Publications by authors named "Roman Sankowski"

30 Publications

  • Page 1 of 1

Mapping the origin and fate of myeloid cells in distinct compartments of the eye by single-cell profiling.

EMBO J 2021 Mar 8;40(6):e105123. Epub 2021 Feb 8.

Institute of Neuropathology, Medical Faculty, University of Freiburg, Freiburg, Germany.

Similar to the brain, the eye is considered an immune-privileged organ where tissue-resident macrophages provide the major immune cell constituents. However, little is known about spatially restricted macrophage subsets within different eye compartments with regard to their origin, function, and fate during health and disease. Here, we combined single-cell analysis, fate mapping, parabiosis, and computational modeling to comprehensively examine myeloid subsets in distinct parts of the eye during homeostasis. This approach allowed us to identify myeloid subsets displaying diverse transcriptional states. During choroidal neovascularization, a typical hallmark of neovascular age-related macular degeneration (AMD), we recognized disease-specific macrophage subpopulations with distinct molecular signatures. Our results highlight the heterogeneity of myeloid subsets and their dynamics in the eye that provide new insights into the innate immune system in this organ which may offer new therapeutic targets for ophthalmological diseases.
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http://dx.doi.org/10.15252/embj.2020105123DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7957431PMC
March 2021

IL-17 controls central nervous system autoimmunity through the intestinal microbiome.

Sci Immunol 2021 Feb;6(56)

Institute for Molecular Medicine, University Medical Center of the Johannes Gutenberg-University Mainz, Mainz, Germany.

Interleukin-17A- (IL-17A) and IL-17F-producing CD4 T helper cells (T17 cells) are implicated in the development of chronic inflammatory diseases, such as multiple sclerosis and its animal model, experimental autoimmune encephalomyelitis (EAE). T17 cells also orchestrate leukocyte invasion of the central nervous system (CNS) and subsequent tissue damage. However, the role of IL-17A and IL-17F as effector cytokines is still confused with the encephalitogenic function of the cells that produce these cytokines, namely, T17 cells, fueling a long-standing debate in the neuroimmunology field. Here, we demonstrated that mice deficient for IL-17A/F lose their susceptibility to EAE, which correlated with an altered composition of their gut microbiota. However, loss of IL-17A/F in T cells did not diminish their encephalitogenic capacity. Reconstitution of a wild-type-like intestinal microbiota or reintroduction of IL-17A specifically into the gut epithelium of IL-17A/F-deficient mice reestablished their susceptibility to EAE. Thus, our data demonstrated that IL-17A and IL-17F are not encephalitogenic mediators but rather modulators of intestinal homeostasis that indirectly alter CNS-directed autoimmunity.
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http://dx.doi.org/10.1126/sciimmunol.aaz6563DOI Listing
February 2021

Microglia facilitate repair of demyelinated lesions via post-squalene sterol synthesis.

Nat Neurosci 2021 01 21;24(1):47-60. Epub 2020 Dec 21.

Department of Neurogenetics, Max Planck Institute of Experimental Medicine, Göttingen, Germany.

The repair of inflamed, demyelinated lesions as in multiple sclerosis (MS) necessitates the clearance of cholesterol-rich myelin debris by microglia/macrophages and the switch from a pro-inflammatory to an anti-inflammatory lesion environment. Subsequently, oligodendrocytes increase cholesterol levels as a prerequisite for synthesizing new myelin membranes. We hypothesized that lesion resolution is regulated by the fate of cholesterol from damaged myelin and oligodendroglial sterol synthesis. By integrating gene expression profiling, genetics and comprehensive phenotyping, we found that, paradoxically, sterol synthesis in myelin-phagocytosing microglia/macrophages determines the repair of acutely demyelinated lesions. Rather than producing cholesterol, microglia/macrophages synthesized desmosterol, the immediate cholesterol precursor. Desmosterol activated liver X receptor (LXR) signaling to resolve inflammation, creating a permissive environment for oligodendrocyte differentiation. Moreover, LXR target gene products facilitated the efflux of lipid and cholesterol from lipid-laden microglia/macrophages to support remyelination by oligodendrocytes. Consequently, pharmacological stimulation of sterol synthesis boosted the repair of demyelinated lesions, suggesting novel therapeutic strategies for myelin repair in MS.
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http://dx.doi.org/10.1038/s41593-020-00757-6DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7116742PMC
January 2021

Single cell RNA sequencing of human microglia uncovers a subset associated with Alzheimer's disease.

Nat Commun 2020 11 30;11(1):6129. Epub 2020 Nov 30.

Center for Translational and Computational Neuroimmunology, Columbia University Medical Center, New York, NY, USA.

The extent of microglial heterogeneity in humans remains a central yet poorly explored question in light of the development of therapies targeting this cell type. Here, we investigate the population structure of live microglia purified from human cerebral cortex samples obtained at autopsy and during neurosurgical procedures. Using single cell RNA sequencing, we find that some subsets are enriched for disease-related genes and RNA signatures. We confirm the presence of four of these microglial subpopulations histologically and illustrate the utility of our data by characterizing further microglial cluster 7, enriched for genes depleted in the cortex of individuals with Alzheimer's disease (AD). Histologically, these cluster 7 microglia are reduced in frequency in AD tissue, and we validate this observation in an independent set of single nucleus data. Thus, our live human microglia identify a range of subtypes, and we prioritize one of these as being altered in AD.
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http://dx.doi.org/10.1038/s41467-020-19737-2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7704703PMC
November 2020

Chitinase 3-like 1 and neurofilament light chain in CSF and CNS atrophy in MS.

