scholarly journals Extracellular vesicles from amyloid-β exposed cell cultures induce severe dysfunction in cortical neurons

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Chiara Beretta ◽  
Elisabeth Nikitidou ◽  
Linn Streubel-Gallasch ◽  
Martin Ingelsson ◽  
Dag Sehlin ◽  
...  
Keyword(s):  
Extracellular Vesicles ◽  
Cortical Neurons ◽  
Amyloid Β ◽  
Neuronal Cell ◽  
Time Lapse ◽  
Cellular Mechanisms ◽  
Axonal Swelling ◽  
Tem Analysis ◽  
Neuronal Cell Bodies ◽  
Time Lapse Imaging

AbstractAlzheimer’s disease (AD) is characterized by a substantial loss of neurons and synapses throughout the brain. The exact mechanism behind the neurodegeneration is still unclear, but recent data suggests that spreading of amyloid-β (Aβ) pathology via extracellular vesicles (EVs) may contribute to disease progression. We have previously shown that an incomplete degradation of Aβ42 protofibrils by astrocytes results in the release of EVs containing neurotoxic Aβ. Here, we describe the cellular mechanisms behind EV-associated neurotoxicity in detail. EVs were isolated from untreated and Aβ42 protofibril exposed neuroglial co-cultures, consisting mainly of astrocytes. The EVs were added to cortical neurons for 2 or 4 days and the neurodegenerative processes were followed with immunocytochemistry, time-lapse imaging and transmission electron microscopy (TEM). Addition of EVs from Aβ42 protofibril exposed co-cultures resulted in synaptic loss, severe mitochondrial impairment and apoptosis. TEM analysis demonstrated that the EVs induced axonal swelling and vacuolization of the neuronal cell bodies. Interestingly, EV exposed neurons also displayed pathological lamellar bodies of cholesterol deposits in lysosomal compartments. Taken together, our data show that the secretion of EVs from Aβ exposed cells induces neuronal dysfunction in several ways, indicating a central role for EVs in the progression of Aβ-induced pathology.

scholarly journals Sindbis Virus-Induced Neuronal Death Is both Necrotic and Apoptotic and Is Ameliorated byN-Methyl-d-Aspartate Receptor Antagonists

2001 ◽  
Vol 75 (15) ◽  
pp. 7114-7121 ◽  
Author(s):  
Jennifer L. Nargi-Aizenman ◽  
Diane E. Griffin
Keyword(s):  
Cell Death ◽  
Cortical Neurons ◽  
Sindbis Virus ◽  
Apoptotic Cell ◽  
Time Lapse ◽  
Apoptotic Cell Death ◽  
Time Lapse Imaging ◽  
Alphavirus Infection

ABSTRACT Virus infection of neurons leads to different outcomes ranging from latent and noncytolytic infection to cell death. Viruses kill neurons directly by inducing either apoptosis or necrosis or indirectly as a result of the host immune response. Sindbis virus (SV) is an alphavirus that induces apoptotic cell death both in vitro and in vivo. However, apoptotic changes are not always evident in neurons induced to die by alphavirus infection. Time lapse imaging revealed that SV-infected primary cortical neurons exhibited both apoptotic and necrotic morphological features and that uninfected neurons in the cultures also died. Antagonists of the N-methyl-d-aspartate (NMDA) subtype of glutamate receptors protected neurons from SV-induced death without affecting virus replication or SV-induced apoptotic cell death. These results provide evidence that SV infection activates neurotoxic pathways that result in aberrant NMDA receptor stimulation and damage to infected and uninfected neurons.

