scholarly journals α-Synuclein polymorphism determines oligodendroglial dysfunction

2021 ◽  
Author(s):  
Benedikt Frieg ◽  
James A Geraets ◽  
Timo Strohaeker ◽  
Christian Dienemann ◽  
Panagiota Mavroeidi ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Brain Tissue ◽  
Ex Vivo ◽  
Neurological Diseases ◽  
Alpha Synuclein ◽  
Published Data ◽  
Cryo Electron Microscopy ◽  
Fibril Structure ◽  
Potential Biomarker

Synucleinopathies, such as Parkinson's disease (PD) and Multiple System Atrophy (MSA) are progressive and unremitting neurological diseases. For both PD and MSA, alpha-synuclein fibril inclusions inside brain cells are neuropathological hallmarks. In addition, amplification of alpha-synuclein fibrils from body fluids is a potential biomarker distinguishing PD from MSA. However, little is known about the structure of alpha-synuclein fibrils amplified from human samples and its connection to alpha-synuclein fibril structure in the human brain. Here we amplified alpha-synuclein fibrils from PD and MSA brain tissue, characterized its seeding potential in oligodendroglia, and determined the 3D structures by cryo-electron microscopy. We show that the alpha-synuclein fibrils from a MSA patient are more potent in recruiting the endogenous alpha-synuclein and evoking a redistribution of TPPP/p25alpha protein in mouse primary oligodendroglial cultures compared to those amplified from a PD patient. Cryo-electron microscopy shows that the PD- and MSA-amplified alpha-synuclein fibrils share a similar protofilament fold but differ in their inter-protofilament interface. The structures of the brain-tissue amplified alpha-synuclein fibrils are also similar to other in vitro and ex vivo alpha-synuclein fibrils. Together with published data, our results suggest that aSyn fibrils differ between PD and MSA in their quaternary arrangement and could further vary between different forms of PD and MSA.

scholarly journals Cryo-EM structure and polymorphism of Aβ amyloid fibrils purified from Alzheimer’s brain tissue

2019 ◽  
Vol 10 (1) ◽  
Author(s):  
Marius Kollmer ◽  
William Close ◽  
Leonie Funk ◽  
Jay Rasmussen ◽  
Aref Bsoul ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Brain Tissue ◽  
Amyloid Fibrils ◽  
Structural Basis ◽  
Amyloid Angiopathy ◽  
Cryo Electron Microscopy ◽  
Aβ Amyloid ◽  
Cerebral Amyloid ◽  
Aβ Fibrils

Abstract The formation of Aβ amyloid fibrils is a neuropathological hallmark of Alzheimer’s disease and cerebral amyloid angiopathy. However, the structure of Aβ amyloid fibrils from brain tissue is poorly understood. Here we report the purification of Aβ amyloid fibrils from meningeal Alzheimer’s brain tissue and their structural analysis with cryo-electron microscopy. We show that these fibrils are polymorphic but consist of similarly structured protofilaments. Brain derived Aβ amyloid fibrils are right-hand twisted and their peptide fold differs sharply from previously analyzed Aβ fibrils that were formed in vitro. These data underscore the importance to use patient-derived amyloid fibrils when investigating the structural basis of the disease.

Monocytes as Carriers of Magnetic Nanoparticles for Tracking Inflammation in the Epileptic Rat Brain

2019 ◽  
Vol 16 (7) ◽  
pp. 637-644 ◽  
Author(s):  
Hadas Han ◽  
Sara Eyal ◽  
Emma Portnoy ◽  
Aniv Mann ◽  
Miriam Shmuel ◽  
...  
Keyword(s):  
Brain Tissue ◽  
Ex Vivo ◽  
In Vivo Studies ◽  
Nanoparticle Distribution ◽  
Spontaneous Seizures ◽  
Systemic Distribution ◽  
Hippocampal Ca1 ◽  
Pilocarpine Model

Background: Inflammation is a hallmark of epileptogenic brain tissue. Previously, we have shown that inflammation in epilepsy can be delineated using systemically-injected fluorescent and magnetite- laden nanoparticles. Suggested mechanisms included distribution of free nanoparticles across a compromised blood-brain barrier or their transfer by monocytes that infiltrate the epileptic brain. Objective: In the current study, we evaluated monocytes as vehicles that deliver nanoparticles into the epileptic brain. We also assessed the effect of epilepsy on the systemic distribution of nanoparticleloaded monocytes. Methods: The in vitro uptake of 300-nm nanoparticles labeled with magnetite and BODIPY (for optical imaging) was evaluated using rat monocytes and fluorescence detection. For in vivo studies we used the rat lithium-pilocarpine model of temporal lobe epilepsy. In vivo nanoparticle distribution was evaluated using immunohistochemistry. Results: 89% of nanoparticle loading into rat monocytes was accomplished within 8 hours, enabling overnight nanoparticle loading ex vivo. The dose-normalized distribution of nanoparticle-loaded monocytes into the hippocampal CA1 and dentate gyrus of rats with spontaneous seizures was 176-fold and 380-fold higher compared to the free nanoparticles (p<0.05). Seizures were associated with greater nanoparticle accumulation within the liver and the spleen (p<0.05). Conclusion: Nanoparticle-loaded monocytes are attracted to epileptogenic brain tissue and may be used for labeling or targeting it, while significantly reducing the systemic dose of potentially toxic compounds. The effect of seizures on monocyte biodistribution should be further explored to better understand the systemic effects of epilepsy.

