ABSTRACT
Despite recent advances in cancer immunotherapy, certain tumor types, such as Glioblastomas, are highly resistant due to their tumor microenvironment disabling the anti-tumor immune response. Here we show, by applying an in-silico multidimensional model integrating spatially resolved and single-cell gene expression data of 45,615 immune cells from 12 tumor samples, that a subset of Interleukin-10-releasing HMOX1+ myeloid cells, spatially localizing to mesenchymal-like tumor regions, drive T-cell exhaustion and thus contribute to the immunosuppressive tumor microenvironment. These findings are validated using a human ex-vivo neocortical glioblastoma model inoculated with patient derived peripheral T-cells to simulate the immune compartment. This model recapitulates the dysfunctional transformation of tumor infiltrating T-cells. Inhibition of the JAK/STAT pathway rescues T-cell functionality both in our model and in-vivo, providing further evidence of IL-10 release being an important driving force of tumor immune escape. Our results thus show that integrative modelling of single cell and spatial transcriptomics data is a valuable tool to interrogate the tumor immune microenvironment and might contribute to the development of successful immunotherapies.
Subject(s)
Brain Neoplasms/immunology , Glioblastoma/immunology , Interleukin-10/metabolism , Myeloid Cells/metabolism , T-Lymphocytes/immunology , Adult , Aged , Brain Neoplasms/drug therapy , Brain Neoplasms/pathology , Cell Communication/immunology , Cell Line, Tumor , Female , Glioblastoma/drug therapy , Glioblastoma/pathology , Healthy Volunteers , Heme Oxygenase-1/metabolism , Humans , Immunotherapy/methods , Janus Kinase Inhibitors/pharmacology , Janus Kinase Inhibitors/therapeutic use , Janus Kinases/antagonists & inhibitors , Janus Kinases/metabolism , Male , Middle Aged , Neocortex/cytology , Neocortex/immunology , Neocortex/pathology , Primary Cell Culture , RNA-Seq , STAT Transcription Factors/metabolism , Signal Transduction/drug effects , Signal Transduction/immunology , Single-Cell Analysis , T-Lymphocytes/drug effects , T-Lymphocytes/metabolism , Tissue Culture Techniques , Tumor Escape , Tumor Microenvironment/immunologyABSTRACT
Genes and environment interact during intrauterine life, and potentially alter the developmental trajectory of the brain. This can result in life-long consequences on brain function. We have previously developed two transgenic mouse lines that suppress Gad1 expression in parvalbumin (PVALB) and neuropeptide Y (NPY) expressing interneuron populations using a bacterial artificial chromosome (BAC)-driven miRNA-based silencing technology. We were interested to assess if maternal immune activation (MIA), genetic interneuronal inhibition, and the combination of these two factors disrupt and result in long-term changes in neuroinflammatory gene expression, sterol biosynthesis, and acylcarnitine levels in the brain of maternally exposed offspring. Pregnant female WT mice were given a single intraperitoneal injection of saline or polyinosinic-polycytidilic acid [poly(I:C)] at E12.5. Brains of offspring were analyzed at postnatal day 90. We identified complex and persistent neuroinflammatory gene expression changes in the hippocampi of MIA-exposed offspring, as well in the hippocampi of Npy/Gad1 and Pvalb/Gad1 mice. In addition, both MIA and genetic inhibition altered the post-lanosterol sterol biosynthesis in the neocortex and disrupted the typical acylcarnitine profile. In conclusion, our findings suggest that both MIA and inhibition of interneuronal function have long-term consequences on critical homeostatic mechanisms of the brain, including immune function, sterol levels, and energy metabolism.
Subject(s)
Inflammation Mediators/immunology , Interneurons/immunology , Neuroimmunomodulation/physiology , Prenatal Exposure Delayed Effects/genetics , Prenatal Exposure Delayed Effects/immunology , Animals , Female , Glutamate Decarboxylase/deficiency , Glutamate Decarboxylase/genetics , Hippocampus/drug effects , Hippocampus/immunology , Hippocampus/metabolism , Inflammation Mediators/metabolism , Interferon Inducers/toxicity , Interneurons/drug effects , Interneurons/metabolism , Male , Mice , Mice, Inbred C57BL , Mice, Transgenic , Neocortex/drug effects , Neocortex/immunology , Neocortex/metabolism , Neuroimmunomodulation/drug effects , Poly I-C/toxicity , Pregnancy , Prenatal Exposure Delayed Effects/chemically induced , Prenatal Exposure Delayed Effects/metabolismSubject(s)
Cytokines/metabolism , Epilepsy, Temporal Lobe/immunology , Microvessels/pathology , Neocortex/blood supply , Nitric Oxide Synthase Type II/metabolism , Adolescent , Adult , Case-Control Studies , Child , Cytokines/analysis , Epilepsy, Temporal Lobe/pathology , Epilepsy, Temporal Lobe/surgery , Female , Humans , Male , Microvessels/immunology , Middle Aged , Neocortex/immunology , Neocortex/pathology , Neocortex/surgery , Nitric Oxide Synthase Type II/analysis , Young AdultABSTRACT
