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1.
Elife ; 122023 12 12.
Article in English | MEDLINE | ID: mdl-38085657

ABSTRACT

Microglial endolysosomal (dys)function is strongly implicated in neurodegenerative disease. Transcriptomic studies show that a microglial state characterised by a set of genes involved in endolysosomal function is induced in both mouse Alzheimer's disease (AD) models and human AD brain, and that the emergence of this state is emphasised in females. Cst7 (encoding cystatin F) is among the most highly upregulated genes in these microglia. However, despite such striking and robust upregulation, the function of Cst7 in neurodegenerative disease is not understood. Here, we crossed Cst7-/- mice with the AppNL-G-F mouse to test the role of Cst7 in a model of amyloid-driven AD. Surprisingly, we found that Cst7 plays a sexually dimorphic role regulating microglia in this model. In females, Cst7-/-AppNL-G-F microglia had greater endolysosomal gene expression, lysosomal burden, and amyloid beta (Aß) burden in vivo and were more phagocytic in vitro. However, in males, Cst7-/-AppNL-G-F microglia were less inflammatory and had a reduction in lysosomal burden but had no change in Aß burden. Overall, our study reveals functional roles for one of the most commonly upregulated genes in microglia across disease models, and the sex-specific profiles of Cst7-/--altered microglial disease phenotypes. More broadly, the findings raise important implications for AD including crucial questions on sexual dimorphism in neurodegenerative disease and the interplay between endolysosomal and inflammatory pathways in AD pathology.


Subject(s)
Alzheimer Disease , Cystatins , Neurodegenerative Diseases , Animals , Female , Humans , Male , Mice , Alzheimer Disease/metabolism , Amyloid beta-Peptides/genetics , Amyloid beta-Peptides/metabolism , Amyloid beta-Protein Precursor/metabolism , Cystatins/metabolism , Disease Models, Animal , Mice, Transgenic , Microglia/metabolism , Neurodegenerative Diseases/pathology
2.
Cell Rep Med ; 4(9): 101175, 2023 09 19.
Article in English | MEDLINE | ID: mdl-37652017

ABSTRACT

Synapse loss correlates with cognitive decline in Alzheimer's disease (AD). Data from mouse models suggests microglia are important for synapse degeneration, but direct human evidence for any glial involvement in synapse removal in human AD remains to be established. Here we observe astrocytes and microglia from human brains contain greater amounts of synaptic protein in AD compared with non-disease controls, and that proximity to amyloid-ß plaques and the APOE4 risk gene exacerbate this effect. In culture, mouse and human astrocytes and primary mouse and human microglia phagocytose AD patient-derived synapses more than synapses from controls. Inhibiting interactions of MFG-E8 rescues the elevated engulfment of AD synapses by astrocytes and microglia without affecting control synapse uptake. Thus, AD promotes increased synapse ingestion by human glial cells at least in part via an MFG-E8 opsonophagocytic mechanism with potential for targeted therapeutic manipulation.


Subject(s)
Alzheimer Disease , Microglia , Animals , Humans , Mice , Astrocytes , Eating , Synapses
3.
Immunology ; 167(4): 558-575, 2022 12.
Article in English | MEDLINE | ID: mdl-35881080

ABSTRACT

Post-stroke infection is a common complication of stroke that is associated with poor outcome. We previously reported that stroke induces an ablation of multiple sub-populations of B cells and reduces levels of immunoglobulin M (IgM) antibody, which coincides with the development of spontaneous bacterial pneumonia. The loss of IgM after stroke could be an important determinant of infection susceptibility and highlights this pathway as a target for intervention. We treated mice with a replacement dose of IgM-enriched intravenous immunoglobulin (IgM-IVIg) prior to and 24 h after middle cerebral artery occlusion (MCAO) and allowed them to recover for 2- or 5-day post-surgery. Treatment with IgM-IVIg enhanced bacterial clearance from the lung after MCAO and improved lung pathology but did not impact brain infarct volume. IgM-IVIg treatment induced immunomodulatory effects systemically, including rescue of splenic plasma B cell numbers and endogenous mouse IgM and IgA circulating immunoglobulin concentrations that were reduced by MCAO. Treatment attenuated MCAO-induced elevation of selected pro-inflammatory cytokines in the lung. IgM-IVIg treatment did not increase the number of lung mononuclear phagocytes or directly modulate macrophage phagocytic capacity but enhanced phagocytosis of Staphylococcus aureus bioparticles in vitro. Low-dose IgM-IVIg contributes to increased clearance of spontaneous lung bacteria after MCAO likely via increasing availability of antibody in the lung to enhance opsonophagocytic activity. Immunomodulatory effects of IgM-IVIg treatment may also contribute to reduced levels of damage in the lung after MCAO. IgM-IVIg shows promise as an antibacterial and immunomodulatory agent to use in the treatment of post-stroke infection.