scholarly journals Alterations in Sub-Axonal Architecture Between Normal Aging and Parkinson’s Diseased Human Brains Using Label-Free Cryogenic X-ray Nanotomography

2020 ◽  
Vol 14 ◽  
Author(s):  
Hung Tri Tran ◽  
Esther H. R. Tsai ◽  
Amanda J. Lewis ◽  
Tim Moors ◽  
J. G. J. M. Bol ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Human Brain ◽  
Brain Tissue ◽  
Multispectral Imaging ◽  
Alpha Synuclein ◽  
Label Free ◽  
Myelinated Axons ◽  
X Ray ◽  
X Ray Imaging ◽  
Non Destructive

Gaining insight to pathologically relevant processes in continuous volumes of unstained brain tissue is important for a better understanding of neurological diseases. Many pathological processes in neurodegenerative disorders affect myelinated axons, which are a critical part of the neuronal circuitry. Cryo ptychographic X-ray computed tomography in the multi-keV energy range is an emerging technology providing phase contrast at high sensitivity, allowing label-free and non-destructive three dimensional imaging of large continuous volumes of tissue, currently spanning up to 400,000 μm3. This aspect makes the technique especially attractive for imaging complex biological material, especially neuronal tissues, in combination with downstream optical or electron microscopy techniques. A further advantage is that dehydration, additional contrast staining, and destructive sectioning/milling are not required for imaging. We have developed a pipeline for cryo ptychographic X-ray tomography of relatively large, hydrated and unstained biological tissue volumes beyond what is typical for the X-ray imaging, using human brain tissue and combining the technique with complementary methods. We present four imaged volumes of a Parkinson’s diseased human brain and five volumes from a non-diseased control human brain using cryo ptychographic X-ray tomography. In both cases, we distinguish neuromelanin-containing neurons, lipid and melanic pigment, blood vessels and red blood cells, and nuclei of other brain cells. In the diseased sample, we observed several swellings containing dense granular material resembling clustered vesicles between the myelin sheaths arising from the cytoplasm of the parent oligodendrocyte, rather than the axoplasm. We further investigated the pathological relevance of such swollen axons in adjacent tissue sections by immunofluorescence microscopy for phosphorylated alpha-synuclein combined with multispectral imaging. Since cryo ptychographic X-ray tomography is non-destructive, the large dataset volumes were used to guide further investigation of such swollen axons by correlative electron microscopy and immunogold labeling post X-ray imaging, a possibility demonstrated for the first time. Interestingly, we find that protein antigenicity and ultrastructure of the tissue are preserved after the X-ray measurement. As many pathological processes in neurodegeneration affect myelinated axons, our work sets an unprecedented foundation for studies addressing axonal integrity and disease-related changes in unstained brain tissues.

scholarly journals Cryo-EM demonstrates the in vitro proliferation of an ex vivo amyloid fibril morphology by seeding

2022 ◽  
Vol 13 (1) ◽  
Author(s):  
Thomas Heerde ◽  
Matthies Rennegarbe ◽  
Alexander Biedermann ◽  
Dilan Savran ◽  
Peter B. Pfeiffer ◽  
...  
Keyword(s):  
Amyloid Fibrils ◽  
Ex Vivo ◽  
Proteinase K ◽  
Seed Structure ◽  
In Vitro Proliferation ◽  
Amyloid A ◽  
Fibril Structure ◽  
Proteolytic Stability ◽  
Fibril Morphology

AbstractSeveral studies showed that seeding of solutions of monomeric fibril proteins with ex vivo amyloid fibrils accelerated the kinetics of fibril formation in vitro but did not necessarily replicate the seed structure. In this research we use cryo-electron microscopy and other methods to analyze the ability of serum amyloid A (SAA)1.1-derived amyloid fibrils, purified from systemic AA amyloidosis tissue, to seed solutions of recombinant SAA1.1 protein. We show that 98% of the seeded fibrils remodel the full fibril structure of the main ex vivo fibril morphology, which we used for seeding, while they are notably different from unseeded in vitro fibrils. The seeded fibrils show a similar proteinase K resistance as ex vivo fibrils and are substantially more stable to proteolytic digestion than unseeded in vitro fibrils. Our data support the view that the fibril morphology contributes to determining proteolytic stability and that pathogenic amyloid fibrils arise from proteolytic selection.

scholarly journals Cryo-EM structure of alpha-synuclein fibrils

eLife ◽  
2018 ◽  
Vol 7 ◽  
Author(s):  
Ricardo Guerrero-Ferreira ◽  
Nicholas MI Taylor ◽  
Daniel Mona ◽  
Philippe Ringler ◽  
Matthias E Lauer ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Parkinson’S Disease ◽  
Parkinson's Disease ◽  
Lewy Bodies ◽  
Alpha Synuclein ◽  
Diagnosis And Treatment ◽  
Lewy Neurites ◽  
Cryo Electron Microscopy ◽  
High Concentrations ◽  
Hydrophobic Cleft

