Generation of a set of isogenic, gene-edited iPSC lines homozygous for all main APOE variants and an APOE knock-out line

Schmid, Benjamin; Prehn, Kennie R; Nimsanor, Natakarn; Garcia, Blanca Irene Aldana; Poulsen, Ulla; Jørring, Ida; Rasmussen, Mette; Clausen, Christian; Mau-Holzmann, Ulrike A.; Ramakrishna, Sarayu; Muddashetty, Ravi; Steeg, Rachel; Bruce, Kevin; Mackintosh, Peter; Ebneth, Andreas; Holst, Bjørn; Cabrera-Socorro, Alfredo

doi:10.1016/j.scr.2018.11.010

Cited by 36 publications

(31 citation statements)

References 4 publications

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“…With the emergence of powerful gene-editing tools such as the CRISPR-Cas system [ 336 – 338 ], generating isogenic iPSC lines from one APOE genotype (normally APOE*ε3/3 ) to other genotypes becomes efficient and cost-effective [ 339 – 341 ]. Compared to APOE*ε3 cells, isogenic APOE*ε4 cells show dramatic phenotypic changes, including increased Aβ42 and phosphorylated tau in neurons, impaired Aβ uptake and cholesterol metabolism in astrocyte, and reduced phagocytosis of Aβ in microglia [ 339 , 340 ].…”

Section: Main Textmentioning

confidence: 99%

APOE2: protective mechanism and therapeutic implications for Alzheimer’s disease

Shue

Zhao

et al. 2020

Mol Neurodegeneration

152

119

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Investigations of apolipoprotein E (APOE) gene, the major genetic risk modifier for Alzheimer’s disease (AD), have yielded significant insights into the pathogenic mechanism. Among the three common coding variants, APOE*ε4 increases, whereas APOE*ε2 decreases the risk of late-onset AD compared with APOE*ε3. Despite increased understanding of the detrimental effect of APOE*ε4, it remains unclear how APOE*ε2 confers protection against AD. Accumulating evidence suggests that APOE*ε2 protects against AD through both amyloid-β (Aβ)-dependent and independent mechanisms. In addition, APOE*ε2 has been identified as a longevity gene, suggesting a systemic effect of APOE*ε2 on the aging process. However, APOE*ε2 is not entirely benign; APOE*ε2 carriers exhibit increased risk of certain cerebrovascular diseases and neurological disorders. Here, we review evidence from both human and animal studies demonstrating the protective effect of APOE*ε2 against AD and propose a working model depicting potential underlying mechanisms. Finally, we discuss potential therapeutic strategies designed to leverage the protective effect of APOE2 to treat AD.

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Section: Main Textmentioning

confidence: 99%

APOE2: protective mechanism and therapeutic implications for Alzheimer’s disease

Shue

Zhao

et al. 2020

Mol Neurodegeneration

152

119

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“…iPSC maintenance and conditioned media collection -The iPSCs (APOE KO, APOE 3/3, and APOE 4/4) were obtained from Bioneer A/S, Denmark. Briefly, the iPSCs from an 18-year old male of APOE 3/4 genotype were subjected to CRISPR-Cas9 gene editing to obtain isogenic iPSC lines of APOE KO, APOE 3/3, and APOE 4/4 genotypes 42 . The iPSCs were maintained in mTeSR1 complete media (72232, Stem Cell Technologies) at 37˚C, 5% CO2 conditions.…”

Section: Methodsmentioning

confidence: 99%

“…We used conditioned or secreted media from human induced pluripotent cells (hiPSCs) as one of the sources of APOE. iPSC lines of different APOE genotypes (APOE3/3, APOE4/4, and APOE KO) were generated by CRISPR-Cas9 based editing of the original APOE3/4 iPSC line 42 . These iPSCs were characterized for the expression of the pluripotency markers (OCT4 and Nanog) and normal karyotype profile ( Fig S1A and S1B).…”

Section: Apoe4 Causes a Reduction Of Protein Synthesis In Neuronsmentioning

confidence: 99%

APOE4 affects basal and NMDAR mediated protein synthesis in neurons by perturbing calcium homeostasis