BACKGROUND: Interferon-γ (IFN-γ, a type II IFN) is present in the central nervous system (CNS) under various conditions. Evidence is emerging that, in addition to its immunological role, IFN-γ modulates neuronal morphology, function, and development in several brain regions. Previously, we have shown that raising levels of IFN-ß (a type I IFN) lead to increased neuronal excitability of neocortical layer 5 pyramidal neurons. Because of shared non-canonical signaling pathways of both cytokines, we hypothesized a similar neocortical role of acutely applied IFN-γ. METHODS: We used semi-quantitative RT-PCR, immunoblotting, and immunohistochemistry to analyze neuronal expression of IFN-γ receptors and performed whole-cell patch-clamp recordings in layer 5 pyramidal neurons to investigate sub- and suprathreshold excitability, properties of hyperpolarization-activated cyclic nucleotide-gated current (Ih), and inhibitory neurotransmission under the influence of acutely applied IFN-γ. RESULTS: We show that IFN-γ receptors are present in the membrane of rat's neocortical layer 5 pyramidal neurons. As expected from this and the putative overlap in IFN type I and II alternative signaling pathways, IFN-γ diminished Ih, mirroring the effect of type I IFNs, suggesting a likewise activation of protein kinase C (PKC). In contrast, IFN-γ did neither alter subthreshold nor suprathreshold neuronal excitability, pointing to augmented inhibitory transmission by IFN-γ. Indeed, IFN-γ increased electrically evoked inhibitory postsynaptic currents (IPSCs) on neocortical layer 5 pyramidal neurons. Furthermore, amplitudes of spontaneous IPSCs and miniature IPSCs were elevated by IFN-γ, whereas their frequency remained unchanged. CONCLUSIONS: The expression of IFN-γ receptors on layer 5 neocortical pyramidal neurons together with the acute augmentation of inhibition in the neocortex by direct application of IFN-γ highlights an additional interaction between the CNS and immune system. Our results strengthen our understanding of the role of IFN-γ in neocortical neurotransmission and emphasize its impact beyond its immunological properties, particularly in the pathogenesis of neuropsychiatric disorders.
Subject(s)
Interferon-gamma/metabolism , Neocortex/metabolism , Neuroimmunomodulation/physiology , Pyramidal Cells/metabolism , Receptors, Interferon/metabolism , Animals , Interferon-gamma/pharmacology , Male , Neocortex/drug effects , Neocortex/immunology , Pyramidal Cells/drug effects , Pyramidal Cells/immunology , Rats , Rats, WistarABSTRACT
Although the potential contribution of the human gastrointestinal (GI) tract microbiome to human health, aging, and disease is becoming increasingly acknowledged, the molecular mechanics and signaling pathways of just how this is accomplished is not well-understood. Major bacterial species of the GI tract, such as the abundant Gram-negative bacilli Bacteroides fragilis (B. fragilis) and Escherichia coli (E. coli), secrete a remarkably complex array of pro-inflammatory neurotoxins which, when released from the confines of the healthy GI tract, are pathogenic and highly detrimental to the homeostatic function of neurons in the central nervous system (CNS). For the first time here we report the presence of bacterial lipopolysaccharide (LPS) in brain lysates from the hippocampus and superior temporal lobe neocortex of Alzheimer's disease (AD) brains. Mean LPS levels varied from two-fold increases in the neocortex to three-fold increases in the hippocampus, AD over age-matched controls, however some samples from advanced AD hippocampal cases exhibited up to a 26-fold increase in LPS over age-matched controls. This "Perspectives" paper will further highlight some very recent research on GI tract microbiome signaling to the human CNS, and will update current findings that implicate GI tract microbiome-derived LPS as an important internal contributor to inflammatory degeneration in the CNS.
Subject(s)
Alzheimer Disease/microbiology , Bacteroides fragilis/metabolism , Escherichia coli/metabolism , Gastrointestinal Microbiome , Gastrointestinal Tract/microbiology , Hippocampus/immunology , Lipopolysaccharides/metabolism , Alzheimer Disease/immunology , Bacteroides fragilis/immunology , Escherichia coli/immunology , Humans , Neocortex/immunologyABSTRACT
Mammalian target of rapamycin (mTOR) regulates cell growth, cell differentiation and protein synthesis. Rapamycin, an inhibitor of mTOR, has been widely used as an immunosuppressant and anti-cancer drug. Recently, mTOR inhibitors have also been reported to be a potential anti-epileptic drug, which may be effective when used in young patients with genetic epilepsy. Thus, a suitable dose of rapamycin which can maintain the normal function of mTOR and has fewer side effects ideally should be identified. In the present study, we first detected changes in marker proteins of mTOR signaling pathway during development. Then we determined the dose of rapamycin by treating rats of 2 weeks of age with different doses of rapamycin for 3 days and detected its effect on mTOR pathway. Young rats were then treated with a suitable dose of rapamycin for 4 weeks and the effect of rapamycin on mTOR, development and immunity were investigated. We found that the expression of the marker proteins of mTOR pathway was changed during development in brain hippocampus and neocortex. After 3 days of treanent, 0.03 mg/kg rapamycin had no effect on phospho-S6, whereas 0.1, 0.3, 1.0 and 3.0 mg/kg rapamycin inhibited phospho-S6 in a dose-dependent manner. However, only 1.0 mg/kg and 3.0 mg/kg rapamycin inhibited phospho-S6 after 4 weeks treatment of rapamycin. Parallel to this result, rats treated with 0.1 and 0.3 mg/kg rapamycin had no obvious adverse effects, whereas rats treated with 1.0 and 3.0 mg/kg rapamycin showed significant decreases in body, spleen and thymus weight. Additionally, rats treated with 1.0 and 3.0 mg/kg rapamycin exhibited cognitive impairment and anxiety as evident by maze and open field experiments. Furthermore, the content of IL-1ß, IL-2, IFN-γ, TNF-α in serum and cerebral cortex were significantly decreased in 1.0 and 3.0 mg/kg rapamycin-treated rats. The expression of DCX was also significantly decreased in 1.0 and 3.0 mg/kg rapamycin-treated rats. However, rats treated with 1.0 mg/kg rapamycin exhibited fewer and milder side effects than those treated with 3.0 mg/kg. In summary, all these data suggest that there is not a rapamycin dose that can inhibit mTOR for epilepsy without causing any side effects, but 1 mg/kg may be the optimal dose for young rats for suppressing mTOR with relatively few side effects.
