Cholesterol and matrisome pathways dysregulated by APOE4 in human glia

Background The association of Apolipoprotein E (APOE) ɛ4 risk with Alzheimer’s disease (AD) is well‐established. However, the impact of APOE ɛ4 on human brain cell function remains unclear. We hypothesized that human APOE ε4/ε4 (APOE 44) genotype contributes to disease risk through cell autonomous and non‐cell autonomous mechanisms in a human cell specific manner. Method Global transcriptomic analyses were performed on astrocytes, microglia, mixed cortical cultures (neurons and astrocytes) and brain microvascular endothelial cells derived from human induced pluripotent stem cells (iPSCs) and post‐mortem AD brains. Lipid pathway dysregulation identified through GSEA, WGCNA and cell type deconvolution was validated in vitro using Gas Chromatography‐Mass Spectrometry (GC‐MS), filipin staining, autophagy fluorescent reporters with immunocytochemistry and western blotting to determine the subcellular compartment that accumulates cholesterol in iPSC‐derived astrocytes, and further confirmed by immunohistochemical analysis in human APOE4 post‐mortem brain. Matrisome pathway dysregulation was validated by Luminex multiplex immunoassays and intracellular protein levels in pure astrocytes and mixed cortical cultures. Result Global transcriptomic analyses reveal that APOE 44 drives cholesterol metabolic dysregulation in astrocytes and microglia from human iPSCs and cell type‐deconvolved post‐mortem AD brain. APOE ε4 leads to elevated de novo cholesterol synthesis despite the intracellular accumulation of free cholesterol in lysosome and cholesterol ester in lipid droplets coupled with reduced efflux due to impaired lipophagy in iPSC‐derived astrocytes. Lysosomal cholesterol accumulation is found in astrocytes from APOE ε4 human brain. Activation of LXRs and induction of lipophagy ameliorates the lipid phenotypes in APOE ε4 astrocytes. Further, matrisome dysregulation is associated with upregulated chemotaxis, glial activation, and lipid biosynthesis in astrocytes co‐cultured with neurons, which recapitulates altered astrocyte matrisome signaling in human brain. Conclusion Thus, human APOE ε4 causes human glia‐specific, cell autonomous and non‐cell autonomous cholesterol and matrisome pathway dysregulation that may increase AD risk.