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Cited by 498Alzheimer's disease research and treatmentsNeuroinflammation and Neurodegeneration MechanismsNeurogenesis and neuroplasticity mechanisms

APOE4 impairs myelination via cholesterol dysregulation in oligodendrocytes

Open access
Blanchard, Joel W., Akay, Leyla Anne, Davila-Velderrain, Jose, von Maydell, Djuna, Mathys, Hansruedi, Davidson, Shawn M., Effenberger, Audrey, Chen, Chih-Yu +20 more·2022·Nature
DOI 10.1038/s41586-022-05439-wPMC PMC9870060
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Abstract

APOE4 is the strongest genetic risk factor for Alzheimer's disease 1-3. However, the effects of APOE4 on the human brain are not fully understood, limiting opportunities to develop targeted therapeutics for individuals carrying APOE4 and other risk factors for Alzheimer's disease 4-8. Here, to gain more comprehensive insights into the impact of APOE4 on the human brain, we performed single-cell transcriptomics profiling of post-mortem human brains from APOE4 carriers compared with non-carriers. This revealed that APOE4 is associated with widespread gene expression changes across all cell types of the human brain. Consistent with the biological function of APOE 2-6, APOE4 significantly altered signalling pathways associated with cholesterol homeostasis and transport. Confirming these findings with histological and lipidomic analysis of the post-mortem human brain, induced pluripotent stem-cell-derived cells and targeted-replacement mice, we show that cholesterol is aberrantly deposited in oligodendrocytes-myelinating cells that are responsible for insulating and promoting the electrical activity of neurons. We show that altered cholesterol localization in the APOE4 brain coincides with reduced myelination. Pharmacologically facilitating cholesterol transport increases axonal myelination and improves learning and memory in APOE4 mice. We provide a single-cell atlas describing the transcriptional effects of APOE4 on the aging human brain and establish a functional link between APOE4, cholesterol, myelination and memory, offering therapeutic opportunities for Alzheimer's disease.

AI snapshot

Written from the paper’s full text.

TL;DR

APOE4 is associated with widespread transcriptomic changes in the aged human brain, including cholesterol dysregulation in oligodendrocytes and reduced myelination. Facilitating cholesterol transport with cyclodextrin increases myelination and improves learning and memory in APOE4/4 targeted-replacement mice.

Key findings

  • Single-nucleus RNA-sequencing of post-mortem prefrontal cortex from 32 subjects (12 APOE3/3, 12 APOE3/4, 8 APOE4/4) yielded 178,789 nuclei (164,741 after QC); APOE4 was associated with 486 candidate perturbed GO Biological Processes, including 404 cell-type-specific and 82 broadly dysregulated pathways.
  • APOE4 oligodendrocytes showed dose-dependent increases in cholesterol-related gene expression (4/4 > 3/4 > 3/3, p = 0.01) and altered cholesterol localization: in APOE4-carriers, Bodipy-cholesterol along neurofilaments decreased from 58% to 19% (p = 0.0106), with higher mean Bodipy-cholesterol intensity (p = 0.0220).
  • APOE4 was associated with reduced myelination: post-mortem APOE3/4 PFC had reduced Black Gold II area (p = 0.0031) and reduced MBP immunoreactivity (p = 0.0027 and p = 0.0294); APOE4/4 corpus callosum had higher g-ratios (p < 0.0001); APOE4/4-TR mice had fewer myelinated axons (p = 0.0265) and higher g-ratios (p < 0.0001).
  • Cyclodextrin treatment increased MBP colocalization with neurofilament in APOE4/4 iPSC co-cultures by 43% ± 5% (p = 0.0274), reduced g-ratio in APOE4/4-TR mice (p < 0.0001), increased MBP immunoreactivity (p = 0.0008), increased novel object preference (p = 0.0466), and decreased Puzzle Box latency (p = 0.02).

Methods

  • Post-mortem human prefrontal cortex from ROSMAP was profiled by 10x Genomics snRNA-seq; nuclei were clustered with ACTIONet, differential expression was modeled with NEBULA negative binomial mixed models, and pathway activity was analyzed with GSVA and Limma.
  • Lipidomic profiling used mass spectrometry on post-mortem corpus callosum (APOE4/4 n = 3, all female; APOE3/3 n = 3, all female) and a separate prefrontal cortex dataset of 314 individuals; isogenic APOE3/3 and APOE4/4 iPSC-derived oligodendroglia and iNeuron co-cultures, plus APOE targeted-replacement mice, were used for functional and histological validation.
  • Pharmacological experiments treated APOE4/4 iPSC-derived oligodendroglia with simvastatin, atorvastatin, or cyclodextrin, and treated APOE4/4-TR mice with subcutaneous cyclodextrin (2 g/kg twice weekly for eight weeks) or saline; myelination was assessed by imaging and TEM, and cognition by novel object recognition and Puzzle Box tasks.

Limitations

  • The paper states that additional well-powered lipidomic analyses designed to quantify cholesteryl esters in post-mortem tissue are needed; the corpus callosum lipidomic comparison had n = 3 per genotype and showed only a trend toward higher cholesteryl esters (p ≥ 0.7).
  • The paper states that genetic diversity may confound APOE4 effects observed in post-mortem human brain tissue, motivating the use of isogenic iPSC lines and APOE-TR mice; it also notes that covariates inherent to post-mortem human samples were controlled by quantifying myelin in inbred APOE-TR mice.
  • The paper states that power analysis to determine sample size was not performed and that experimenters were not blinded to drug injections or behavioral trials (TEM analysis was blinded); evident from the text, several human and mouse experiments used small samples, such as n = 3 per genotype for human TEM and Black Gold II staining and n = 4–5 mice per group for in vivo cyclodextrin histology/TEM.

Model-written summary. Check it against the paper before relying on it.

Integrity signals

Registry

Crossref · 10.1038/s41586-022-05439-w

Cites

52 of 52 references verified0 citation errors · 0 not checkable

Statistics

Nothing to recompute.

Claims the paper makes

32 of 43 cited sources checked · 10 supported · 0 flagged · accuracy not yet measured

  • Findingcitation
    § Author contributions

    APOE4 significantly alters signaling pathways associated with cholesterol homeostasis and transport.

    “Consistent with APOE’s biological function2–6, APOE4 significantly altered signaling pathways associated with cholesterol homeostasis and transport.”
    Quote in the paperSubstantial claim4 of 5 sources checked
    • cites [2] Corder, E. H. et al. Gene dose of apolipoprotein E type 4 allele and the risk of Alzheimer’s disease in late onset families. Science 261, 921–923 (1993). source
    • cites [3] Strittmatter, W. J. et al. Apolipoprotein E: high-avidity binding to beta-amyloid and increased frequency of type 4 allele in late-onset familial Alzheimer disease. Proc. Natl Acad. Sci. USA 90, 1977–1981 (1993). source
    • cites [4] Liu, C.-C. et al. ApoE4 accelerates early seeding of amyloid pathology. Neuron 96, 1024–1032 (2017). source
    • cites [5] Shi, Y. et al. ApoE4 markedly exacerbates tau-mediated neurodegeneration in a mouse model of tauopathy. Nature 549, 523–527 (2017). source

      The paper’s results discuss ApoE4-associated changes in inflammatory and normal-cell-function gene expression in tauopathy mice, but do not identify cholesterol homeostasis or transport pathways. Searches for “cholesterol homeostasis and transport,” “cholesterol,” and related lipid-pathway terms found no passage supporting the attributed claim. (LLM, conf=0.73)

      Experimental — accuracy not yet measured
    • cites [6] Castellano, J. M. et al. Human apoE isoforms differentially regulate brain amyloid-β peptide clearance. Sci. Transl. Med. 3, 89ra57 (2011). source

      The citation is attached to the background phrase about APOE’s biological function, not directly to the citing authors’ finding that APOE4 significantly altered signaling pathways. The source’s Results and Discussion discuss apoE lipidation and amyloid-β clearance, but the passages found for cholesterol homeostasis and transport do not address cholesterol-related signaling pathways. (LLM, conf=0.72)

      Experimental — accuracy not yet measured
  • Premisecitation
    § Introduction

    The APOE4 polymorphism alters the accessibility of the lipid-binding region, interfering with APOE's cholesterol and lipid transport capacity.

