APOE traffics to astrocyte lipid droplets and modulates triglyceride saturation and droplet size

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

Astrocytes store neuron-derived lipids in lipid droplets, which protect against neurotoxicity.

“Other studies demonstrated that lipid peroxides formed in neurons under oxidative or excitotoxic stress can be transported by APOE-lipoproteins to astrocytes; astrocytes store these neuron-derived lipids in LDs, which protect against neurotoxicity and may play a role in detoxifying these deleterious lipid species (Walker et al., 2006; Liu et al., 2015, 2017; Ioannou et al., 2019; Moulton et al., 2”

• cites Walker et al., 2006 sourceNot checked — no full text of the source
• cites Liu et al., 2015 sourceDisputed

  The Results sections report lipid droplets accumulating in glia after neuronal mitochondrial defects, but do not establish that astrocytes store lipids transported from neurons. More importantly, the Results report that reducing glial lipid-droplet accumulation delays neurodegeneration—the opposite of the claimed protection against neurotoxicity. The indexed passages have no page numbers. (LLM, conf=0.88)

  Checked by a reasoning model — may be wrong.

  Experimental — accuracy not yet measured
• cites Ioannou et al., 2019 sourceNot checked — no full text of the source

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

APOE4 iPSCs exhibit defects in endocytosis.

“APOE4 iPSCs also exhibit defects in endocytosis (Narayan et al., 2020).”

• cites Narayan et al., 2020 sourcePartly supported

  The Results section (‘APOE4 disrupts endocytosis in iPSC-derived astrocytes’) reports reduced early endocytosis in APOE4 astrocytes compared with APOE3 astrocytes. The tested cells were derived from iPSCs; the cited passages do not establish an endocytic defect in undifferentiated APOE4 iPSCs themselves. (LLM, conf=0.87)

  Experimental — accuracy not yet measured

May combine two claims.

Blocking fatty acid desaturation through inhibition of stearoyl-CoA desaturase 1 or supplementing cells with choline reduces lipid droplet accumulation in APOE4 iPSCs.

“Blocking fatty acid desaturation through inhibition of stearoyl-CoA desaturase 1, an enzyme elevated in Alzheimer's disease patient brains, or supplementing cells with choline reduced LD accumulation in APOE4 iPSCs (Astarita et al., 2011; Sienski et al., 2021).”

• cites Astarita et al., 2011 sourceSupported
• cites Sienski et al., 2021 sourcePartly supported

  The Results sections on lipid desaturation and choline supplementation report that SCD1 inhibition and CDP-choline treatment each reduced lipid droplet numbers. The experiments used APOE4-expressing human iPSC-derived astrocytes, not undifferentiated APOE4 iPSCs as the claim states. (LLM, conf=0.86)

  Experimental — accuracy not yet measured

Classified as probably not a substantial claim.

Proximity ligation identified APOE as a protein on the cytoplasmic face of lipid droplets in Huh7 hepatocarcinoma cells.

“A proximity ligation strategy identified APOE as a protein on the cytoplasmic face of LDs in Huh7 hepatocarcinoma cells (Bersuker et al., 2018).”

• cites Bersuker et al., 2018 sourcePartly supported

  The Results section, “An Integrated U2OS and Huh7 High-Confidence LD Proteome” (page unavailable), reports using APEX2 proximity biotinylation to identify lipid-droplet proteins in Huh7 cells. Neither that passage nor searches for APOE and apolipoprotein E identify APOE as one of those proteins or establish its location on the cytoplasmic face. The paper describes proximity labeling, not proximity ligation. (LLM, conf=0.85)

  Experimental — accuracy not yet measured

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

APOE was identified in subsequent studies of the lipid droplet proteome in mouse liver and THP-1 macrophages.

“APOE was identified in subsequent studies of the LD proteome in mouse liver and THP-1 macrophages (Krahmer et al., 2018; Mejhert et al., 2020).”

