Neuronal apolipoprotein E4 increases cell death and phosphorylated tau release in alzheimer disease

Abstract

Objective:The apolipoprotein E (APOE) E4 isoform is the strongest genetic risk factor for sporadic Alzheimer disease (AD). Although APOE is predominantly expressed by astrocytes in the central nervous system, neuronal expression of APOE is of increasing interest in age-related cognitive impairment, neurological injury, and neurodegeneration. Here, we show that endogenous expression of E4 in stem-cell-derived neurons predisposes them to injury and promotes the release of phosphorylated tau. Methods: Induced pluripotent stem cells from 2 unrelated AD patients carrying the E4 allele were corrected to the E3/E3 genotype with the CRISPR/Cas9 system and differentiated into pure cultures of forebrain excitatory neurons without contamination from other cells types. Results: Compared to unedited E4 neurons, E3 neurons were less susceptible to ionomycin-induced cytotoxicity. Biochemically, E4 cells exhibited increased tau phosphorylation and ERK1/2 phosphoactivation. Moreover, E4 neurons released increased amounts of phosphorylated tau extracellularly in an isoform-dependent manner by a heparin sulfate proteoglycan-dependent mechanism. Interpretation: Our results demonstrate that endogenous expression of E4 by stem-cell-derived forebrain excitatory neurons predisposes neurons to calcium dysregulation and ultimately cell death. This change is associated with increased cellular tau phosphorylation and markedly enhanced release of phosphorylated tau. Importantly, these effects are independent of glial APOE. These findings suggest that E4 accelerates spreading of tau pathology and neuron death in part by neuron-specific, glia-independent mechanisms. ANN NEUROL 2019;85:726-739 T he apolipoprotein E (APOE) E4 allele, the strongest genetic risk factor for sporadic Alzheimer disease (AD), differs from the risk-neutral E3 allele by a single nucleotide polymorphism (SNP). 1,2 E4 patients exhibit greater brain atrophy, accumulation of hyperphosphorylated tau protein, and deposition of amyloid, albeit by unclear mechanisms. 3-6 Although most APOE is expressed by astrocytes in the brain, 7-9 neuronal APOE is of increasing interest in age-related cognitive impairment, neurological injury, and neurodegeneration. 10-12 Disease modeling using isogenic stem cells has demonstrated that, compared to E3, expression of E4 leads to distinct transcriptomic differences in multiple neural cell types and increases tau phosphorylation in neurons. 13,14 However, the effects of APOE genotype on neuronal viability and tau release are unknown. Using genome editing and a reductionist human stem cell culture approach, we show that endogenous expression of E4 predisposes pure cultures of forebrain excitatory neurons to injury and promotes release of phosphorylated tau (p-tau). These findings suggest that neuronal APOE can accelerate brain atrophy and the spreading of tau pathology in E4-carrying AD patients independently of glia. Subjects and Methods Reprogramming and Culture of Stem CellsFibroblast lines from patients diagnosed with AD (AD1, AG11414; AD...

Microglia prevent beta-amyloid plaque formation in the early stage of an Alzheimer’s disease mouse model with suppression of glymphatic clearance

“…Apart from Aβ, evidence from recent studies suggests that apoE in uencing tau pathology and taumediated neurodegeneration is also in an isoform-dependent manner [60,61]. Furthermore, apoE ε4 accumulation impairs glial responsiveness, lipid transport and cerebrovascular integrity and function, which might be independent of Aβ-related pathways [31,62,63].…”

Modeling Alzheimer’s disease with iPSC-derived brain cells

“…The first identified SAD risk factor, APOE4 has been the most studied risk mutation in iPSC-derived cells, as it has been in rodent and other models. iPSCderived neurons carrying APOE4 produce more Aβ and have higher levels of tau phosphorylation compared to APOE3 cells [35,38,39,110,114]. Surprisingly, elevated p-tau levels were not dependent on Aβ in APOE4 cells, indicating perturbation of additional pathways regulating tau phosphorylation [39].…”

Therapeutic approaches targeting Apolipoprotein E function in Alzheimer’s disease

“…CRISPR/Cas9 has already been proven successful in iPS cells, where cells derived from a healthy E3/E4 individual were converted into E2/E2, E3/E3, E4/E4, or an APOE KO genotype [157]. A second group used iPS cells derived neurons from an APOE4 carrier and found that CRISPR-editing the APOE4 reduced tau phosphorylation and inomycin-induced cell death [158]. Interestingly, though in the CNS APOE is mostly synthesized by astrocytes, this study showed that editing neuronal APOE to the E3 isoform in these iPS-derived neurons is sufficient to protect them from cytotoxic injury [158].…”

“…A second group used iPS cells derived neurons from an APOE4 carrier and found that CRISPR-editing the APOE4 reduced tau phosphorylation and inomycin-induced cell death [158]. Interestingly, though in the CNS APOE is mostly synthesized by astrocytes, this study showed that editing neuronal APOE to the E3 isoform in these iPS-derived neurons is sufficient to protect them from cytotoxic injury [158]. Another study generated different brain cell types and organoids from iPS cells derived from a human subjecton editing the APOE4 allele to APOE3 in these iPS-derived cells increased Aβ clearance and reduced Aβ in the organoid cultures [49].…”