Abnormal accumulation of Aβ (amyloid β) within AEL (autophagy–endosomal–lysosomal) vesicles is a prominent neuropathological feature of AD (Alzheimer's disease), but the mechanism of accumulation within vesicles is not clear. We express secretory forms of human Aβ 1–40 or Aβ 1–42 in Drosophila neurons and observe preferential localization of Aβ 1–42 within AEL vesicles. In young animals, Aβ 1–42 appears to associate with plasma membrane, whereas Aβ 1–40 does not, suggesting that recycling endocytosis may underlie its routing to AEL vesicles. Aβ 1–40 , in contrast, appears to partially localize in extracellular spaces in whole brain and is preferentially secreted by cultured neurons. As animals become older, AEL vesicles become dysfunctional, enlarge and their turnover appears delayed. Genetic inhibition of AEL function results in decreased Aβ 1–42 accumulation. In samples from older animals, Aβ 1–42 is broadly distributed within neurons, but only the Aβ 1–42 within dysfunctional AEL vesicles appears to be in an amyloid-like state. Moreover, the Aβ 1–42 -containing AEL vesicles share properties with AD-like extracellular plaques. They appear to be able to relocate to extracellular spaces either as a consequence of age-dependent neurodegeneration or a non-neurodegenerative separation from host neurons by plasma membrane infolding. We propose that dysfunctional AEL vesicles may thus be the source of amyloid-like plaque accumulation in Aβ 1–42 -expressing Drosophila with potential relevance for AD.
We describe the construction and phenotypic analysis of a human embryonic stem cell model of progressive Aβ-dependent neurodegeneration (ND) with potential relevance to Alzheimer's disease (AD). We modified one allele of the normal APP locus to directly express a secretory form of Aβ40 or Aβ42, enabling expression from this edited allele to bypass the normal amyloidogenic APP processing pathway. Following neuronal differentiation, edited cell lines specifically accumulate intracellular aggregated/oligomeric Aβ, exhibit a synaptic deficit, and have an abnormal accumulation of endolysosomal vesicles. Edited cultures progress to a stage of overt ND. All phenotypes appear at earlier culture times for Aβ42 relative to Aβ40. Whole transcriptome RNA-Seq analysis identified 23 up and 70 down regulated genes (differentially expressed genes) with similar directional fold change but larger absolute values in the Aβ42 samples suggesting common underlying pathogenic mechanisms. Pathway/annotation analysis suggested that down regulation of extracellular matrix and cilia functions is significantly overrepresented. This cellular model could be useful for uncovering mechanisms directly linking Aβ to neuronal death and as a tool to screen for new therapeutic agents that slow or prevent human ND.
Abstract We describe construction and phenotypic analysis of a human embryonic stem cell model of progressive Aβ-dependent neurodegeneration (ND) with potential relevance to Alzheimer’s disease (AD). We modified one allele of the normal APP locus to directly express a secretory form of Aβ40 or Aβ42, eliminating the need for amyloidogenic APP proteolysis. Following neuronal differentiation edited cell lines specifically accumulate aggregated/oligomeric Aβ, exhibit a synaptic deficit and have an abnormal accumulation of endolysosomal vesicles. Edited cultures progress to a stage of overt ND. All phenotypes appear at earlier culture times for Aβ42 relative to Aβ40. Whole transcriptome RNA-Seq analysis identified 23 up and 70 down regulated genes (DEGs) with similar directional fold change but larger absolute values in the Aβ42 samples suggesting common underlying pathogenic mechanisms. Pathway/annotation analysis suggested that down regulation of extracellular matrix and cilia functions are significantly overrepresented. This cellular model could be useful for uncovering mechanisms directly linking Aβ to neuronal death and as a tool to screen for new therapeutic agents that slow or prevent human ND.
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