Neurol Neuroimmunol Neuroinflamm 2021 01 10;8(1). Epub 2020 Nov 10.

From the Department of Neurology (R. Schneider, B.G., S.H., R.G., A.H.), Institute of Neuroradiology (R. Schneider, B.B., C.L.), and Department of Radiology and Nuclear Medicine (C.L.), St. Josef Hospital, Ruhr University Bochum; Institute of Neuropathology (R. Sankowski, M.P.), Medical Faculty, Signalling Research Centers BIOSS and CIBSS (M.P.), and Center for Basics in NeuroModulation (NeuroModulBasics) (M.P.), Faculty of Medicine, University of Freiburg, Germany.

Objective: To investigate cross-sectional associations of CSF levels of neurofilament light chain (NfL) and of the newly emerging marker chitinase 3-like protein 1 (CHI3L1) with brain and spinal cord atrophy, which are established MRI markers of disease activity in MS, to study CHI3L1 and NfL in relapsing (RMS) and progressive MS (PMS), and to assess the expression of CHI3L1 in different cell types.

Methods: In a single-center study, 131 patients with MS (42 RMS and 89 PMS) were assessed for NfL and CHI3L1 concentrations in CSF, MRI-based spinal cord and brain volumetry, MS subtype, age, disease duration, and disability. We included 42 matched healthy controls receiving MRI. CHI3L1 expression of human brain cell types was examined in 2 published single-cell RNA sequencing data sets.

Results: CHI3L1 was associated with spinal cord volume (B = -1.07, 95% CI -2.04 to -0.11, = 0.029) but not with brain volumes. NfL was associated with brain gray matter (B = -7.3, 95% CI -12.0 to -2.7, = 0.003) but not with spinal cord volume. CHI3L1 was suitable to differentiate between progressive or relapsing MS ( = 0.015, OR 1.0103, CI for OR 1.002-1.0187), and its gene expression was found in MS-associated microglia and macrophages and in astrocytes of MS brains.

Conclusions: NfL and CHI3L1 in CSF were differentially related to brain and spinal cord atrophy. CSF CHI3L1 was associated with spinal cord volume loss and was less affected than NfL by disease duration and age, whereas CSF NfL was associated with brain gray matter atrophy. CSF NfL and CHI3L1 measurement provides complementary information regarding brain and spinal cord volumes.

Classification Of Evidence: This study provides Class II evidence that CSF CHI3L1 is associated with spinal cord volume loss and that CSF NfL is associated with gray matter atrophy.
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http://dx.doi.org/10.1212/NXI.0000000000000906DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7713721PMC
January 2021

Characterization of longitudinal transformation of T2-hyperintensity in oligodendroglioma.

BMC Cancer 2020 Aug 27;20(1):818. Epub 2020 Aug 27.

Translational NeuroOncology Research Group, Medical Center, University of Freiburg, Freiburg, Germany.

Background: Oligodendroglioma (ODG) are CNS resistant tumors characterized by their unique molecular signature, namely a combined deletion of 1p and 19q simultaneously to an IDH1/2 mutation. These tumors have a more favorable clinical outcome compared to other gliomas and a long-time survival that ranges between 10 and 20 years. However, during the course of the disease, multiple recurrences occur and the optimal treatment at each stage of the disease remains unclear. Here we report a retrospective longitudinal observation study of 836 MRI examinations in 44 ODG patients.

Methods: We quantified the volume of T2-hyperintensity to compute growth behavior in dependence of different treatment modalities, using various computational models.

Results: The identified growth pattern revealed dynamic changes, which were found to be patient-specific an did not correlate with clinical parameter or therapeutic interventions. Further, we showed that, surgical resection is beneficial for overall survival regardless the WHO grad or timepoint of surgery. To improve overall survival, an extent of resection above 50% is required. Multiple resections do not generally improve overall survival, except a greater extent of resection than in previous surgeries was achieved.

Conclusions: Our data aids to improve the interpretation of MRI images in clinical practice.
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http://dx.doi.org/10.1186/s12885-020-07290-6DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7450792PMC
August 2020

Author Correction: Novel Hexb-based tools for studying microglia in the CNS.

Nat Immunol 2020 10;21(10):1302

Institute of Neuropathology, Faculty of Medicine, University of Freiburg, Freiburg, Germany.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.
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http://dx.doi.org/10.1038/s41590-020-0774-6DOI Listing
October 2020

Novel Hexb-based tools for studying microglia in the CNS.

Nat Immunol 2020 07 15;21(7):802-815. Epub 2020 Jun 15.

Institute of Neuropathology, Faculty of Medicine, University of Freiburg, Freiburg, Germany.

Microglia and central nervous system (CNS)-associated macrophages (CAMs), such as perivascular and meningeal macrophages, are implicated in virtually all diseases of the CNS. However, little is known about their cell-type-specific roles in the absence of suitable tools that would allow for functional discrimination between the ontogenetically closely related microglia and CAMs. To develop a new microglia gene targeting model, we first applied massively parallel single-cell analyses to compare microglia and CAM signatures during homeostasis and disease and identified hexosaminidase subunit beta (Hexb) as a stably expressed microglia core gene, whereas other microglia core genes were substantially downregulated during pathologies. Next, we generated Hexb mice to stably monitor microglia behavior in vivo. Finally, the Hexb locus was employed for tamoxifen-inducible Cre-mediated gene manipulation in microglia and for fate mapping of microglia but not CAMs. In sum, we provide valuable new genetic tools to specifically study microglia functions in the CNS.
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http://dx.doi.org/10.1038/s41590-020-0707-4DOI Listing
July 2020

Key role of MIF-related neuroinflammation in neurodegeneration and cognitive impairment in Alzheimer's disease.