• cites Krahmer et al., 2018 sourcePartly supported

  The Results and Discussion (pages unavailable) describe a mouse-liver lipid-droplet proteome, supporting the identification of Krahmer et al. as a study of that proteome. The Results also report increased levels of several apolipoproteins, but the copy does not name APOE or identify it as a lipid-droplet protein. The THP-1 macrophage portion appears to be attributed to the other citation, not this paper. (LLM, conf=0.77)

  Experimental — accuracy not yet measured
• cites Mejhert et al., 2020 sourcePartly supported

  The Results section, “Identification of LD Proteins in Human THP-1 Macrophages” (page unavailable), reports an LD-enriched proteomic analysis of THP-1 cells. The passages name several identified proteins but do not name APOE, so our copy does not establish that this study identified APOE in the LD proteome. (LLM, conf=0.77)

  Experimental — accuracy not yet measured

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

After secretion, APOE-coated lipoproteins are lipidated via reverse cholesterol transport from the plasma membrane mediated by ABC transporters, primarily ABCA1.

“After secretion, APOE-coated lipoproteins are lipidated via reverse cholesterol transport from the plasma membrane mediated by ABC transporters, primarily ABCA1 (Wahrle et al., 2004).”

• cites Wahrle et al., 2004 sourcePartly supported

  The Discussion (page unavailable in our copy) says ABCA1 is required for normal lipidation of astrocyte-secreted apoE and is consistent with a role in transporting cellular cholesterol onto lipoproteins. It explicitly leaves the location unresolved: the function may occur in the astrocyte secretory pathway or extracellularly. It therefore does not establish that lipidation occurs after secretion through reverse cholesterol transport from the plasma membrane. (LLM, conf=0.88)

  Experimental — accuracy not yet measured

May combine two claims.

Multiple transcriptomic studies of APOE4-expressing iPSC-derived astrocytes and microglia demonstrate alterations in expression of lipid metabolic genes and defects in cholesterol trafficking and metabolism.

“Multiple transcriptomic studies of APOE4-expressing iPSC-derived astrocytes and microglia demonstrate alterations in the expression of lipid metabolic genes as well as defects in cholesterol trafficking and metabolism (Lin et al., 2018; de Leeuw et al., 2022; Tcw et al., 2022).”

• cites Lin et al., 2018 sourcePartly supported

  The Results section on APOE4 astrocytes reports altered expression of lipid-metabolism and transport genes and increased intracellular and secreted cholesterol. The microglia Results section reports transcriptomic changes, but the paper describes lipid-metabolism abnormalities in microglia as only potential; it does not demonstrate cholesterol-trafficking or metabolism defects in those cells. (LLM, conf=0.87)

  Experimental — accuracy not yet measured
• cites de Leeuw et al., 2022 sourcePartly supported

  The Discussion section, “Lipid and cholesterol metabolism and the endolysosomal system” (page unavailable), reports altered lipid-metabolism pathways and reduced cholesterol efflux in APOE4 iPSC-derived astrocytes, along with increased cholesterol localization in lysosomes. But this paper describes proteomic analysis, not a transcriptomic study of lipid metabolic gene expression, and it does not study microglia. (LLM, conf=0.88)

  Experimental — accuracy not yet measured
• cites Tcw et al., 2022 sourceSupported

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

The cleaved C-terminus of APOE can exist in the cytoplasm and associate with mitochondria in neurons.

“Studies on the trafficking of APOE have also suggested that the cleaved C-terminus of APOE can exist in the cytoplasm and associate with mitochondria in neurons (Chang et al., 2005).”

• cites Chang et al., 2005 sourcePartly supported

  The Results report that apoE4(1–272) fragments escaped the secretory pathway and were observed at mitochondria in Neuro-2a cells; the Discussion describes their entry into the cytosol. The paper studies C-terminally truncated apoE4, however, not an isolated cleaved C-terminus. That distinction limits support for the claim as worded. (LLM, conf=0.82)

  Experimental — accuracy not yet measured

Classified as probably not a substantial claim.

Cytoplasmic lipid droplets are pivotal components of astrocyte lipid homeostasis.

“Cytoplasmic lipid droplets (LDs) are pivotal components of astrocyte lipid homeostasis (Ralhan et al., 2021).”

