Transcriptomic and morphological maturation of human astrocytes in cerebral organoids.

Cerebral organoids (CerOrgs) derived from human induced pluripotent stem cells (iPSCs) are a valuable tool to study human astrocytes and their interaction with neurons and microglia. The timeline of astrocyte development and maturation in this model is currently unknown and this limits the value and applicability of the model. Therefore, we generated CerOrgs from three healthy individuals and assessed astrocyte maturation after 5, 11, 19, and 37 weeks in culture. At these four time points, the astrocyte lineage was isolated based on the expression of integrin subunit alpha 6 (ITGA6). Based on the transcriptome of the isolated ITGA6-positive cells, astrocyte development started between 5 and 11 weeks in culture and astrocyte maturation commenced after 11 weeks in culture. After 19 weeks in culture, the ITGA6-positive astrocytes had the highest expression of human mature astrocyte genes, and the predicted functional properties were related to brain homeostasis. After 37 weeks in culture, a subpopulation of ITGA6-negative astrocytes appeared, highlighting the heterogeneity within the astrocytes. The morphology shifted from an elongated progenitor-like morphology to the typical bushy astrocyte morphology. Based on the morphological properties, predicted functional properties, and the similarities with the human mature astrocyte transcriptome, we concluded that ITGA6-positive astrocytes have developed optimally in 19-week-old CerOrgs.

Early amyloid-induced changes in microglia gene expression in male APP/PS1 mice.

Alzheimer's disease (AD) is a progressive neurodegenerative disease and the most common cause of dementia, characterized by deposition of extracellular amyloid-beta (Aß) aggregates and intraneuronal hyperphosphorylated Tau. Many AD risk genes, identified in genome-wide association studies (GWAS), are expressed in microglia, the innate immune cells of the central nervous system. Specific subtypes of microglia emerged in relation to AD pathology, such as disease-associated microglia (DAMs), which increased in number with age in amyloid mouse models and in human AD cases. However, the initial transcriptional changes in these microglia in response to amyloid are still unknown. Here, to determine early changes in microglia gene expression, hippocampal microglia from male APPswe/PS1dE9 (APP/PS1) mice and wild-type littermates were isolated and analyzed by RNA sequencing (RNA-seq). By bulk RNA-seq, transcriptomic changes were detected in hippocampal microglia from 6-months-old APP/PS1 mice. By performing single-cell RNA-seq of CD11c-positive and negative microglia from 6-months-old APP/PS1 mice and analysis of the transcriptional trajectory from homeostatic to CD11c-positive microglia, we identified a set of genes that potentially reflect the initial response of microglia to Aß.

Prevention of microgliosis halts early memory loss in a mouse model of Alzheimer's disease.

Alzheimer's disease (AD) is a neurodegenerative disorder characterized by cognitive decline, the neuropathological formation of amyloid-beta (Aß) plaques and neurofibrillary tangles. The best cellular correlates of the early cognitive deficits in AD patients are synapse loss and gliosis. In particular, it is unclear whether the activation of microglia (microgliosis) has a neuroprotective or pathological role early in AD. Here we report that microgliosis is an early mediator of synaptic dysfunction and cognitive impairment in APP/PS1 mice, a mouse model of increased amyloidosis. We found that the appearance of microgliosis, synaptic dysfunction and behavioral impairment coincided with increased soluble Aß42 levels, and occurred well before the presence of Aß plaques. Inhibition of microglial activity by treatment with minocycline (MC) reduced gliosis, synaptic deficits and cognitive impairments at early pathological stages and was most effective when provided preventive, i.e., before the onset of microgliosis. Interestingly, soluble Aß levels or Aß plaques deposition were not affected by preventive MC treatment at an early pathological stage (4 months) whereas these were reduced upon treatment at a later stage (6 months). In conclusion, this study demonstrates the importance of early-stage prevention of microgliosis on the development of cognitive impairment in APP/PS1 mice, which might be clinically relevant in preventing memory loss and delaying AD pathogenesis.

Aß Pathology and Neuron-Glia Interactions: A Synaptocentric View.