Mol Med 2020 04 17;26(1):34. Epub 2020 Apr 17.

Institute of Immunology, Philipps University Marburg, Marburg, Germany.

Background: Macrophage Migration Inhibitory Factor (MIF) is a potent proinflammatory cytokine that promotes the production of other immune mediators. MIF is produced by most cell types in the brain including microglia, astrocytes and neurons. Enhanced expression of MIF might contribute to the persistent activation of glial, chronic neuroinflammation and neurodegeneration. Here, we investigated the effect of MIF on inflammatory markers and spatial learning in a mouse model of sporadic AD and on tau pathology in AD patients.

Methods: We examined the effects of MIF deficiency and pharmacological MIF inhibition in vitro and in vivo. In vitro, quantitative PCR and ELISA were used to assess cytokine production of STZ-treated glial cells. In vivo, C57BL/6 mice were subjected to intracerebroventricular streptozotocin injection (3 mg/kg, ICV-STZ). Neuroinflammation and contextual learning performance were assessed using quantitative PCR and fear conditioning, respectively. Pharmacological MIF inhibition was achieved with intraperitoneal injections of ISO-1 (daily, IP, 20 mg/kg in 5% DMSO in 0.9% NaCl) for 4 weeks following ICV-STZ injection. The findings from ISO-1 treated mice were confirmed in MIF knockout C57BL/6. To assess the role of MIF in human AD, cerebrospinal fluid levels of MIF and hyperphosphorylated tau were measured using ELISA.

Results: Administration ICV-STZ resulted in hippocampal dependent cognitive impairment. MIF inhibition with ISO-1 significantly improved the STZ-induced impairment in contextual memory performance, indicating MIF-related inflammation as a major contributor to ICV-STZ-induced memory deficits. Furthermore, inhibition of the MIF resulted in reduced cytokine production in vitro and in vivo. In human subjects with AD at early clinical stages, cerebrospinal fluid levels of MIF were increased in comparison with age-matched controls, and correlated with biomarkers of tau hyper-phosphorylation and neuronal injury hinting at MIF levels as a potential biomarker for early-stage AD.

Conclusions: The present study indicates the key role of MIF in controlling the chronic cytokine release in neuroinflammation related to tau hyperphosphorylation, neurodegeneration, and clinical manifestations of AD, suggesting the potential of MIF inhibition as therapeutic strategy to slow down neurodegeneration and clinical disease progression.
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http://dx.doi.org/10.1186/s10020-020-00163-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7164357PMC
April 2020

Persistent inflammatory states and their implications in brain disease.

Curr Opin Neurol 2020 06;33(3):341-346

Institute of Neuropathology.

Purpose Of Review: Apart from mental, motor and sensory functions, the human central nervous system (CNS) regulates a plethora of homeostatic (autonomic and hormonal) bodily functions. These functions are dependent on specialized neuronal networks. To ensure connectivity of these networks, they are continuously refined and supported by glial cells that outnumber neurons by, according to some accounts, an order of magnitude. Among glial cells, microglia - the brain resident macrophages - plays a crucial role in maintaining neuronal networks. However, in their concomitant role as brain immune cells microglia also engage in inflammatory signaling that may disrupt neuronal networks. Here, we review novel insights for molecular pathways involved in the protective functions of microglia and other immune cells in response to systemic signals and stimuli.

Recent Findings: Recent evidence suggests that aging and systemic disease push individual microglia toward proinflammatory phenotypes compromising the connectivity of neuronal networks, resulting in neuropsychiatric disease. Furthermore, cells (self as well as the microbiome) outside the CNS have been shown to affect neuronal function.

Summary: These recent findings have critical implications for mental health, particularly of an aging population, in particular for the development of novel immunomodulatory therapies for brain disease.
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http://dx.doi.org/10.1097/WCO.0000000000000809DOI Listing
June 2020

Microglia Heterogeneity in the Single-Cell Era.

Cell Rep 2020 02;30(5):1271-1281

Institute of Neuropathology, Faculty of Medicine, University of Freiburg, Freiburg, Germany; Signalling Research Centres BIOSS and CIBSS, University of Freiburg, Freiburg, Germany; Center for Basics in NeuroModulation (NeuroModulBasics), Faculty of Medicine, University of Freiburg, Freiburg, Germany. Electronic address:

Microglia are resident immune cells in the central nervous system (CNS) that are capable of carrying out prominent and various functions during development and adulthood under both homeostatic and disease conditions. Although microglia are traditionally thought to be heterogeneous populations, which potentially allows them to achieve a wide range of responses to environmental changes for the maintenance of CNS homeostasis, a lack of unbiased and high-throughput methods to assess microglia heterogeneity has prevented the study of spatially and temporally distributed microglia subsets. The recent emergence of novel single-cell techniques, such as cytometry by time-of-flight mass spectrometry (CyTOF) and single-cell RNA sequencing, enabled scientists to overcome such limitations and reveal the surprising context-dependent heterogeneity of microglia. In this review, we summarize the current knowledge about the spatial, temporal, and functional diversity of microglia during development, homeostasis, and disease in mice and humans.
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http://dx.doi.org/10.1016/j.celrep.2020.01.010DOI Listing
February 2020

Resection of recurrent glioblastoma multiforme in elderly patients: a pseudo-randomized analysis revealed clinical benefit.

J Neurooncol 2020 Jan 13;146(2):381-387. Epub 2020 Jan 13.

Translational NeuroOncology Research Group, Medical Center, University of Freiburg, Freiburg, Germany.