    “The APOE4 polymorphism alters the accessibility of the lipid-binding region, which interferes with its cholesterol and lipid transport capacity7.”
    Quote in the paperSubstantial claim1 of 1 sources checked
    • cites [7] Yamazaki, Y., Zhao, N., Caulfield, T. R., Liu, C.-C. & Bu, G. Apolipoprotein E and Alzheimer disease: pathobiology and targeting strategies. Nat. Rev. Neurol. 15, 501–518 (2019). source

      The Biology of APOE section says APOE polymorphisms alter structure and lipid binding, and the Lipid transport section says APOE4 is less efficient at transporting lipids to neurons and reports impaired cholesterol efflux in APOE*ε4 homozygotes. But the paper does not say APOE4 alters access to the lipid-binding region or that this causes the transport impairment. Its Biology of APOE section instead describes lipid binding as increasing access to the receptor-binding region in a model of APOE3; page numbers are unavailable in our copy. (LLM, conf=0.88)

      Experimental — accuracy not yet measured
  • Premisecitation
    § Introduction

    Previous single-cell analysis uncovered AD-associated oligodendrocyte subpopulations highlighting a role of myelin-related processes in AD pathogenesis.

    “Our previous single-cell analysis uncovered AD-associated oligodendrocyte subpopulations highlighting a role of myelin-related processes in AD pathogenesis12, consistent with reports of hypomyelination and white matter changes in AD23,24.”
    Quote in the paperNot a substantial claim1 of 1 sources checked
    • cites [12] Mathys, H. et al. Single-cell transcriptomic analysis of Alzheimer’s disease. Nature 570, 332–337 (2019). source
  • Findingcitation
    § Introduction

    Post-mortem oligodendrocytes had low but detectable APOE expression.

    “Post-mortem oligodendrocytes had low but detectable APOE expression, consistent with previous single-cell resolution reports25 (Extended Data Fig. 5k, l).”
    Quote in the paperSubstantial claim1 of 1 sources checked
    • cites [25] Jäkel, S. et al. Altered human oligodendrocyte heterogeneity in multiple sclerosis. Nature 566, 543–547 (2019). source

      The Methods (Human Donor tissue; page not given) identify the samples as post-mortem tissue, and the paper identifies APOE as an RNA marker of an oligodendroglial subcluster. It supports detectable APOE expression in that subcluster, but does not establish that expression was low across post-mortem oligodendrocytes generally. (LLM, conf=0.83)

      Experimental — accuracy not yet measured
  • Premisecitation
    § Discussion

    Infant APOE4-carriers exhibit alterations in myelin volume and cognitive maturation trajectories relative to APOE3/3-carriers.

    “Infant APOE4-carriers exhibit alterations in myelin volume and cognitive maturation trajectories relative to APOE3/3-carriers34,35.”
    Quote in the paperSubstantial claim2 of 2 sources checked
    • cites [34] Dean, D. C. 3rd et al. Brain differences in infants at differential genetic risk for late-onset Alzheimer disease: a cross-sectional imaging study. JAMA Neurol. 71, 11–22 (2014). source

      The Results (2- to 25-Month-Old and 2- to 6-Month-Old MWF Comparisons) report regional myelin water fraction differences between infant ε4 carriers and noncarriers; the Results (Associations Between Brain Imaging Measurements and Age) also report different age-related myelin trajectories. The Discussion says cognitive differences remain unresolved and calls for studies of cognitive performance starting in infancy. This source therefore does not establish altered cognitive maturation trajectories. (LLM, conf=0.90)

      Experimental — accuracy not yet measured
    • cites [35] Remer, J. et al. Longitudinal white matter and cognitive development in pediatric carriers of the apolipoprotein ε4 allele. Neuroimage 222, 117243 (2020). source

      The Background and Discussion report differences in white-matter myelin and cognitive-development trajectories in infants and young children with the APOE ε4 allele. But the study compares ε4 carriers with all non-carriers, not specifically ε3/3 children, and uses myelin water fraction as a surrogate for myelin content rather than measuring myelin volume. No page numbers are available in our copy. (LLM, conf=0.86)

      Experimental — accuracy not yet measured
  • Classified as probably not a substantial claim. 40
  • May combine two claims. 59
  • The quote only partly states this claim. 1

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Interpretationcitation
    § Introduction

    These APOE4 pathway effects are consistent with reported roles of APOE4 in inflammation, synaptic impairment, MAPK1 signaling and downstream APP transcription, and early endosomes.

    “All these effects are consistent with reported roles of APOE4 in inflammation, synaptic impairment, MAPK1 signaling and downstream APP transcription, and early endosomes7,18.”
    Quote in the paperNot a substantial claim2 of 2 sources checked
    • cites [7] Yamazaki, Y., Zhao, N., Caulfield, T. R., Liu, C.-C. & Bu, G. Apolipoprotein E and Alzheimer disease: pathobiology and targeting strategies. Nat. Rev. Neurol. 15, 501–518 (2019). source

      The Neuroinflammation and Synaptic function sections report APOE4-associated inflammatory activation and synaptic impairment. The Aβ production section reports greater APP transcription in human stem-cell-derived neurons exposed to APOE4, while noting conflicting mouse findings. The paper discusses intracellular endosomal compartments but does not identify early endosomes specifically, and the searched passages do not establish MAPK1 signaling as the pathway to APP transcription (pages unavailable in our copy). (LLM, conf=0.86)

      Experimental — accuracy not yet measured
    • cites [18] Huang, Y.-W. A., Zhou, B., Wernig, M. & Südhof, T. C. ApoE2, ApoE3, and ApoE4 differentially stimulate APP transcription and Aβ secretion. Cell 168, 427–441 (2017). source

      The Discussion (page unavailable) reports that ApoE4 is more potent than other ApoE isoforms in activating an ERK1/2 MAP-kinase pathway that stimulates APP transcription. The passages do not establish the other attributed roles—inflammation, synaptic impairment, or early endosomes—and the paper describes ERK1/2 rather than naming MAPK1 specifically. (LLM, conf=0.86)

      Experimental — accuracy not yet measured

  • Classified as probably not a substantial claim. May combine two claims.

    Premisecitation
    § Introduction

    APOE is a soluble factor expressed by many cell types.

    “Insight into APOE4-mediated pathogenesis is complicated by the fact that APOE is a soluble factor expressed by many cell types12,13.”
    Quote in the paperSubstantial claim1 of 2 sources checked
    • cites [12] Mathys, H. et al. Single-cell transcriptomic analysis of Alzheimer’s disease. Nature 570, 332–337 (2019). source

      The section on cell-type-specific alterations reports APOE expression changes in both microglia and astrocytes, supporting expression in multiple cell types. Our copy does not say that APOE is a soluble factor, so that part of the claim is unsupported. (LLM, conf=0.82)

      Experimental — accuracy not yet measured
    • cites [13] Xu, Q. et al. Profile and regulation of apolipoprotein E (ApoE) expression in the CNS in mice with targeting of green fluorescent protein gene to the ApoE locus. J. Neurosci. 26, 4985–4994 (2006). source

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Study design and conductcitation
    § Introduction

    Cell type annotations were supported by expression patterns of curated marker genes.

    “Annotations were supported by expression patterns of curated marker genes16,17 (Extended Data Fig. 1e; Supplementary Table S2).”
    Quote in the paperNot a substantial claim2 of 2 sources checked
    • cites [16] Wang, D. et al. Comprehensive functional genomic resource and integrative model for the human brain. Science 362, eaat8464 (2018). source

      In “Transcriptome analysis: Bulk and single cell” (page not provided), the paper describes brain cell-type profiles and expression signatures comprising marker genes, and reports that biomarker-gene expression for the same cell type correlated across two data formats. Our copy does not say the marker genes were curated or that their expression patterns were used to support the cell-type annotations. (LLM, conf=0.78)

      Experimental — accuracy not yet measured
    • cites [17] Franzén, O., Gan, L.-M. & Björkegren, J. L. M. PanglaoDB: a web server for exploration of mouse and human single-cell RNA sequencing data. Database 2019, baz046 (2019). source

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Premisecitation
    § Author contributions

    APOE4 is the strongest genetic risk factor for Alzheimer's disease.

