Neuronal Exosome-Derived Human Tau is Toxic to Recipient Mouse Neurons in vivo.
ABSTRACT: Progressive accumulation of aggregation-prone proteins, amyloid-β (Aβ) and hyperphosphorylated tau (p-tau), are the defining hallmarks of Alzheimer's disease (AD). The mechanisms by which Aβ and p-tau are transmitted throughout the diseased brain are not yet completely understood. Interest in exosome research has grown dramatically over the past few years, specifically due to their potential role as biomarkers for staging of neurodegenerative diseases, including AD. Despite their diagnostic utility, the pathogenic potential of exosomes has yet to be fully elucidated. In this study, we use a series of recombinant tau antibodies to characterize a new model of human tau in vivo. Exosome suspensions derived from neuronally-differentiated, human induced pluripotent stem cells that express the repeat domain of tau P301L and V337M mutations (NiPSCEs) were injected into the wild-type mouse brain and pathological changes were characterized by immunostaining at one- (1 m) and two-month (2 m) post-injection. We found that tau inclusions were present throughout the brain at 2 m post-injection, which were detectable using antibodies raised against full-length tau (K9JA) and misfolded tau (MC1). Furthermore, we found that phosphorylated tau immunoreactivity was elevated 1 m post-injection, which was surprisingly normalized after 2 m. Finally, we observed extensive degeneration of neuronal dendrites in both ipsilateral and contralateral hippocampi in NiPSCE treated mice. In summary, we demonstrate that exosomes are sufficient to cause long-distance propagation of tau pathology and neurodegeneration in vivo. These novel findings support an active role of exosomes in AD pathogenesis.
Project description:Accumulation and propagation of hyperphosphorylated Tau (p-Tau) is a common neuropathological hallmark associated with neurodegeneration of Alzheimer's disease (AD), frontotemporal dementia and parkinsonism linked to chromosome 17 (FTDP-17), and related tauopathies. Extracellular vesicles, specifically exosomes, have recently been demonstrated to participate in mediating Tau propagation in brain. Exosomes produced by human induced pluripotent stem cell (iPSC)-derived neurons expressing mutant Tau (mTau), containing the P301L and V337M Tau mutations of FTDP-17, possess the ability to propagate p-Tau pathology after injection into mouse brain. To gain an understanding of the mTau exosome cargo involved in Tau pathogenesis, these pathogenic exosomes were analyzed by proteomics and bioinformatics. The data showed that mTau expression dysregulates the exosome proteome to result in 1) proteins uniquely present only in mTau, and not control exosomes, 2) the absence of proteins in mTau exosomes, uniquely present in control exosomes, and 3) shared proteins which were significantly upregulated or downregulated in mTau compared with control exosomes. Notably, mTau exosomes (not control exosomes) contain ANP32A (also known as I1PP2A), an endogenous inhibitor of the PP2A phosphatase which regulates the phosphorylation state of p-Tau. Several of the mTau exosome-specific proteins have been shown to participate in AD mechanisms involving lysosomes, inflammation, secretases, and related processes. Furthermore, the mTau exosomes lacked a substantial portion of proteins present in control exosomes involved in pathways of localization, vesicle transport, and protein binding functions. The shared proteins present in both mTau and control exosomes represented exosome functions of vesicle-mediated transport, exocytosis, and secretion processes. These data illustrate mTau as a dynamic regulator of the biogenesis of exosomes to result in acquisition, deletion, and up- or downregulation of protein cargo to result in pathogenic mTau exosomes capable of <i>in vivo</i> propagation of p-Tau neuropathology in mouse brain.
