Cellular perturbations underlying Alzheimer’s disease (AD) are primarily studied in human postmortem samples and model organisms. Here, we generated a single-nucleus atlas from a rare cohort of ...cortical biopsies from living individuals with varying degrees of AD pathology. We next performed a systematic cross-disease and cross-species integrative analysis to identify a set of cell states that are specific to early AD pathology. These changes—which we refer to as the early cortical amyloid response—were prominent in neurons, wherein we identified a transitional hyperactive state preceding the loss of excitatory neurons, which we confirmed by acute slice physiology on independent biopsy specimens. Microglia overexpressing neuroinflammatory-related processes also expanded as AD pathology increased. Finally, both oligodendrocytes and pyramidal neurons upregulated genes associated with β-amyloid production and processing during this early hyperactive phase. Our integrative analysis provides an organizing framework for targeting circuit dysfunction, neuroinflammation, and amyloid production early in AD pathogenesis.
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•Single-nucleus profiling of human cortex biopsies uncovers amyloid-associated states•Upper-layer pyramidal neurons show hyperactivity prior to degeneration•Microglial states correlate with pathological and clinical progression•Signatures of amyloid production identified in both neurons and oligodendrocytes
Generating single-nucleus atlas from cortical biopsies of living individuals at early stage of Alzheimer’s disease, cell states of neurons, microglia, and oligodendrocytes associated with AD pathology are identified.
β-amyloid (Aβ) and α-synuclein (α-syn) are aggregation-prone proteins typically associated with two distinct neurodegenerative disorders: Alzheimer's disease (AD) and Parkinson's disease. Yet α-syn ...was first found in association with AD plaques several years before being linked to Parkinson's disease or Lewy body formation. Nowadays, a large subset of AD patients (~50%) is well recognized to co-exhibit significant α-syn Lewy body pathology. Unfortunately, these AD Lewy body variant patients suffer from additional symptoms and an accelerated disease course. Basic research has begun to show that Aβ and α-syn may act synergistically to promote the aggregation and accumulation of each other. While the exact mechanisms by which these proteins interact remain unclear, growing evidence suggests that Aβ may drive α-syn pathology by impairing protein clearance, activating inflammation, enhancing phosphorylation, or directly promoting aggregation. This review examines the interactions between Aβ and α-syn and proposes potential mechanistic links between Aβ accumulation and α-syn pathogenesis.
Making the most of high‐dimensional cytometry data Marsh‐Wakefield, Felix MD; Mitchell, Andrew J; Norton, Samuel E ...
Immunology and cell biology,
August 2021, Letnik:
99, Številka:
7
Journal Article
Recenzirano
Odprti dostop
High‐dimensional cytometry represents an exciting new era of immunology research, enabling the discovery of new cells and prediction of patient responses to therapy. A plethora of analysis and ...visualization tools and programs are now available for both new and experienced users; however, the transition from low‐ to high‐dimensional cytometry requires a change in the way users think about experimental design and data analysis. Data from high‐dimensional cytometry experiments are often underutilized, because of both the size of the data and the number of possible combinations of markers, as well as to a lack of understanding of the processes required to generate meaningful data. In this article, we explain the concepts behind designing high‐dimensional cytometry experiments and provide considerations for new and experienced users to design and carry out high‐dimensional experiments to maximize quality data collection.
High‐dimensional cytometry represents an exciting new era of immunology research; however, the transition from low‐ to high‐dimensional cytometry requires a change in the way users think about experimental design and data analysis. We explain the concepts behind designing high‐dimensional cytometry experiments and provide considerations for new and experienced users to design and carry out high‐dimensional experiments.