Subject(s)
Gene Expression Regulation, Developmental/drug effects , Hippocampus/drug effects , Immunosuppressive Agents/adverse effects , Neocortex/drug effects , Sirolimus/adverse effects , Age Factors , Animals , Anxiety/chemically induced , Anxiety/genetics , Anxiety/pathology , Body Weight/drug effects , Dose-Response Relationship, Drug , Doublecortin Domain Proteins , Doublecortin Protein , Exploratory Behavior/drug effects , Female , Hippocampus/growth & development , Hippocampus/immunology , Interleukins/genetics , Interleukins/immunology , Male , Maze Learning/drug effects , Microtubule-Associated Proteins/genetics , Microtubule-Associated Proteins/immunology , Neocortex/growth & development , Neocortex/immunology , Neuropeptides/genetics , Neuropeptides/immunology , Organ Size/drug effects , Phosphorylation , Rats , Rats, Sprague-Dawley , Ribosomal Protein S6 Kinases/genetics , Ribosomal Protein S6 Kinases/immunology , Spleen/drug effects , Spleen/growth & development , Spleen/immunology , TOR Serine-Threonine Kinases/antagonists & inhibitors , TOR Serine-Threonine Kinases/genetics , TOR Serine-Threonine Kinases/immunology , Thymus Gland/drug effects , Thymus Gland/growth & development , Thymus Gland/immunology , Tumor Necrosis Factor-alpha/genetics , Tumor Necrosis Factor-alpha/immunologyABSTRACT
Amyloid ß peptide (Aß) immunization of Alzheimer's disease (AD) patients has been reported to induce amyloid plaque removal, but with little impact on cognitive decline. We have explored the consequences of Aß immunotherapy on neurons in post mortem brain tissue. Eleven immunized (AN1792, Elan Pharmaceuticals) AD patients were compared to 28 non-immunized AD cases. Immunohistochemistry on sections of neocortex was performed for neuron-specific nuclear antigen (NeuN), neurofilament protein (NFP) and phosphorylated-(p)PKR (pro-apoptotic kinase detected in degenerating neurons). Quantification was performed for pPKR and status spongiosis (neuropil degeneration), NeuN-positive neurons/field, curvature of the neuronal processes and interneuronal distance. Data were corrected for age, gender, duration of dementia and APOE genotype and also assessed in relation to Aß42 and tau pathology and key features of AD. In non-immunized patients, the degree of neuritic curvature correlated with spongiosis and pPKR, and overall the neurodegenerative markers correlated better with tau pathology than Aß42 load. Following immunization, spongiosis increased, interneuronal distance increased, while the number of NeuN-positive neurons decreased, consistent with enhanced neuronal loss. However, neuritic curvature was reduced and pPKR was associated with Aß removal in immunized patients. In AD, associations of spongiosis status, curvature ratio and pPKR load with microglial markers Iba1, CD68 and CD32 suggest a role for microglia in neurodegeneration. After immunization, correlations were detected between the number of NeuN-positive neurons and pPKR with Iba1, CD68 and CD64, suggesting that microglia are involved in the neuronal loss. Our findings suggest that in established AD this form of active Aß immunization may predominantly accelerate loss of damaged degenerating neurons. This interpretation is consistent with in vivo imaging indicating an increased rate of cerebral atrophy in immunized AD patients.