• cites Ralhan et al., 2021 sourceSupported

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

Lipid peroxides formed in neurons under oxidative or excitotoxic stress can be transported by APOE-lipoproteins to astrocytes.

“Other studies demonstrated that lipid peroxides formed in neurons under oxidative or excitotoxic stress can be transported by APOE-lipoproteins to astrocytes; astrocytes store these neuron-derived lipids in LDs, which protect against neurotoxicity and may play a role in detoxifying these deleterious lipid species (Walker et al., 2006; Liu et al., 2015, 2017; Ioannou et al., 2019; Moulton et al., 2”

• cites Walker et al., 2006 sourceNot checked — no full text of the source
• cites Liu et al., 2015 sourcePartly supported

  The Discussion (page not provided) links neuronal mitochondrial defects and elevated ROS to lipid synthesis in neurons and lipid droplets in glia. The Results section on lipid peroxidation (page not provided) reports peroxidated lipids in this setting. Neither passage shows that lipid peroxides form in neurons and are transported to astrocytes by APOE-lipoproteins. (LLM, conf=0.88)

  Experimental — accuracy not yet measured
• cites Ioannou et al., 2019 sourceNot checked — no full text of the source

May combine two claims.

Mouse astrocytes expressing human APOE4 have defects in lipid droplet metabolism as well as in fatty acid uptake and oxidation.

“Mouse astrocytes expressing human APOE4 have defects in LD metabolism as well as in fatty acid uptake and oxidation (Farmer et al., 2019; Qi et al., 2021).”

• cites Farmer et al., 2019 sourceSupported
• cites Qi et al., 2021Not matched

  Couldn’t be matched to a paper in Crossref, so it can’t be checked.

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

APOE and APOCIII were identified on nuclear lipid droplets in HepG2 cells, and APOAV has been reported on the surface of cytoplasmic lipid droplets in adipocytes.

“APOE and APOCIII were identified on nuclear LDs in HepG2 cells, and APOAV has previously been reported on the surface of cytoplasmic LDs in adipocytes (Shu et al., 2010; Gao et al., 2012; Sołtysik et al., 2019).”

• cites Shu et al., 2010 sourcePartly supported

  The Introduction and Results §3.3 report apoA-V associated with cytosolic lipid droplets in transfected hepatoma cells and mouse liver. They do not place it on droplet surfaces in adipocytes, and the paper does not report APOE or APOCIII on nuclear lipid droplets in HepG2 cells. (LLM, conf=0.86)

  Experimental — accuracy not yet measured
• cites Gao et al., 2012 sourcePartly supported

  The Discussion (page unavailable) reports apoA-V on cytosolic lipid droplets in hepatocyte-derived cells, supporting that part of the claim. It does not place apoA-V on droplets in adipocytes or report APOE and APOCIII on nuclear lipid droplets in HepG2 cells. (LLM, conf=0.88)

  Experimental — accuracy not yet measured
• cites Sołtysik et al., 2019 sourcePartly supported

  The Results section (‘NR-lumenal LDs are converted to nucleoplasmic LDs,’ Fig. 3a) reports ApoE and ApoCIII on some nuclear lipid droplets, but its figure caption identifies the cells as Huh7, not HepG2. The APOAV finding in adipocytes is not reported in this paper and appears to be covered by the other citations in the sentence. (LLM, conf=0.90)

  Experimental — accuracy not yet measured

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

The ER membrane complex (EMC) is involved in the topological mediation of integral ER proteins.

“The ER membrane complex (EMC) is involved in the topological mediation of integral ER proteins (Hegde 2022).”

• cites Hegde 2022 sourceSupported

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

APOV and APOCIII have been shown to go to lipid droplets.

“APOV and APOCIII have been shown to go to LDs (Shu et al., 2010; Gao et al., 2012; Sołtysik et al., 2019).”

