Abstract
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.
Highlights
Despite intensive research efforts, no effective treatment or preventative strategies for Alzheimer’s disease (AD) have yet been identified
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)
The double strand break (DSB) was repaired by homologous recombination in the presence of donor plasmids that contained a secretory signal sequence derived from the rat preproenkephalin gene (PENK, Rattus norvegicus) fused in frame to either a human
Summary
No effective treatment or preventative strategies for Alzheimer’s disease (AD) have yet been identified Reasons for this include the relative experimental inaccessibility of human brain, a lengthy preclinical symptom free stage, as well as the complexity of the disorder. Transgenic rodent models have been dominant for preclinical studies which have largely failed in human trials (Cummings et al, 2014; Sasaguri et al, 2017). The reasons for these failures are unknown but often attributed to technical explanations such as the stage of AD during treatment, unknown target engagement, improper dosage, or off-target “side effects.”. The reasons for these failures are unknown but often attributed to technical explanations such as the stage of AD during treatment, unknown target engagement, improper dosage, or off-target “side effects.” An alternative explanation could be more generally related to well-known phenotypic deficiencies of
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