Alzheimer's disease (AD) is the leading cause of age‐related neurodegeneration and is characterized neuropathologically by the accumulation of insoluble beta‐amyloid (Aβ) peptides. In AD brains, ...plaque‐associated myeloid (PAM) cells cluster around Aβ plaques but fail to effectively clear Aβ by phagocytosis. PAM cells were originally thought to be brain‐resident microglia. However, several studies have also suggested that Aβ‐induced inflammation causes peripheral monocytes to enter the otherwise immune‐privileged brain. The relationship between AD progression and inflammation in the brain remains ambiguous because microglia and monocyte‐derived macrophages are extremely difficult to distinguish from one another in an inflamed brain. Whether PAM cells are microglia, peripheral macrophages, or a mixture of both remains unclear. CD11a is a component of the β2 integrin LFA1. We have determined that CD11a is highly expressed on peripheral immune cells, including macrophages, but is not expressed by mouse microglia. These expression patterns remain consistent in LPS‐treated inflamed mice, as well as in two mouse models of AD. Thus, CD11a can be used as a marker to distinguish murine microglia from infiltrating peripheral immune cells. Using CD11a, we show that PAM cells in AD transgenic brains are comprised entirely of microglia. We also demonstrate a novel fluorescence‐assisted quantification technique (FAQT), which reveals a significant increase in T lymphocytes, especially in the brains of female AD mice. Our findings support the notion that microglia are the lead myeloid players in AD and that rejuvenating their phagocytic potential may be an important therapeutic strategy.
Main Points
CD11a expression can distinguish microglia (CD11a−) from peripheral immune cells (CD11a+) in homeostasis, neuroinflammation, and Alzheimer's disease mice.
Plaque‐associated myeloid (PAM) cells do not express CD11a and are microglia.
Fluorescence‐assisted quantification technique (FAQT) reveals a significant increase in microglia numbers and in infiltrating T cells in the brains of AD female mice.
Transplantation of neural stem cells (NSCs) can improve cognition in animal models of Alzheimer's disease (AD). However, AD is a protracted disorder, and prior studies have examined only short-term ...effects. We therefore used an immune-deficient model of AD (Rag-5xfAD mice) to examine long-term transplantation of human NSCs (StemCells Inc.; HuCNS-SCs). Five months after transplantation, HuCNS-SCs had engrafted and migrated throughout the hippocampus and exhibited no differences in survival or migration in response to β-amyloid pathology. Despite robust engraftment, HuCNS-SCs failed to terminally differentiate and over a quarter of the animals exhibited ectopic human cell clusters within the lateral ventricle. Unlike prior short-term experiments with research-grade HuCNS-SCs, we also found no evidence of improved cognition, no changes in brain-derived neurotrophic factor, and no increase in synaptic density. These data, while disappointing, reinforce the notion that individual human NSC lines need to be carefully assessed for efficacy and safety in appropriate long-term models.
•Human neural stem cells (HuCNS-SC) have been used in multiple human clinical trials•HuCNS-SC originally derived under GMP conditions did not improve cognition in AD mice•HuCNS-SC failed to differentiate, improve synaptic density, or increase BDNF levels•HuCNS-SC formed ectopic ventricular clusters in a quarter of transplanted mice
In this article, Blurton-Jones and colleagues demonstrate that StemCells, Inc. human neural stem cells (HuCNS-SC), which were originally derived under GMP conditions, failed to improve cognition, synaptic density, or increase BDNF in an immune-deficient AD mouse model. Furthermore, cells formed ectopic ventricular clusters in over a quarter of transplanted animals.
Disruption of retromer-dependent endosomal trafficking is considered pathogenic in late-onset Alzheimer's disease (AD). Here, to investigate this disruption in the intact brain, we turn to a genetic ...mouse model where the retromer core protein VPS35 is depleted in hippocampal neurons, and then we replete VPS35 using an optimized viral vector protocol. The VPS35 depletion-repletion studies strengthen the causal link between the neuronal retromer and AD-associated neuronal phenotypes, including the acceleration of amyloid precursor protein cleavage and the loss of synaptic glutamate receptors. Moreover, the studies show that the neuronal retromer can regulate a distinct, dystrophic, microglia morphology, phenotypic of hippocampal microglia in AD. Finally, the neuronal and, in part, the microglia responses to VPS35 depletion were found to occur independent of tau. Showing that the neuronal retromer can regulate AD-associated pathologies in two of AD's principal cell types strengthens the link, and clarifies the mechanism, between endosomal trafficking and late-onset sporadic AD.