Alzheimer's disease (AD) causes the majority of dementia cases worldwide. Early pathological hallmarks include the accumulation of amyloid-ß (Aß) and activation of both astrocytes and microglia. Neurons form the building blocks of the central nervous system, and astrocytes and microglia provide essential input for its healthy functioning. Their function integrates at the level of the synapse, which is therefore sometimes referred to as the "quad-partite synapse". Increasing evidence puts AD forward as a disease of the synapse, where pre- and postsynaptic processes, as well as astrocyte and microglia functioning progressively deteriorate. Here, we aim to review the current knowledge on how Aß accumulation functionally affects the individual components of the quad-partite synapse. We highlight a selection of processes that are essential to the healthy functioning of the neuronal synapse, including presynaptic neurotransmitter release and postsynaptic receptor functioning. We further discuss how Aß affects the astrocyte's capacity to recycle neurotransmitters, release gliotransmitters, and maintain ion homeostasis. We additionally review literature on how Aß changes the immunoprotective function of microglia during AD progression and conclude by summarizing our main findings and highlighting the challenges in current studies, as well as the need for further research.

Amyloid-ß plaques affect astrocyte Kir4.1 protein expression but not function in the dentate gyrus of APP/PS1 mice.

Alzheimer pathology is accompanied by astrogliosis. Reactive astrocytes surrounding amyloid plaques may directly affect neuronal communication, and one of the mechanisms by which astrocytes impact neuronal function is by affecting K+ homeostasis. Here we studied, using hippocampal slices from 9-month-old Alzheimer mice (APP/PS1) and wild-type littermates, whether astrocyte function is changed by analyzing Kir4.1 expression and function and astrocyte coupling in astrocytes surrounding amyloid-ß plaques. Immunohistochemical analysis of Kir4.1 protein in the dentate gyrus revealed localized increases in astrocytes surrounding amyloid-ß plaque deposits. We subsequently focused on changes in astrocyte function by using patch-clamp slice electrophysiology on both plaque- and non-plaque associated astrocytes to characterize general membrane properties. We found that Ba2+ -sensitive Kir4.1 conductance in astrocytes surrounding plaques was not affected by changes in Kir4.1 protein expression. Additional analysis of astrocyte gap junction coupling efficiency in the dentate gyrus revealed no apparent changes. Quantification of basic features of glutamatergic transmission to granule cells did not indicate disturbed neuronal communication in the dentate gyrus of APP/PS1 mice. Together, these results suggest that astrocytes in the dentate gyrus of APP/PS1 mice maintain their ability to buffer extracellular K+ and attempt to rectify imbalances in K+ concentration to maintain normal neuronal and synaptic function, possibly by localized increases in Kir4.1 protein expression. Our earlier transcriptomic data indicated that chronically activated astrocytes lose their neuronal support function. Here we show that, despite localized increased Kir4.1 protein expression, astrocyte Kir4.1 channel dysfunction is likely not involved in the pathogenesis of Alzheimer's disease.

Current perspectives on diffuse midline glioma and a different role for the immune microenvironment compared to glioblastoma.

Diffuse midline glioma (DMG), formerly called diffuse intrinsic pontine glioma (DIPG), is a high-grade malignant pediatric brain tumor with a near-zero survival rate. To date, only radiation therapy provides marginal survival benefit; however, the median survival time remains less than a year. Historically, the infiltrative nature and sensitive location of the tumor rendered surgical removal and biopsies difficult and subsequently resulted in limited knowledge of the disease, as only post-mortem tissue was available. Therefore, clinical decision-making was based upon experience with the more frequent and histologically similar adult glioblastoma (GBM). Recent advances in tissue acquisition and molecular profiling revealed that DMG and GBM are distinct disease entities, with separate tissue characteristics and genetic profiles. DMG is characterized by heterogeneous tumor tissue often paired with an intact blood-brain barrier, possibly explaining its resistance to chemotherapy. Additional profiling shed a light on the origin of the disease and the influence of several mutations such as a highly recurring K27M mutation in histone H3 on its tumorigenesis. Furthermore, early evidence suggests that DMG has a unique immune microenvironment, characterized by low levels of immune cell infiltration, inflammation, and immunosuppression that may impact disease development and outcome. Within the tumor microenvironment of GBM, tumor-associated microglia/macrophages (TAMs) play a large role in tumor development. Interestingly, TAMs in DMG display distinct features and have low immune activation in comparison to other pediatric gliomas. Although TAMs have been investigated substantially in GBM over the last years, this has not been the case for DMG due to the lack of tissue for research. Bit by bit, studies are exploring the TAM-glioma crosstalk to identify what factors within the DMG microenvironment play a role in the recruitment and polarization of TAMs. Although more research into the immune microenvironment is warranted, there is evidence that targeting or stimulating TAMs and their factors provide a potential treatment option for DMG. In this review, we provide insight into the current status of DMG research, assess the knowledge of the immune microenvironment in DMG and GBM, and present recent findings and therapeutic opportunities surrounding the TAM-glioma crosstalk.