Subject(s)
Alzheimer Disease/drug therapy , Alzheimer Vaccines/therapeutic use , Amyloid beta-Peptides/therapeutic use , Neocortex/drug effects , Neurons/drug effects , Aged , Aged, 80 and over , Alzheimer Disease/diagnosis , Alzheimer Disease/immunology , Alzheimer Disease/metabolism , Amyloid beta-Peptides/analysis , Antigens, Nuclear/analysis , Autopsy , Biomarkers/analysis , Case-Control Studies , Female , Humans , Male , Middle Aged , Neocortex/chemistry , Neocortex/immunology , Neocortex/pathology , Nerve Degeneration , Nerve Tissue Proteins/analysis , Neurofibrillary Tangles/drug effects , Neurofibrillary Tangles/immunology , Neurofibrillary Tangles/pathology , Neurofilament Proteins/analysis , Neurons/chemistry , Neurons/immunology , Neurons/pathology , Peptide Fragments/analysis , Phosphorylation , Plaque, Amyloid , Treatment Outcome , eIF-2 Kinase/analysis , tau Proteins/analysisABSTRACT
Narcolepsy is a chronic sleep disorder, likely with an autoimmune component. During 2009 and 2010, a link between A(H1N1)pdm09 Pandemrix vaccination and onset of narcolepsy was suggested in Scandinavia. In this study, we searched for autoantibodies related to narcolepsy using a neuroanatomical array: rat brain sections were processed for immunohistochemistry/double labeling using patient sera/cerebrospinal fluid as primary antibodies. Sera from 89 narcoleptic patients, 52 patients with other sleep-related disorders (OSRDs), and 137 healthy controls were examined. Three distinct patterns of immunoreactivity were of particular interest: pattern A, hypothalamic melanin-concentrating hormone and proopiomelanocortin but not hypocretin/orexin neurons; pattern B, GABAergic cortical interneurons; and pattern C, mainly globus pallidus neurons. Altogether, 24 of 89 (27%) narcoleptics exhibited pattern A or B or C. None of the patterns were exclusive for narcolepsy but were also detected in the OSRD group at significantly lower numbers. Also, some healthy controls exhibited these patterns. The antigen of pattern A autoantibodies was identified as the common C-terminal epitope of neuropeptide glutamic acid-isoleucine/α-melanocyte-stimulating hormone (NEI/αMSH) peptides. Passive transfer experiments on rat showed significant effects of pattern A human IgGs on rapid eye movement and slow-wave sleep time parameters in the inactive phase and EEG θ-power in the active phase. We suggest that NEI/αMSH autoantibodies may interfere with the fine regulation of sleep, contributing to the complex pathogenesis of narcolepsy and OSRDs. Also, patterns B and C are potentially interesting, because recent data suggest a relevance of those brain regions/neuron populations in the regulation of sleep/arousal.
Subject(s)
Autoantibodies/blood , Brain/immunology , Brain/pathology , Narcolepsy/immunology , Narcolepsy/pathology , Sleep/physiology , Adolescent , Adult , Animals , Autoantibodies/immunology , Colchicine/analogs & derivatives , Colchicine/pharmacology , Electroencephalography , Globus Pallidus/immunology , Globus Pallidus/pathology , Hippocampus/immunology , Hippocampus/pathology , Humans , Immunoglobulin G/blood , Interneurons/immunology , Interneurons/pathology , Male , Mice , Mice, Inbred C57BL , Mice, Transgenic , Neocortex/immunology , Neocortex/pathology , Nerve Tissue Proteins/metabolism , Olfactory Bulb/immunology , Olfactory Bulb/pathology , Rats , Rats, Wistar , Young AdultABSTRACT
The CD200/CD200R inhibitory immune ligand-receptor system regulates microglial activation/quiescence in adult brain. Here, we investigated CD200/CD200R at different stages of postnatal development, when microglial maturation takes place. We characterized the spatiotemporal, cellular, and quantitative expression pattern of CD200 and CD200R in the developing and adult C57/BL6 mice brain by immunofluorescent labeling and Western blotting. CD200 expression increased from postnatal day 1 (P1) to P5-P7, when maximum levels were found, and decreased to adulthood. CD200 was located surrounding neuronal bodies, and very prominently in cortical layer I, where CD200(+) structures included glial fibrillary acidic protein (GFAP)(+) astrocytes until P7. In the hippocampus, CD200 was mainly observed in the hippocampal fissure, where GFAP(+) /CD200(+) astrocytes were also found until P7. CD200(+) endothelium was seen in the hippocampal fissure and cortical blood vessels, notably from P14, showing maximum vascular CD200 in adults. CD200R(+) cells were a population of ameboid/pseudopodic Iba1(+) microglia/macrophages observed at all ages, but significantly decreasing with increasing age. CD200R(+) /Iba1(+) macrophages were prominent in the pial meninges and ventricle lining, mainly at P1-P5. CD200R(+) /Iba1(+) perivascular macrophages were observed in cortical and hippocampal fissure blood vessels, showing maximum density at P7, but being prominent until adulthood. CD200R(+) /Iba1(+) ameboid microglia in the cingulum at P1-P5 were the only CD200R(+) cells in the nervous tissue. In conclusion, the main sites of CD200/CD200R interaction seem to include the molecular layer and pial surface in neonates and blood vessels from P7 until adulthood, highlighting the possible role of the CD200/CD200R system in microglial development and renewal.