Young’s Modulus of Cortical and P19 Derived Neurons Measured by Atomic Force Microscopy

2012 ◽  
Vol 1420 ◽  
Author(s):  
Elise Spedden ◽  
James D. White ◽  
David Kaplan ◽  
Cristian Staii
Keyword(s):  
Young’S Modulus ◽  
Cortical Neurons ◽  
Young's Modulus ◽  
Neuronal Cell ◽  
Short Term ◽  
Force Microscopy ◽  
Protein Substrates ◽  
Atomic Force ◽  
Neuronal Cell Bodies ◽  
Bulk Tissue

ABSTRACTIn this paper we use the Atomic Force Microscope to measure the Young’s modulus for two types of neuronal cell bodies: cortical neurons obtained from rat embryos and neurons derived from P19 mouse embryonic carcinoma stem cells. The neurons are plated on different substrates coated with two types of protein growth factors, poly-D-lysine and laminin. We report on the Young’s modulus of each type of neuron as well as the variation of modulus between cells plated on different protein substrates. We compare these results to various individual cell and bulk tissue measurements reported in literature. We additionally report on an observed change in the Young’s modulus of cortical neurons when subjected to a short-term reduction in ambient temperature.

scholarly journals The schizophrenia susceptibility factor dysbindin and its associated complex sort cargoes from cell bodies to the synapse

2011 ◽  
Vol 22 (24) ◽  
pp. 4854-4867 ◽  
Author(s):  
Jennifer Larimore ◽  
Karine Tornieri ◽  
Pearl V. Ryder ◽  
Avanti Gokhale ◽  
Stephanie A. Zlatic ◽  
...  
Keyword(s):  
Membrane Protein ◽  
Cortical Neurons ◽  
Neuronal Cells ◽  
Neuronal Cell ◽  
Adaptor Protein ◽  
Nerve Terminals ◽  
Endosome Trafficking ◽  
Neuronal Cell Bodies ◽  
Cultured Cortical Neurons ◽  
Lysosome Related Organelles

Dysbindin assembles into the biogenesis of lysosome-related organelles complex 1 (BLOC-1), which interacts with the adaptor protein complex 3 (AP-3), mediating a common endosome-trafficking route. Deficiencies in AP-3 and BLOC-1 affect synaptic vesicle composition. However, whether AP-3-BLOC-1–dependent sorting events that control synapse membrane protein content take place in cell bodies upstream of nerve terminals remains unknown. We tested this hypothesis by analyzing the targeting of phosphatidylinositol-4-kinase type II α (PI4KIIα), a membrane protein present in presynaptic and postsynaptic compartments. PI4KIIα copurified with BLOC-1 and AP-3 in neuronal cells. These interactions translated into a decreased PI4KIIα content in the dentate gyrus of dysbindin-null BLOC-1 deficiency and AP-3–null mice. Reduction of PI4KIIα in the dentate reflects a failure to traffic from the cell body. PI4KIIα was targeted to processes in wild-type primary cultured cortical neurons and PC12 cells but failed to reach neurites in cells lacking either AP-3 or BLOC-1. Similarly, disruption of an AP-3–sorting motif in PI4KIIα impaired its sorting into processes of PC12 and primary cultured cortical neuronal cells. Our findings indicate a novel vesicle transport mechanism requiring BLOC-1 and AP-3 complexes for cargo sorting from neuronal cell bodies to neurites and nerve terminals.

scholarly journals Loss of function of the mitochondrial peptidase PITRM1 induces proteotoxic stress and Alzheimer’s disease-like pathology in human cerebral organoids

2020 ◽  
Author(s):  
Dina Ivanyuk ◽  
María José Pérez ◽  
Vasiliki Panagiotakopoulou ◽  
Gabriele Di Napoli ◽  
Dario Brunetti ◽  
...  
Keyword(s):  
Alzheimer’S Disease ◽  
Alzheimer's Disease ◽  
Cell Death ◽  
Cortical Neurons ◽  
Amyloid Β ◽  
Neuronal Cell ◽  
Protein Aggregates ◽  
Neuronal Cultures ◽  
Loss Of Function ◽  
Proteotoxic Stress