Calcium signaling in individual APP/PS1 mouse dentate gyrus astrocytes increases ex vivo with Aß pathology and age without affecting astrocyte network activity.

Astrocytes are critical for healthy brain function. In Alzheimer's disease, astrocytes become reactive, which affects their signaling properties. Here, we measured spontaneous calcium transients ex vivo in hippocampal astrocytes in brain slices containing the dentate gyrus of 6- (6M) and 9-month-old (9M) APPswe/PSEN1dE9 (APP/PS1) mice. We investigated the frequency and duration of calcium transients in relation to aging, amyloid-ß (Aß) pathology, and the proximity of the astrocyte to Aß plaques. The 6M APP/PS1 astrocytes showed no change in spontaneous calcium-transient properties compared to wild-type (WT) astrocytes. 9M APP/PS1 astrocytes, however, showed more hyperactivity compared to WT, characterized by increased spontaneous calcium transients that were longer in duration. Our data also revealed an effect of aging, as 9M astrocytes overall showed an increase in calcium activity compared to 6M astrocytes. Subsequent calcium-wave analysis showed an increase in sequential calcium transients (i.e., calcium waves) in 9M astrocytes, suggesting increased network activity ex vivo. Further analysis using null models revealed that this network effect is caused by chance, due to the increased number of spontaneous transients. Our findings show that alterations in calcium signaling in individual hippocampal astrocytes of APP/PS1 mice are subject to both aging and Aß pathology but these do not lead to a change in astrocyte network activity. These alterations in calcium dynamics of astrocytes may help to understand changes in neuronal physiology leading to cognitive decline and ultimately dementia.

New GFAP splice isoform (GFAPµ) differentially expressed in glioma translates into 21 kDa N-terminal GFAP protein.

The glial fibrillary acidic protein (GFAP) is a type III intermediate filament (IF) protein that is highly expressed in astrocytes, neural stem cells, and in gliomas. Gliomas are a heterogeneous group of primary brain tumors that arise from glia cells or neural stem cells and rely on accurate diagnosis for prognosis and treatment strategies. GFAP is differentially expressed between glioma subtypes and, therefore, often used as a diagnostic marker. However, GFAP is highly regulated by the process of alternative splicing; many different isoforms have been identified. Differential expression of GFAP isoforms between glioma subtypes suggests that GFAP isoform-specific analyses could benefit diagnostics. In this study we report on the differential expression of a new GFAP isoform between glioma subtypes, GFAPµ. A short GFAP transcript resulting from GFAP exon 2 skipping was detected by RNA sequencing of human glioma. We show that GFAPµ mRNA is expressed in healthy brain tissue, glioma cell lines, and primary glioma cells and that it translates into a ~21 kDa GFAP protein. 21 kDa GFAP protein was detected in the IF protein fraction isolated from human spinal cord as well. We further show that induced GFAPµ expression disrupts the GFAP IF network. The characterization of this new GFAP isoform adds on to the numerous previously identified GFAP splice isoforms. It emphasizes the importance of studying the contribution of IF splice variants to specialized functions of the IF network and to glioma research.

Transcriptomic and functional analysis of Aß1-42 oligomer-stimulated human monocyte-derived microglia-like cells.