Subject(s)
Antigens, CD/metabolism , Brain Chemistry/immunology , Membrane Glycoproteins/metabolism , Neural Inhibition/immunology , Aging/genetics , Aging/immunology , Animals , Animals, Newborn , Antibody Specificity/genetics , Antigen-Antibody Reactions/genetics , Antigens, CD/immunology , Brain Chemistry/genetics , Female , Hippocampus/blood supply , Hippocampus/growth & development , Hippocampus/immunology , Macrophages/cytology , Macrophages/immunology , Macrophages/metabolism , Male , Membrane Glycoproteins/immunology , Mice , Mice, Inbred C57BL , Microglia/cytology , Microglia/immunology , Microglia/metabolism , Neocortex/blood supply , Neocortex/growth & development , Neocortex/immunology , Neural Inhibition/genetics , Neurogenesis/genetics , Neurogenesis/immunologyABSTRACT
Alzheimer's disease (AD) pathology is characterised by aggregation in the brain of amyloid-beta (Abeta) peptide and hyperphosphorylated tau (phospho-tau), although how these proteins interact in disease pathogenesis is unclear. Abeta immunisation results in removal of Abeta from the brain but cognitive decline continues to progress, possibly due to persistent phospho-tau. We quantified phospho-tau and Abeta42 in the brains of 10 AD patients (iAD) who were actively immunised with Abeta42 (AN1792, Elan Pharmaceuticals) compared with 28 unimmunised AD cases (cAD). The phospho-tau load was lower in the iAD than the cAD group in the cerebral cortex (cAD 1.08% vs. iAD 0.72%, P = 0.048), CA1 hippocampus (cAD 2.26% vs. iAD 1.05%; P = 0.001), subiculum (cAD 1.60% vs. iAD 0.31%; P = 0.001) and entorhinal cortex (cAD 1.10% vs. iAD 0.18%; P < 0.001). Assessment of the localisation within neurons of phospho-tau indicated that the Abeta immunotherapy-associated reduction was confined to neuronal processes, i.e. neuropil threads and dystrophic neurites. However, the phospho-tau accumulation in the neuronal cell bodies, contributing to neurofibrillary tangles, appeared not to be affected. In showing that Abeta immunisation can influence phospho-tau pathology, we confirm the position of Abeta as a target for modifying tau accumulation in AD and demonstrate a link between these proteins. However, the continuing progression of cognitive decline in AD patients after Abeta immunisation may be explained by its lack of apparent effect on tangles.
Subject(s)
Alzheimer Disease/immunology , Alzheimer Disease/metabolism , Amyloid beta-Peptides/immunology , Neurons/immunology , Neurons/metabolism , Peptide Fragments/immunology , tau Proteins/metabolism , Aged , Aged, 80 and over , Alzheimer Disease/pathology , Alzheimer Disease/therapy , CA1 Region, Hippocampal/immunology , CA1 Region, Hippocampal/metabolism , Entorhinal Cortex/immunology , Entorhinal Cortex/metabolism , Female , Follow-Up Studies , Humans , Immunization , Immunohistochemistry , Male , Middle Aged , Neocortex/immunology , Neocortex/metabolism , Neurites/immunology , Neurites/metabolism , Neurofibrillary Tangles/immunology , Neurofibrillary Tangles/metabolism , Phosphorylation , Treatment OutcomeABSTRACT
Various immune responses have been described in epileptic patients and animal models of epilepsy, but immune responses in brain after a single seizure are poorly understood. We studied immune responses in brain after a single brief generalized tonic-clonic seizure in mice. C57bl/6 mice, either unanesthetized or anesthetized (pentobarbital, ethyl chloride) received either electrical (15-30 mA, 100 Hz, 1s) or sham stimulation (subcutaneous electrodes over frontal lobe, no current). Electrical stimulation of unanesthetized mice resulted in tonic-clonic convulsions with hind-limb extension (maximal seizure), tonic-clonic convulsions without hind-limb extension (submaximal seizure), or no seizure. In contrast, such stimulation of anesthetized mice did not result in seizure. Mice were killed at 1h-7 days after seizure. Brains or regions dissected from brain (neocortex, hippocampus, midbrain, cerebellum) of each group were pooled, single cell suspensions prepared, and cells separated according to density. CD4(+) (CD3(+)CD45(Hi)) and CD8(+) (CD3(+)CD45(Hi)) T cell and CD45R(+) (CD45(Hi)) B cell numbers were determined by flow cytometry. At 24h after a maximal seizure, CD4(+) and CD8(+) T cells and CD45R(+) B cells appeared in brain, reaching peak numbers at 48 h, but were no longer detected at 7days. CD4(+) T cells and CD45R(+) B cells were preferentially found in neocortex compared with hippocampus, whereas CD8(+) T cells were preferentially found in hippocampus at 24h after a maximal seizure. In contrast, virtually no lymphocytes were detected in brains of unstimulated or sham stimulated mice, unanesthetized stimulated mice after submaximal or no seizure, and anesthetized stimulated mice at 1 h-7 day. Neither Ly6-G+ neutrophils nor erythrocytes were detected in brains of any animals, nor was there any detectable increase of blood-brain barrier permeability by uptake of Evans Blue dye. The results indicate that lymphocyte entry into brain after a single brief seizure is due to a selective process of recruitment into cortical regions.