• cites Shu et al., 2010 sourceSupported
• cites Gao et al., 2012 sourcePartly supported

  The Discussion (page unavailable) says apoA-V accrues on cytosolic lipid droplets, supporting the apoA-V part of the sentence. The paper does not address apoC-III localization, so it cannot support that part of the claim. (LLM, conf=0.86)

  Experimental — accuracy not yet measured
• cites Sołtysik et al., 2019 sourcePartly supported

  The Results section (‘NR-lumenal LDs are converted to nucleoplasmic LDs’; page unavailable) reports that ApoCIII binds lumenal lipid droplets and was occasionally found in nucleoplasmic lipid droplets. It reports this for ApoE, not APOV, so the APOV part of the claim is not supported by our copy. (LLM, conf=0.88)

  Experimental — accuracy not yet measured

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

The N-terminal domain of APOE consists of four amphipathic alpha helices that form a bundle in solution.

“The N-terminal domain consists of four amphipathic alpha helices that form a bundle in solution (Chen et al., 2021).”

• cites Chen et al., 2021 sourcePartly supported

  The Historical Progress in Understanding ApoE Structure section (page unavailable) describes the N-terminal domain as a four-helix bundle and discusses its structure before lipid association. Our copy does not establish that all four N-terminal helices are amphipathic or explicitly say that the bundle forms in solution; its explicit discussion of amphipathic helices concerns the C-terminal domain. (LLM, conf=0.83)

  Experimental — accuracy not yet measured

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

A recent study identified both early and late ERTOLD pathways for protein trafficking from the ER to lipid droplets.

“A recent study of the mechanism of protein trafficking from the ER to LDs identified both “early” and “late” ERTOLD pathways (Song et al., 2022).”

• cites Song et al., 2022 sourceSupported

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

The presence of lipid-laden microglia correlates with an impaired response to stroke in aged mice.

“The presence of these lipid-laden microglia correlates with an impaired response to stroke in aged mice (Arbaizar-Rovirosa et al., 2023).”

• cites Arbaizar-Rovirosa et al., 2023 sourceSupported

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

EMC has been implicated in topological decision-making of ER hairpin proteins that target lipid droplets.

“EMC has also been implicated in topological decision-making of ER hairpin proteins that target LDs (Leznicki et al., 2022).”

• cites Leznicki et al., 2022 sourcePartly supported

  The Results section (‘Co-translational insertion of LD membrane proteins relies on the EMC’; page unavailable) implicates EMC in inserting lipid-droplet proteins into the ER with a transient membrane-spanning orientation. The Discussion says EMC may not discriminate between proteins that retain that orientation and those that later form hairpins, and leaves the mechanism of that transition unresolved. Thus, the paper supports EMC involvement in an initial topological state, but not a demonstrated role in deciding the final hairpin topology. (LLM, conf=0.83)

  Experimental — accuracy not yet measured

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

Microglia near amyloid beta plaques accumulate lipid droplets in vivo.

“More recent work has also shown that microglia near amyloid beta plaques accumulate LDs in vivo (Haney et al., 2023, Preprint; Prakash et al., 2023, Preprint).”

• cites Haney et al., 2023, Preprint sourcePartly supported

  In “Intracellular lipid accumulation is linked to AD pathology” (page unavailable), the paper reports lipid-positive cells near amyloid-beta plaques and, separately, lipid-droplet-positive microglia in mouse AD models. It suggests that the lipid-containing cells near plaques could be microglia, but says their cell type was not confirmed. The claim states that identification more definitively than this source supports. (LLM, conf=0.88)

  Experimental — accuracy not yet measured
• cites Prakash et al., 2023, Preprint sourceSupported

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

Lipid droplets rich in cholesterol esters drive the accumulation of phosphorylated tau in iPSC-derived neurons.

“LDs rich in cholesterol esters drive the accumulation of phosphorylated tau in iPSC-derived neurons (van der Kant et al., 2019).”

• cites van der Kant et al., 2019 sourcePartly supported

  The Results section, “The Effect of Statins on pTau Is Mediated by Cholesteryl Esters” (page unavailable), reports that cholesteryl esters regulate phosphorylated tau levels in human iPSC-derived familial Alzheimer’s neurons. The Discussion, “Pathways that Mediate the Effect of CE on pTau and Aβ” (page unavailable), identifies lipid droplets as cholesteryl-ester storage sites, but does not establish that the droplets themselves drive tau accumulation. (LLM, conf=0.87)

  Experimental — accuracy not yet measured

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

Proteins with uncleaved signal anchors can adopt orientations in which their N-terminus faces the cytoplasm or the ER lumen, a process regulated by EMC.

