune cells into the CNS, T cell responses and viral clearance. Over the course of infection, microglia transitioned from a homeostatic to an anti-viral and then into an immune cell-recruiting phenotype. Conversely, MC adopted antigen-presenting, immune cell-recruiting and NO-producing phenotypes, which all had anti-viral function. Overall, this study defines for the first time the single-cell transcriptomic responses of microglia and MCs over the course of WNE, demonstrating both protective and pathological roles of these cells that could potentially be targeted for differential therapeutic intervention to dampen immune-mediated pathology, while maintaining viral clearance functions.

PLX5622 Reduces Disease Severity in Lethal CNS Infection by Off-Target Inhibition of Peripheral Inflammatory Monocyte Production.

PLX5622 is a CSF-1R inhibitor and microglia-depleting reagent, widely used to investigate the biology of this central nervous system (CNS)-resident myeloid population, but the indirect or off-target effects of this agent remain largely unexplored. In a murine model of severe neuroinflammation induced by West Nile virus encephalitis (WNE), we showed PLX5622 efficiently depleted both microglia and a sub-population of border-associated macrophages in the CNS. However, PLX5622 also significantly depleted mature Ly6Chi monocytes in the bone marrow (BM), inhibiting their proliferation and lethal recruitment into the infected brain, reducing neuroinflammation and clinical disease scores. Notably, in addition, BM dendritic cell subsets, plasmacytoid DC and classical DC, were depleted differentially in infected and uninfected mice. Confirming its protective effect in WNE, cessation of PLX5622 treatment exacerbated disease scores and was associated with robust repopulation of microglia, rebound BM monopoiesis and markedly increased inflammatory monocyte infiltration into the CNS. Monoclonal anti-CSF-1R antibody blockade late in WNE also impeded BM monocyte proliferation and recruitment to the brain, suggesting that the protective effect of PLX5622 is via the inhibition of CSF-1R, rather than other kinase targets. Importantly, BrdU incorporation in PLX5622-treated mice, suggest remaining microglia proliferate independently of CSF-1 in WNE. Our study uncovers significantly broader effects of PLX5622 on the myeloid lineage beyond microglia depletion, advising caution in the interpretation of PLX5622 data as microglia-specific. However, this work also strikingly demonstrates the unexpected therapeutic potential of this molecule in CNS viral infection, as well as other monocyte-mediated diseases.

Impact of Dietary Fiber on West Nile Virus Infection.

Dietary fiber supports healthy gut bacteria and their production of short-chain fatty acids (SCFA), which promote anti-inflammatory cell development, in particular, regulatory T cells. It is thus beneficial in many diseases, including influenza infection. While disruption of the gut microbiota by antibiotic treatment aggravates West Nile Virus (WNV) disease, whether dietary fiber is beneficial is unknown. WNV is a widely-distributed neurotropic flavivirus that recruits inflammatory monocytes into the brain, causing life-threatening encephalitis. To investigate the impact of dietary fiber on WNV encephalitis, mice were fed on diets deficient or enriched with dietary fiber for two weeks prior to inoculation with WNV. To induce encephalitis, mice were inoculated intranasally with WNV and maintained on these diets. Despite increased fecal SCFA acetate and changes in gut microbiota composition, dietary fiber did not affect clinical scores, leukocyte infiltration into the brain, or survival. After the brain, highest virus loads were measured in the colon in neurons of the submucosal and myenteric plexuses. Associated with this, there was disrupted gut homeostasis, with shorter colon length and higher local inflammatory cytokine levels, which were not affected by dietary fiber. Thus, fiber supplementation is not effective in WNV encephalitis.

Ocular tropism of SARS-CoV-2 in animal models with retinal inflammation via neuronal invasion following intranasal inoculation.

Although ocular manifestations are reported in patients with COVID-19, consensus on ocular tropism of SARS-CoV-2 is lacking. Here, we infect K18-hACE2 transgenic mice with SARS-CoV-2 using various routes. We observe ocular manifestation and retinal inflammation with production of pro-inflammatory cytokines in the eyes of intranasally (IN)-infected mice. Intratracheal (IT) infection results in dissemination of the virus from the lungs to the brain and eyes via trigeminal and optic nerves. Ocular and neuronal invasions are confirmed using intracerebral (IC) infection. Notably, the eye-dropped (ED) virus does not cause lung infection and becomes undetectable with time. Ocular and neurotropic distribution of the virus in vivo is evident in fluorescence imaging with an infectious clone of SARS-CoV-2-mCherry. The ocular tropic and neuroinvasive characteristics of SARS-CoV-2 are confirmed in wild-type Syrian hamsters. Our data can improve the understanding regarding viral transmission and clinical characteristics of SARS-CoV-2 and help in improving COVID-19 control procedures.

Immovable Object Meets Unstoppable Force? Dialogue Between Resident and Peripheral Myeloid Cells in the Inflamed Brain.

Inflammation of the brain parenchyma is characteristic of neurodegenerative, autoimmune, and neuroinflammatory diseases. During this process, microglia, which populate the embryonic brain and become a permanent sentinel myeloid population, are inexorably joined by peripherally derived monocytes, recruited by the central nervous system. These cells can quickly adopt a morphology and immunophenotype similar to microglia. Both microglia and monocytes have been implicated in inducing, enhancing, and/or maintaining immune-mediated pathology and thus disease progression in a number of neuropathologies. For many years, experimental and analytical systems have failed to differentiate resident microglia from peripherally derived myeloid cells accurately. This has impeded our understanding of their precise functions in, and contributions to, these diseases, and hampered the development of novel treatments that could target specific cell subsets. Over the past decade, microglia have been investigated more intensively in the context of neuroimmunological research, fostering the development of more precise experimental systems. In light of our rapidly growing understanding of these cells, we discuss the differential origins of microglia and peripherally derived myeloid cells in the inflamed brain, with an analysis of the problems resolving these cell types phenotypically and morphologically, and highlight recent developments enabling more precise identification.