Project description:The accumulation and propagation of hyperphosphorylated tau (p-Tau) is a neuropathological hallmark occurring with neurodegeneration of Alzheimer's disease (AD). Extracellular vesicles, exosomes, have been shown to initiate tau propagation in the brain. Notably, exosomes from human-induced pluripotent stem cell (iPSC) neurons expressing the AD familial A246E mutant form of presenilin 1 (mPS1) are capable of inducing tau deposits in the mouse brain after <i>in vivo</i> injection. To gain insights into the exosome proteome cargo that participates in propagating tau pathology, this study conducted proteomic analysis of exosomes produced by human iPSC neurons expressing A246E mPS1. Significantly, mPS1 altered the profile of exosome cargo proteins to result in (1) proteins present only in mPS1 exosomes and not in controls, (2) the absence of proteins in the mPS1 exosomes which were present only in controls, and (3) shared proteins which were upregulated or downregulated in the mPS1 exosomes compared to controls. These results show that mPS1 dysregulates the proteome cargo of exosomes to result in the acquisition of proteins involved in the extracellular matrix and protease functions, deletion of proteins involved in RNA and protein translation systems along with proteasome and related functions, combined with the upregulation and downregulation of shared proteins, including the upregulation of amyloid precursor protein. Notably, mPS1 neuron-derived exosomes displayed altered profiles of protein phosphatases and kinases involved in regulating the status of p-tau. The dysregulation of exosome cargo proteins by mPS1 may be associated with the ability of mPS1 neuron-derived exosomes to propagate tau pathology.
Project description:Amyloid β (Aβ) peptides generated from the amyloid precursor protein (APP) play a critical role in the development of Alzheimer's disease (AD) pathology. Aβ-containing neuronal exosomes, which represent a novel form of intercellular communication, have been shown to influence function/vulnerability of neurons in AD. Unlike neurons, the significance of exosomes derived from astrocytes remains unclear. In this study, we evaluated the significance of exosomes derived from U18666A-induced cholesterol-accumulated astrocytes in the development of AD pathology. Our results show that cholesterol accumulation decreases exosome secretion, whereas lowering cholesterol level increases exosome secretion from cultured astrocytes. Interestingly, exosomes secreted from U18666A-treated astrocytes contain higher levels of APP, APP-CTFs, soluble APP, APP secretases and Aβ1-40 than exosomes secreted from control astrocytes. Furthermore, we show that exosomes derived from U18666A-treated astrocytes can lead to neurodegeneration, which is attenuated by decreasing Aβ production or by neutralizing exosomal Aβ peptide with an Aβ antibody. These results, taken together, suggest that exosomes derived from cholesterol-accumulated astrocytes can play an important role in trafficking APP/Aβ peptides and influencing neuronal viability in the affected regions of the AD brain.
Project description:In Alzheimer's disease (AD), deposition of pathological tau and amyloid-β (Aβ) drive synaptic loss and cognitive decline. The injection of misfolded tau aggregates extracted from human AD brains drives templated spreading of tau pathology within WT mouse brain. Here, we assessed the impact of Aβ copathology, of deleting loci known to modify AD risk (Ptk2b, Grn, and Tmem106b) and of pharmacological intervention with an Fyn kinase inhibitor on tau spreading after injection of AD tau extracts. The density and spreading of tau inclusions triggered by human tau seed were unaltered in the hippocampus and cortex of APPswe/PSEN1ΔE9 transgenic and App<sup>NL-F/NL-F</sup> knock-in mice. In mice with human tau sequence replacing mouse tau, template matching enhanced neuritic tau burden. Human AD brain tau-enriched preparations contained aggregated Aβ, and the Aβ coinjection caused a redistribution of Aβ aggregates in mutant AD model mice. The injection-induced Aβ phenotype was spatially distinct from tau accumulation and could be ameliorated by depleting Aβ from tau extracts. These data suggest that Aβ and tau pathologies propagate by largely independent mechanisms after their initial formation. Altering the activity of the Fyn and Pyk2 (Ptk2b) kinases involved in Aβ-oligomer-induced signaling, or deleting expression of the progranulin and TMEM106B lysosomal proteins, did not alter the somatic tau inclusion burden or spreading. However, mouse aging had a prominent effect to increase the accumulation of neuritic tau after injection of human AD tau seeds into WT mice. These studies refine our knowledge of factors capable of modulating tau spreading.