Introduction: Elderly patients constitute an expanding part of our society. Due to a continuously increasing life expectancy, an optimal quality of life is expected even into advanced age. Glioblastoma (GBM) is more common in older patients, but they are still often withheld from efficient treatment due to worry of worse tolerance and have a significantly worse prognosis compared to younger patients. Our retrospective observational study aimed to investigate the therapeutic benefit from a second resection in recurrent glioblastoma of elderly patients.

Materials And Methods: We included a cohort of 39 elderly patients (> 65 years) with a second resection as treatment option in the case of a tumor recurrence. A causal inference model was built by multiple non- and semiparametric models, which was used to identify matched patients from our elderly GBM database which comprises 538 patients. The matched cohorts were analyzed by a Cox-regression model adjusted by time-dependent covariates.

Results: The Cox-regression analysis showed a significant survival benefit (Hazard Ratio: 0.6, 95% CI 0.36-0.9, p-value = 0.0427) for the re-resected group (18.0 months, 95% CI 13.97-23.2 months) compared to the group without re-resection (10.1 months, 95% CI 8.09-20.9 months). No differences in the co-morbidities or hemato-oncological side effects during chemotherapy could be detected. Anesthetic- and surgical complications were rare and comparable to the complication rate of patients undergoing the first-line resection.

Conclusion: Taken together, in elderly patients, re-resection is an acceptable treatment option in the recurrent state of a glioblastoma. The individual evaluation of the patients' medical status as well as the chances of withstanding general anesthesia needs to be done in close interdisciplinary consultation. If these requirements are met, elderly patients benefit from a re-resection.
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http://dx.doi.org/10.1007/s11060-020-03393-zDOI Listing
January 2020

Endogenous retroviruses are associated with hippocampus-based memory impairment.

Proc Natl Acad Sci U S A 2019 12 2;116(51):25982-25990. Epub 2019 Dec 2.

Center for Molecular Innovation, Feinstein Institutes for Medical Research, Manhasset, NY 11030;

Retrotransposons compose a staggering 40% of the mammalian genome. Among them, endogenous retroviruses (ERV) represent sequences that closely resemble the proviruses created from exogenous retroviral infection. ERVs make up 8 to 10% of human and mouse genomes and range from evolutionarily ancient sequences to recent acquisitions. Studies in have provided a causal link between genomic retroviral elements and cognitive decline; however, in mammals, the role of ERVs in learning and memory remains unclear. Here we studied 2 independent murine models for ERV activation: muMT strain (lacking B cells and antibody production) and intracerebroventricular injection of streptozotocin (ICVI-STZ). We conducted behavioral assessments (contextual fear memory and spatial learning), as well as gene and protein analysis (RNA sequencing, PCR, immunohistochemistry, and western blot assays). Mice lacking mitochondrial antiviral-signaling protein (MAVS) and mice lacking stimulator of IFN genes protein (STING), 2 downstream sensors of ERV activation, provided confirmation of ERV impact. We found that muMT mice and ICVI-STZ mice induced hippocampal ERV activation, as shown by increased gene and protein expression of the Gag sequence of the transposable element intracisternal A-particle. ERV activation was accompanied by significant hippocampus-related memory impairment in both models. Notably, the deficiency of the MAVS pathway was protective against ICVI-STZ-induced cognitive pathology. Overall, our results demonstrate that ERV activation is associated with cognitive impairment in mice. Moreover, they provide a molecular target for strategies aimed at attenuating retroviral element sensing, via MAVS, to treat dementia and neuropsychiatric disorders.
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http://dx.doi.org/10.1073/pnas.1822164116DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6925997PMC
December 2019

Mapping microglia states in the human brain through the integration of high-dimensional techniques.

Nat Neurosci 2019 12 18;22(12):2098-2110. Epub 2019 Nov 18.

Institute of Neuropathology, Faculty of Medicine, University of Freiburg, Freiburg, Germany.

Microglia are tissue-resident macrophages of the CNS that orchestrate local immune responses and contribute to several neurological and psychiatric diseases. Little is known about human microglia and how they orchestrate their highly plastic, context-specific adaptive responses during pathology. Here we combined two high-dimensional technologies, single-cell RNA-sequencing and time-of-flight mass cytometry, to identify microglia states in the human brain during homeostasis and disease. This approach enabled us to identify and characterize a previously unappreciated spectrum of transcriptional states in human microglia. These transcriptional states are determined by their spatial distribution, and they further change with aging and brain tumor pathology. This description of multiple microglia phenotypes in the human CNS may open promising new avenues for subset-specific therapeutic interventions.
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http://dx.doi.org/10.1038/s41593-019-0532-yDOI Listing
December 2019

Astrogliosis Releases Pro-Oncogenic Chitinase 3-Like 1 Causing MAPK Signaling in Glioblastoma.

Cancers (Basel) 2019 Sep 26;11(10). Epub 2019 Sep 26.

Translational NeuroOncology Research Group, Medical Center, University of Freiburg, 79106 Freiburg, Germany.

Although reactive astrocytes constitute a major component of the cellular environment in glioblastoma, their function and crosstalk to other components of the environment is still poorly understood. Gene expression analysis of purified astrocytes from both the tumor core and non-infiltrated cortex reveals a tumor-related up-regulation of Chitinase 3-like 1 (CHI3L1), a cytokine which is related to inflammation, extracellular tissue remodeling, and fibrosis. Further, we established and validated a co-culture model to investigate the impact of reactive astrocytes within the tumor microenvironment. Here we show that reactive astrocytes promote a subtype-shift of glioblastoma towards the mesenchymal phenotype, driving mitogen-activated protein kinases (MAPK) signaling as well as increased proliferation and migration. In addition, we demonstrate that MAPK signaling is directly caused by a CHI3L1-IL13RA2 co-binding, which leads to increased downstream MAPK and AKT signaling. This novel microenvironmental crosstalk highlights the crucial role of non-neoplastic cells in malignant brain tumors and opens up new perspectives for targeted therapies in glioblastoma.
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http://dx.doi.org/10.3390/cancers11101437DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6826948PMC
September 2019

Age-Related Gliosis Promotes Central Nervous System Lymphoma through CCL19-Mediated Tumor Cell Retention.