Dysregulation of microglial function contributes to Alzheimer's disease (AD) pathogenesis. Several genetic and transcriptome studies have revealed microglia specific genetic risk factors, and changes in microglia expression profiles in AD pathogenesis, viz. the human-Alzheimer's microglia/myeloid (HAM) profile in AD patients and the disease-associated microglia profile (DAM) in AD mouse models. The transcriptional changes involve genes in immune and inflammatory pathways, and in pathways associated with Aß clearance. Aß oligomers have been suggested to be the initial trigger of microglia activation in AD. To study the direct response to Aß oligomers exposure, we assessed changes in gene expression in an in vitro model for microglia, the human monocyte-derived microglial-like (MDMi) cells. We confirmed the initiation of an inflammatory profile following LPS stimulation, based on increased expression of IL1B, IL6, and TNFα. In contrast, the Aß1-42 oligomers did not induce an inflammatory profile or a classical HAM profile. Interestingly, we observed a specific increase in the expression of metallothioneins in the Aß1-42 oligomer treated MDMi cells. Metallothioneins are involved in metal ion regulation, protection against reactive oxygen species, and have anti-inflammatory properties. In conclusion, our data suggests that exposure to Aß1-42 oligomers may initially trigger a protective response in vitro.

Distinct non-inflammatory signature of microglia in post-mortem brain tissue of patients with major depressive disorder.

Findings from epidemiological studies, biomarker measurements and animal experiments suggest a role for aberrant immune processes in the pathogenesis of major depressive disorder (MDD). Microglia, the resident immune cells of the brain, are likely to play a key role in these processes. Previous post-mortem studies reported conflicting findings regarding microglial activation and an in-depth profiling of those cells in MDD is lacking. The aim of this study was therefore to characterize the phenotype and function of microglia in MDD. We isolated microglia from post-mortem brain tissue of patients with MDD (n = 13-19) and control donors (n = 12-25). Using flow cytometry and quantitative Polymerase Chain Reaction (qPCR), we measured protein and mRNA levels of a panel of microglial markers across four different brain regions (medial frontal gyrus, superior temporal gyrus, thalamus, and subventricular zone). In MDD cases, we found a significant upregulation of CX3CR1 and TMEM119 mRNA expression and a downregulation of CD163 mRNA expression and CD14 protein expression across the four brain regions. Expression levels of microglial activation markers, such as HLA-DRA, IL6, and IL1ß, as well as the inflammatory responses to lipopolysaccharide and dexamethasone were unchanged. Our findings suggest that microglia enhance homeostatic functions in MDD but are not immune activated.

Investigation of glial fibrillary acidic protein (GFAP) in body fluids as a potential biomarker for glioma: a systematic review and meta-analysis.

INTRODUCTION: Liquid biopsies are promising diagnostic tools for glioma. In this quantitative systematic review, we investigate whether the detection of intermediate filaments (IF) in body fluids can be used as a tool for glioma diagnosis and prognosis. MATERIALS AND METHODS: We included all studies in which IF-levels were determined in patients with glioma and healthy controls. Of the 28 identified eligible studies, 12 focussed on levels of GFAP in serum (sGFAP) and were included for metadata analysis. RESULTS: In all studies combined, 62.7% of all grade-IV patients had detectable levels of sGFAP compared to 12.7% of healthy controls. sGFAP did not surpass the limit of detection in lower-grade patients or healthy controls, but sGFAP was significantly elevated in grade-IV glioma (0.12 ng/mL (0.06 - 0.18), P < 0.001) and showed an average median difference of 0.15 ng/mL (0.04 - 0.25, P < 0.01) compared to healthy controls. sGFAP levels were linked to tumour volume, but not to patient outcome. CONCLUSION: The presence of sGFAP is indicative of grade-IV glioma, but additional studies are necessary to fully determine the usefulness of GFAP in body fluids as a tool for grade-IV glioma diagnosis and follow-up.

Reactive astrocytes as treatment targets in Alzheimer's disease-Systematic review of studies using the APPswePS1dE9 mouse model.