AbstractMutations in pitrilysin metallopeptidase 1 (PITRM1), a mitochondrial protease involved in mitochondrial precursor processing and degradation, result in a slow-progressive syndrome, characterized by cerebellar ataxia, psychotic episodes and obsessive behavior as well as cognitive decline. To investigate the pathogenetic mechanisms of mitochondrial presequence processing, we employed cortical neurons and cerebral organoids generated from PITRM1 knockout human induced pluripotent stem cells (iPSCs). PITRM1 deficiency strongly induced mitochondrial unfolded protein response (UPRmt) and enhanced mitochondrial clearance in iPSC-derived neurons. Furthermore, we observed increased levels of amyloid precursor protein and amyloid β in PITRM1 knockout neurons. However, neither cell death nor protein aggregates were observed in 2D iPSC-derived cortical neuronal cultures. On the contrary, cerebral organoids generated from PITRM1 knockout iPSCs spontaneously developed over time pathological features of Alzheimer’s disease (AD), including accumulation of protein aggregates, tau pathology, and neuronal cell death. Importantly, we provide evidence for a protective role of UPRmt and mitochondrial clearance against impaired mitochondrial presequence processing and proteotoxic stress. In summary, we propose a novel concept of PITRM1-linked neurological syndrome whereby defects of mitochondrial presequence processing induce an early activation of UPRmt that, in turn, modulates cytosolic quality control pathways. Thus our work supports a mechanistic link between mitochondrial function and common neurodegenerative proteinopathies.

scholarly journals Ischemic Cerebral Endothelial Cell–Derived Exosomes Promote Axonal Growth

Stroke ◽  
2020 ◽  
Vol 51 (12) ◽  
pp. 3701-3712
Author(s):  
Yi Zhang ◽  
Yi Qin ◽  
Michael Chopp ◽  
Chao Li ◽  
Amy Kemper ◽  
...  
Keyword(s):  
Cortical Neurons ◽  
Neuronal Cell ◽  
Axonal Growth ◽  
Mature Mirnas ◽  
Bioinformatic Analysis ◽  
Axonal Outgrowth ◽  
Ischemic Brain ◽  
Neuronal Homeostasis ◽  
Neuronal Cell Bodies

Background and Purpose: Cerebral endothelial cells (CECs) and axons of neurons interact to maintain vascular and neuronal homeostasis and axonal remodeling in normal and ischemic brain, respectively. However, the role of exosomes in the interaction of CECs and axons in brain under normal conditions and after stroke is unknown. Methods: Exosomes were isolated from CECs of nonischemic rats and is chemic rats (nCEC-exos and isCEC-exos), respectively. A multicompartmental cell culture system was used to separate axons from neuronal cell bodies. Results: Axonal application of nCEC-exos promotes axonal growth of cortical neurons, whereas isCEC-exos further enhance axonal growth than nCEC-exos. Ultrastructural analysis revealed that CEC-exos applied into distal axons were internalized by axons and reached to their parent somata. Bioinformatic analysis revealed that both nCEC-exos and isCEC-exos contain abundant mature miRNAs; however, isCEC-exos exhibit more robust elevation of select miRNAs than nCEC-exos. Mechanistically, axonal application of nCEC-exos and isCEC-exos significantly elevated miRNAs and reduced proteins in distal axons and their parent somata that are involved in inhibiting axonal outgrowth. Blockage of axonal transport suppressed isCEC-exo–altered miRNAs and proteins in somata but not in distal axons. Conclusions: nCEC-exos and isCEC-exos facilitate axonal growth by altering miRNAs and their target protein profiles in recipient neurons.

scholarly journals Transforming Growth Factor β2 Is a Neuronal Death-Inducing Ligand for Amyloid-β Precursor Protein

2005 ◽  
Vol 25 (21) ◽  
pp. 9304-9317 ◽  
Author(s):  
Yuichi Hashimoto ◽  
Tomohiro Chiba ◽  
Marina Yamada ◽  
Mikiro Nawa ◽  
Kohsuke Kanekura ◽  
...  
Keyword(s):  
Growth Factor ◽  
Binding Affinity ◽  
Neuronal Death ◽  
Cortical Neurons ◽  
Transforming Growth Factor ◽  
Neuronal Cell Death ◽  
Amyloid Β ◽  
Neuronal Cell ◽  
Precursor Protein ◽  
Transforming Growth Factor Β2

