Microglia have critical roles not only in neural development and homeostasis, but also in neurodegenerative and neuroinflammatory diseases of the central nervous system
. These highly diverse and ...specialized functions may be executed by subsets of microglia that already exist in situ, or by specific subsets of microglia that develop from a homogeneous pool of cells on demand. However, little is known about the presence of spatially and temporally restricted subclasses of microglia in the central nervous system during development or disease. Here we combine massively parallel single-cell analysis, single-molecule fluorescence in situ hybridization, advanced immunohistochemistry and computational modelling to comprehensively characterize subclasses of microglia in multiple regions of the central nervous system during development and disease. Single-cell analysis of tissues of the central nervous system during homeostasis in mice revealed specific time- and region-dependent subtypes of microglia. Demyelinating and neurodegenerative diseases evoked context-dependent subtypes of microglia with distinct molecular hallmarks and diverse cellular kinetics. Corresponding clusters of microglia were also identified in healthy human brains, and the brains of patients with multiple sclerosis. Our data provide insights into the endogenous immune system of the central nervous system during development, homeostasis and disease, and may also provide new targets for the treatment of neurodegenerative and neuroinflammatory pathologies.
Transmission of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) from humans to farmed mink has been observed in Europe and the US. In the infected animals, viral variants arose that ...harbored mutations in the spike (S) protein, the target of neutralizing antibodies, and these variants were transmitted back to humans. This raised concerns that mink might become a constant source of human infection with SARS-CoV-2 variants associated with an increased threat to human health and resulted in mass culling of mink. Here, we report that mutations frequently found in the S proteins of SARS-CoV-2 from mink are mostly compatible with efficient entry into human cells and its inhibition by soluble angiotensin-converting enzyme 2 (ACE2). In contrast, mutation Y453F reduces neutralization by an antibody with emergency use authorization for coronavirus disease 2019 (COVID-19) therapy and sera/plasma from COVID-19 patients. These results suggest that antibody responses induced upon infection or certain antibodies used for treatment might offer insufficient protection against SARS-CoV-2 variants from mink.
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•SARS-CoV-2 from mink harbor up to five mutations in the spike protein•Entry inhibitors under clinical evaluation block mink spike proteins•Mutation Y453F confers partial escape from a therapeutic antibody•Y453F allows evasion of antibodies induced by SARS-CoV-2 infection of humans
Transmission of SARS-CoV-2 between humans and farmed mink has caused concern because viruses from mink have acquired mutations in the spike protein. Hoffmann et al. show that these mink-specific mutations do not increase entry into human cells but do cause partial evasion from neutralization by a therapeutic antibody and convalescent plasma/sera.
Low female mating frequencies often appear to be cases of direct male induction that can oppose female interests. Mating plugs are most obvious means leading to low degrees of multiple mating in ...females. In spiders, mating plugs are formed by a variety of amorphous materials, by the breakage of the male sperm transferring organ, or by the whole male that functions as a mating barrier. Our compilation of the available information on the presence of the various types of mating plugs suggests that plugs predominantly occur in entelegyne spiders. In this group, plugs do not interfere with oviposition since separate openings for insemination and oviposition are present. In contrast, mating plugs seem to be rare in haplogyne spiders that do not possess separate openings. The available experimental studies on the function of the different types of plugs suggest that plugs can be considered as male adaptations to avoid sperm competition. However, females in some cases were shown to have evolved means to prevent or control male manipulation or may selectively favour plug production in specific males, an aspect which has largely been neglected. In order to understand plug evolution and function we need to explore the morphological, behavioural and biochemical aspects involved and extend our approach to interactions between the sexes.
Objective
The serum of most neuromyelitis optica (NMO) patients contains autoantibodies (NMO‐IgGs) directed against the aquaporin‐4 (AQP4) water channel located on astrocyte foot processes in the ...perivessel and subpial areas of the brain. Our objectives were to determine the source of central nervous system (CNS) NMO‐IgGs and their role in disease pathogenesis.
Methods
Fluorescence‐activated cell sorting and single‐cell reverse transcriptase polymerase chain reaction were used to identify overrepresented plasma cell immunoglobulin (Ig) sequences in the cerebrospinal fluid (CSF) of an NMO patient after a first clinical attack. Monoclonal recombinant antibodies (rAbs) were generated from the paired heavy and light chain sequences and tested for target specificity and Fc effector function. The effect of CSF rAbs on CNS immunopathology was investigated by delivering single rAbs to rats with experimental autoimmune encephalomyelitis (EAE).