Project description:Tau, the main component of the neurofibrillary tangles (NFTs), is an attractive target for immunotherapy in Alzheimer's disease (AD) and other tauopathies. MC1/Alz50 are currently the only antibodies targeting a disease-specific conformational modification of tau. Passive immunization experiments using intra-peritoneal injections have previously shown that MC1 is effective at reducing tau pathology in the forebrain of tau transgenic JNPL3 mice. In order to reach a long-term and sustained brain delivery, and avoid multiple injection protocols, we tested the efficacy of the single-chain variable fragment of MC1 (scFv-MC1) to reduce tau pathology in the same animal model, with focus on brain regional differences. ScFv-MC1 was cloned into an AAV delivery system and was directly injected into the hippocampus of adult JNPL3 mice. Specific promoters were employed to selectively target neurons or astrocytes for scFv-MC1 expression. ScFv-MC1 was able to decrease soluble, oligomeric and insoluble tau species, in our model. The effect was evident in the cortex, hippocampus and hindbrain. The astrocytic machinery appeared more efficient than the neuronal, with significant reduction of pathology in areas distant from the site of injection. To our knowledge, this is the first evidence that an anti-tau conformational scFv antibody, delivered directly into the mouse adult brain, is able to reduce pathological tau, providing further insight into the nature of immunotherapy strategies.
Project description:To assess the efficacy of [18F]AV1451 PET in visualizing tau pathology in vivo in a patient with frontotemporal dementia (FTD) associated with the V337M microtubule-associated protein tau (MAPT) mutation.MAPT mutations are associated with the deposition of hyperphosphorylated tau protein in neurons and glia. The PET tracer [18F]AV1451 binds with high affinity to paired helical filaments tau that comprises neurofibrillary tangles in Alzheimer disease (AD), while postmortem studies suggest lower or absent binding to the tau filaments of the majority of non-AD tauopathies. We describe clinical, structural MRI, and [18F]AV1451 PET findings in a V337M MAPT mutation carrier affected by FTD and pathologic findings in his affected mother and in an unrelated V337M MAPT carrier also affected with FTD. The biochemical similarity between paired helical filament tau in AD and MAPT V337M predicts that the tau pathology associated with this mutation constitutes a compelling target for [18F]AV1451 imaging.We found a strong association between topography and degree of [18F]AV1451 tracer retention in the proband and distribution of tau pathology in the brain of the proband's mother and the unrelated V337M mutation carrier. We also found a significant correlation between the degree of regional MRI brain atrophy and the extent of [18F]AV1451 binding in the proband and a strong association between the proband's clinical presentation and the extent of regional brain atrophy and tau accumulation as assessed by structural brain MRI and [18F]AV1451PET.Our study supports the usefulness of [18F]AV1451 to characterize tau pathology in at least a subset of pathogenic MAPT mutations.
Project description:Exosomes are bilipid layer-enclosed vesicles derived from endosomes and are released from neural cells. They contain a diversity of proteins, mRNAs, and microRNAs (miRNAs) that are delivered to neighboring cells and/or are transported to distant sites. miRNAs released from exosomes appear to be associated with multiple neurodegenerative conditions linking to Alzheimer's disease (AD) which is marked by hyperphosphorylated tau proteins and accumulation of Aβ plaques. Exciting findings reveal that miRNAs released from exosomes modulate the expression and function of amyloid precursor proteins (APP) and tau proteins. These open up the possibility that dysfunctional exosomal miRNAs may influence AD progression. In addition, it has been confirmed that the interaction between miRNAs released by exosomes and Toll-like receptors (TLR) initiates inflammation. In exosome support-deprived neurons, exosomal miRNAs may regulate neuroplasticity to relieve neurological damage. In this review, we summarize the literature on the function of exosomal miRNAs in AD pathology, the potential of these miRNAs as diagnostic biomarkers in AD, and the use of exosomes in the delivery of miRNAs which may lead to major advances in the field of macromolecular drug delivery.