    “APOE4 is the strongest genetic risk factor for Alzheimer’s disease (AD)1–3.”
    Quote in the paperNot a substantial claim2 of 3 sources checked
    • cites [1] Lambert, J. C. et al. Meta-analysis of 74,046 individuals identifies 11 new susceptibility loci for Alzheimer’s disease. Nat. Genet. 45, 1452–1458 (2013). source

      The paper states that the APOE gene is a major genetic risk factor for Alzheimer’s disease (Abstract; page not available), and its references identify the APOE type 4 allele as associated with risk (References; page not available). Our copy does not establish that APOE4, specifically, is the strongest genetic risk factor. (LLM, conf=0.83)

      Experimental — accuracy not yet measured
    • cites [2] Corder, E. H. et al. Gene dose of apolipoprotein E type 4 allele and the risk of Alzheimer’s disease in late onset families. Science 261, 921–923 (1993). source
    • cites [3] Strittmatter, W. J. et al. Apolipoprotein E: high-avidity binding to beta-amyloid and increased frequency of type 4 allele in late-onset familial Alzheimer disease. Proc. Natl Acad. Sci. USA 90, 1977–1981 (1993). source

      The paper reports a significant association between APOE-e4 and late-onset familial Alzheimer’s disease: its allele frequency was 0.50 in affected patients versus 0.16 in controls. It does not compare APOE4 with other genetic risk factors to establish that it is the strongest, or establish the claim for Alzheimer’s disease generally. (LLM, conf=0.87)

      Experimental — accuracy not yet measured

  • Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

    Premisecitation
    § Introduction

    APOE4 increases cholesterol sequestration in astrocytes leading to changes in the brain matrisome.

    “and increases cholesterol sequestration in astrocytes leading to changes in the brain matrisome11.”
    Quote in the paperNot a substantial claim1 of 1 sources checked
    • cites [11] Tcw, J. et al. Cholesterol and matrisome pathways dysregulated in astrocytes and microglia. Cell 185, 2213–2233 (2022). source

      The Results section, “Matrisome exacerbates lipid dysregulation in APOE4 astrocytes” (page unavailable), states that APOE4 drives lysosomal cholesterol sequestration in astrocytes and that matrisome dysregulation occurs. It does not say sequestration leads to matrisome changes; instead, it links those changes partly to astrocyte–neuron communication and says they worsen lipid dysregulation. (LLM, conf=0.88)

      Experimental — accuracy not yet measured

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Premisecitation
    § Introduction

    APOE4 promotes the accumulation of unsaturated triglycerides and lipid droplets in iPSC-derived astrocytes.

    “Recent in vitro studies discovered that APOE4 promotes the accumulation of unsaturated triglycerides and lipid droplets in iPSC-derived astrocytes10”
    Quote in the paperNot a substantial claim1 of 1 sources checked
    • cites [10] Sienski, G. et al. APOE4 disrupts intracellular lipid homeostasis in human iPSC-derived glia. Sci. Transl. Med. 13, eaaz4564 (2021). source

  • Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

    Premisecitation
    § Introduction

    APOE4 is associated with increased amyloid-β deposition, hyperphosphorylation and aggregation of tau, and accelerated cognitive decline.

    “APOE4 is associated with increased amyloid-β deposition, hyperphosphorylation and aggregation of tau, and accelerated cognitive decline4–7.”
    Quote in the paperNot a substantial claim4 of 4 sources checked
    • cites [4] Liu, C.-C. et al. ApoE4 accelerates early seeding of amyloid pathology. Neuron 96, 1024–1032 (2017). source

      The Introduction states that Aβ plaques are more abundant in APOE4 carriers, and the Discussion describes apoE4’s role in early amyloid plaque formation (page numbers unavailable). The passages do not link APOE4 to tau hyperphosphorylation or aggregation, or to accelerated cognitive decline; the Discussion mentions cognitive decline only as a later clinical event relative to Aβ deposition. (LLM, conf=0.88)

      Experimental — accuracy not yet measured
    • cites [5] Shi, Y. et al. ApoE4 markedly exacerbates tau-mediated neurodegeneration in a mouse model of tauopathy. Nature 549, 523–527 (2017). source
    • cites [6] Castellano, J. M. et al. Human apoE isoforms differentially regulate brain amyloid-β peptide clearance. Sci. Transl. Med. 3, 89ra57 (2011). source

      The Results section on Aβ and amyloid deposition reports greater deposition in apoE4 mice than in apoE3 or apoE2 mice; the human biomarker Results section also links APOE ε4 to greater amyloid burden. The paper does not establish the attributed association with tau hyperphosphorylation or aggregation, or accelerated cognitive decline. Its Introduction describes an earlier age of Alzheimer’s onset, which is not the same as a faster rate of cognitive decline. Page numbers were not available in our copy. (LLM, conf=0.88)

      Experimental — accuracy not yet measured
    • cites [7] Yamazaki, Y., Zhao, N., Caulfield, T. R., Liu, C.-C. & Bu, G. Apolipoprotein E and Alzheimer disease: pathobiology and targeting strategies. Nat. Rev. Neurol. 15, 501–518 (2019). source

      The ‘APOE4 and Aβ pathology’ section (page unavailable) reports greater Aβ deposition in APOE4 carriers. The ‘Tau, α-synuclein and TDP43 proteinopathies’ section (page unavailable) reports increased tau phosphorylation in experimental models and more tau aggregates in people with two APOE4 alleles when Aβ pathology is present. The copy links APOE4 to earlier symptom onset, but the passages found do not establish accelerated cognitive decline; the claim also omits the condition on the human tau-aggregation finding. (LLM, conf=0.84)

      Experimental — accuracy not yet measured

  • Classified as probably not a substantial claim.

    Premisecitation
    § Introduction

    The APOE genotype has a widespread impact on cell-autonomous and non-autonomous biological processes.

    “The APOE genotype has a widespread impact on cell-autonomous and non-autonomous biological processes14.”
    Quote in the paperSubstantial claim1 of 1 sources checked
    • cites [14] Lin, Y.-T. et al. APOE4 causes widespread molecular and cellular alterations associated with Alzheimer’s disease phenotypes in human iPSC-derived brain cell types. Neuron 98, 1141–1154 (2018). source

  • Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

    Premisecitation
    § Introduction

    Altered amyloid-β processing and DNA damage have been associated with APOE4 and Alzheimer's disease.

    “Altered amyloid-β processing and DNA damage have been associated with APOE4 and AD19–21.”
    Quote in the paperNot a substantial claim3 of 3 sources checked
    • cites [19] Zalocusky, K. A. et al. Neuronal ApoE upregulates MHC-I expression to drive selective neurodegeneration in Alzheimer’s disease. Nat. Neurosci. 24, 786–798 (2021). source

      The Abstract and Results report correlations between neuronal apoE expression and DNA-damage-and-repair pathways in an Alzheimer’s disease context, but do not establish that association specifically for APOE4. The passages do not support the other part of the claim, altered amyloid-β processing; the paper’s reported findings instead focus on MHC-I and Tau pathology. (LLM, conf=0.82)

      Experimental — accuracy not yet measured
    • cites [20] Thadathil, N. et al. DNA double-strand break accumulation in Alzheimer’s disease: evidence from experimental models and postmortem human brains. Mol. Neurobiol. 58, 118–131 (2021). source

      The supplied passages and the retrieved passages are labeled as a Parkinson’s disease paper, not the cited Alzheimer’s disease paper. Searches for APOE4 and altered amyloid-β processing did not yield passages from our copy of the cited paper, so the attributed association cannot be assessed. (LLM, conf=0.72)

      Experimental — accuracy not yet measured
    • cites [21] Ye, S. et al. Apolipoprotein (apo) E4 enhances amyloid beta peptide production in cultured neuronal cells: apoE structure as a potential therapeutic target. Proc. Natl Acad. Sci. USA 102, 18700–18705 (2005). source

      The Discussion (p. 18703) reports that apoE4 enhances amyloid-β production more than apoE3 and relates this finding to amyloid deposition in Alzheimer’s disease. The paper does not address DNA damage, so it supports only the amyloid-β part of the claim. (LLM, conf=0.88)

      Experimental — accuracy not yet measured

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Premisecitation
    § Introduction

    LPC and LPA are lipid classes known to promote brain inflammation and demyelination.

    “two lipid classes known to promote brain inflammation and demyelination (Extended Data Fig. 6; Supplementary Table S11)26.”
    Quote in the paperNot a substantial claim1 of 1 sources checked
    • cites [26] Law, S.-H. et al. An updated review of lysophosphatidylcholine metabolism in human diseases. Int. J. Mol. Sci. 20, 1149 (2019). source

      Section 1.3 reports that LPC induces demyelination in the central nervous system and inflammation in the brain microvasculature. Section 5.2 discusses LPA in inflammatory disorders and the central nervous system, but does not say that LPA promotes brain inflammation or demyelination. The claim is therefore supported for LPC, not for both lipid classes. (LLM, conf=0.88)

      Experimental — accuracy not yet measured

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Premisecitation
    § Introduction

    Accumulation of lipid droplets is a common hallmark of ER stress.