Results
Repertoire analysis revealed a dynamic, clonally expanded plasma cell population with features of an antigen‐targeted response. Using multiple independent assays, 6 of 11 rAbs generated from CSF plasma cell clones specifically bound to AQP4. AQP4‐specific rAbs recognized conformational epitopes and mediated both AQP4‐directed antibody‐dependent cellular cytotoxicity and complement‐mediated lysis. When administered to rats with EAE, an AQP4‐specific NMO CSF rAb induced NMO immunopathology: perivascular astrocyte depletion, myelinolysis, and complement and Ig deposition.
Interpretation
Molecular characterization of the CSF plasma cell repertoire in an early NMO patient demonstrates that AQP4‐specfic Ig is synthesized intrathecally at disease onset and directly contributes to CNS pathology. AQP4 is now the first confirmed antigenic target in human demyelinating disease. Ann Neurol 2009;66:617–629
Approximately 80% of neuromyelitis optica spectrum disorder (NMOSD) patients harbor serum anti-aquaporin-4 autoantibodies targeting astrocytes in the CNS. Crucial for NMOSD lesion initiation is ...disruption of the blood-brain barrier (BBB), which allows the entrance of Abs and serum complement into the CNS and which is a target for new NMOSD therapies. Astrocytes have important functions in BBB maintenance; however, the influence of their loss and the role of immune cell infiltration on BBB permeability in NMOSD have not yet been investigated. Using an experimental model of targeted NMOSD lesions in rats, we demonstrate that astrocyte destruction coincides with a transient disruption of the BBB and a selective loss of occludin from tight junctions. It is noteworthy that BBB integrity is reestablished before astrocytes repopulate. Rather than persistent astrocyte loss, polymorphonuclear leukocytes (PMNs) are the main mediators of BBB disruption, and their depletion preserves BBB integrity and prevents astrocyte loss. Inhibition of PMN chemoattraction, activation, and proteolytic function reduces lesion size. In summary, our data support a crucial role for PMNs in BBB disruption and NMOSD lesion development, rendering their recruitment and activation promising therapeutic targets.
Magnetic resonance imaging (MRI) is the gold standard for the detection of multiple sclerosis (MS) lesions. However, current MRI techniques provide little information about the structural features of ...a brain lesion with inflammatory cell infiltration, demyelination, gliosis, acute axonal damage and axonal loss. To identify methods for a differentiation of demyelination, inflammation, and axonal damage we developed a novel mouse model combining cuprizone-induced demyelination and experimental autoimmune encephalomyelitis. MS-like brain lesions were assessed by T1-weighted, T2-weighted, and magnetization transfer MRI as well as by diffusion tensor imaging (DTI). T2-weighted MRI differentiated control and diseased mice, while T1-weighted MRI better reflected the extent of inflammation and axonal damage. In DTI, axonal damage and cellular infiltration led to a reduction of the axial diffusivity, whereas primary demyelination after cuprizone treatment was reflected by changes in radial but not axial diffusivity. Importantly, alterations in radial diffusivity were less pronounced in mice with demyelination, inflammation, and acute axonal damage, indicating that radial diffusivity may underestimate demyelination in acute MS lesions. In conclusion, the combined information from different DTI parameters allows for a more precise identification of solely demyelinated lesions versus demyelinated and acutely inflamed lesions. These findings are of relevance for offering individualized, stage-adapted therapies for MS patients.
Modeling chronic cortical demyelination allows the study of long-lasting pathological changes observed in multiple sclerosis such as failure of remyelination, chronically disturbed functions of ...oligodendrocytes, neurons and astrocytes, brain atrophy and cognitive impairments. We aimed at generating an animal model for studying the consequences of chronic cortical demyelination and meningeal inflammation. To induce long-lasting cortical demyelination and chronic meningeal inflammation, we immunized female Lewis rats against myelin oligodendrocyte glycoprotein (MOG) and injected lentiviruses for continuing overexpression of the cytokines TNFα and IFNγ in the cortical brain parenchyma. Immunization with MOG and overexpression of TNFα and IFNγ led to widespread subpial demyelination and meningeal inflammation that were stable for at least 10 weeks. We demonstrate here that immunization with MOG is necessary for acute as well as chronic cortical demyelination. In addition, long-lasting overexpression of TNFα and IFNγ in the brain parenchyma is sufficient to induce chronic meningeal inflammation. Our model simulates key features of chronic cortical demyelination and inflammation, reminiscent of human multiple sclerosis pathology. This will allow molecular, cellular and functional investigations for a better understanding of the adaptation mechanisms of the cerebral cortex in multiple sclerosis.