“Proteins with uncleaved signal anchors can adopt orientations in which their N-terminus faces the cytoplasm or the ER lumen, a process regulated by EMC (Wu and Hegde 2023).”

• cites Wu and Hegde 2023 sourceSupported

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

Instances of ischemia are a major risk factor for Alzheimer's disease.

“Instances of ischemia are a major risk factor for Alzheimer's disease, and increased sensitivity to hypoxic insult may play a role in driving pathology in APOE4 (Elman-Shina and Efrati 2022).”

• cites Elman-Shina and Efrati 2022 sourceSupported

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

FRAP experiments were analyzed as described in Day et al. (2012).

“FRAP experiments were analyzed as described in Day et al. (2012).”

• cites Day et al. (2012) sourceSupported

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

Aged microglia accumulate lipid droplets and adopt a proinflammatory state.

“Aged microglia accumulate LDs and adopt a proinflammatory state (Marschallinger et al., 2020).”

• cites Marschallinger et al., 2020 sourceSupported

Classified as probably not a substantial claim.

In the absence of APOE, higher surface tension would make lowering the surface area to volume ratio of lipid droplets via Oswalt ripening more thermodynamically favorable.

“In the absence of APOE, higher surface tension would make lowering the surface area to volume ratio of LDs via Oswalt ripening more thermodynamically favorable (Thiam et al., 2013).”

• cites Thiam et al., 2013 sourcePartly supported

  In “Mechanisms and Consequences of Emulsion Destabilization” (page unavailable), the paper says ripening minimizes interfacial surface energy and that greater surface tension increases the Laplace-pressure difference driving Ostwald ripening. It does not say that absence of APOE raises lipid-droplet surface tension; it also cautions that conventional Ostwald ripening is difficult to envisage between lipid droplets because their core lipids are nearly insoluble in water. (LLM, conf=0.86)

  Experimental — accuracy not yet measured

May combine two claims.

The C-terminal domain of APOE bound to LDs with or without a signal peptide, although its binding to LDs was less efficient than the full-length protein.

“The C-terminal domain of APOE bound to LDs with or without a signal peptide, although its binding to LDs was less efficient than the full-length protein (Fig. 4, C and D).”

May combine two claims.

Only segments of APOE rings colocalized with the ER at a contact site, while other segments localized to the LD in the absence of ER.

“By contrast, only segments of APOE rings colocalized with the ER at a contact site, while other segments localized to the LD in the absence of ER.”

May combine two claims.

The N-terminal domain Δss was soluble in the cytoplasm but did not exhibit LD surface enrichment.

“The N-terminal domain Δss was soluble in the cytoplasm but did not exhibit LD surface enrichment (Fig. 4, C and D).”

May combine two claims.

At baseline, TRAE3-H astrocytes contain few lipid droplets and APOE localizes largely to the Golgi apparatus.

“At their baseline state, TRAE3-H cells contained few LDs, and APOE localized largely to the Golgi apparatus, as expected (Fig. 1, A–C).”

May combine two claims.

Linoleic acid and arachidonic acid, like oleic acid, induce lipid droplet formation and cause APOE to traffic to lipid droplets in TRAE3-H cells.

“Like OA (18:1), LA and ARA also induced LD formation and caused APOE to traffic to LDs.”

May combine two claims.

In the absence of oleic acid, APOE3-mEm localizes exclusively to the secretory pathway and never coats lipid droplets, matching endogenous localization.

“In the absence of OA, APOE3-mEm localized exclusively to the secretory pathway and never coated LDs, matching the endogenous localization pattern.”

May combine two claims.

TRAE3-H cells at baseline exhibit few lipid droplets and no lipid-droplet-associated APOE.

“This is in contrast to the TRAE3-H cells, which, at baseline, exhibit few LDs and no LD-associated APOE (Fig. 1, D and E).”