    “Accumulation of lipid droplets is a common hallmark of ER stress27,28.”
    Quote in the paperNot a substantial claim1 of 2 sources checked
    • cites [27] Ye, J. et al. ER stress induces cleavage of membrane-bound ATF6 by the same proteases that process SREBPs. Mol. Cell 6, 1355–1364 (2000). source
    • cites [28] Hapala, I., Marza, E. & Ferreira, T. Is fat so bad? Modulation of endoplasmic reticulum stress by lipid droplet formation. Biol. Cell 103, 271–285 (2011). source

  • Classified as probably not a substantial claim.

    Premisecitation
    § Introduction

    Cyclodextrin is a molecule known to reduce intracellular cholesterol accumulation in Niemann-Pick disease type C.

    “Cyclodextrin is a molecule known to reduce intracellular cholesterol accumulation in Niemann-Pick disease type C30.”
    Quote in the paperSubstantial claim1 of 1 sources checked
    • cites [30] Ottinger, E. A. et al. Collaborative development of 2-hydroxypropyl-β-cyclodextrin for the treatment of Niemann-Pick type C1 disease. Curr. Top. Med. Chem. 14, 330–339 (2014). source

  • May combine two claims.

    Premisecitation
    § Discussion

    Reduced myelin volume and integrity often occur prior to the onset of dementia and are predictive of future cognitive impairment.

    “Reduced myelin volume and integrity often occur prior to the onset of dementia and are predictive of future cognitive impairment23,24,33,36.”
    Quote in the paperSubstantial claim2 of 4 sources checked
    • cites [23] Brun, A. & Englund, E. A white matter disorder in dementia of the Alzheimer type: a pathoanatomical study. Ann. Neurol. 19, 253–262 (1986). source
    • cites [24] Bartzokis, G. Age-related myelin breakdown: a developmental model of cognitive decline and Alzheimer’s disease. Neurobiol. Aging 25, 5–18 (2004). source
    • cites [33] Dean, D. C. 3rd et al. Association of amyloid pathology with myelin alteration in preclinical Alzheimer disease. JAMA Neurol. 74, 41–49 (2017). source

      The Conclusions (page not provided) suggest that myelin alterations may be among the earliest features of Alzheimer disease, and the study examined asymptomatic adults. But the Discussion’s Limitations describe the analyses as cross-sectional and call for longitudinal studies; this paper does not show that reduced myelin predicts future cognitive impairment. It also does not establish how often the alterations occur before dementia. (LLM, conf=0.88)

      Experimental — accuracy not yet measured
    • cites [36] Gold, B. T., Powell, D. K., Andersen, A. H. & Smith, C. D. Alterations in multiple measures of white matter integrity in normal women at high risk for Alzheimer’s disease. Neuroimage 52, 1487–1494 (2010). source

      The Discussion (page unavailable) reports reduced white-matter integrity in cognitively normal women at high risk for Alzheimer’s disease and says the imaging pattern is consistent with early myelin loss. It does not establish reduced myelin volume or show that these changes predict future cognitive impairment; the authors say longitudinal follow-up is needed to test prediction. (LLM, conf=0.87)

      Experimental — accuracy not yet measured

  • Classified as probably not a substantial claim. The quote only partly states this claim.

    Study design and conductcitation
    § Methods

    Differential gene expression by NBMM modeled per-gene expression levels per cell type with random effects for within-individual and between-individual effects and fixed effects including APOE4, controlling for amyloid, nft, age_death, and batch.

    “Per-gene expression levels per cell type were modeled as a negative binomial model with random effects that model both within-individual and between-individual effects, as implemented in the R package NEBULA (version 1.2.1)40”
    Quote in the paperSubstantial claim1 of 1 sources checked
    • cites [40] He, L. et al. NEBULA is a fast negative binomial mixed model for differential or co-expression analysis of large-scale multi-subject single-cell data. Commun. Biol. 4, 629 (2021). source

  • May combine two claims.

    Findingown data
    § Author contributions

    Pharmacologically facilitating cholesterol transport increases axonal myelination and improves learning and memory in APOE4 mice.

    “Pharmacologically facilitating cholesterol transport increases axonal myelination and improves learning and memory in APOE4 mice.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data
    § Introduction

    Pharmacologically facilitating cholesterol transport improves myelination and learning and memory in aged APOE4/4 mice.

    “We establish that pharmacologically facilitating cholesterol transport improves myelination and learning and memory in aged APOE4/4 mice, thus providing new insight into the mechanistic connection between APOE4, myelination, and AD pathogenesis.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data
    § Introduction

    Associated pathways included amyloid-β and protein processing, lipid metabolism, and synaptic-related processes.

    “As expected, associated pathways included amyloid-β and protein processing, lipid metabolism, and synaptic-related processes.”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Findingown data
    § Introduction

    Decreased glycosyltransferase activity in APOE4 neurons and widespread alterations to amyloid-β metabolism genes in excitatory neurons, oligodendrocytes, and OPCs were identified.

    “We also identified decreased glycosyltransferase activity (e.g., ALG6, ALG3) in APOE4 neurons and widespread alterations to amyloid-β metabolism genes in excitatory neurons, oligodendrocytes, and OPCs (amyloid-β formation, e.g., BIN1, ROCK1; Fig. 1a).”
    Quote in the paperNot a substantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Findingown data
    § Introduction

    Cell type expression profiles were strongly correlated between individual donors.

    “Cell type expression profiles were strongly correlated between individual donors (average Pearson r=0.94 and r=0.65 for high and low-abundant cell types, respectively; Extended Data Fig. 1h, i; Supplementary Table S3).”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Findingown data
    § Introduction

    Chaperone-mediated protein folding was increased in most cell types, and ATF-mediated unfolded protein response was increased in neurons and oligodendrocytes.

    “Chaperone-mediated protein folding was increased in most cell types (e.g., DNAJB14, HSPA1A), and ATF-mediated unfolded protein response was increased in neurons and oligodendrocytes.”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Findingown data
    § Introduction

    In APOE4 excitatory neurons, ERK1/2 signaling and endosomal-related transport genes were upregulated.

    “In APOE4 excitatory neurons, we identified upregulation of ERK1/2 signaling (e.g., MAPK1, MAP2K1, and MAP2K2) and endosomal-related transport genes (e.g., RAB21 and SNX12) (Supplementary Table S4).”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Findingown data
    § Introduction

    APOE4 is associated with endoplasmic reticulum stress and reduced myelination.

    “and (2) is associated with endoplasmic reticulum stress and reduced myelination.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data
    § Introduction

    Punctate, perinuclear PLIN1 stained lipid droplets appeared in both APOE3/3 and APOE3/4 tissue; staining was more prominent around distinct nuclei in APOE4-carriers.

    “Punctate, perinuclear PLIN1 stained lipid droplets appeared in both APOE3/3 and APOE3/4 tissue; staining was more prominent around distinct nuclei in APOE4-carriers (Extended Data Fig. 4a).”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data
    § Introduction

    APOE4-carriers have decreased localization of cholesterol along neurofilaments and increased intracellular accumulation or storage of cholesterol compared to age-matched APOE3/3 individuals.

    “We find that APOE4-carriers have decreased localization of cholesterol along neurofilaments and increased intracellular accumulation or storage of cholesterol as compared to brains from age-matched APOE3/3 individuals.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data
    § Introduction

    APOE mRNA and protein were expressed at low to moderate levels in iPSC-derived oligodendroglia, with no significant difference between isogenic APOE3/3 and APOE4/4 (p = 0.2635).

    “APOE mRNA and protein were expressed at low to moderate levels in iPSC-derived oligodendroglia, with no significant (p = 0.2635) difference between isogenic APOE3/3 and APOE4/4 iPSC-derived oligodendroglia (Extended Data Fig. 5i, j).”
    Quote in the paperSubstantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Findingown data
    § Introduction

    PLIN1 immunoreactivity around OLIG2-positive nuclei in APOE3/4 human PFC from AD individuals exhibited a minimal but non-significant increase compared to APOE3/3 (p = 0.1521, n = 3 per genotype).

    “PLIN1 immunoreactivity around OLIG2-positive nuclei in APOE3/4 human PFC from AD individuals also exhibited a minimal but non-significant increase compared to APOE3/3 (p = 0.1521, n = 3 per genotype) (Extended Data Fig. 4d, e).”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Findingown data
    § Introduction

    Genes associated with cholesterol biosynthesis and droplet formation were upregulated, while cholesterol transport genes were downregulated.