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•Endosomal trafficking is a pathogenic biological pathway in Alzheimer's disease (AD)•Retromer conducts specific endosomal recycling routes found defective in AD•Neuronal and microglial phenotypes characterize AD-associated hippocampal pathology•Retromer in hippocampal neurons regulates neuronal and microglial phenotypes
Alzheimer's cellular pathologies in the hippocampal formation are typified by a neuronal phenotype and a distinct, dystrophic, microglial phenotype. Qureshi et al. show that retromer-dependent endosomal recycling in hippocampal neurons, a pathogenic biological pathway, can regulate both cellular phenotypes and find that this regulation is largely independent of tau.
Synucleinopathies are a group of neurodegenerative disorders sharing the common feature of misfolding and accumulation of the presynaptic protein α‐synuclein (α‐syn) into insoluble aggregates. Within ...this diverse group, Dementia with Lewy Bodies (DLB) is characterized by the aberrant accumulation of α‐syn in cortical, hippocampal, and brainstem neurons, resulting in multiple cellular stressors that particularly impair dopamine and glutamate neurotransmission and related motor and cognitive function. Recent studies show that murine neural stem cell (NSC) transplantation can improve cognitive or motor function in transgenic models of Alzheimer's and Huntington's disease, and DLB. However, examination of clinically relevant human NSCs in these models is hindered by the challenges of xenotransplantation and the confounding effects of immunosuppressant drugs on pathology and behavior. To address this challenge, we developed an immune‐deficient transgenic model of DLB that lacks T‐, B‐, and NK‐cells, yet exhibits progressive accumulation of human α‐syn (h‐α‐syn)‐laden inclusions and cognitive and motor impairments. We demonstrate that clinically relevant human neural progenitor cells (line CNS10‐hNPCs) survive, migrate extensively and begin to differentiate preferentially into astrocytes following striatal transplantation into this DLB model. Critically, grafted CNS10‐hNPCs rescue both cognitive and motor deficits after 1 and 3 months and, furthermore, restore striatal dopamine and glutamate systems. These behavioral and neurochemical benefits are likely achieved by reducing α‐syn oligomers. Collectively, these results using a new model of DLB demonstrate that hNPC transplantation can impact a broad array of disease mechanisms and phenotypes and suggest a cellular therapeutic strategy that should be pursued. Stem Cells Translational Medicine 2017;6:1477–1490
Background
Microglia have emerged as key players in the pathogenesis of neurodegenerative conditions such as Alzheimer’s disease (AD), taking on distinct transcriptional and functional states. While ...the ability to profile complex tissues at single‐cell resolution in postmortem brain tissue provides insight into disease‐associated cellular states within the brain, more is required to understand how these changes modulate disease pathogenesis. Building on foundational transcriptomics, the advances in multi‐modal profiling of genomics and proteomics allows for a greater understanding of neuroimmune dysfunction in neurodegenerative disease.
Method
Transcriptomic datasets are often confounded by variability in collection and sequencing methodologies. We have optimized tissue dissociation and cell isolation protocols to avoid many of the pitfalls commonly found in bulk and single‐cell transcriptomic studies. Using these optimized protocols, we have begun assessing paired samples (blood, brain biopsy, and cerebrospinal fluid (CSF), from patients at risk for AD to understand neuroimmune changes in early‐phases of AD. We have also characterized the transcriptional and functional responses of human induced pluripotent stem cell (iPSC) microglia in response to a variety of neurodegenerative brain‐relevant challenges, including amyloid, apoptotic neurons and synaptic debris.
Result
Using scRNA‐seq on fresh mouse and human tissue we have identified a dissociation‐induced signature in microglia that is highly prevalent in current literature, and developed an experimental methodology to prevent this artifact. We have also identified a similar signature in post‐mortem tissue via snRNA‐seq that may be the result of acute‐pre/post‐mortem processes (Marsh et al., 2022). Powerfully, we also have cerebrospinal fluid (CSF) from the same patients, allowing us to correlate proteomic and transcriptomic analyses to determine the connections between disease‐associated transcriptomic cell states and analyte biomarkers. We can then use the iPSC microglia (iMGLs) to understand how altered transcriptomic states and biomarker profiles may alter microglial functions to elucidate mechanisms by which microglia contribute to disease pathogenesis.
Conclusion
Optimization of experimental and analysis methods along with extensive multi‐modal profiling of the same patients will lead to greater understanding of the neuroimmune landscape of neurodegenerative disease to better enable development of novel predictive biomarkers and therapeutic targets.
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