Cancer Cell 2019 09;36(3):250-267.e9

Institute of Virology, Technical University of Munich, 81675 Munich, Germany; Helmholtz Center Munich, 85764 Neuherberg, Germany; Institute of Molecular Immunology and Experimental Oncology, Technical University of Munich, Ismaningerstraße 22, 81675 Munich, Germany; Division of Chronic Inflammation and Cancer, German Cancer Research Center (DKFZ), Im Neuenheimer Feld 242, 69120 Heidelberg, Germany. Electronic address:

How lymphoma cells (LCs) invade the brain during the development of central nervous system lymphoma (CNSL) is unclear. We found that NF-κB-induced gliosis promotes CNSL in immunocompetent mice. Gliosis elevated cell-adhesion molecules, which increased LCs in the brain but was insufficient to induce CNSL. Astrocyte-derived CCL19 was required for gliosis-induced CNSL. Deleting CCL19 in mice or CCR7 from LCs abrogated CNSL development. Two-photon microscopy revealed LCs transiently entering normal brain parenchyma. Astrocytic CCL19 enhanced parenchymal CNS retention of LCs, thereby promoting CNSL formation. Aged, gliotic wild-type mice were more susceptible to forming CNSL than young wild-type mice, and astrocytic CCL19 was observed in both human gliosis and CNSL. Therefore, CCL19-CCR7 interactions may underlie an increased age-related risk for CNSL.
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http://dx.doi.org/10.1016/j.ccell.2019.08.001DOI Listing
September 2019

Giant Osteolytic Schwannoma from the Sixth Thoracic Nerve Root - An Interesting Case.

Rofo 2019 Dec 1;191(12):1123-1124. Epub 2019 Aug 1.

Neurosurgery, University of Freiburg, Freiburg im Breisgau, Germany.

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http://dx.doi.org/10.1055/a-0969-2254DOI Listing
December 2019

Human organotypic brain slice culture: a novel framework for environmental research in neuro-oncology.

Life Sci Alliance 2019 08 27;2(4). Epub 2019 Jun 27.

Translational NeuroOncology Research Group, Medical Center, University of Freiburg, Freiburg im Breisgau, Germany

When it comes to the human brain, models that closely mimic in vivo conditions are lacking. Living neuronal tissue is the closest representation of the in vivo human brain outside of a living person. Here, we present a method that can be used to maintain therapeutically resected healthy neuronal tissue for prolonged periods without any discernible changes in tissue vitality, evidenced by immunohistochemistry, genetic expression, and electrophysiology. This method was then used to assess glioblastoma (GBM) progression in its natural environment by microinjection of patient-derived tumor cells into cultured sections. The result closely resembles the pattern of de novo tumor growth and invasion, drug therapy response, and cytokine environment. Reactive transformation of astrocytes, as an example of the cellular nonmalignant tumor environment, can be accurately simulated with transcriptional differences similar to those of astrocytes isolated from acute GBM specimens. In a nutshell, we present a simple method to study GBM in its physiological environment, from which valuable insights can be gained. This technique can lead to further advancements in neuroscience, neuro-oncology, and pharmacotherapy.
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http://dx.doi.org/10.26508/lsa.201900305DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6599970PMC
August 2019

Large-Scale Validation of the Paddling Pool Task in the Clockmaze for Studying Hippocampus-Based Spatial Cognition in Mice.

Front Behav Neurosci 2019 7;13:121. Epub 2019 Jun 7.

Laboratory of Immune & Neural Networks, Institute of Molecular Medicine, Feinstein Institute for Medical Research, Northwell Health, Manhasset, NY, United States.

Rationally designed behavioral tests are important tools to assess the function of specific brain regions. The hippocampus is a crucial neural substrate for spatial cognition, and many studies have linked hippocampal dysfunction with defects on spatial learning and memory in neurological conditions ranging from Alzheimer's disease to autoimmune syndromes, such as neuropsychiatric lupus. While our understanding of hippocampal function, from the molecular to the system levels, has increased dramatically over the last decades, this effort has not yet translated into efficacious therapies for cognitive impairment. We think that the availability of highly validated behavioral paradigms to measure cognition in mouse models is likely to enhance the potential success of preclinical therapeutic modalities. Here, we present an extensive study of the paddling pool task (PPT), first reported by Deacon and Rawlins, in which mice learn to escape from shallow water through a peripheral exit in a circular arena dubbed the clockmaze. We show that the PPT provides highly reliable results when assaying spatial cognition in C57/BL6 mice (120 males, 40 females) and BALB/c mice (40 males, 90 females). Additionally, we develop a robust algorithm for the assessment of escape strategies with clearly quantifiable readouts, enabling fine-granular phenotyping. Notably, the use of spatial strategy increases linearly across trials in the PPT. In a separate cohort of mice, we apply muscimol injections to silence the dorsal CA1 region of the hippocampus and show that the use of the spatial strategy in the PPT relies on the integrity of the dorsal hippocampus. Additionally, we compare directly the PPT and the Morris water maze (MWM) task in C57/BL6 mice (20 males, 20 females) and BALB/c mice (20 males, 20 females) and we find that the PPT induces significantly lower anxiety, exhaustion and hypothermia than the MWM. We conclude that the PPT provides a robust assessment of spatial cognition in mice, which can be applied in conjunction with other tests, to facilitate hypothesis testing and drug development to combat cognitive impairment.
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http://dx.doi.org/10.3389/fnbeh.2019.00121DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6568215PMC
June 2019

Tumor-associated reactive astrocytes aid the evolution of immunosuppressive environment in glioblastoma.