    “Genes associated with cholesterol biosynthesis and droplet formation (DHCR24, LPIN2, IRS2, NR1H2, LBR, LPIN1, and MBTPS1) were upregulated, while cholesterol transport genes were downregulated (PCYT1B, SEC23A, PRKN, SCP2, and LPCAT3; Fig. 2c).”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data
    § Introduction

    APOE3/3 subjects with and without pathology exhibited the lowest cholesterol biosynthesis gene expression, and those with both APOE4 and AD pathology the largest.

    “APOE3/3 subjects with and without pathology exhibited the lowest cholesterol biosynthesis gene expression, and those with both APOE4 and AD pathology the largest (Extended Data Fig. 2k).”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data
    § Introduction

    Approximately 2% of Bodipy-cholesterol particles localized to the endosome, 18% to the lysosome, and 80% to the ER in APOE4/4 oligodendroglia.

    “Approximately ~2% of Bodipy-cholesterol particles localized to the endosome, ~18% to the lysosome, and ~80% to the ER”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data
    § Introduction

    PFC from APOE3/4 had a significant reduction in the area positive for Black Gold II staining, and axonal fasciculation was largely absent.

    “In contrast, PFC from APOE3/4 had a significant (p = 0.0031) reduction in the area positive for Black Gold II staining, and axonal fasciculation was largely absent”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data
    § Introduction

    88 lipid species were increased (adjusted p-value < 0.05) in APOE4/4 oligodendroglia compared to APOE3/3 controls, and only one species decreased.

    “Mass-spectrometry-based lipidomic profiling of APOE3/3 and APOE4/4 isogenic iPSC-derived oligodendroglia detected 88 lipid species increased (adjusted p-value < 0.05) in APOE4/4 oligodendroglia compared to APOE3/3 controls, and only one species decreased (Fig. 3a; Extended Data Fig. 6; Supplementary table S11).”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data
    § Introduction

    PFC white matter regions from APOE4-carriers exhibited significantly decreased MBP immunoreactivity compared to APOE3/3 non-carriers, with fewer MBP regions surrounding neurofilament-positive axons.

    “In contrast, PFC white matter regions from APOE4-carriers exhibited significantly (p = 0.0027, p = 0.0294) decreased MBP immunoreactivity as compared to APOE3/3 non-carriers, with fewer MBP regions surrounding neurofilament-positive axons (Fig. 4c and Extended Data Fig. 8b).”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data
    § Introduction

    The area within a 2 μm radius of OLIG2-positive nuclei exhibited significantly more Bodipy-cholesterol staining, with 77% of OLIG2-positive nuclei surrounded by intense Bodipy-cholesterol signal.

    “In contrast, the area within a 2 μm radius of OLIG2-positive nuclei exhibited significantly (p < 0.0001) more Bodipy-cholesterol staining, with 77% of OLIG2-positive nuclei surrounded by intense Bodipy-cholesterol signal, suggesting that cholesterol may aberrantly accumulate in APOE4 oligodendrocytes (Extended Data Fig. 4c).”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data
    § Introduction

    Cholesterol biosynthesis gene expression increased in individuals without AD pathology and showed a non-significant increase in individuals with AD.

    “Cholesterol biosynthesis gene expression increased in individuals without AD pathology (Extended Data Fig. 2e, f; lipid storage pathways also increased) and a non-significant increase in individuals with AD (Extended Data Fig. 2g), indicating an APOE4 effect independent of AD pathology.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data
    § Introduction

    Levels of MBP surrounding neurofilament in APOE4/4 oligodendroglia co-cultured with APOE3 iNeurons were similar to all-APOE4/4 co-cultures (p > 0.9999), with only ~20% of neurofilament-positive axons surrounded by MBP staining.

    “The levels of MBP surrounding neurofilament in APOE4/4 oligodendroglia co-cultured with APOE3 iNeurons were similar (p > 0.9999) to all-APOE4/4 co-cultures, with only ~20% of neurofilament-positive axons surrounded by MBP staining (Fig. 5b).”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data
    § Introduction

    Total distance traveled and time spent in the center of the home cage were similar in control (n = 12) and cyclodextrin-treated (n = 14) APOE4/4-TR mice (p = 0.1275 and p = 0.4478).

    “Total distance traveled and time spent in the center of the home cage (p = 0.1275 and p = 0.4478, respectively) was similar in control (n = 12) and cyclodextrin-treated (n = 14) APOE4/4-TR mice”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data
    § Introduction

    APOE4/4-TR mice had fewer myelinated axons (p = 0.0265, 140 average per field, n=3) and higher g-ratios compared to APOE3/3-TR mice (p < 0.0001).

    “APOE4/4-TR mice had fewer myelinated axons (p = 0.0265, 140 myelinated axons in average per field of view, n=3 mice), and higher g-ratios compared to APOE3/3-TR mice (p < 0.0001; Fig. 4e).”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data
    § Introduction

    MBP immunoreactivity and axonal localization in APOE−/− co-cultures were significantly lower than APOE3/3 co-cultures (p = 0.0005) and were similar to APOE4/4 co-cultures.

    “MBP immunoreactivity and axonal localization in the APOE−/− co-cultures were significantly (p = 0.0005) lower than APOE3/3 co-cultures and were similar to APOE4/4 co-cultures (Extended Data Fig. 9j).”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data
    § Introduction

    High-magnification imaging confirmed that cells surrounding neurofilaments were positive for MBP, and TEM showed axonal segments surrounded by electron-dense membrane rings at low frequency.

    “High-magnification imaging confirmed that the cells surrounding neurofilaments were positive for MBP (Extended Data Fig. 9e, Supplemental Video 1), and transmission electron microscopy (TEM) showed axonal segments surrounded by electron-dense membrane rings at a low frequency”
    Quote in the paperSubstantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Findingown data
    § Methods

    Lipid-related genesets with evidence of having an APOE4 effect (p-value < 0.05) are shown in Extended Fig. 2d.

    “Lipid-related genesets with evidence of having an APOE4 effect (p-value < 0.05) are shown in Extended Fig. 2d.”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Findingown data
    § Introduction

    Cyclodextrin-treated APOE4/4-TR mice exhibited a decrease in latency to enter the goal box (p-value = 0.02), while saline-treated APOE4/4-TR mice did not improve between trial days (p-value > 0.6).

    “Cyclodextrin-treated APOE4/4-TR mice exhibited a decrease in latency to enter the goal box (p-value = 0.02), while saline-treated APOE4/4-TR mice (control) did not improve between the trial days (p-value > 0.6; Fig. 6f).”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data
    § Introduction

    In cyclodextrin-treated APOE4/4 oligodendroglia, there was a significant reduction in the number of intracellular droplets of Bodipy-cholesterol and a significant reduction of neutral lipid droplets stained with Bodipy.

    “In cyclodextrin-treated APOE4/4 oligodendroglia, we observed a significant (p = 0.0092) reduction in the number of intracellular droplets of Bodipy-cholesterol, as well as a significant (p < 0.0001) reduction of neutral lipid droplets stained with Bodipy.”
    Quote in the paperSubstantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Findingreasoning
    § Methods

    This procedure resulted in prioritized candidate pathways for all major cell types.

    “This procedure resulted in prioritized candidate pathways for all major cell types.”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Findingown data
    § Introduction

    After two weeks in 3D co-cultures, MBP-positive oligodendroglia spread and engaged neurofilament-positive axons.

    “After two weeks in these 3D co-cultures, MBP-positive oligodendroglia spread and engaged neurofilament-positive axons (Extended Data Fig. 9b).”
    Quote in the paperSubstantial claim

  • Classified as probably not a substantial claim.

    Study design and conductown data
    § Introduction

    The differential pathway activity analysis controlled for sex, postmortem interval, amyloid, neurofibrillary tangles, and age at death.

    “We compared APOE3/4 and APOE4/4 vs APOE3/3 scores using a multivariate linear model controlling for sex, postmortem interval (PMI), amyloid, neurofibrillary tangles, and age at death.”
    Quote in the paperSubstantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Study design and conductown data
    § Introduction

    A total of 11 cell types were supported by gene marker expression and significant enrichment of marker sets.

