Huang, Yiyao ORCID: 0000-0003-1749-963X, Driedonks, Tom AP, Cheng, Lesley ORCID: 0000-0002-8075-6144, Rajapaksha, Harinda, Turchinovich, Andrey, Routenberg, David A, Nagaraj, Rajini, Redding-Ochoa, Javier, Arab, Tanina ORCID: 0000-0003-4485-1096, Powell, Bonita H, Pletniková, Olga, Troncoso, Juan C, Zheng, Lei, Hill, Andrew F ORCID: 0000-0001-5581-2354, Mahairaki, Vasiliki and Witwer, Kenneth W ORCID: 0000-0003-1664-4233 (2022) Relationships of APOE genotypes with small RNA and protein cargo of brain tissue extracellular vesicles from patients with late-stage AD. Neurology: Genetics, 8 (6). e200026-e200026. ISSN 2376-7839

Abstract

Background and Objectives Variants of the apolipoprotein E (APOE) gene are the greatest known risk factors for sporadic Alzheimer disease (AD). Three major APOE isoform alleles, ϵ2, ϵ3, and ϵ4, encode and produce proteins that differ by only 1-2 amino acids but have different binding partner interactions. Whereas APOE ϵ2 is protective against AD relative to ϵ3, ϵ4 is associated with an increased risk for AD development. However, the role of APOE in gene regulation in AD pathogenesis has remained largely undetermined. Extracellular vesicles (EVs) are lipid bilayer-delimited particles released by cells to dispose of unwanted materials and mediate intercellular communication, and they are implicated in AD pathophysiology. Brain-derived EVs (bdEVs) could act locally in the tissue and reflect cellular changes. To reveal whether APOE genotype affects EV components in AD brains, bdEVs were separated from patients with AD with different APOE genotypes for parallel small RNA and protein profile. Methods bdEVs from late-stage AD brains (BRAAK stages 5-6) from patients with APOE genotypes ϵ2/3 (n = 5), ϵ3/3 (n = 5), ϵ3/4 (n = 6), and ϵ4/4 (n = 6) were separated using our published protocol into a 10,000g pelleted extracellular fraction (10K) and a further purified EV fraction. Counting, sizing, and multiomic characterization by small RNA sequencing and proteomic analysis were performed for 10K, EVs, and source tissue. Results Comparing APOE genotypes, no significant differences in bdEV total particle concentration or morphology were observed. Overall small RNA and protein profiles of 10K, EVs, and source tissue also did not differ substantially between different APOE genotypes. However, several differences in individual RNAs (including miRNAs and tRNAs) and proteins in 10K and EVs were observed when comparing the highest and lowest risk groups (ϵ4/4 and ϵ2/3). Bioinformatic analysis and previous publications indicate a potential regulatory role of these molecules in AD. Discussion For patients with late-stage AD in this study, only a few moderate differences were observed for small RNA and protein profiles between APOE genotypes. Among these, several newly identified 10K and EV-associated molecules may play roles in AD progression. Possibly, larger genotype-related differences exist and are more apparent in or before earlier disease stages.

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