Parkinson’s disease is a progressive neuropathological disorder that belongs to the class of synucleinopathies, in which the protein alpha-synuclein is found at abnormally high concentrations in affected neurons. Its hallmark are intracellular inclusions called Lewy bodies and Lewy neurites. We here report the structure of cytotoxic alpha-synuclein fibrils (residues 1–121), determined by cryo-electron microscopy at a resolution of 3.4 Å. Two protofilaments form a polar fibril composed of staggered β-strands. The backbone of residues 38 to 95, including the fibril core and the non-amyloid component region, are well resolved in the EM map. Residues 50–57, containing three of the mutation sites associated with familial synucleinopathies, form the interface between the two protofilaments and contribute to fibril stability. A hydrophobic cleft at one end of the fibril may have implications for fibril elongation, and invites for the design of molecules for diagnosis and treatment of synucleinopathies.

scholarly journals Iroquois Homeobox Protein 2 Identified as a Potential Biomarker for Parkinson’s Disease

2020 ◽  
Vol 21 (10) ◽  
pp. 3455
Author(s):  
Hyuna Sim ◽  
Joo-Eun Lee ◽  
Hee Min Yoo ◽  
Sunwha Cho ◽  
Hana Lee ◽  
...  
Keyword(s):  
Parkinson’S Disease ◽  
Parkinson's Disease ◽  
Ex Vivo ◽  
Expression Patterns ◽  
Cell Types ◽  
Motor Symptoms ◽  
Intestinal Organoids ◽  
Potential Biomarker ◽  
Homeobox Protein

The diagnosis of Parkinson’s disease (PD) is initiated after the occurrence of motor symptoms, such as resting tremors, rigidity, and bradykinesia. According to previous reports, non-motor symptoms, notably gastrointestinal dysfunction, could potentially be early biomarkers in PD patients as such symptoms occur earlier than motor symptoms. However, connecting PD to the intestine is methodologically challenging. Thus, we generated in vitro human intestinal organoids from PD patients and ex vivo mouse small intestinal organoids from aged transgenic mice. Both intestinal organoids (IOs) contained the human LRRK2 G2019S mutation, which is the most frequent genetic cause of familial and sporadic PD. By conducting comprehensive genomic comparisons with these two types of IOs, we determined that a particular gene, namely, Iroquois homeobox protein 2 (IRX2), showed PD-related expression patterns not only in human pluripotent stem cell (PSC)-derived neuroectodermal spheres but also in human PSC-derived neuronal cells containing dopaminergic neurons. We expected that our approach of using various cell types presented a novel technical method for studying the effects of multi-organs in PD pathophysiology as well as for the development of diagnostic markers for PD.

scholarly journals Antioxidant and Neuroprotective Effects Induced by Cannabidiol and Cannabigerol in Rat CTX-TNA2 Astrocytes and Isolated Cortexes

2020 ◽  
Vol 21 (10) ◽  
pp. 3575 ◽  
Author(s):  
Viviana di Giacomo ◽  
Annalisa Chiavaroli ◽  
Lucia Recinella ◽  
Giustino Orlando ◽  
Amelia Cataldi ◽  
...  
Keyword(s):  
Kynurenic Acid ◽  
Cannabis Sativa ◽  
Ex Vivo ◽  
Neurological Diseases ◽  
Docking Studies ◽  
Experimental Models ◽  
Neuroprotective Effects ◽  
Rat Astrocytes ◽  
Species Production

Cannabidiol (CBD) and cannabigerol (CBG) are Cannabis sativa terpenophenols. Although CBD’s effectiveness against neurological diseases has already been demonstrated, nothing is known about CBG. Therefore, a comparison of the effects of these compounds was performed in two experimental models mimicking the oxidative stress and neurotoxicity occurring in neurological diseases. Rat astrocytes were exposed to hydrogen peroxide and cell viability, reactive oxygen species production and apoptosis occurrence were investigated. Cortexes were exposed to K+ 60 mM depolarizing stimulus and serotonin (5-HT) turnover, 3-hydroxykinurenine and kynurenic acid levels were measured. A proteomic analysis and bioinformatics and docking studies were performed. Both compounds exerted antioxidant effects in astrocytes and restored the cortex level of 5-HT depleted by neurotoxic stimuli, whereas sole CBD restored the basal levels of 3-hydroxykinurenine and kynurenic acid. CBG was less effective than CBD in restoring the levels of proteins involved in neurotransmitter exocytosis. Docking analyses predicted the inhibitory effects of these compounds towards the neurokinin B receptor. Conclusion: The results in the in vitro system suggest brain non-neuronal cells as a target in the treatment of oxidative conditions, whereas findings in the ex vivo system and docking analyses imply the potential roles of CBD and CBG as neuroprotective agents.

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