    “A total of 11 cell types supported by gene marker expression and significant enrichment (FDR < 0.01, permutation test) of marker sets (Extended Data Fig. 1g) were annotated.”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Study design and conductown data
    § Introduction

    High-abundant cell types were well-represented across all donors, independent of AD diagnosis or genotypic background, with reproducible proportions in most cases.

    “High-abundant cell types were well-represented across all donors, independent of AD diagnosis or genotypic background, with reproducible proportions in most cases (Extended Data Fig. 1l, m).”
    Quote in the paperSubstantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

    Premisecitation
    § Introduction

    One copy of the APOE4 allele increases the risk of developing Alzheimer's disease 3 to 4 fold, while two copies increase it by 8 to 12 fold.

    “One copy of the APOE4 allele, which differs from the major allele (APOE3) by a single amino acid change from Cys112 to Arg112, increases the risk of developing AD 3 to 4 fold, while two copies increase it by 8 to 12 fold2.”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [2] Corder, E. H. et al. Gene dose of apolipoprotein E type 4 allele and the risk of Alzheimer’s disease in late onset families. Science 261, 921–923 (1993). source

  • May combine two claims.

    Study design and conductown data
    § Introduction

    The APOE3/3 and APOE3/4 subgroups were balanced by AD pathological diagnosis and gender.

    “The APOE3/3 and APOE3/4 subgroups were balanced by AD pathological diagnosis (n=6 each for positive and negative diagnoses of AD) and gender (3 male and 3 female each; Extended Data Fig. 1a; Supplementary Table S1).”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Study design and conductown data
    § Introduction

    There were no differences in age or post-mortem intervals between APOE3/3 and APOE3/4 groups and AD status.

    “All AD subjects presented high levels of both amyloid and neurofibrillary tangle (Tau) pathology, and there were no differences in age or post-mortem intervals between APOE3/3 and APOE3/4 groups and AD status (Extended Data Fig. 1b).”
    Quote in the paperSubstantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Interpretationown data
    § Introduction

    APOE4 may promote DNA damage in specific cell types.

    “DNA damage response was also altered in APOE4 microglia and OPCs, suggesting APOE4 may promote DNA damage in specific cell types.”
    Quote in the paperNot a substantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Premisecitation
    § Introduction

    Reports exist of hypomyelination and white matter changes in Alzheimer's disease.

    “consistent with reports of hypomyelination and white matter changes in AD23,24.”
    Quote in the paperNot a substantial claim0 of 2 sources checked
    • cites [23] Brun, A. & Englund, E. A white matter disorder in dementia of the Alzheimer type: a pathoanatomical study. Ann. Neurol. 19, 253–262 (1986). source
    • cites [24] Bartzokis, G. Age-related myelin breakdown: a developmental model of cognitive decline and Alzheimer’s disease. Neurobiol. Aging 25, 5–18 (2004). source

  • May combine two claims.

    Interpretationown data
    § Introduction

    These data support an association between APOE4 and brain cholesterol potentially indicating increased cholesterol storage in lipid droplets.

    “these further support an association between APOE4 and brain cholesterol potentially indicating increased cholesterol storage in lipid droplets (Supplementary Tables S7 and S8).”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Interpretationown data
    § Introduction

    iPSC-derived oligodendroglia are transcriptionally similar to ex vivo human oligodendrocytes and OPCs, providing a reliable proxy for investigating the effect of APOE4 on human oligodendrocytes.

    “This demonstrates that iPSC-derived oligodendroglia are transcriptionally similar to ex vivo human oligodendrocytes and OPCs, providing a reliable proxy for investigating the effect of APOE4 on human oligodendrocytes.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Interpretationown data
    § Introduction

    APOE4 is associated with increased cholesteryl esters and lipid droplets.

    “Together, these results demonstrate that APOE4 is associated with reduced localization of cholesterol in the plasma membrane of oligodendrocytes, increased cholesteryl esters and lipid droplets, and upregulation of ATF6-mediated ER stress response.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Study design and conductown data
    § Introduction

    65-81% of cells were immunoreactive for MBP and 69-82% were immunoreactive for PLP1 across cultures.

    “Following completion of the differentiation protocol, 65–81% of cells were immunoreactive for MBP and 69–82% were immunoreactive for PLP1 across cultures (Extended Data Fig. 5c).”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Interpretationown data
    § Introduction

    The increased Bodipy-cholesterol in APOE4 oligodendroglia was not entirely localized to lysosome, suggesting cholesterol may be accumulating in other organelles.

    “However, the increased bodipy-cholesterol in APOE4 oligodendroglia was not entirely localized to lysosome, suggesting cholesterol may be accumulating in other organelles.”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Premisecitation
    § Introduction

    Oligodendrocytes myelinate the central nervous system and the bioavailability of cholesterol is rate-limiting for the formation and maintenance of myelin.

    “Oligodendrocytes myelinate the central nervous system and the bioavailability of cholesterol is rate-limiting for the formation and maintenance of myelin29.”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [29] Saher, G., Quintes, S. & Nave, K.-A. Cholesterol: a novel regulatory role in myelin formation. Neuroscientist 17, 79–93 (2011). source

  • May combine two claims.

    Interpretationown data
    § Introduction

    ER stress and impaired cholesterol transport could have a central role in the observed intracellular cholesterol accumulation.

    “suggesting ER stress and impaired cholesterol transport could have a central role in the observed intracellular cholesterol accumulation.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Interpretationown data
    § Introduction

    Repeated cyclodextrin treatment did not affect general locomotion or induce anxiety-like behavior.

    “suggesting that repeated cyclodextrin treatment did not affect general locomotion or induce anxiety-like behavior (Extended Data Fig. 10c).”
    Quote in the paperSubstantial claim

  • Classified as probably not a substantial claim.

    Study design and conductown data
    § Methods

    Differential pathway activity analysis modeled pathway activity scores using a multivariate linear model including APOE4, amyloid, nft, age_death, msex, and pmi.

    “For each pathway-cell type combination, pathway activity scores were modeled using a multivariate linear model as follows: pathway activity ~ β0 * APOE4 + β1 * amyloid + β2 * nft + β3 * age_death + β4 * msex + β5 * pmi.”
    Quote in the paperSubstantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Study design and conductown data
    § Methods

    After QC filtering, the dataset included 17,915 genes profiled in 164,741 nuclei.

    “After applying QC filtering steps, the dataset included 17,915 genes profiled in 164,741 nuclei.”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Interpretationown data
    § Discussion

    These effects are not observed when targeting biosynthesis, suggesting APOE4 plays a role in cholesterol trafficking and increased biosynthesis might be a compensatory mechanism.

    “These effects are not observed when targeting biosynthesis, suggesting that APOE4 plays a role in cholesterol trafficking and that increased biosynthesis might be a compensatory mechanism.”
    Quote in the paperSubstantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Study design and conductreasoning
    § Methods

    A batch correction step considering sequencing batch as indicator vector was performed as part of the initial dimensionality reduction in all analyses.

    “To avoid biased cell mixing due to independent sequencing batches or potential technical artifacts, a batch correction step considering sequencing batch as indicator vector was performed as part of the initial dimensionality reduction in all analyses.”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Interpretationown data
    § Discussion

    Cholesterol gene expression and localization are dysregulated in oligodendrocytes, leading to impaired myelination.

    “Cholesterol gene expression and localization are dysregulated in oligodendrocytes, leading to impaired myelination.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Interpretationown data
    § Introduction

    Cyclodextrin reduced both intracellular cholesterol and accumulation of neutral lipids such as triacylglycerides.

    “This suggests that cyclodextrin reduced both intracellular cholesterol and accumulation of neutral lipids such as triacylglycerides (Fig. 6b and Extended Data Fig. 10a).”
    Quote in the paperSubstantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

    Study design and conductnone given
    § Methods

    Experimenters were not blinded to drug injections and when executing behavioral trials.

    “Experimenters were not blinded to drug injections and when executing behavioral trials.”
    Quote in the paperNot a substantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing), S4 (based).

    Study design and conductnone given
    § Methods

    Sample size was determined from previous publications from our lab.

    “Sample size was determined from previous publications from our lab.”
    Quote in the paperNot a substantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Study design and conductown data
    § Methods

    Pseudo-bulk differential gene expression fitted individual-level normalized gene expression averages with a linear model including APOE4, amyloid, nft, age_death, msex, and pmi.

    “Individual-level normalized gene expression averages were fitted with the following linear model: gene expression ~ β0*APOE4 + β1*amyloid + β2*nft + β3*age_death + β4*msex + β5*pmi.”
    Quote in the paperNot a substantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Study design and conductnone given
    § Methods

    Data analysis of the TEM images was done manually under blinded conditions.