Individuals with impaired immunity are more susceptible to infections than immunocompetent subjects. No vaccines are currently available to induce protection against E. coli meningoencephalitis. This ...study evaluated the potential of poly(I:C) pre-treatment to induce trained immunity. Poly(I:C) was administered as a non-specific stimulus of innate immune responses to protect immunocompetent and neutropenic wild-type mice from a subsequent challenge by the intracranial injection of E. coli K1.
Three days prior to infection, mice received an intraperitoneal injection of poly(I:C) or vehicle. Kaplan-Meier survival curves were analyzed. In short-term experiments, bacterial titers and the inflammatory response were characterized in the blood, cerebellum, and spleen homogenates. NK cell subpopulations in the brain and spleen were analyzed by flow cytometry. Numbers of microglia and activation scores were evaluated by histopathology.
Pre-treatment with 200 μg poly(I:C) increased survival time, reduced mortality, and enhanced bacterial clearance in the blood, cerebellum, and spleen at early infection in neutropenic mice. Poly(I:C)-mediated protection correlated with an augmented number of NK cells (CD45
NK1.1
CD3
) and Iba-1
microglial cells and a higher production of IFN-γ in the brain. In the spleen, levels of CCL5/RANTES and IFN-γ were increased and sustained in surviving poly(I:C)-treated animals for 14 days after infection. In immunocompetent animals, survival time was not significantly prolonged in poly(I:C)-treated animals although poly(I:C) priming reduced brain bacterial concentrations compared with vehicle-injected animals at early infection.
Pre-treatment with the viral TLR3 agonist poly(I:C) modulated innate immune responses and strengthened the resistance of neutropenic mice against E. coli K1 meningoencephalitis.
Multiple sclerosis (MS) is a chronic inflammatory disease of the central nervous system. Although the cause of MS is still uncertain, many findings point toward an ongoing autoimmune response to ...myelin antigens. Because of its location on the outer surface of the myelin sheath and its pathogenicity in the experimental autoimmune encephalomyelitis model, myelin oligodendrocyte glycoprotein (MOG) is one of the potential disease-causing self antigens in MS. However, the role of MOG in the pathogenesis of MS has remained controversial. In this study we addressed the occurrence of autoantibodies to native MOG and its implication for demyelination and axonal loss in MS. We applied a high-sensitivity bioassay, which allowed detecting autoantibodies that bind to the extracellular part of native MOG. Antibodies, mostly IgG, were found in sera that bound with high affinity to strictly conformational epitopes of the extracellular domain of MOG. IgG but not IgM antibody titers to native MOG were significantly higher in MS patients compared with different control groups with the highest prevalence in primary progressive MS patients. Serum autoantibodies to native MOG induced death of MOGexpressing target cells in vitro. Serum from MS patients with high anti-MOG antibody titers stained white matter myelin in rat brain and enhanced demyelination and axonal damage when transferred to autoimmune encephalomyelitis animals. Overall these findings suggest a pathogenic antibody response to native MOG in a subgroup of MS patients.
Quinoline-3-carboxamides, such as laquinimod, ameliorate CNS autoimmunity in patients and reduce tumor cell metastasis experimentally. Previous studies have focused on the immunomodulatory effect of ...laquinimod on myeloid cells. The data contained herein suggest that quinoline-3-carboxamides improve the immunomodulatory and anti-tumor effects of NK cells by upregulating the adhesion molecule DNAX accessory molecule-1 (DNAM-1).
We explored how NK cell activation by laquinimod inhibits CNS autoimmunity in experimental autoimmune encephalomyelitis (EAE), the most utilized model of MS, and improves immunosurveillance of experimental lung melanoma metastasis. Functional manipulations included in vivo NK and DC depletion experiments and in vitro assays of NK cell function. Clinical, histological, and flow cytometric read-outs were assessed.
We demonstrate that laquinimod activates natural killer (NK) cells via the aryl hydrocarbon receptor and increases their DNAM-1 cell surface expression. This activation improves the cytotoxicity of NK cells against B16F10 melanoma cells and augments their immunoregulatory functions in EAE by interacting with CD155
dendritic cells (DC). Noteworthy, the immunosuppressive effect of laquinimod-activated NK cells was due to decreasing MHC class II antigen presentation by DC and not by increasing DC killing.
This study clarifies how DNAM-1 modifies the bidirectional crosstalk of NK cells with CD155
DC, which can be exploited to suppress CNS autoimmunity and strengthen tumor surveillance.
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