Nat Commun 2019 06 11;10(1):2541. Epub 2019 Jun 11.

Translational NeuroOncology Research Group, Medical Center, University of Freiburg, 79106, Freiburg, Germany.

Reactive astrocytes evolve after brain injury, inflammatory and degenerative diseases, whereby they undergo transcriptomic re-programming. In malignant brain tumors, their function and crosstalk to other components of the environment is poorly understood. Here we report a distinct transcriptional phenotype of reactive astrocytes from glioblastoma linked to JAK/STAT pathway activation. Subsequently, we investigate the origin of astrocytic transformation by a microglia loss-of-function model in a human organotypic slice model with injected tumor cells. RNA-seq based gene expression analysis of astrocytes reveals a distinct astrocytic phenotype caused by the coexistence of microglia and astrocytes in the tumor environment, which leads to a large release of anti-inflammatory cytokines such as TGFβ, IL10 and G-CSF. Inhibition of the JAK/STAT pathway shifts the balance of pro- and anti-inflammatory cytokines towards a pro-inflammatory environment. The complex interaction of astrocytes and microglia cells promotes an immunosuppressive environment, suggesting that tumor-associated astrocytes contribute to anti-inflammatory responses.
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http://dx.doi.org/10.1038/s41467-019-10493-6DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6559986PMC
June 2019

Author Correction: Spatial and temporal heterogeneity of mouse and human microglia at single-cell resolution.

Nature 2019 Apr;568(7751):E4

Institute of Neuropathology, Medical Faculty, University of Freiburg, Freiburg, Germany.

In this Letter, Dominic Grün and Sagar have been added to the author list (affiliated with Max-Planck-Institute of Immunology and Epigenetics (MPI-IE), Freiburg, Germany). The author list, 'Author contribution' and 'Acknowledgements' sections have been corrected online. See accompanying Amendment.
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http://dx.doi.org/10.1038/s41586-019-1045-2DOI Listing
April 2019

Spatial and temporal heterogeneity of mouse and human microglia at single-cell resolution.

Nature 2019 02 13;566(7744):388-392. Epub 2019 Feb 13.

Institute of Neuropathology, Medical Faculty, University of Freiburg, Freiburg, Germany.

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.
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http://dx.doi.org/10.1038/s41586-019-0924-xDOI Listing
February 2019

Single-cell profiling identifies myeloid cell subsets with distinct fates during neuroinflammation.

Science 2019 01;363(6425)

Institute of Neuropathology, Faculty of Medicine, University of Freiburg, Freiburg, Germany.

The innate immune cell compartment is highly diverse in the healthy central nervous system (CNS), including parenchymal and non-parenchymal macrophages. However, this complexity is increased in inflammatory settings by the recruitment of circulating myeloid cells. It is unclear which disease-specific myeloid subsets exist and what their transcriptional profiles and dynamics during CNS pathology are. Combining deep single-cell transcriptome analysis, fate mapping, in vivo imaging, clonal analysis, and transgenic mouse lines, we comprehensively characterized unappreciated myeloid subsets in several CNS compartments during neuroinflammation. During inflammation, CNS macrophage subsets undergo self-renewal, and random proliferation shifts toward clonal expansion. Last, functional studies demonstrated that endogenous CNS tissue macrophages are redundant for antigen presentation. Our results highlight myeloid cell diversity and provide insights into the brain's innate immune system.
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http://dx.doi.org/10.1126/science.aat7554DOI Listing
January 2019

The multi-target effects of CNI-1493: convergence of anti-amylodogenic and anti-inflammatory properties in animal models of Alzheimer's disease.

Mol Med 2016 Dec 15;22:776-788. Epub 2016 Nov 15.

Center for Molecular Innovation, The Feinstein Institute for Medical Research, 350 Community drive, Manhasset, NY 11030 USA.

After several decades of Alzheimer's disease (AD) research and failed clinical trials, one can speculate that targeting a single pathway is not sufficient. However, a cocktail of novel therapeutics will constitute a challenging clinical trial. A more plausible approach will capitalize on a drug that has relevant and synergistic multiple-target effects in AD. We have previously demonstrated the efficacy of CNI-1493 in the CRND8 transgenic AD mouse model. Similar to many anti-inflammatory drugs that were tested in preclinical model of AD, it was speculated that the significant effect of CNI-1493 is due to its established anti-inflammatory properties in rodents and humans. In the present study, we set out to elucidate the protective mechanism of CNI-1493 as a drug simultaneously targeting several aspects of AD pathology. Using C1213, a highly similar analogue of CNI-1493 that lacks anti-inflammatory properties, we show that both compounds directly interact with soluble and insoluble Amyloid β (Aβ) aggregates and attenuate Aβ cytotoxicity in vitro. Additionally, CNI-1493 and C1213 ameliorated Aβ-induced behavioral deficits in nematodes. Finally, C1213 reduced Aβ plaque burden and cognitive deficits in transgenic CRND8 mice to a similar extent as previously shown with CNI-1493. Taken together, our findings suggest anti-amyloidogenic activity as a relevant component for the in-vivo efficacy of CNI-1493 and its analogue C1213. Thus, CNI-1493, a drug with proven safety in humans, is a viable candidate for novel multi-target therapeutic approaches to AD.
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http://dx.doi.org/10.2119/molmed.2016.00163DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5193463PMC
December 2016

Rodent models of neuroinflammation for Alzheimer's disease.