ABSTRACT APP, amyloid β precursor protein, is linked to the onset of Alzheimer's disease (AD). We have here found that transforming growth factor β2 (TGFβ2), but not TGFβ1, binds to APP. The binding affinity of TGFβ2 to APP is lower than the binding affinity of TGFβ2 to the TGFβ receptor. On binding to APP, TGFβ2 activates an APP-mediated death pathway via heterotrimeric G protein Go, c-Jun N-terminal kinase, NADPH oxidase, and caspase 3 and/or related caspases. Overall degrees of TGFβ2-induced death are larger in cells expressing a familial AD-related mutant APP than in those expressing wild-type APP. Consequently, superphysiological concentrations of TGFβ2 induce neuronal death in primary cortical neurons, whose one allele of the APP gene is knocked in with the V642I mutation. Combined with the finding indicated by several earlier reports that both neural and glial expression of TGFβ2 was upregulated in AD brains, it is speculated that TGFβ2 may contribute to the development of AD-related neuronal cell death.

scholarly journals Spatial constraints dictate glial territories at murine neuromuscular junctions

2011 ◽  
Vol 195 (2) ◽  
pp. 293-305 ◽  
Author(s):  
Monika S. Brill ◽  
Jeff W. Lichtman ◽  
Wesley Thompson ◽  
Yi Zuo ◽  
Thomas Misgeld
Keyword(s):  
Nervous System ◽  
Glial Cells ◽  
Neuromuscular Junctions ◽  
Spatial Competition ◽  
Time Lapse ◽  
Axon Degeneration ◽  
Spatial Constraints ◽  
Cellular Mechanisms ◽  
Synapse Loss ◽  
Time Lapse Imaging

Schwann cells (SCs), the glial cells of the peripheral nervous system, cover synaptic terminals, allowing them to monitor and modulate neurotransmission. Disruption of glial coverage leads to axon degeneration and synapse loss. The cellular mechanisms that establish and maintain this coverage remain largely unknown. To address this, we labeled single SCs and performed time-lapse imaging experiments. Adult terminal SCs are arranged in static tile patterns, whereas young SCs dynamically intermingle. The mechanism of developmental glial segregation appears to be spatial competition, in which glial–glial and axonal–glial contacts constrain the territory of single SCs, as shown by four types of experiments: (1) laser ablation of single SCs, which led to immediate territory expansion of neighboring SCs; (2) axon removal by transection, resulting in adult SCs intermingling dynamically; (3) axotomy in mutant mice with blocked axon fragmentation in which intermingling was delayed; and (4) activity blockade, which had no immediate effects. In summary, we conclude that glial cells partition synapses by competing for perisynaptic space.

scholarly journals Regulation of Neuronal Cell Cycle and Apoptosis by miR-34a

2015 ◽  
pp. MCB.00589-15 ◽  
Author(s):  
Prashant Kumar Modi ◽  
Surbhi Jaiswal ◽  
Pushkar Sharma
Keyword(s):  
Cell Cycle ◽  
Mouse Model ◽  
Neurodegenerative Disorders ◽  
Cortical Neurons ◽  
Neuronal Apoptosis ◽  
Amyloid Β ◽  
Neuronal Cell ◽  
Erk Pathway ◽  
Cycle Arrest ◽  
Cell Cycle Machinery

The cell cycle of neurons remains suppressed to maintain the state of differentiation and aberrant cell cycle re-entry results in loss of neurons, which is a feature in neurodegenerative disorders like Alzheimer's disease (AD). Present studies revealed that the expression of microRNA-34a (miR-34a) needs to be optimal in neurons as aberrant increase or decrease in its expression causes apoptosis. miR-34a keeps the neuronal cell cycle under check by preventing the expression of cyclin D1 and promotes cell cycle arrest. Neurotoxic Amyloid β1-42peptide (Aβ42) treatment of cortical neurons suppressed miR-34a resulting in unscheduled cell cycle re-entry, which resulted in apoptosis. The repression of miR-34a was a result of degradation of TAp73, which was mediated by aberrant activation of MEK-ERK pathway by Aβ42. A significant decrease in miR-34a and TAp73 was observed in the cortex of >12m old transgenic (Tg) mouse model of AD, which corroborated well with cell cycle re-entry observed in the neurons of these animals. Importantly, the overexpression of TAp73α and miR-34a reversed the Cell cycle Related Neuronal Apoptosis (CRNA). These studies provide novel insights into how modulation of neuronal cell cycle machinery may lead to neurodegeneration and may contribute to the understanding of disorders like AD.