Subject(s)
Hippocampus/pathology , Lymphocytes/physiology , Neocortex/pathology , Neutrophil Infiltration/physiology , Seizures/pathology , Anesthesia , Animals , Antibodies, Monoclonal , B-Lymphocytes/physiology , CD4-CD8 Ratio , Cell Movement , Cerebellum/pathology , Coloring Agents , Electrodes, Implanted , Electroshock , Erythrocytes/physiology , Evans Blue , Flow Cytometry , Hippocampus/immunology , Male , Mesencephalon/pathology , Mice , Mice, Inbred C57BL , Neocortex/immunology , Seizures/immunologySubject(s)
Neocortex/pathology , Neurons/immunology , Neurons/pathology , Paraneoplastic Cerebellar Degeneration/pathology , Adenocarcinoma/complications , Aged , Autoantibodies/immunology , Autoantigens/immunology , Blotting, Western , Carcinoma, Non-Small-Cell Lung/complications , Fluorescent Antibody Technique , Humans , Lung Neoplasms/complications , Male , Neocortex/immunology , Paraneoplastic Cerebellar Degeneration/etiology , Paraneoplastic Cerebellar Degeneration/immunologyABSTRACT
Recent studies revealed an important involvement of the cerebral cortex in multiple sclerosis (MS) patients. Cortical lesions in MS were reported to be less inflammatory and to show less structural damage than white matter lesions. Animal models reflecting the histopathological hallmarks of cortical demyelinated lesions in MS are sparse. Induction of experimental autoimmune encephalomyelitis (EAE) in the common marmoset has turned out to be an attractive non-human-primate model for MS. In the present study we investigated the presence and detailed cellular composition of cortical inflammatory demyelinating pathology in the common marmoset upon immunization with myelin oligodendrocyte glycoprotein (MOG). Extensive cortical demyelination reflecting the topographically distinct cortical lesion types in MS patients was revealed by immunohistochemistry for myelin basic protein (MBP). We explored the density of T- and B-lymphocytes, MHC-II expressing macrophages/microglia cells and early activated macrophages (MRP14) at perivascular and parenchymal lesions sites in neocortex and subcortical white matter. Despite a similar density of perivascular inflammatory infiltrates in the demyelinated neocortex, a considerable lower fraction of macrophages was found to express MRP14 in the neocortex indicating a different activation pattern in cortical compared with white matter lesions. Furthermore, cortical EAE lesions in marmoset monkeys revealed immunoglobulin leakage and complement component C9 deposition in intracortical but not subpial demyelination. Our findings indicate that the inflammatory response, especially macrophage and microglia activation, may be regulated differently in gray matter areas in primate brain.
Subject(s)
Demyelinating Diseases/immunology , Encephalomyelitis, Autoimmune, Experimental/immunology , Macrophages/immunology , Microglia/immunology , Neocortex/immunology , Animals , Callithrix , Demyelinating Diseases/etiology , Demyelinating Diseases/pathology , Disease Models, Animal , Encephalomyelitis, Autoimmune, Experimental/complications , Encephalomyelitis, Autoimmune, Experimental/pathology , Female , Immunization , Macrophages/metabolism , Male , Microglia/metabolism , Multiple Sclerosis/immunology , Multiple Sclerosis/pathology , Myelin Basic Protein/metabolism , Myelin Proteins , Myelin-Associated Glycoprotein/immunology , Myelin-Oligodendrocyte Glycoprotein , Neocortex/metabolism , Neocortex/pathology , Neutrophil Infiltration/immunology , Tissue DistributionABSTRACT
The use of immunohistochemical methods has led to a new understanding of the prevalence and significance of cortical lesions in multiple sclerosis but these lesions have not yet been formally described in an animal model. In this study we have set out to use immunohistochemical techniques to identify and describe cortical lesions in marmosets with experimental autoimmune encephalomyelitis (EAE). Using antibodies to proteolipid protein (PLP), we found a total of 70 cortical lesions in 11 tissue blocks from 6 animals. These lesions were subdivided into leucocortical (40), intracortical (12) and subpial lesions (18). We quantified the density of inflammatory cells within lesions using a double labelling protocol which employed anti-PLP in addition to antibodies against markers of B-lymphocytes (CD20), T-lymphocytes (CD3), macrophages (MAC387) and MHC-II expressing cells (CR3/43). This analysis revealed that the large subpial lesions accounted for the majority of demyelinated cortex (88%) despite possessing the lowest density of inflammatory cells. This study has shown that lesions in this model share many of the major features of cortical lesions in multiple sclerosis both in terms of morphology and inflammatory cell content. We believe that this tool can be exploited in future studies to investigate the aetiology, development and clinical significance of cortical lesions in demyelinating disease.
Subject(s)
Encephalomyelitis, Autoimmune, Experimental/pathology , Multiple Sclerosis/pathology , Neocortex/pathology , Animals , Callithrix , Disease Models, Animal , Encephalomyelitis, Autoimmune, Experimental/immunology , Encephalomyelitis, Autoimmune, Experimental/metabolism , Female , Male , Multiple Sclerosis/immunology , Multiple Sclerosis/metabolism , Myelin Proteolipid Protein/immunology , Myelin Proteolipid Protein/metabolism , Neocortex/immunology , Neocortex/metabolismABSTRACT
Alzheimer's disease (AD) is characterized by a progressive cognitive decline leading to dementia and involves the deposition of amyloid-beta (Abeta) peptides into senile plaques. Other neuropathological features that accompany progression of the disease include a decrease in synaptic density, neurofibrillary tangles, dystrophic neurites, inflammation, and neuronal cell loss. In this study, we report the early kinetics of brain amyloid deposition and its associated inflammation in an early onset transgenic mouse model of AD (TgCRND8) harboring the human amyloid precursor protein gene with the Indiana and Swedish mutations. Both diffuse and compact plaques were detected as early as 9-10 weeks of age. Abeta-immunoreactive (Abeta-IR) plaques (4G8-positive) appeared first in the neocortex and amygdala, then in the hippocampal formation, and lastly in the thalamus. Compact plaques (ThioS-positive) with an amyloid core were observed as early as diffuse plaques were detected, but in lower numbers. Amyloid deposition increased progressively with age. The formation of plaques was concurrent with the appearance of activated microglial cells and shortly followed by the clustering of activated astrocytes around plaques at 13-14 weeks of age. This TgCRND8 mouse model allows for a rapid, time-dependent study of the relationship between the fibrillogenic process and the inflammatory response during the brain amyloidogenic process.