    “Data analysis of the TEM images was done manually under blinded conditions.”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Interpretationnone given
    § Introduction

    APOE4 expression in oligodendrocytes promotes both cholesterol accumulation and impaired myelination.

    “APOE4 expression in oligodendrocytes promotes both cholesterol accumulation and impaired myelination.”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Interpretationown data
    § Introduction

    APOE4/4 neurons do not impair the ability of APOE3/3 oligodendroglia to produce MBP or engage neuronal axons.

    “suggesting that APOE4/4 neurons do not impair the ability of APOE3/3 oligodendroglia to produce MBP or engage neuronal axons (Fig. 5b).”
    Quote in the paperSubstantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Study design and conductnone given
    § Methods

    At least four images for each animal were analyzed.

    “At least four images for each animal were analyzed.”
    Quote in the paperNot a substantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Study design and conductown data
    § Methods

    4 out of 17 peaks passed this quality control cutoff (Supplementary Table S8).

    “4 out of 17 peaks passed this quality control cutoff (Supplementary Table S8).”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Interpretationown data
    § Introduction

    Myelination defects are observed in APOE4/4 and APOE−/− oligodendrocytes and can be rescued with the addition of APOE3 protein.

    “Therefore, myelination defects are observed in APOE4/4 and APOE−/− oligodendrocytes and can be rescued with the addition of APOE3 protein.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Interpretationown data
    § Introduction

    APOE4/4 oligodendrocytes were producing less MBP and engaging less with neuronal processes than isogenic APOE3/3 co-cultures.

    “suggesting that APOE4/4 oligodendrocytes were producing less MBP and engaging less with neuronal processes than isogenic APOE3/3 co-cultures (Fig. 5a).”
    Quote in the paperSubstantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing), S4 (based).

    Study design and conductnone given
    § Methods

    Activity scores computed this way accurately recovered cell type signatures not recovered by equally-sized randomly sampled genesets.

    “Activity scores computed this way accurately recovered cell type signatures not recovered by equally-sized randomly sampled genesets (data not shown)”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Interpretationown data
    § Discussion

    Facilitating cholesterol transport increases myelination and improves cognitive function in APOE4.

    “We demonstrate facilitating cholesterol transport increases myelination and improves cognitive function in APOE4.”
    Quote in the paperSubstantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

    Study design and conductown data
    § Methods

    For APOE3/3 and APOE3/4-carriers, subjects were balanced between sexes and matched for age.

    “For APOE3/3 and APOE3/4-carriers, subjects were balanced between sexes (12 each) and matched for age (median=84.6 for APOE3/3 and 85.2 for APOE3/4).”
    Quote in the paperNot a substantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Study design and conductnone given
    § Methods

    Power analysis to determine sample size was not performed.

    “Power analysis to determine sample size was not performed.”
    Quote in the paperNot a substantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Study design and conductnone given
    § Methods

    Pathway activity scores were computed using GSVA with parameters mx.diff=TRUE, kcdf=c(Gaussian), min.sz=5, max.sz=150.

    “The following parameters were used to evaluate the GSVA function: mx.diff=TRUE, kcdf=c(“Gaussian”), min.sz=5, max.sz=150.”
    Quote in the paperNot a substantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Study design and conductnone given
    § Methods

    Treatment and genotype groups were matched on age as much as possible.

    “Treatment and genotype groups were matched on age as much as possible.”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Interpretationown data
    § Discussion

    Pharmacologically promoting cholesterol efflux increases myelination in vitro and in vivo, and improves cognition in APOE4/4-TR mice.

    “We observed that pharmacologically promoting cholesterol efflux increases myelination in vitro and in vivo, and improves cognition in APOE4/4-TR mice.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Interpretationown data
    § Introduction

    Corpus callosum from aged APOE4/4 individuals contains fewer myelinated axons and thinner overall myelin sheaths compared to APOE3/3 individuals.

    “suggesting that corpus callosum from aged APOE4/4 individuals contains fewer myelinated axons and thinner overall myelin sheaths compared to APOE3/3 individuals (Fig. 4d).”
    Quote in the paperSubstantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

    Study design and conductnone given
    § Methods

    Both males and females were included in the experiments and sex was balanced across treatment and genotype conditions.

    “Both males and females were included in the experiments and sex was balanced across treatment and genotype conditions.”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Interpretationown data
    § Discussion

    These results establish a causal link between cholesterol dysregulation and myelination in APOE4-carriers, which may influence the onset and severity of cognitive decline in AD.

    “These results establish a causal link between cholesterol dysregulation and myelination in APOE4-carriers, which may influence the onset and severity of cognitive decline in AD.”
    Quote in the paperSubstantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

    Study design and conductnone given
    § Methods

    Outlier cells with less than 500 or more than 10,000 genes detected were excluded, and only genes detected in at least 10 cells were considered.

    “Outlier cells with less than 500 or more than 10,000 genes detected were excluded, and only genes detected in at least 10 cells were considered.”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Interpretationown data
    § Introduction

    Cyclodextrin treatment increased the number of myelinated axons and thicker myelin sheaths.

    “indicating an increased number of myelinated axons and thicker myelin sheaths (Fig 6d).”
    Quote in the paperSubstantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Study design and conductnone given
    § Methods

    Analysis was performed blinded.

    “Analysis was performed blinded.”
    Quote in the paperNot a substantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Study design and conductnone given
    § Methods

    Each experiment consisted of at least 3 mice per a treatment group.

    “Each experiment consisted of at least 3 mice per a treatment group.”
    Quote in the paperNot a substantial claim
90 of 90

  • The paper relies on this without support.

    The mechanisms by which APOE4 mediates its effects on amyloid-β deposition, tau pathology, and cognitive decline are not fully understood.

    “However, the mechanisms by which APOE4 mediates these effects are not fully understood.”

  • The paper relies on this without support.

    How cholesterol and lipid abnormalities impair learning and memory in the APOE4 brain, and whether such effects can be mitigated, remains unclear.

    “Despite these longstanding associations, how cholesterol and lipid abnormalities impair learning and memory in the APOE4 brain, and whether such effects can be mitigated, remains unclear.”

  • The paper relies on this without support.

    APOE is a lipid and cholesterol transporter.

    “APOE is a lipid and cholesterol transporter.”

  • The paper relies on this without support.

    Genetic diversity may confound the effects of APOE4 observed on human post-mortem brain tissue.

    “Genetic diversity may confound the effects of APOE4 observed on human post-mortem brain tissue.”

  • The paper relies on this without support.

    APOE-TR mice recapitulate human APOE4 effects on myelination sufficiently for conclusions about human APOE4.

    “To control for covariates inherent to post-mortem human samples, we also quantified myelin levels in inbred APOE-TR mice (9 months old, male and female mice; n = 4 per genotype).”

  • The paper relies on this without support.

    Isogenic iPSC-derived oligodendroglia and iNeuron co-cultures model human oligodendrocyte myelination sufficiently for conclusions about human APOE4.

    “To investigate this, we employed an in vitro myelination assay using isogenic iPSC-derived oligodendroglia co-cultured with NGN2-induced neurons derived from the same iPSCs (iNeurons) in a three-dimensional extracellular matrix that allows cells to interact with each other and self-assemble into tissue-like structures (Extended Data Fig. 9a).”

  • The paper relies on this without support.

    MBP immunoreactivity and g-ratio are valid measures of myelin levels.

    “To quantify relative myelin levels per axon, we calculated the g-ratio (the inner axonal diameter divided by the outer diameter with myelin sheath) for 150 neurons per condition.”

  • The paper relies on this without support.

    Bodipy-cholesterol and filipin staining reflect intracellular cholesterol localization and storage.

    “To further characterize APOE4-mediated differences in cholesterol storage, we stained isogenic APOE3/3 and APOE4/4 oligodendroglia with the cholesterol dye filipin, Bodipy-cholesterol, and lipid-droplet associated PLIN1.”

  • The paper relies on this without support.

    Cyclodextrin’s beneficial effects on myelination and cognition in APOE4 mice are mediated by facilitating cholesterol transport.

    “Collectively, these data demonstrate that pharmacological enhancement of cholesterol transport can improve APOE4 myelination in vitro and in vivo.”
9 of 9

Claims other papers make about this one

Each is checked against this paper’s own text: does it say what the citing paper says it does?

  • APOE4 is known to disrupt oligodendrocyte differentiation and contribute to myelin breakdown in mouse models without any Alzheimer's disease pathology.

