9 research outputs found
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Suppression of tau propagation using an inhibitor that targets the DK-switch of nSMase2.
Targeting of molecular pathways involved in the cell-to-cell propagation of pathological tau species is a novel approach for development of disease-modifying therapies that could block tau pathology and attenuate cognitive decline in patients with Alzheimer's disease and other tauopathies. We discovered cambinol through a screening effort and show that it is an inhibitor of cell-to-cell tau propagation. Our in vitro data demonstrate that cambinol inhibits neutral sphingomyelinase 2 (nSMase2) enzyme activity in dose response fashion, and suppresses extracellular vesicle (EV) production while reducing tau seed propagation. Our in vivo testing with cambinol shows that it can reduce the nSMase2 activity in the brain after oral administration. Our molecular docking and simulation analysis reveals that cambinol can target the DK-switch in the nSMase2 active site
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Pharmacological inhibition of nSMase2 reduces brain exosome release and α-synuclein pathology in a Parkinson’s disease model
AimWe have previously reported that cambinol (DDL-112), a known inhibitor of neutral sphingomyelinase-2 (nSMase2), suppressed extracellular vesicle (EV)/exosome production in vitro in a cell model and reduced tau seed propagation. The enzyme nSMase2 is involved in the production of exosomes carrying proteopathic seeds and could contribute to cell-to-cell transmission of pathological protein aggregates implicated in neurodegenerative diseases such as Parkinson's disease (PD). Here, we performed in vivo studies to determine if DDL-112 can reduce brain EV/exosome production and proteopathic alpha synuclein (αSyn) spread in a PD mouse model.MethodsThe acute effects of single-dose treatment with DDL-112 on interleukin-1β-induced extracellular vesicle (EV) release in brain tissue of Thy1-αSyn PD model mice and chronic effects of 5 week DDL-112 treatment on behavioral/motor function and proteinase K-resistant αSyn aggregates in the PD model were determined.Results/discussionIn the acute study, pre-treatment with DDL-112 reduced EV/exosome biogenesis and in the chronic study, treatment with DDL-112 was associated with a reduction in αSyn aggregates in the substantia nigra and improvement in motor function. Inhibition of nSMase2 thus offers a new approach to therapeutic development for neurodegenerative diseases with the potential to reduce the spread of disease-specific proteopathic proteins
AD synapses contain abundant Aβ monomer and multiple soluble oligomers, including a 56-kDa assembly
Co-Localization of Amyloid Beta and Tau Pathology in Alzheimer’s Disease Synaptosomes
The amyloid cascade hypothesis proposes that amyloid β (Aβ) pathology precedes and induces tau pathology, but the neuropathological connection between these two lesions has not been demonstrated. We examined the regional distribution and co-localization of Aβ and phosphorylated tau (p-tau) in synaptic terminals of Alzheimer’s disease brains. To quantitatively examine large populations of individual synaptic terminals, flow cytometry was used to analyze synaptosomes prepared from cryopreserved Alzheimer’s disease tissue. An average 68.4% of synaptic terminals in the Alzheimer’s disease cohort (n = 11) were positive for Aβ, and 32.3% were positive for p-tau; Aβ and p-tau fluorescence was lowest in cerebellum. In contrast to synaptic p-tau, which was highest in the entorhinal cortex and hippocampus (P = 0.004), synaptic Aβ fluorescence was significantly lower in the entorhinal cortex and hippocampus relative to neocortical regions (P = 0.0003). Synaptic Aβ and p-tau fluorescence was significantly correlated (r = 0.683, P < 0.004), and dual-labeling experiments demonstrated that 24.1% of Aβ-positive terminals were also positive for p-tau, with the highest fraction of dual labeling (39.3%) in the earliest affected region, the entorhinal cortex. Western blotting experiments show a significant correlation between synaptic Aβ levels measured by flow cytometry and oligomeric Aβ species (P < 0.0001). These results showing overlapping Aβ and tau pathology are consistent with a model in which both synaptic loss and dysfunction are linked to a synaptic amyloid cascade within the synaptic compartment
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Exosomal tau with seeding activity is released from Alzheimer's disease synapses, and seeding potential is associated with amyloid beta.
Synaptic transfer of tau has long been hypothesized from the human pathology pattern and has been demonstrated in vitro and in vivo, but the precise mechanisms remain unclear. Extracellular vesicles such as exosomes have been suggested as a mechanism, but not all tau is exosomal. The present experiments use a novel flow cytometry assay to quantify depolarization of synaptosomes by KCl after loading with FM2-10, which induces a fluorescence reduction associated with synaptic vesicle release; the degree of reduction in cryopreserved human samples equaled that seen in fresh mouse synaptosomes. Depolarization induced the release of vesicles in the size range of exosomes, along with tetraspanin markers of extracellular vesicles. A number of tau peptides were released, including tau oligomers; released tau was primarily unphosphorylated and C-terminal truncated, with Aβ release just above background. When exosomes were immunopurified from release supernatants, a prominent tau band showed a dark smeared appearance of SDS-stable oligomers along with the exosomal marker syntenin-1, and these exosomes induced aggregation in the HEK tau biosensor assay. However, the flow-through did not seed aggregation. Size exclusion chromatography of purified released exosomes shows faint signals from tau in the same fractions that show a CD63 band, an exosomal size signal, and seeding activity. Crude synaptosomes from control, tauopathy, and AD cases demonstrated lower seeding in tauopathy compared to AD that is correlated with the measured Aβ42 level. These results show that AD synapses release exosomal tau that is C-terminal-truncated, oligomeric, and with seeding activity that is enhanced by Aβ. Taken together with previous findings, these results are consistent with a direct prion-like heterotypic seeding of tau by Aβ within synaptic terminals, with subsequent loading of aggregated tau onto exosomes that are released and competent for tau seeding activity