scholarly journals E2-25K/Hip-2 regulates caspase-12 in ER stress–mediated Aβ neurotoxicity

2008 ◽  
Vol 182 (4) ◽  
pp. 675-684 ◽  
Author(s):  
Sungmin Song ◽  
Huikyong Lee ◽  
Tae-In Kam ◽  
Mei Ling Tai ◽  
Joo-Yong Lee ◽  
...  
Keyword(s):  
Cell Death ◽  
Er Stress ◽  
Cortical Neurons ◽  
Neuronal Cell Death ◽  
Amyloid Β ◽  
Neuronal Cell ◽  
Ubiquitin Proteasome System ◽  
Proteolytic Activation ◽  
Upstream Regulator ◽  
Caspase 12

Amyloid-β (Aβ) neurotoxicity is believed to contribute to the pathogenesis of Alzheimer's disease (AD). Previously we found that E2-25K/Hip-2, an E2 ubiquitin-conjugating enzyme, mediates Aβ neurotoxicity. Here, we report that E2-25K/Hip-2 modulates caspase-12 activity via the ubiquitin/proteasome system. Levels of endoplasmic reticulum (ER)–resident caspase-12 are strongly up-regulated in the brains of AD model mice, where the enzyme colocalizes with E2-25K/Hip-2. Aβ increases expression of E2-25K/Hip-2, which then stabilizes caspase-12 protein by inhibiting proteasome activity. This increase in E2-25K/Hip-2 also induces proteolytic activation of caspase-12 through its ability to induce calpainlike activity. Knockdown of E2-25K/Hip-2 expression suppresses neuronal cell death triggered by ER stress, and thus caspase-12 is required for the E2-25K/Hip-2–mediated cell death. Finally, we find that E2-25K/Hip-2–deficient cortical neurons are resistant to Aβ toxicity and to the induction of ER stress and caspase-12 expression by Aβ. E2-25K/Hip-2 is thus an essential upstream regulator of the expression and activation of caspase-12 in ER stress–mediated Aβ neurotoxicity.

scholarly journals Adaptive prospective optical gating enables day-long 3D time-lapse imaging of the beating embryonic zebrafish heart

2019 ◽  
Vol 10 (1) ◽  
Author(s):  
Jonathan M. Taylor ◽  
Carl J. Nelson ◽  
Finnius A. Bruton ◽  
Aryan Kaveh ◽  
Charlotte Buckley ◽  
...  
Keyword(s):  
Time Course ◽  
Cardiac Development ◽  
Three Dimensional ◽  
Time Lapse ◽  
Beating Heart ◽  
Cellular Mechanisms ◽  
Phase Lock ◽  
Optical Gating ◽  
Time Lapse Imaging ◽  
Zebrafish Heart

AbstractThree-dimensional fluorescence time-lapse imaging of the beating heart is extremely challenging, due to the heart’s constant motion and a need to avoid pharmacological or phototoxic damage. Although real-time triggered imaging can computationally “freeze” the heart for 3D imaging, no previous algorithm has been able to maintain phase-lock across developmental timescales. We report a new algorithm capable of maintaining day-long phase-lock, permitting routine acquisition of synchronised 3D + time video time-lapse datasets of the beating zebrafish heart. This approach has enabled us for the first time to directly observe detailed developmental and cellular processes in the beating heart, revealing the dynamics of the immune response to injury and witnessing intriguing proliferative events that challenge the established literature on cardiac trabeculation. Our approach opens up exciting new opportunities for direct time-lapse imaging studies over a 24-hour time course, to understand the cellular mechanisms underlying cardiac development, repair and regeneration.

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