    “APOE4 is known to disrupt oligodendrocyte differentiation and contribute to myelin breakdown in mouse models without any AD pathology. 26 , 27 , 28”

    From Apolipoprotein ε4 exacerbates white matter impairment in a mouse model of Aβ amyloidosis by decreasing actively myelinating oligodendrocytes (2025) · cites this paper as [28]

    The Introduction reports fewer myelinated axons and thinner myelin sheaths in APOE4 targeted-replacement mice than in APOE3 mice. It does not establish that APOE4 disrupted oligodendrocyte differentiation or caused breakdown of existing myelin in those mice. The finding in individuals without AD pathology concerns human myelin-associated gene expression, not a documented absence of AD pathology in the mice. (LLM, conf=0.86)

    Experimental — accuracy not yet measured

  • Previous studies using humanized APOE3 and APOE4 mouse models without amyloid pathology revealed inconsistent findings regarding myelination.

    “Previous studies using humanized APOE3 and APOE4 mouse models without amyloid pathology revealed inconsistent findings regarding myelination. One study found no significant difference in myelination, while others observed reduced myelination from electron microscopy images. 27 , 28”

    From Apolipoprotein ε4 exacerbates white matter impairment in a mouse model of Aβ amyloidosis by decreasing actively myelinating oligodendrocytes (2025) · cites this paper as [28]

    The Introduction reports that electron microscopy found fewer myelinated axons and thinner myelin sheaths in humanized APOE4/4 mice than in APOE3/3 mice. The Methods identify these as mice with human APOE replacing the mouse gene. Our copy does not report a mouse study finding no significant myelination difference, so it does not establish the claimed inconsistency across studies. (LLM, conf=0.86)

    Experimental — accuracy not yet measured

  • APOE4-associated glial LD accumulation modulates neuron function and brain memory.

    “and the APOE4-associated glial LD accumulation modulates neuron function and brain memory17–19.”

    From PICALM Alzheimer’s risk allele causes aberrant lipid droplets in microglia (2025) · cites this paper as [17]

    The Introduction (page unavailable) reports increased lipid droplets in APOE4 oligodendroglia and reduced droplets, improved myelination, and improved learning and memory after treatment in APOE4 mice. The Discussion links cholesterol dysregulation to impaired myelination and cognition, but the paper does not establish that lipid-droplet accumulation itself modulates neuronal function or memory; the intervention also changes cholesterol transport. (LLM, conf=0.84)

    Experimental — accuracy not yet measured

  • Elevated CE levels have been reported in AD brains and AD models.

    “Elevated CE levels have been reported in AD brains as well as AD models12,14,15.”

    From Decreased lipidated ApoE‐receptor interactions confer protection against pathogenicity of ApoE and its lipid cargoes in lysosomes (2025) · cites this paper as [12]

    The Introduction reports significantly increased cholesteryl ester species in APOE4/4 iPSC-derived oligodendroglia, supporting the AD-model portion. In human brain tissue, it reports only a nonsignificant trend in a small APOE4/4 comparison; its separate significant finding was in people without AD pathology. It therefore does not establish elevated CE levels in AD brains generally. (LLM, conf=0.83)

    Experimental — accuracy not yet measured

  • CE is upregulated in AD patient brain and various AD models, and its downregulation is protective in various AD models.

    “based on its upregulation in AD patient brain and various AD models, along with the fact that its downregulation is protective in various AD models10,12,15,64.”

    From Decreased lipidated ApoE‐receptor interactions confer protection against pathogenicity of ApoE and its lipid cargoes in lysosomes (2025) · cites this paper as [12]

    The Introduction (page unavailable) reports increased cholesteryl esters in APOE4-derived oligodendroglia and evidence of an increase in APOE4-carrier brain tissue, but it does not establish a general increase in AD patient brains or various AD models. It also reports that cyclodextrin reduced cholesterol droplets and improved myelination and cognition in APOE4 models; it does not show that downregulating cholesteryl esters was protective across various AD models. (LLM, conf=0.87)

    Experimental — accuracy not yet measured

Self-check

Percentages, sums and reported differences re-computed from the paper’s own numbers.

Each contradiction between two passages listed here was confirmed by an independent check.

  • Mouse age range excludes the two-month-old mice studied

    Minor

    The Methods state that mice in all mouse-model experiments were 6 to 9 months old, but the results report a TEM experiment on two-month-old mice.

1 of 1

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Authors
Blanchard, J. W., Akay, L. A., Davila-Velderrain, J., von Maydell, D., Mathys, H., Davidson, S. M., Effenberger, A., Chen, C.-Y., Maner-Smith, K., Hajjar, I., Ortlund, E. A., Bula, M., Agbas, E., Ng, A., Jiang, X., Kahn, M., Blanco-Duque, C., Lavoie, N., Liu, L., Reyes, R., Lin, Y.-T., Ko, T., R’Bibo, L., Ralvenius, W. T., Bennett, D. A., Cam, H. P., Kellis, M., & Tsai, L.-H.
Affiliations
  • Massachusetts Institute of Technology (US)
  • MIT Computer Science and Artificial Intelligence Laboratory (US)
  • University of Pittsburgh (US)
  • Princeton University (US)
  • Emory University (US)
  • Icahn School of Medicine at Mount Sinai (US)
  • Rush University Medical Center (US)
  • Broad Institute (US)
Corresponding author
  • Kellis, Manolis
  • Tsai, Li-Huei
ORCID
Venue
Nature
Publisher
Springer Science and Business Media LLC
Published
2022-11-16
Volume
611
Issue
7937
Pages
769-779
ISSN
  • 0028-0836 (ISSN-L)
  • 1476-4687
DOI
10.1038/s41586-022-05439-w
PMC
PMC9870060
OpenAlex
W4309507742
Cited by
493
References
48
Citations by year
2026: 81 · 2025: 165 · 2024: 128 · 2023: 109 · 2022: 7 · 2012: 1
FWCI
53.36
Type
journal article
Language
en
Update notices
correction (2024-12-04)
Open access
Licence
Locations
  • Nature (publishedVersion)
  • PubMed (publishedVersion)
  • PubMed Central (submittedVersion)
  • RePEc: Research Papers in Economics (submittedVersion)
  • DSpace@MIT (Massachusetts Institute of Technology) (submittedVersion, cc-by-nc-sa)
Funders
  • National Institute on Aging (P30 AG072975, RF1AG054012, RF1 AG051633, U01AG061356, U01AG046152, RF1 AG057470, R01 AG058002, R01 AG017917, K24 AG062786, RF1 AG062377, P30AG010161, R01-AG015819)
  • National Human Genome Research Institute (U54-HG008097)
  • National Institute of Neurological Disorders and Stroke (UH3 NS115064, U01 NS110453, R01NS114239, UG3 NS115064)
Topics
  • Health Sciences › Medicine › Physiology › Alzheimer's disease research and treatments
  • Life Sciences › Neuroscience › Neurology › Neuroinflammation and Neurodegeneration Mechanisms
  • Life Sciences › Neuroscience › Developmental Neuroscience › Neurogenesis and neuroplasticity mechanisms
Keywords
APOE ε4, Alzheimer's disease, myelination, oligodendrocytes, cholesterol homeostasis, cholesterol transport, single-cell transcriptomics, postmortem brain, brain aging, learning and memory, axonal myelination, lipidomics, induced pluripotent stem cell-derived cells, targeted replacement mice
MeSH terms
Aging, Alzheimer Disease, Animals, Autopsy, Biological Transport, Brain, Cholesterol, Heterozygote, Homeostasis, Humans, Memory, Myelin Sheath, Nerve Fibers, Myelinated, Neurons, Oligodendroglia, Gene Expression Profiling, Mice, Apolipoprotein E4, Induced Pluripotent Stem Cells, Single-Cell Analysis

About these checks

Registry, retraction, Cites and statistics results are computed, not judged, with one exception: a printed reference that no index lists is read once by a model before it counts as a citation error. Claims are extracted by a model and kept only when their quote appears verbatim in the text; each cited source is then checked, with a model’s help, for whether it says what the claim relies on. Those verdicts can be wrong — read the claim’s quote and the cited source before relying on one.

Claims read by deepseek-v4.1-flash under definition version 3 (extract-v3) on 2026-10-11.

Classified by jev-1.13-20260917 (gold-guideline-v0.2) on 2026-10-11.

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Statistics and arithmetic are recomputed, not judged.

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