J Neuroinflammation 2015 Apr 17;12:74. Epub 2015 Apr 17.

Center for Molecular Innovation, The Feinstein Institute for Medical Research, 350 Community drive, Manhasset, NY, 11030, USA.

Alzheimer's disease remains incurable, and the failures of current disease-modifying strategies for Alzheimer's disease could be attributed to a lack of in vivo models that recapitulate the underlying etiology of late-onset Alzheimer's disease. The etiology of late-onset Alzheimer's disease is not based on mutations related to amyloid-β (Aβ) or tau production which are currently the basis of in vivo models of Alzheimer's disease. It has recently been suggested that mechanisms like chronic neuroinflammation may occur prior to amyloid-β and tau pathologies in late-onset Alzheimer's disease. The aim of this study is to analyze the characteristics of rodent models of neuroinflammation in late-onset Alzheimer's disease. Our search criteria were based on characteristics of an idealistic disease model that should recapitulate causes, symptoms, and lesions in a chronological order similar to the actual disease. Therefore, a model based on the inflammation hypothesis of late-onset Alzheimer's disease should include the following features: (i) primary chronic neuroinflammation, (ii) manifestations of memory and cognitive impairment, and (iii) late development of tau and Aβ pathologies. The following models fit the pre-defined criteria: lipopolysaccharide- and PolyI:C-induced models of immune challenge; streptozotocin-, okadaic acid-, and colchicine neurotoxin-induced neuroinflammation models, as well as interleukin-1β, anti-nerve growth factor and p25 transgenic models. Among these models, streptozotocin, PolyI:C-induced, and p25 neuroinflammation models are compatible with the inflammation hypothesis of Alzheimer's disease.
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http://dx.doi.org/10.1186/s12974-015-0291-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4404276PMC
April 2015

Systemic inflammation and the brain: novel roles of genetic, molecular, and environmental cues as drivers of neurodegeneration.

Front Cell Neurosci 2015 2;9:28. Epub 2015 Feb 2.

Elmezzi Graduate School of Molecular Medicine , Manhasset, NY , USA ; Feinstein Institute for Medical Research , Manhasset, NY , USA ; Department of Neurology, Instituto Nacional de Ciencias Médicas y Nutrición Salvador Zubirán , México City , Mexico.

The nervous and immune systems have evolved in parallel from the early bilaterians, in which innate immunity and a central nervous system (CNS) coexisted for the first time, to jawed vertebrates and the appearance of adaptive immunity. The CNS feeds from, and integrates efferent signals in response to, somatic and autonomic sensory information. The CNS receives input also from the periphery about inflammation and infection. Cytokines, chemokines, and damage-associated soluble mediators of systemic inflammation can also gain access to the CNS via blood flow. In response to systemic inflammation, those soluble mediators can access directly through the circumventricular organs, as well as open the blood-brain barrier. The resulting translocation of inflammatory mediators can interfere with neuronal and glial well-being, leading to a break of balance in brain homeostasis. This in turn results in cognitive and behavioral manifestations commonly present during acute infections - including anorexia, malaise, depression, and decreased physical activity - collectively known as the sickness behavior (SB). While SB manifestations are transient and self-limited, under states of persistent systemic inflammatory response the cognitive and behavioral changes can become permanent. For example, cognitive decline is almost universal in sepsis survivors, and a common finding in patients with systemic lupus erythematosus. Here, we review recent genetic evidence suggesting an association between neurodegenerative disorders and persistent immune activation; clinical and experimental evidence indicating previously unidentified immune-mediated pathways of neurodegeneration; and novel immunomodulatory targets and their potential relevance for neurodegenerative disorders.
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http://dx.doi.org/10.3389/fncel.2015.00028DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4313590PMC
February 2015

Genetic determinants of obesity and related vascular diseases.

Vitam Horm 2013 ;91:29-48

Department of Neurology, Philipps-University, Marburg Germany.

Obesity is one of the major risk factors of vascular diseases, and its prevalence is increasing worldwide. In the past decade, progress has been made in the understanding of genetic determinants of obesity and obesity-associated diseases. Genome-wide association studies identified a number of genetic variants associated with obesity. In addition to common variants, FTO and MC4R, new loci, such as TMEM18, KCTD15, GNPDA2, SH2B1, MTCH2, and NEGR1 have been detected. In the past years, abdominal obesity has been shown to be a more important vascular risk factor than the body mass index. In the context of vascular risk assessment, identification of genetic polymorphisms associated with accumulation of visceral fat is of special importance. Some polymorphisms associated with abdominal obesity, such as variants of gene encoding microsomal triglyceride transfer protein, have been already discovered. In this chapter, we provide a review of genetic determinants of obesity and discuss their role in obesity-related vascular diseases.
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http://dx.doi.org/10.1016/B978-0-12-407766-9.00002-XDOI Listing
December 2013

CNI-1493 attenuates neuroinflammation and dopaminergic neurodegeneration in the acute MPTP mouse model of Parkinson's disease.

Neurodegener Dis 2013 7;12(2):103-10. Epub 2012 Nov 7.

Department of Neurology, Philipps University Marburg, Marburg, Germany.

