Modelling APOE ɛ3/4 allele-associated sporadic Alzheimer’s disease in an induced neuron

Kim, Hongwon; Yoo, Jang-Yong; Shin, Jaein; Chang, Yu‐Jung; Jung, Junghyun; Jo, Dong-Gyu; Kim, Jang Hwan; Jang, Wonhee; Lengner, Christopher J.; Kim, Byung Soo; Kim, Jong-Pil

doi:10.1093/brain/awx144

Cited by 21 publications

(35 citation statements)

References 47 publications

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“…26 Neurons derived from these hiPSCs were previously shown to exhibit increased amyloid processing, tau phosphorylation, and vulnerability to calcium dysregulation compared to neurons derived from control patient hiPSCs. 15,27 To determine whether the E4 allele influences this cellular phenotype, both E4/E3 hiPSC lines were corrected to E3/E3 genotype by overexpression of Cas9 nuclease, a single guide RNA sequence (sgRNA) targeted to the terminal exon of the gene, and a homologydirected repair template encoding a segment of the E3 allele (Fig 1). Genotypes of the untargeted and edited cell lines were confirmed by PCR amplification and direct Sanger sequencing.…”

Section: Resultsmentioning

confidence: 99%

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

Wadhwani

Affaneh

Gulden

et al. 2019

Annals of Neurology

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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...

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Section: Resultsmentioning

confidence: 99%

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

Wadhwani

Affaneh

Gulden

et al. 2019

Annals of Neurology

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“…One of the challenges to date has been modeling sporadic AD in both rodent and human models of disease, with familial AD mutations accounting for only 5–10% of all AD cases (Kim et al, 2017 ). Excitingly, Lin et al ( 2018 ) describes the first experiments in which CRISPR/Cas9 technology has been used to generate isogenic APOE4 iPSC-derived microglia.…”

Section: Modeling Microglial Involvementmentioning

confidence: 99%

Microglia in Alzheimer's Disease: A Role for Ion Channels

et al. 2018

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Alzheimer's disease is the most common form of dementia, it is estimated to affect over 40 million people worldwide. Classically, the disease has been characterized by the neuropathological hallmarks of aggregated extracellular amyloid-β and intracellular paired helical filaments of hyperphosphorylated tau. A wealth of evidence indicates a pivotal role for the innate immune system, such as microglia, and inflammation in the pathology of Alzheimer's disease. The over production and aggregation of Alzheimer's associated proteins results in chronic inflammation and disrupts microglial clearance of these depositions. Despite being non-excitable, microglia express a diverse array of ion channels which shape their physiological functions. In support of this, there is a growing body of evidence pointing to the involvement of microglial ion channels contributing to neurodegenerative diseases such as Alzheimer's disease. In this review, we discuss the evidence for an array of microglia ion channels and their importance in modulating microglial homeostasis and how this process could be disrupted in Alzheimer's disease. One promising avenue for assessing the role that microglia play in the initiation and progression of Alzheimer's disease is through using induced pluripotent stem cell derived microglia. Here, we examine what is already understood in terms of the molecular underpinnings of inflammation in Alzheimer's disease, and the utility that inducible pluripotent stem cell derived microglia may have to advance this knowledge. We outline the variability that occurs between the use of animal and human models with regards to the importance of microglial ion channels in generating a relevant functional model of brain inflammation. Overcoming these hurdles will be pivotal in order to develop new drug targets and progress our understanding of the pathological mechanisms involved in Alzheimer's disease.

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“…Subsequent additions of other transcription factors, including NEUROD1 and SOX2 have improved the conversion efficiency resulted in complex neuronal morphologies and expression of neuron‐specific proteins, for example β‐III tubulin and MAP2 (Karow et al, 2012; Pang et al, 2011; Wang et al, 2014). Nanopattern topography has also been shown to improve direct conversion efficiency in mouse and human fibroblast cells, in which the cells are plated on defined nanoscale grooved patterns during the direct conversion process (Kim et al, 2017), and combinations of miRNAs can also directly convert fibroblasts (Yoo et al, 2011). Finally, knockdown of human polypyrimidine tract binding protein (PTBP), an RNA‐binding protein that can block neuronal differentiation, has also been shown to directly convert fibroblasts to iNs (Liu et al, 2014).…”

Section: Induced Neuronsmentioning

confidence: 99%

“…Furthermore, gene expression profiling and network analysis of these iNs indicated dysregulation of desmoglein 2 ( DSG2 ), a putative AD risk gene (Karch & Goate, 2015). Functional studies of DSG2 knockdown demonstrated that inhibition of DSG2 reduced Aβ aggregation induced by overexpression of APP in APOE 3/4 iNs (Kim et al, 2017). There are fewer studies using iNs from sporadic AD subjects than familial genetic AD subjects, in part due to immense genetic and phenotypic variability between sporadic AD subjects.…”

Section: Induced Neuronsmentioning

confidence: 99%

The application of in vitro‐derived human neurons in neurodegenerative disease modeling

D’Souza

Rose

Knupp

et al. 2020

J of Neuroscience Research

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The development of safe and effective treatments for age‐associated neurodegenerative disorders is an on‐going challenge faced by the scientific field. Key to the development of such therapies is the appropriate selection of modeling systems in which to investigate disease mechanisms and to test candidate interventions. There are unique challenges in the development of representative laboratory models of neurodegenerative diseases, including the complexity of the human brain, the cumulative and variable contributions of genetic and environmental factors over the course of a lifetime, inability to culture human primary neurons, and critical central nervous system differences between small animal models and humans. While traditional rodent models have advanced our understanding of neurodegenerative disease mechanisms, key divergences such as the species‐specific genetic background can limit the application of animal models in many cases. Here we review in vitro human neuronal systems that employ stem cell and reprogramming technology and their application to a range of neurodegenerative diseases. Specifically, we compare human‐induced pluripotent stem cell‐derived neurons to directly converted, or transdifferentiated, induced neurons, as both model systems can take advantage of patient‐derived human tissue to produce neurons in culture. We present recent technical developments using these two modeling systems, as well as current limitations to these systems, with the aim of advancing investigation of neuropathogenic mechanisms using these models.

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Modelling APOE ɛ3/4 allele-associated sporadic Alzheimer’s disease in an induced neuron

Cited by 21 publications

References 47 publications

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

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

Microglia in Alzheimer's Disease: A Role for Ion Channels

The application of in vitro‐derived human neurons in neurodegenerative disease modeling

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