The abnormal assembly of tau protein in neurons is a pathological hallmark of multiple neurodegenerative diseases, including Alzheimer’s disease (AD). Assembled tau associates with extracellular vesicles (EVs) in the central nervous system of individuals with AD, which is linked to its clearance and prion-like propagation. However, the identities of the assembled tau species and EVs, as well as how they associate, are not known. Here, we combined quantitative mass spectrometry, cryo-electron tomography and single-particle cryo-electron microscopy to study brain EVs from individuals with AD. We found tau filaments composed mainly of truncated tau that were enclosed within EVs enriched in endo-lysosomal proteins. We observed multiple filament interactions, including with molecules that tethered filaments to the EV limiting membrane, suggesting selective packaging. Our findings will guide studies into the molecular mechanisms of EV-mediated secretion of assembled tau and inform the targeting of EV-associated tau as potential therapeutic and biomarker strategies for AD.

Electronic nicotine delivery systems (ENDS) are a rapidly growing global market advertised as a safer alternative to combustible cigarettes. However, comprehensive investigations of END aerosol physicochemical and toxicological properties have not been fully explored across brands to assess relative safety. In this study, we evaluated aerosols collected from three ENDS – Juul Fruit Medley (5% nicotine), Logic Power (2.4% nicotine), and Mistic (1.8% nicotine). ENDS aerosols were generated using standard machine puffing regimen and collected with a novel fluoropolymer condensation trap. Triple quadrupole-inductively coupled plasma-mass determined the presence of heavy metals in collected aerosols. The toxicological effects of ENDS aerosols on normal human bronchial epithelial cells (NHBE) were investigated using cellular viability, reactive oxygen species, oxidative stress assays, along with DNA damage assessments using the CometChip©. Results indicated the total metal concentrations within collected ENDS aerosols were higher for Mistic and Logic compared to Juul. Logic Power aerosols elicited higher reactive oxygen species levels than Mistic and Juul in NHBE after 24-hr exposure. Similar dose-dependent reductions of cellular viability and total glutathione were found for each exposure. However, Logic and Juul aerosols caused greater single stranded DNA damage compared to Mistic. Our study indicates that regardless of brand, ENDS aerosols are toxic to upper airway epithelial cells and may pose a potential respiratory hazard to occasional and frequent users.

Reactive astrocytes are associated with Alzheimer's disease (AD), and several AD genetic risk variants are associated with genes highly expressed in astrocytes. However, the contribution of genetic risk within astrocytes to cellular processes relevant to the pathogenesis of AD remains ill‐defined. Here, we present a resource for studying AD genetic risk in astrocytes using a large collection of induced pluripotent stem cell (iPSC) lines from deeply phenotyped individuals with a range of neuropathological and cognitive outcomes. IPSC lines from 44 individuals were differentiated into astrocytes followed by unbiased molecular profiling using RNA sequencing and tandem mass tag‐mass spectrometry. We demonstrate the utility of this resource in examining gene‐ and pathway‐level associations with clinical and neuropathological traits, as well as in analyzing genetic risk and resilience factors through parallel analyses of iPSC‐astrocytes and brain tissue from the same individuals. Our analyses reveal that genes and pathways altered in iPSC‐derived astrocytes from individuals with AD are concordantly dysregulated in AD brain tissue. This includes increased levels of prefoldin proteins, extracellular matrix factors, COPI‐mediated trafficking components and reduced levels of proteins involved in cellular respiration and fatty acid oxidation. Additionally, iPSC‐derived astrocytes from individuals resilient to high AD neuropathology show elevated basal levels of interferon response proteins and increased secretion of interferon gamma. Correspondingly, higher polygenic risk scores for AD are associated with lower levels of interferon response proteins in astrocytes. This study establishes an experimental system that integrates genetic information with a matched iPSC lines and brain tissue data from a large cohort of individuals to identify genetic contributions to molecular pathways affecting AD risk and resilience.

Alzheimer’s disease, the leading cause of dementia in the elderly, is a neurodegenerative disorder that has been studied to uncover therapeutic pathways through its molecular and cellular hallmarks. Here, we present a comprehensive investigation of cellular heterogeneity from the temporal cortex region of 40 individuals, comprising healthy donors and individuals with differing AD pathology. Using single-nucleus transcriptomic analysis of 430,271 nuclei from both gray and white matter of these individuals, we identified cell type-specific subclusters in both neuronal and glial cell types with varying degrees of association with AD pathology. We extended this analysis by performing multiplexed in situ hybridization using the CARTANA platform, capturing 155 genes in 13 individuals with differing tau pathology. We not only replicated snRNA data key findings from our spatial data analysis but also identified a set of cell type-specific genes that show selective enrichment or depletion near pathological inclusions.

SUMMARY: The abnormal assembly of tau protein in neurons is the pathological hallmark of multiple neurodegenerative diseases, including Alzheimer’s disease (AD). In addition, assembled tau associates with extracellular vesicles (EVs) in the central nervous system of patients with AD, which is linked to its clearance and prion-like propagation between neurons. However, the identities of the assembled tau species and the EVs, as well as how they associate, are not known. Here, we combined quantitative mass spectrometry, cryo-electron tomography and single-particle cryo-electron microscopy to study brain EVs from AD patients. We found filaments of truncated tau enclosed within EVs enriched in endo-lysosomal proteins. We observed multiple filament interactions, including with molecules that tethered filaments to the EV limiting membrane, suggesting selective packaging. Our findings will guide studies into the molecular mechanisms of EV-mediated secretion of assembled tau and inform the targeting of EV-associated tau as potential therapeutic and biomarker strategies for AD.