Subject(s)
Bacterial Infections , Stroke , Mice , Animals , Immunoglobulins, Intravenous/therapeutic use , Immunologic Factors , Immunoglobulin M , Stroke/complications , Stroke/therapy , Bacteria , Lung
4.
J Neurosci ; 41(13): 3025-3038, 2021 03 31.
Article in English | MEDLINE | ID: mdl-33597269

ABSTRACT

Alzheimer's disease is a devastating neurodegenerative disease with a dramatically increasing prevalence and no disease-modifying treatment. Inflammatory lifestyle factors increase the risk of developing Alzheimer's disease. Zinc deficiency is the most prevalent malnutrition in the world and may be a risk factor for Alzheimer's disease potentially through enhanced inflammation, although evidence for this is limited. Here we provide epidemiological evidence suggesting that zinc supplementation was associated with reduced risk and slower cognitive decline, in people with Alzheimer's disease and mild cognitive impairment. Using the APP/PS1 mouse model of Alzheimer's disease fed a control (35 mg/kg zinc) or diet deficient in zinc (3 mg/kg zinc), we determined that zinc deficiency accelerated Alzheimer's-like memory deficits without modifying amyloid ß plaque burden in the brains of male mice. The NLRP3-inflammasome complex is one of the most important regulators of inflammation, and we show here that zinc deficiency in immune cells, including microglia, potentiated NLRP3 responses to inflammatory stimuli in vitro, including amyloid oligomers, while zinc supplementation inhibited NLRP3 activation. APP/PS1 mice deficient in NLRP3 were protected against the accelerated cognitive decline with zinc deficiency. Collectively, this research suggests that zinc status is linked to inflammatory reactivity and may be modified in people to reduce the risk and slow the progression of Alzheimer's disease.SIGNIFICANCE STATEMENT Alzheimer's disease is a common condition mostly affecting the elderly. Zinc deficiency is also a global problem, especially in the elderly and also in people with Alzheimer's disease. Zinc deficiency contributes to many clinical disorders, including immune dysfunction. Inflammation is known to contribute to the risk and progression of Alzheimer's disease; thus, we hypothesized that zinc status would affect Alzheimer's disease progression. Here we show that zinc supplementation reduced the prevalence and symptomatic decline in people with Alzheimer's disease. In an animal model of Alzheimer's disease, zinc deficiency worsened cognitive decline because of an enhancement in NLRP3-driven inflammation. Overall, our data suggest that zinc status affects Alzheimer's disease progression, and that zinc supplementation could slow the rate of cognitive decline.


Subject(s)
Alzheimer Disease/blood , Cognitive Dysfunction/blood , Disease Progression , NLR Family, Pyrin Domain-Containing 3 Protein/metabolism , Zinc/blood , Adult , Aged , Alzheimer Disease/diagnostic imaging , Alzheimer Disease/diet therapy , Animals , Cells, Cultured , Cognitive Dysfunction/diagnostic imaging , Cognitive Dysfunction/diet therapy , Dietary Supplements , Female , Follow-Up Studies , Humans , Inflammation Mediators/metabolism , Male , Mice , Mice, Inbred C57BL , Mice, Transgenic , Middle Aged , Zinc/administration & dosage , Zinc/deficiency
5.
J Biol Chem ; 294(21): 8325-8335, 2019 05 24.
Article in English | MEDLINE | ID: mdl-30940725

ABSTRACT

Interleukin (IL)-1 family cytokines potently regulate inflammation, with the majority of the IL-1 family proteins being secreted from immune cells via unconventional pathways. In many cases, secretion of IL-1 cytokines appears to be closely coupled to cell death, yet the secretory mechanisms involved remain poorly understood. Here, we studied the secretion of the three best-characterized members of the IL-1 superfamily, IL-1α, IL-1ß, and IL-18, in a range of conditions and cell types, including murine bone marrow-derived and peritoneal macrophages, human monocyte-derived macrophages, HeLa cells, and mouse embryonic fibroblasts. We discovered that IL-1ß and IL-18 share a common secretory pathway that depends upon membrane permeability and can operate in the absence of complete cell lysis and cell death. We also found that the pathway regulating the trafficking of IL-1α is distinct from the pathway regulating IL-1ß and IL-18. Although the release of IL-1α could also be dissociated from cell death, it was independent of the effects of the membrane-stabilizing agent punicalagin, which inhibited both IL-1ß and IL-18 release. These results reveal that in addition to their role as danger signals released from dead cells, IL-1 family cytokines can be secreted in the absence of cell death. We propose that models used in the study of IL-1 release should be considered context-dependently.