Background: Parkinson's disease (PD) is associated with neurodegeneration of dopaminergic neurons in the substantia nigra. Neuroinflammatory processes have been shown to be a key component of this neurodegeneration and, as such, small molecule compounds which inhibit these inflammatory events are a critical research focus.

Objective: CNI-1493 is an anti-inflammatory compound that strongly inhibits macrophages and also stimulates the cholinergic anti-inflammatory pathway. We have examined whether CNI-1493 has a neuroprotective effect in the acute 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) mouse model of PD.

Methods: CNI-1493 (8 mg/kg i.p.) or placebo administration was started 1 day before MPTP intoxication and repeated daily until sacrifice after MPTP intoxication. C57/Bl6 mice - either treated with CNI-1493 or with placebo - were injected intraperitoneally 4 times at 2-hour intervals with either 20 mg/kg MPTP-HCl or a corresponding volume of saline. Two or 7 days after the end of the MPTP intoxication, the animals were killed and their brains were processed for further analysis.

Results: Administration of CNI-1493 markedly protected tyrosine hydroxylase-positive substantia nigra neurons against MPTP neurotoxicity. CNI-1493 treatment in the MPTP model was also accompanied by a profound reduction of activated microglia within the substantia nigra, as measured by ionized calcium-binding adapter molecule-1 staining.

Conclusions: These findings support that CNI-1493 could reduce the MPTP-induced toxicity likely by inhibition of neuroinflammatory responses. The neuroprotective effect of CNI-1493 suggests that CNI-1493 might be a valuable neuroprotective candidate in the future treatment of PD.
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http://dx.doi.org/10.1159/000342714DOI Listing
January 2014

Health-related quality of life and its determinants in the urban Russian population with major depressive disorder: a cross-sectional study.

Int J Psychiatry Med 2012 ;43(1):35-49

Dept of Neurology, Philipps-University Marburg, Germany.

Objective: Depressive disorders pose a major challenge to healthcare in the countries of the former Soviet Union. The objective of the current study was to evaluate health-related quality of life (HrQoL) and its determinants in outpatients with major depressive disorder in an urban Russian population.

Methods: We consecutively recruited 100 urban Russian outpatients with major depression and 100 non-depressed controls who were matched for age and sex. The severity of their depression was assessed using the Hamilton Depression Rating Scale (HDRS). HrQoL was evaluated using the EuroQol (the EQ-5D and the visual analogue scale, EQ VAS). Independent determinants of HrQoL were identified using multiple regression analysis.

Results: The mean EQ VAS score was 43.0 +/- 27.4 in patients with depression compared to 81.4 +/- 14.7 in the controls (p < 0.01). Out of the domains of the EQ-5D, "anxiety/depression," "usual activities," and "self-care" were the most impaired. Independent determinants of reduced HrQoL were: severity of depression according to the HDRS; violent suicide attempts; suicide attempts in the past; and drug addiction.

Conclusions: HrQoL is considerably reduced in Russians with major depression. The disease-specific patterns of HrQoL impairment and the independent determinants of HrQoL identified in our study could be addressed in focused healthcare programs and clinical trials. Comorbid drug addiction as a determinant of HrQoL should receive greater attention in the management of depressive disorders in urban Russian populations.
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http://dx.doi.org/10.2190/PM.43.1.cDOI Listing
June 2012

Resveratrol mitigates lipopolysaccharide- and Aβ-mediated microglial inflammation by inhibiting the TLR4/NF-κB/STAT signaling cascade.

J Neurochem 2012 Feb 16;120(3):461-72. Epub 2011 Dec 16.

The Litwin-Zucker Research Center for the Study of Alzheimer's Disease, The Feinstein Institute for Medical Research, Manhasset, New York, USA.

Activation of microglia, the resident macrophages of the brain, around the amyloid plaques is a key hallmark of Alzheimer's disease (AD). Recent evidence in mouse models indicates that microglia are required for the neurodegenerative process of AD. Amyloid-β (Aβ) peptides, the core components of the amyloid plaques, can trigger microglial activation by interacting with several Toll-like receptors (TLRs), including TLR4. In this study, we show that resveratrol, a natural polyphenol associated with anti-inflammatory effects and currently in clinical trials for AD, prevented the activation of murine RAW 264.7 macrophages and microglial BV-2 cells treated with the TLR4 ligand, lipopolysaccharide (LPS). Resveratrol preferentially inhibited nuclear factor κ-light-chain-enhancer of activated B cells (NF-κB) activation upon LPS stimulation by interfering with IKK and IκB phosphorylation, an effect that potently reduced the transcriptional stimulation of several NF-κB target genes, including tumor necrosis factor-α and interleukin-6. Consequently, downstream phosphorylation of signal transducer and activator of transcription (STAT)1 and STAT3 upon LPS stimulation was also inhibited by resveratrol. We found that resveratrol acted upstream in the activation cascade by interfering with TLR4 oligomerization upon receptor stimulation. Resveratrol treatment also prevented the pro-inflammatory effect of fibrillar Aβ on macrophages by potently inhibiting the effect of Aβ on IκB phosphorylation, activation of STAT1 and STAT3, and on tumor necrosis factor-α and interleukin-6 secretion. Importantly, orally administered resveratrol in a mouse model of cerebral amyloid deposition lowered microglial activation associated with cortical amyloid plaque formation. Together this work provides strong evidence that resveratrol has in vitro and in vivo anti-inflammatory effects against Aβ-triggered microglial activation. Further studies in cell culture systems showed that resveratrol acted via a mechanism involving the TLR4/NF-κB/STAT signaling cascade.
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http://dx.doi.org/10.1111/j.1471-4159.2011.07594.xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3253186PMC
February 2012