The relationship between genetic variation and gene expression in individual brain cell types and subtypes has remained elusive. Here, we generated single-nucleus RNA sequencing data from the dorsolateral prefrontal cortex of 424 individuals of advanced age; analyzing 1.5 million nuclear transcriptomes, we assessed the effect of genetic variants on RNA expression incis(cis-eQTL) for 7 cell types and 81 cell subtypes. This effort identified 10,004 eGenes at the cell type level and 8,138 eGenes at the cell subtype level. Many eGenes are only detected within cell subtypes. A new variant influencesAPOEexpression only in microglia and is associated with greater cerebral amyloid angiopathy but not Alzheimer pathology, accounting for the effect ofAPOEε4, providing mechanistic insights into both pathologies. While eQTLs are readily detected, only aTMEM106Bvariant robustly affects the proportion of cell subtypes. Integration of these results with GWAS highlighted the targeted cell type and likely causal gene within susceptibility loci for Alzheimer's, Parkinson's, schizophrenia, and educational attainment.

Alzheimer's Disease (AD) is a progressive neurodegenerative disease seen with advancing age. Recent studies have revealed diverse AD-associated cell states, yet when and how they impact the causal chain leading to AD remains unknown. To reconstruct the dynamics of the brain's cellular environment along the disease cascade and to distinguish between AD and aging effects, we built a comprehensive cell atlas of the aged prefrontal cortex from 1.64 million single-nucleus RNA-seq profiles. We associated glial, vascular and neuronal subpopulations with AD-related traits for 424 aging individuals, and aligned them along the disease cascade using causal modeling. We identified two distinct lipid-associated microglial subpopulations, one contributed to amyloid-β proteinopathy while the other mediated the effect of amyloid-beta in accelerating tau proteinopathy, as well as an astrocyte subpopulation that mediated the effect of tau on cognitive decline. To model the coordinated dynamics of the entire cellular environment we devised the BEYOND methodology which uncovered two distinct trajectories of brain aging that are defined by distinct sequences of changes in cellular communities. Older individuals are engaged in one of two possible trajectories, each associated with progressive changes in specific cellular communities that end with: (1) AD dementia or (2) alternative brain aging. Thus, we provide a cellular foundation for a new perspective of AD pathophysiology that could inform the development of new therapeutic interventions targeting cellular communities, while designing a different clinical management for those individuals on the path to AD or to alternative brain aging.

BackgroundAlzheimer’s Disease (AD) is a fatal neurodegenerative disease where abnormal accumulation of amyloid‐β and tau aggregates drive neurodegeneration and leads to cognitive impairment, dementia and eventually death. Combining results from recent studies, each finding a novel AD‐associated cell state of different non‐neuronal cell types, suggests a system level change of the cellular environment of the AD brain and its potential role in disease outcome.MethodTo better understand the dynamics of the cellular environment in aging human brains and AD, we applied single‐nucleus RNA‐sequencing to profile 424 aging brains of individuals across different clinicopathological characteristics. We characterized the cellular diversity within the aging human prefrontal cortex, resulting in a detailed atlas of ∼1.7 million cells classified into 95 cellular subsets of different cell types, including rare populations of microglia and astrocytes.ResultsQuantifying variations across individuals, we uncovered multiple cell subsets associated with disease pathologies and cognitive impairment. Of these, we prioritize an uncommon Microglial subset 13 (Mic.13, average 3.2% of microglia) and Astrocyte subset 10 (Ast.10, average 3.8% of astrocytes). Mic.13 expresses AD risk gene TREM2 and microglial markers linked to AD (e.g. APOE, SPP1), and its proportion is strongly associated with AD pathologies, amyloid‐β plaques and neurofibrillary tangles, and cognitive decline. Applying a mediation model, we showed that Mic.13 partially mediates the effect of amyloid‐β on tau (proportion mediated [p.m.] = 18%). While the impact of Mic.13 on cognitive decline is largely mediated by tau (p.m. = 43%), an additional subset highlighted by our analysis Ast.10, associated with both tau and cognitive decline, partially mediates the impact of tau and Mic.13 on cognitive decline (p.m. = 7.2% and p.m. = 12% respectively). Integration of all cellular states revealed an intricate network of multi‐cellular communities, each composed of multiple states of different cell types. Investigating the dynamics of the cellular communities in the aging brain and along disease progression, uncovered unique trajectories spanning from a healthy to diseased brains.ConclusionOur charted atlas of the cellular environment and modelling of the cellular dynamics supports a system level change underlying AD, with an active role for multiple cell types in disease progression, and highlights novel research directions and potential therapeutic targets.