Subject(s)
Alzheimer Disease/metabolism , Amyloid beta-Protein Precursor/metabolism , Brain/metabolism , Inflammation/metabolism , Plaque, Amyloid/metabolism , Age Factors , Alzheimer Disease/genetics , Alzheimer Disease/immunology , Alzheimer Disease/pathology , Amygdala/immunology , Amygdala/metabolism , Amygdala/pathology , Amyloid beta-Protein Precursor/genetics , Amyloid beta-Protein Precursor/immunology , Animals , Benzothiazoles , Brain/immunology , Brain/pathology , CD11b Antigen/immunology , CD11b Antigen/metabolism , Disease Models, Animal , Glial Fibrillary Acidic Protein/metabolism , Hippocampus/immunology , Hippocampus/metabolism , Hippocampus/pathology , Immunohistochemistry , Inflammation/immunology , Inflammation/pathology , Mice , Mice, Inbred C57BL , Mice, Transgenic , Microglia/immunology , Microglia/metabolism , Neocortex/immunology , Neocortex/metabolism , Neocortex/pathology , Plaque, Amyloid/genetics , Plaque, Amyloid/immunology , Plaque, Amyloid/pathology , Thiazoles/metabolismABSTRACT
The nervous system and the immune system share several functional molecules involved in various cell-cell interaction events. In this study, we used in situ hybridization to identify immune molecules that are expressed by a restricted population of neurons in the mouse brain and found that mRNA for the beta subunit of T-cell receptor (TCRbeta) was predominantly and strongly localized to neurons in deep layers of the cerebral neocortex and weakly expressed in the thalamus. Developmentally, TCRbeta mRNA expression started at embryonic day 15 in the thalamic nuclei and at postnatal day 1 in the cerebral neocortex. The level of TCRbeta mRNA in the neocortex subsequently increased until postnatal day 21, and it remained high in the adult. Detailed analysis revealed that only the Cbeta2 segment of TCRbeta, not the Cbeta1 or Vbeta segments, was expressed by the brain neurons. By the 5' rapid amplification of cDNA ends method, we determined a brain-specific transcription start site in the Jbeta2 region locus, not in the Vbeta region locus. Furthermore, we confirmed that the aberrant transcription around the Jbeta2 region took place only in neurons and lymphocytes in transgenic mice. These results demonstrate that the transcriptional machinery for unrearranged TCRbeta expression is shared by the nervous and immune systems and raise a possibility of gene rearrangement in neurons under certain circumstances.
Subject(s)
Brain/immunology , Cell Differentiation/immunology , Gene Expression Regulation, Developmental/immunology , Gene Rearrangement, beta-Chain T-Cell Antigen Receptor , Receptors, Antigen, T-Cell, alpha-beta/genetics , Transcription, Genetic/immunology , Alternative Splicing/genetics , Alternative Splicing/immunology , Animals , Base Sequence/genetics , Brain/cytology , Brain/metabolism , Cell Communication/immunology , Gene Expression Regulation, Developmental/genetics , Mice , Mice, Inbred BALB C , Mice, Transgenic , Molecular Sequence Data , Neocortex/cytology , Neocortex/immunology , Neocortex/metabolism , Neural Pathways/cytology , Neural Pathways/immunology , Neural Pathways/metabolism , Neuroimmunomodulation/genetics , Neuroimmunomodulation/immunology , Promoter Regions, Genetic/genetics , Promoter Regions, Genetic/immunology , Protein Isoforms/genetics , Protein Isoforms/metabolism , Protein Structure, Tertiary/genetics , RNA, Messenger/metabolism , Receptors, Antigen, T-Cell, alpha-beta/biosynthesis , Receptors, Antigen, T-Cell, alpha-beta/deficiency , Thalamus/cytology , Thalamus/immunology , Thalamus/metabolism , Transcription Initiation Site/physiology , Transcription, Genetic/geneticsABSTRACT
OBJECTIVES: Inflammation contributes to degeneration in Alzheimer's disease (AD), not simply as a secondary phenomenon, but primarily as a significant source of pathology. [(123)I]iodo-PK11195 is a single photon emission computed tomography (SPECT) ligand for the peripheral benzodiazepine receptor, the latter being expressed on microglia (brain resident macrophages) and upregulated under inflammatory circumstances. The objectives were to assess AD inflammation by detecting [(123)I]iodo-PK11195 uptake changes and investigate how uptake values relate with perfusion SPECT and neuropsychological findings. METHODS: Ten AD and 9 control subjects were included. [(123)I]iodo-PK11195 SPECT images were realigned into stereotactic space where binding indices, normalized on cerebellar uptake, were calculated. RESULTS: The mean [(123)I]iodo-PK11195 uptake was increased in AD patients compared with controls in nearly all neocortical regions; however, statistical significance was only reached in the frontal and right mesotemporal regions. Significant correlations were found between regional increased [(123)I]iodo-PK11195 uptake and cognitive deficits. CONCLUSIONS: [(123)I]iodo-PK11195 is a cellular disease activity marker and allows in vivo assessment of microglial inflammation in AD.