Ramakrishna

Jhaveri

Konings

et al. 2020

Preprint

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Apolipoprotein E (APOE), one of the primary lipoproteins in the brain has three isoforms in humans – APOE2, APOE3, and APOE4. APOE4 is the most well-established risk factor increasing the pre-disposition for Alzheimer’s disease. The presence of the APOE4 allele alone is shown to cause synaptic defects in neurons and recent studies have identified multiple pathways directly influenced by APOE4. However, the mechanisms underlying APOE4 induced synaptic dysfunction remain elusive. Here, we report that the acute exposure of primary cortical neurons to APOE4 leads to a significant decrease in global protein synthesis. APOE4 treatment also abrogates the NMDA mediated translation response indicating an impairment of synaptic signaling. Importantly, we demonstrate that both APOE3 and APOE4 generate a distinct translation response which is closely linked to their respective calcium signature. Acute exposure to APOE3 causes a short burst of calcium through NMDARs in neurons leading to an initial decrease in protein synthesis which quickly recovers. Contrarily, APOE4 leads to a sustained increase in calcium levels by activating both NMDARs and L-VGCCs, thereby causing sustained translation inhibition through eEF2 phosphorylation, which in turn disrupts NMDAR response. Thus, we show that APOE4 affects basal and activity mediated protein synthesis response in neurons by affecting calcium homeostasis. We propose this as a possible mechanism to explain the synaptic dysfunction caused by APOE4.Highlights / SummaryAPOE3 treatment causes a short burst of calcium through NMDARs, leading to an acute increase in eEF2 phosphorylation which eventually recovers to basal levels.Global translation follows a similar temporal profile of initial inhibition followed by recovery in APOE3 treated neurons, thus unaffecting the NMDA mediated translation response.APOE4 treatment activates both NMDARs and L-VGCCs leading to a marked elevation in calcium levels, thus causing sustained increase in eEF2 phosphorylation as well as global translation inhibition.Hence, the NMDA mediated response is perturbed, potentially causing a stress-related phenotype in APOE4 treated neurons.Thus, different calcium signatures and sources lead to distinct temporal profiles of translation.

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“…CRISPR/Cas9 basically functions like a pair of molecular scissors where an editable guide RNA leads the Cas9 'scissor' to a specific site of the genome to cut where a different nucleotide sequence can then be inserted to correct a genetic defect [156]. 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].…”

Section: Crispr/cas9 Mediated Gene Editingmentioning

confidence: 99%

Therapeutic approaches targeting Apolipoprotein E function in Alzheimer’s disease

Williams

Borchelt

Chakrabarty

2020

Mol Neurodegeneration

118

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One of the primary genetic risk factors for Alzheimer's disease (AD) is the presence of the Ɛ4 allele of apolipoprotein E (APOE). APOE is a polymorphic lipoprotein that is a major cholesterol carrier in the brain. It is also involved in various cellular functions such as neuronal signaling, neuroinflammation and glucose metabolism. Humans predominantly possess three different allelic variants of APOE, termed E2, E3, and E4, with the E3 allele being the most common. The presence of the E4 allele is associated with increased risk of AD whereas E2 reduces the risk. To understand the molecular mechanisms that underlie APOE-related genetic risk, considerable effort has been devoted towards developing cellular and animal models. Data from these models indicate that APOE4 exacerbates amyloid β plaque burden in a dose-dependent manner. and may also enhance tau pathogenesis in an isoform-dependent manner. Other studies have suggested APOE4 increases the risk of AD by mechanisms that are distinct from modulation of Aβ or tau pathology. Further, whether plasma APOE, by influencing systemic metabolic pathways, can also possibly alter CNS function indirectly is not complete;y understood. Collectively, the available studies suggest that APOE may impact multiple signaling pathways and thus investigators have sought therapeutics that would disrupt pathological functions of APOE while preserving or enhancing beneficial functions. This review will highlight some of the therapeutic strategies that are currently being pursued to target APOE4 towards preventing or treating AD and we will discuss additional strategies that holds promise for the future.

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Generation of a set of isogenic, gene-edited iPSC lines homozygous for all main APOE variants and an APOE knock-out line

Cited by 36 publications

References 4 publications

APOE2: protective mechanism and therapeutic implications for Alzheimer’s disease

APOE2: protective mechanism and therapeutic implications for Alzheimer’s disease

APOE4 affects basal and NMDAR mediated protein synthesis in neurons by perturbing calcium homeostasis

Therapeutic approaches targeting Apolipoprotein E function in Alzheimer’s disease

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