Subject(s)
Bone Marrow Cells/metabolism , Interleukin-18/metabolism , Interleukin-1alpha/metabolism , Interleukin-1beta/metabolism , Macrophages, Peritoneal/metabolism , Animals , Bone Marrow Cells/cytology , Fibroblasts/cytology , Fibroblasts/metabolism , HeLa Cells , Humans , Hydrolyzable Tannins/pharmacology , Macrophages, Peritoneal/cytology , Mice , Protein Transport/drug effects
6.
Nat Commun ; 9(1): 1156, 2018 03 20.
Article in English | MEDLINE | ID: mdl-29559685

ABSTRACT

The interleukin-1 (IL-1) receptor and ligand families are components of the immune system. Knowledge of their evolutionary history is essential to understand their function. Using chromosomal anatomy and sequence similarity, we show that IL-1 receptor family members are related and nine members are likely formed from duplication and modification of a proto-IL-1R1 receptor. The IL-1 ligands have a different evolutionary history. The first proto-IL-1ß gene coincided with proto-IL-1R1 and duplication events resulted in the majority of IL-1 ligand family members. However, large evolutionary distances are observed for IL-1α, IL-18 and IL-33 proteins. Further analysis show that IL-33 and IL-18 have poor sequence similarity and no chromosomal evidence of common ancestry with the IL-1ß cluster and therefore should not be included in the IL-1 ligand ancestral family. IL-1α formed from the duplication of IL-1ß, and moonlighting functions of pro-IL-1α acted as divergent selection pressures for the observed sequence dissimilarity.


Subject(s)
Evolution, Molecular , Interleukin-1beta/genetics , Animals , Humans , Interleukin-18/genetics , Interleukin-18/metabolism , Interleukin-1beta/metabolism , Interleukin-33/genetics , Interleukin-33/metabolism , Multigene Family , Phylogeny , Receptors, Interleukin-1 Type I/genetics , Receptors, Interleukin-1 Type I/metabolism , Vertebrates/classification , Vertebrates/genetics
7.
Sci Rep ; 7(1): 17077, 2017 12 06.
Article in English | MEDLINE | ID: mdl-29213066

ABSTRACT

Inflammation is a host defense process against infection. Inflammatory mediators include cytokines of the interleukin-1 family, such as IL-1α and IL-1ß. Unlike IL-1ß, IL-1α carries an N-terminal nuclear localisation sequence (NLS) and is trafficked to the nucleus. The importance of IL-1α nuclear localisation is poorly understood. Here, we used CRISPR/Cas9 to make inactivating mutations to the NLS on the Il1a gene. A colony of NLS mutant mice was successfully generated with precise knock-in mutations to incapacitate NLS function. NLS mutant mice had no gross changes in immunophenotype or inflammatory response but, surprisingly, failed to express IL-1α. We deduced that, in making specific mutations in the Il1a gene, we also mutated a long-noncoding (lnc)RNA in the complementary strand which has cis-regulatory transcriptional control of the Il1a gene itself. The mutations generated in the Il1a gene also result in mutation of the lncRNA sequence and a predicted alteration of its secondary structure, potentially explaining a subsequent failure to function as a transcriptional activator of Il1a expression. Thus, lncRNA secondary structure may regulate IL-1α expression. Our results serve as a cautionary note that CRISPR -mediated genome editing without full knowledge of genomic context can result in unexpected, yet potentially informative observations.


Subject(s)
CRISPR-Cas Systems/genetics , Cell Nucleus/metabolism , Interleukin-1beta/genetics , Animals , Blood Platelets/cytology , Blood Platelets/metabolism , Bone Marrow Cells/cytology , Bone Marrow Cells/drug effects , Bone Marrow Cells/metabolism , Cytokines/metabolism , Gene Editing , Ionomycin/pharmacology , Lipopolysaccharides/pharmacology , Macrophages/cytology , Macrophages/immunology , Macrophages/metabolism , Mice , Mice, Inbred C57BL , Mutation , Nigericin/pharmacology , Nucleic Acid Conformation , Phenotype , RNA, Long Noncoding/genetics , RNA, Long Noncoding/metabolism , Spleen/metabolism , T-Lymphocytes/immunology , T-Lymphocytes/metabolism
8.
Cell Chem Biol ; 24(11): 1321-1335.e5, 2017 Nov 16.
Article in English | MEDLINE | ID: mdl-28943355