Astrocytes regulate synaptic communication and are essential for proper brain functioning. In Alzheimer's disease (AD) astrocytes become reactive, which is characterized by an increased expression of intermediate filament proteins and cellular hypertrophy. Reactive astrocytes are found in close association with amyloid-beta (Aß) deposits. Synaptic communication and neuronal network function could be directly modulated by reactive astrocytes, potentially contributing to cognitive decline in AD. In this review, we focus on reactive astrocytes as treatment targets in AD in the APPswePS1dE9 AD mouse model, a widely used model to study amyloidosis and gliosis. We first give an overview of the model; that is, how it was generated, which cells express the transgenes, and the effect of its genetic background on Aß pathology. Subsequently, to determine whether modifying reactive astrocytes in AD could influence pathogenesis and cognition, we review studies using this mouse model in which interventions were directly targeted at reactive astrocytes or had an indirect effect on reactive astrocytes. Overall, studies specifically targeting astrocytes to reduce astrogliosis showed beneficial effects on cognition, which indicates that targeting astrocytes should be included in developing novel therapies for AD.

A loss of mature microglial markers without immune activation in schizophrenia.

Microglia, the immune cells of the brain, are important for neurodevelopment and have been hypothesized to play a role in the pathogenesis of schizophrenia (SCZ). Although previous postmortem studies pointed toward presence of microglial activation, this view has been challenged by more recent hypothesis-driven and hypothesis-free analyses. The aim of the present study is to further understand the observed microglial changes in SCZ. We first performed a detailed meta-analysis on studies that analyzed microglial cell density, microglial morphology, and expression of microglial-specific markers. We then further explored findings from the temporal cortex by performing immunostainings and qPCRs on an additional dataset. A random effect meta-analysis showed that the density of microglial cells was unaltered in SCZ (ES: 0.144 95% CI: 0.102 to 0.390, p = .250), and clear changes in microglial morphology were also absent. The expression of several microglial specific genes, such as CX3CR1, CSF1R, IRF8, OLR1, and TMEM119 was decreased in SCZ (ES: -0.417 95% CI: -0.417 to -0.546, p < .0001), consistent with genome-wide transcriptome meta-analysis results. These results indicate a change in microglial phenotype rather than density, which was validated with the use of TMEM119/Iba1 immunostainings on temporal cortex of a separate cohort. Changes in microglial gene expression were overlapping between SCZ and other psychiatric disorders, but largely opposite from changes reported in Alzheimer's disease. This distinct microglial phenotype provides a crucial molecular hallmark for future research into the role of microglia in SCZ and other psychiatric disorders.

Physiological and Pathological Ageing of Astrocytes in the Human Brain.

Ageing is the greatest risk factor for dementia, although physiological ageing by itself does not lead to cognitive decline. In addition to ageing, APOE ε4 is genetically the strongest risk factor for Alzheimer's disease and is highly expressed in astrocytes. There are indications that human astrocytes change with age and upon expression of APOE4. As these glial cells maintain water and ion homeostasis in the brain and regulate neuronal transmission, it is likely that age- and APOE4-related changes in astrocytes have a major impact on brain functioning and play a role in age-related diseases. In this review, we will discuss the molecular and morphological changes of human astrocytes in ageing and the contribution of APOE4. We conclude this review with a discussion on technical issues, innovations, and future perspectives on how to gain more knowledge on astrocytes in the human ageing brain.

GFAP alternative splicing regulates glioma cell-ECM interaction in a DUSP4-dependent manner.

Gliomas are the most common primary brain tumors. Their highly invasive character and the heterogeneity of active oncogenic pathways within single tumors complicate the development of curative therapies and cause poor patient prognosis. Glioma cells express the intermediate filament protein glial fibrillary acidic protein (GFAP), and the level of its alternative splice variant GFAP-δ, relative to its canonical splice variant GFAP-α, is higher in grade IV compared with lower-grade and lower malignant glioma. In this study we show that a high GFAP-δ/α ratio induces the expression of the dual-specificity phosphatase 4 (DUSP4) in focal adhesions. By focusing on pathways up- and downstream of DUSP4 that are involved in the cell-extracellular matrix interaction, we show that a high GFAP-δ/α ratio equips glioma cells to better invade the brain. This study supports the hypothesis that glioma cells with a high GFAP-δ/α ratio are highly invasive and more malignant cells, thus making GFAP alternative splicing a potential therapeutic target.-Van Bodegraven, E. J., van Asperen, J. V., Sluijs, J. A., van Deursen, C. B. J., van Strien, M. E., Stassen, O. M. J. A., Robe, P. A. J., Hol, E. M. GFAP alternative splicing regulates glioma cell-ECM interaction in a DUSP4-dependent manner.

