Regulation of diverse microglial phenotypes in neurodegeneration

University Of Washington, Seattle WA

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Abstract

Alzheimer disease is a multifactorial process that has thus far been challenging to therapeutically target. A large body of literature supports a key role for microglia, the innate immune cells of the brain, in AD pathogenesis. Manipulating microglial activity may be one tractable AD intervention however microglia roles in AD are complex. Microglia orchestrate a suite of functions in the homeostatic, aged and diseased brain. Understanding which of these microglial phenotypes contribute to AD and how these phenotypes are regulated is necessary to develop precise therapies for AD. By performing single nucleus RNAseq (snRNAseq) on more than 5,000 microglia per individual from a cohort of 22 late-onset AD (LOAD) and age-matched control autopsy samples we have identified microglia clusters significantly associated with AD. One of these is specifically enriched in genes in the endolysosomal network (ELN), DNA damage response pathways and Type 1 interferon signaling. While these pathologies have been documented in AD, they are also influenced by aging, highlighting the complication of studying a late age neurodegenerative disease. In contrast, individuals with early onset familial AD (EOFAD), driven by genetic variants develop AD early in life and at time of death, EOFAD individuals have fewer systemic and brain aging changes than those with LOAD. We hypothesize that response to DNA damage exacerbated by a dysfunctional ELN contributes to AD pathogenesis independent of age. In this study we will leverage our unique cohorts of frozen brain tissue and human induced pluripotent stem cell (hiPSC) in 3 Aims. We will determine microglial molecular profiles (Aim 1) and spatial organization (Aim 2) with and without AD across the spectrum of young to older adult age. We will functionally test our hypothesis using hiPSC models, which lack age related changes (Aim 3). We propose to generate microglial transcriptomic, open chromatin and DNA methylation profiles building gene networks from frozen cortical samples of 19 EOFAD individuals who carry pathogenic variants in PSEN1/PSEN2 or APP and compare them to cortical samples from 35 age-matched unaffected controls aged 25 to 60 years and 10 controls aged 65-75 years. We will map the changes associated with age alone in microglia profiles across the adult age in the 25yr-75 yr control cohort. We will leverage our existing older control and LOAD snRNAseq dataset as another comparison and nominate which microglial changes appear in the presence of AD pathology, age alone, and those that are exacerbated by the presence of both AD and age. To functionally measure the significance of the AD phenotypes we have identified we will use hiPSC derived microglia and neurons to test the hypothesis that ELN dysfunction contributes to damage and accumulation of nucleic acids contributing to neuronal toxicity. These integrated omics and in vitro studies evaluating human microglia across the adult age span and AD pathology will identify and functionally test factors regulating microglial phenotypes that can inform effective therapeutic targeting for both EOFAD and LOAD.