Subject(s)
Alzheimer Disease/diagnostic imaging , Encephalitis/diagnostic imaging , Image Processing, Computer-Assisted , Iodine Radioisotopes , Isoquinolines , Microglia/diagnostic imaging , Neocortex/diagnostic imaging , Receptors, GABA-A/physiology , Tomography, Emission-Computed, Single-Photon/methods , Aged , Aged, 80 and over , Alzheimer Disease/immunology , Alzheimer Disease/pathology , Brain Mapping , Dominance, Cerebral/physiology , Encephalitis/immunology , Encephalitis/pathology , Female , Humans , Male , Mental Status Schedule , Microglia/immunology , Microglia/pathology , Middle Aged , Neocortex/immunology , Neocortex/pathology , Pilot Projects , Radioligand Assay , Reference Values , Up-Regulation/physiologyABSTRACT
The effect of subchronic intracerebroventricular injection of the human immunodeficiency virus type 1 (HIV-1) recombinant protein gp120 (100 ng, given daily for up to seven consecutive days) on interleukin-1beta expression was studied by immunohistochemistry in the brain of adult rats. In comparison to control, bovine serum albumin (300 ng, given intracerebroventricularly for up to seven days) -treated animals (n=6), interleukin-1beta immunoreactivity increased in the brain cortex and hippocampus of rats (n=6) receiving a single injection of the viral protein 24 h before analysis with more substantial increases being observed in these regions of the brain (n=6) after seven days treatment. Double-labelling immunofluorescence experiments support a neuronal and, possibly, a microglial cell origin for gp120-enhanced interleukin-1beta expression. Transmission electron microscopy analysis of brain tissue sections revealed that combination treatments (given intracerebroventricularly daily for seven days) with gp120 (100 ng) and interleukin-1 receptor antagonist (80 ng) or with the interleukin converting enzyme inhibitor II (100 pmol), but not with leupeptin (100 pmol), prevented apoptotic death of rat (n=6/group) brain cortical cells typically elicited by the viral protein. These data demonstrate that gp120 enhances interleukin-1beta expression in the brain and this may be involved in the mechanism underlying apoptosis induced by gp120 in the brain cortex of rat. Further support to this hypothesis comes from the evidence that intracerebroventricular injection of murine recombinant interleukin-1beta (200 U, given daily for seven consecutive days) produces DNA fragmentation in the brain cortex of rat (n=6). Interestingly, the latter treatment enhanced nerve growth factor level in the hippocampus but not in the cerebral cortex and this coincides with a similar effect recently reported in identical brain areas of rats treated likewise with gp120. In conclusion, the present data demonstrate that treatment with gp120 enhances interleukin-1beta expression and this participates in the mechanism of apoptotic cell death in the brain cortex of rat. By contrast, in the hippocampus, gp120-enhanced interleukin-1beta expression elevates nerve growth factor that may prevent or delay apoptosis in this plastic region of the rat brain.
Subject(s)
Apoptosis/physiology , HIV Envelope Protein gp120/pharmacology , Interleukin-1/genetics , Interleukin-1/pharmacology , Neocortex/pathology , Animals , Apoptosis/drug effects , Body Temperature/drug effects , Cattle , Citrulline/metabolism , Gene Expression Regulation , HIV Envelope Protein gp120/administration & dosage , Humans , In Situ Nick-End Labeling , Injections, Intraventricular , Interleukin 1 Receptor Antagonist Protein , Interleukin-1/biosynthesis , Kinetics , Leupeptins/pharmacology , Male , Microglia/drug effects , Microglia/immunology , Neocortex/drug effects , Neocortex/immunology , Rats , Rats, Wistar , Recombinant Proteins/administration & dosage , Recombinant Proteins/pharmacology , Serum Albumin, Bovine/administration & dosage , Serum Albumin, Bovine/pharmacology , Sialoglycoproteins/pharmacology , Time FactorsABSTRACT
1. The authors studied the morphology of CalbindinD28K (CaBp) immunoreactive cells and processes in the hippocampal formation and the prefrontal cortex of schizophrenics using the immunohistochemical technique of avidin-biotin-complex method (ABC method), and the results were compared with those from normal human brains. 2. In the hippocampal formation area CA2 of schizophrenics, many CaBp-immunopositive cell bodies and fibers were disordered in their arrangement compared to normal control brains. 3. In the prefrontal cortex (Brodmann area 9) of schizophrenics, many immunopositive cell bodies were exhibited irregular axis arrangement and fiber disarray. 4. The altered distribution pattern of CaBp-immunopositive structures in the hippocampal formation and the prefrontal cortex might indicate the existence of GABA(gamma-aminobutyric acid)ergic dysfunction in the brain of schizophrenic patients.