A characterization of the molecular phenotype and inflammatory response of schizophrenia patient-derived microglia-like cells.

Different lines of evidence support a causal role for microglia in the pathogenesis of schizophrenia. However, how schizophrenia patient-derived microglia are affected at the phenotypic and functional level is still largely unknown. We used a recently described model to induce patient-derived microglia-like cells and used this to analyze changes in the molecular phenotype and function of myeloid cells in schizophrenia. We isolated monocytes from twenty recent-onset schizophrenia patients and twenty non-psychiatric controls. We cultured the cells towards an induced microglia-like phenotype (iMG), analyzed the phenotype of the cells by RNA sequencing and mass cytometry, and their response to LPS. Mass cytometry showed a high heterogeneity of iMG in cells derived from patients as well as controls. The prevalence of two iMG clusters was significantly higher in schizophrenia patients (adjusted p-value < 0.001). These subsets are characterized by expression of ApoE, Ccr2, CD18, CD44, and CD95, as well as IRF8, P2Y12, Cx3cr1 and HLA-DR. In addition, we found that patient-derived iMG show an enhanced response to LPS, with increased secretion of TNF-α. Further studies are needed to replicate these findings, to determine whether similar subclusters are present in schizophrenia patients in vivo, and to address how these subclusters are related to the increased response to LPS, as well as other microglial functions.

Cannabinoids and psychotic symptoms: A potential role for a genetic variant in the P2X purinoceptor 7 (P2RX7) gene.

To investigate the biological mechanisms underlying the higher risk for psychosis in those that use cannabis, we conducted a genome-wide environment-interaction study (GWEIS). In a sample of individuals without a psychiatric disorder (N = 1262), we analyzed the interactions between regular cannabis use and genotype with psychotic-like experiences (PLE) as outcome. PLE were measured using the Community Assessment of Psychic Experiences (CAPE). The sample was enriched for those at the extremes of both cannabis use and PLE to increase power. A single nucleotide polymorphism in the P2RX7 gene (rs7958311) was associated with risk for a high level of psychotic experiences in regular cannabis users (p = 1.10 x10-7) and in those with high levels of lifetime cannabis use (p = 4.5 × 10-6). This interaction was replicated in individuals with high levels of lifetime cannabis use in the IMAGEN cohort (N = 1217, p = 0.020). Functional relevance of P2RX7 in cannabis users was suggested by in vitro experiments on activated monocytes. Exposure of these cells to tetrahydrocannabinol (THC) or cannabidiol (CBD) reduced the immunological response of the P2X7 receptor, which was dependent on the identified genetic variant. P2RX7 variants have been implicated in psychiatric disorders before and the P2X7 receptor is involved in pathways relevant to psychosis, such as neurotransmission, synaptic plasticity and immune regulation. We conclude that P2RX7 plays a role in vulnerability to develop psychotic symptoms when using cannabis and point to a new pathway that can potentially be targeted by newly developed P2X7 antagonists.

Importance of GFAP isoform-specific analyses in astrocytoma.

Gliomas are a heterogenous group of malignant primary brain tumors that arise from glia cells or their progenitors and rely on accurate diagnosis for prognosis and treatment strategies. Although recent developments in the molecular biology of glioma have improved diagnosis, classical histological methods and biomarkers are still being used. The glial fibrillary acidic protein (GFAP) is a classical marker of astrocytoma, both in clinical and experimental settings. GFAP is used to determine glial differentiation, which is associated with a less malignant tumor. However, since GFAP is not only expressed by mature astrocytes but also by radial glia during development and neural stem cells in the adult brain, we hypothesized that GFAP expression in astrocytoma might not be a direct indication of glial differentiation and a less malignant phenotype. Therefore, we here review all existing literature from 1972 up to 2018 on GFAP expression in astrocytoma patient material to revisit GFAP as a marker of lower grade, more differentiated astrocytoma. We conclude that GFAP is heterogeneously expressed in astrocytoma, which most likely masks a consistent correlation of GFAP expression to astrocytoma malignancy grade. The GFAP positive cell population contains cells with differences in morphology, function, and differentiation state showing that GFAP is not merely a marker of less malignant and more differentiated astrocytoma. We suggest that discriminating between the GFAP isoforms GFAPδ and GFAPα will improve the accuracy of assessing the differentiation state of astrocytoma in clinical and experimental settings and will benefit glioma classification.