Project description:Dystrophic neuronal processes harboring neuritic plaque (NP) tau pathology are found in association with Aβ plaques in Alzheimer's disease (AD) brain. Microglia are also in proximity to these plaques and microglial gene variants are known risk factors in AD, including loss-of-function variants of TREM2. We have further investigated the role of Aβ plaque-associated microglia in 5XFAD mice in which NP tau pathology forms after intracerebral injection of AD brain-derived pathologic tau (AD-tau), focusing on the consequences of reduced TREM2 expression and microglial depletion after treatment with the colony-stimulating factor 1 (CSFR1) inhibitor, PLX3397. Young 5XFAD mice treated with PLX3397 had a large reduction of brain microglia, including cortical plaque-associated microglia, with a significant reduction of Aβ plaque burden in the cortex. A corresponding decrease in cortical APP-positive dystrophic processes and NP tau pathology were observed after intracerebral AD-tau injection in the PLX3397-treated 5XFAD mice. Consistent with prior reports, 5XFAD × TREM2<sup>-/-</sup> mice showed a significant reduction of plaque-associated microglial, whereas 5XFAD × TREM2<sup>+/-</sup> mice had significantly more plaque-associated microglia than 5XFAD × TREM2<sup>-/-</sup> mice. Nonetheless, AD-tau injected 5XFAD × TREM2<sup>+/-</sup> mice showed greatly increased AT8-positive NP tau relative to 5XFAD × TREM2<sup>+/+</sup> mice. Expression profiling revealed that 5XFAD × TREM2<sup>+/-</sup> mice had a disease-associated microglial (DAM) gene expression profile in the brain that was generally intermediate between 5XFAD × TREM2<sup>+/+</sup> and 5XFAD × TREM2<sup>-/-</sup> mice. Microarray analysis revealed significant differences in cortical and hippocampal gene expression between AD-tau injected 5XFAD × TREM2<sup>+/-</sup> and 5XFAD × TREM2<sup>-/-</sup> mice, including pathways linked to microglial function. These data suggest there is not a simple correlation between the extent of microglia plaque interaction and plaque-associated neuritic damage. Moreover, the differences in gene expression and microglial phenotype between TREM2<sup>+/-</sup> and TREM2<sup>-/-</sup> mice suggest that the former may better model the single copy TREM2 variants associated with AD risk.
Project description:Extracellular vesicles (ECV), like exosomes, gained recently a lot of attention as potentially playing a significant role in neurodegenerative diseases, particularly in Aβ pathology. While there are a lot of reports on ECV/exosomes derived from a variety of cell types, there is limited information on ECV/exosomes originated from brain microvascular endothelial cells forming the blood-brain barrier (BBB). In this review, we summarize the literature data on brain endothelial ECV/exosomes and present our own data on BBB-derived ECV and their possible involvement in the brain's Aβ pathology. We propose that ECV/exosome release from brain endothelial cells associated with Aβ affects different cells of the neurovascular unit and may be an important contributor to the Aβ deposition in the central nervous system.
Project description:Mutations in the presenilin (PS/PSEN) genes encoding the catalytic components of γ-secretase accelerate amyloid-β (Aβ) and tau pathologies in familial Alzheimer's disease (AD). Although the mechanisms by which these mutations affect Aβ are well defined, the precise role PS/γ-secretase on tau pathology in neurodegeneration independently of Aβ is largely unclear. Here we report that neuronal PS deficiency in conditional knockout (cKO) mice results in age-dependent brain atrophy, inflammatory responses and accumulation of pathological tau in neurons and glial cells. Interestingly, genetic inactivation of presenilin 1 (PS1) or both PS genes in mutant human Tau transgenic mice exacerbates memory deficits by accelerating phosphorylation and aggregation of tau in excitatory neurons of vulnerable AD brain regions (e.g., hippocampus, cortex and amygdala). Remarkably, neurofilament (NF) light chain (NF-L) and phosphorylated NF are abnormally accumulated in the brain of Tau mice lacking PS. Synchrotron infrared microspectroscopy revealed aggregated and oligomeric β-sheet structures in amyloid plaque-free PS-deficient Tau mice. Hippocampal-dependent memory deficits are associated with synaptic tau accumulation and reduction of pre- and post-synaptic proteins in Tau mice. Thus, partial loss of PS/γ-secretase in neurons results in temporal- and spatial-dependent tau aggregation associated with memory deficits and neurodegeneration. Our findings show that tau phosphorylation and aggregation are key pathological processes that may underlie neurodegeneration caused by familial AD-linked PSEN mutations.