ABSTRACT

NLRP3 is a receptor important for host responses to infection, yet is also known to contribute to devastating diseases such as Alzheimer's disease, diabetes, atherosclerosis, and others, making inhibitors for NLRP3 sought after. One of the inhibitors currently in use is 2-aminoethoxy diphenylborinate (2APB). Unfortunately, in addition to inhibiting NLRP3, 2APB also displays non-selective effects on cellular Ca2+ homeostasis. Here, we use 2APB as a chemical scaffold to build a series of inhibitors, the NBC series, which inhibit the NLRP3 inflammasome in vitro and in vivo without affecting Ca2+ homeostasis. The core chemical insight of this work is that the oxazaborine ring is a critical feature of the NBC series, and the main biological insight the use of NBC inhibitors led to was that NLRP3 inflammasome activation was independent of Ca2+. The NBC compounds represent useful tools to dissect NLRP3 function, and may lead to oxazaborine ring-containing therapeutics.


Subject(s)
Boron/chemistry , Inflammasomes/metabolism , NLR Family, Pyrin Domain-Containing 3 Protein/antagonists & inhibitors , Animals , Bone Marrow Cells/cytology , Boron/pharmacology , Boron Compounds/chemistry , Boron Compounds/metabolism , Boron Compounds/pharmacology , Calcium/metabolism , Cells, Cultured , Crystallography, X-Ray , Humans , Inhibitory Concentration 50 , Interleukin-1beta/metabolism , Lipopolysaccharides/toxicity , Macrophages/cytology , Macrophages/drug effects , Macrophages/metabolism , Mice , Molecular Conformation , NLR Family, Pyrin Domain-Containing 3 Protein/metabolism , Structure-Activity Relationship
9.
Int J Mol Sci ; 18(1)2017 Jan 05.
Article in English | MEDLINE | ID: mdl-28067797

ABSTRACT

In the conventional pathway of protein secretion, leader sequence-containing proteins leave the cell following processing through the endoplasmic reticulum (ER) and Golgi body. However, leaderless proteins also enter the extracellular space through mechanisms collectively known as unconventional secretion. Unconventionally secreted proteins often have vital roles in cell and organism function such as inflammation. Amongst the best-studied inflammatory unconventionally secreted proteins are interleukin (IL)-1ß, IL-1α, IL-33 and high-mobility group box 1 (HMGB1). In this review we discuss the current understanding of the unconventional secretion of these proteins and highlight future areas of research such as the role of nuclear localisation.


Subject(s)
HMGB1 Protein/metabolism , Inflammation/metabolism , Interleukin-1alpha/metabolism , Interleukin-1beta/metabolism , Interleukin-33/metabolism , Animals , Extracellular Space/metabolism , Humans , Models, Biological , Protein Transport
10.
Nat Commun ; 7: 12504, 2016 08 11.
Article in English | MEDLINE | ID: mdl-27509875

ABSTRACT

Non-steroidal anti-inflammatory drugs (NSAIDs) inhibit cyclooxygenase-1 (COX-1) and COX-2 enzymes. The NLRP3 inflammasome is a multi-protein complex responsible for the processing of the proinflammatory cytokine interleukin-1ß and is implicated in many inflammatory diseases. Here we show that several clinically approved and widely used NSAIDs of the fenamate class are effective and selective inhibitors of the NLRP3 inflammasome via inhibition of the volume-regulated anion channel in macrophages, independently of COX enzymes. Flufenamic acid and mefenamic acid are efficacious in NLRP3-dependent rodent models of inflammation in air pouch and peritoneum. We also show therapeutic effects of fenamates using a model of amyloid beta induced memory loss and a transgenic mouse model of Alzheimer's disease. These data suggest that fenamate NSAIDs could be repurposed as NLRP3 inflammasome inhibitors and Alzheimer's disease therapeutics.


Subject(s)
Alzheimer Disease/prevention & control , Anti-Inflammatory Agents, Non-Steroidal/pharmacology , Flufenamic Acid/pharmacology , Inflammasomes/metabolism , Mefenamic Acid/pharmacology , NLR Family, Pyrin Domain-Containing 3 Protein/metabolism , Alzheimer Disease/metabolism , Animals , Bone Marrow Cells/metabolism , Cell Death , Chloride Channels/metabolism , Cysteine/metabolism , Female , Genotype , Inflammation , Interleukin-1beta/metabolism , Macrophages/metabolism , Memory Disorders/drug therapy , Mice , Mice, Inbred C57BL , Mice, Knockout , Mice, Transgenic , Pattern Recognition, Visual/drug effects , Rats
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