Clinical and immunological characteristics of the spectrum of GFAP autoimmunity: a case series of 22 patients.

OBJECTIVE: To report the clinical and immunological characteristics of 22 new patients with glial fibrillar acidic protein (GFAP) autoantibodies. METHODS: From January 2012 to March 2017, we recruited 451 patients with suspected neurological autoimmune disease at the Catholic University of Rome. Patients' serum and cerebrospinal fluid (CSF) samples were tested for neural autoantibodies by immunohistochemistry on mouse and rat brain sections, by cell-based assays (CBA) and immunoblot. GFAP autoantibodies were detected by immunohistochemistry and their specificity confirmed by CBA using cells expressing human GFAPα and GFAPδ proteins, by immunoblot and immunohistochemistry on GFAP-/- mouse brain sections. RESULTS: Serum and/or CSF IgG of 22/451 (5%) patients bound to human GFAP, of which 22/22 bound to GFAPα, 14/22 to both GFAPα and GFAPδ and none to the GFAPδ isoform only. The neurological presentation was: meningoencephalomyelitis or encephalitis in 10, movement disorder (choreoathetosis or myoclonus) in 3, anti-epileptic drugs (AED)-resistant epilepsy in 3, cerebellar ataxia in 3, myelitis in 2, optic neuritis in 1 patient. Coexisting neural autoantibodies were detected in five patients. Six patients had other autoimmune diseases. Tumours were found in 3/22 patients (breast carcinoma, 1; ovarian carcinoma, 1; thymoma, 1). Nineteen patients were treated with immunotherapy and 16 patients (84%) improved. Histopathology analysis of the leptomeningeal biopsy specimen from one patient revealed a mononuclear infiltrate with macrophages and CD8+ T cells. CONCLUSIONS: GFAP autoimmunity is not rare. The clinical spectrum encompasses meningoencephalitis, myelitis, movement disorders, epilepsy and cerebellar ataxia. Coexisting neurological and systemic autoimmunity are relatively common. Immunotherapy is beneficial in most cases.

Transcriptional profiling of CD11c-positive microglia accumulating around amyloid plaques in a mouse model for Alzheimer's disease.

Amyloid plaques in Alzheimer's disease (AD) mice are surrounded by activated microglia. The functional role of microglia activation in AD is not well understood; both detrimental and beneficial effects on AD progression have been reported. Here we show that the population of activated microglia in the cortex of the APPswe/PS1dE9 mouse AD model is divided into a CD11c-positive and a CD11c-negative subpopulation. Cd11c transcript levels and number of CD11c-positive microglia increase sharply when plaques start to occur and both parameters continue to rise in parallel with the age-related increasing plaque load. CD11c cells are localized near plaques at all stages of the disease development and constitute 23% of all activated microglia. No differences between these two populations were found in terms of proliferation, immunostaining intensity of Iba1, MHC class II, CD45, or immunoproteasome subunit LMP7/ß5i. Comparison of the transcriptome of isolated CD11c-positive and CD11c-negative microglia from the cortex of aged APPswe/PS1dE9 with WT microglia showed that gene expression changes had a similar general pattern. However, a differential expression was found for genes involved in immune signaling (Il6, S100a8/Mrp8, S100a9/Mrp14, Spp1, Igf1), lysosome activation, and carbohydrate- and cholesterol/lipid-metabolism (Apoe). In addition, the increased expression of Gpnmb/DC-HIL, Tm7sf4/DC-STAMP, and Gp49a/Lilrb4, suggests a suppressive/tolerizing influence of CD11c cells. We show that amyloid plaques in the APP/PS1 model are associated with two distinct populations of activated microglia: CD11c-positive and CD11c-negative cells. Our findings imply that CD11c-positive microglia can potentially counteract amyloid deposition via increased Aß-uptake and degradation, and by containing the inflammatory response.