May combine two claims.

Neither DBeQ nor cycloheximide affected OA-induced LD biogenesis.

“Neither drug affected OA-induced LD biogenesis (Fig. S3, D and G).”

May combine two claims.

In both iAstros and HMC3 cells, APOE localization to lipid droplets correlates with lipid droplet abundance; cells with APOE-coating lipid droplets contain an abundance of lipid droplets, while cells with APOE in the secretory pathway have few or no lipid droplets.

“In both cell types, APOE localization to LDs correlated with LD abundance; cells with APOE-coating LDs contained an abundance of LDs, while cells with APOE in the secretory pathway had few or no LDs (Fig. S2, C–F).”

May combine two claims.

APOE in the secretory pathway is Emerald but not FLAG positive, indicating the signal peptide was cleaved in the luminal pool.

“By contrast, APOE in the secretory pathway was Emerald but not FLAG positive, indicating the signal peptide was cleaved in the luminal pool.”

May combine two claims.

Full-length APOE Δss localized to the cytoplasm and was enriched on the surface of LDs.

“Full-length APOE Δss localized to the cytoplasm and was enriched on the surface of LDs (Fig. 4, C and D).”

May combine two claims.

During OA pulse, APOE had both a significantly higher rate constant of recovery and mobile fraction than after pulse-chase.

“During the OA pulse, APOE had both a significantly higher rate constant of recovery and mobile fraction (Fig. 3, E and F).”

May combine two claims.

Loss of APOE or expression of APOE4 causes the accumulation of polyunsaturated triglyceride molecules within lipid droplets.

“LD-associated APOE is required during lipogenesis for maintaining LD size, while the loss of APOE, or expression of APOE4, causes the accumulation of polyunsaturated TG molecules within LDs.”

• Also stated in: § Discussion · plain

  “APOE4 promotes the accumulation of unsaturated triglycerides in LDs.”

May combine two claims.

When treated with EtOH, both APOE3 and APOE4 cells exhibit minimal lipid peroxidation with no significant difference between genotypes.

“When treated with EtOH, both genotypes exhibited minimal lipid peroxidation with no significant difference between E3 and E4.”

May combine two claims.

After the OA pulse, APOE knockdown cells had larger but fewer LDs.

“After the OA pulse, we observed larger but fewer LDs in APOE knockdown cells.”

May combine two claims.

After OA pulse, E4 LDs were larger and fewer than E3 LDs.

“After OA pulse, E4 LDs were larger and fewer than E3 LDs, yet the total LD area remained the same, similar to the knockdown phenotype (Fig. 7, A–D).”

May combine two claims.

After an OA pulse, there is a pronounced increase in the total abundance of triglyceride (TG) and diglyceride (DG), and a subsequent decrease after the chase.

“As expected, we observed a pronounced increase in the total abundance of triglyceride (TG) and diglyceride (DG) after an OA pulse, and a subsequent decrease in TG and DG after the chase (Fig. 5, F and G).”

May combine two claims.

After OA pulse-chase, E4 cells had a greater total LD area and larger LDs than E3 cells.

“However, after OA pulse-chase, E4 cells had a greater total LD area and larger LDs than E3, but no difference in LD number.”

May combine two claims.

There were no significant differences in total LD area or LD number between TRAE3-H and TRAE4-H astrocytes at baseline.

“There were no significant differences in total LD area or LD number between TRAE3-H and TRAE4-H astrocytes at baseline, although E4 LDs were slightly larger (Fig. 7, A–D).”

May combine two claims.

Global depletion of APOE via siRNA abrogates both LD-associated APOE and APOE in the secretory pathway, which is still observed in a subset of cells upon OA treatment.

“However, global depletion of APOE via siRNA abrogates both LD-associated APOE and APOE in the secretory pathway, which is still observed in a subset of cells upon OA treatment (Fig. 1 C).”

May combine two claims.

APOE depletion and APOE4 expression both cause a smaller number of large lipid droplets to form upon OA loading.

“APOE depletion and APOE4 expression both caused a smaller number of large LDs to form upon OA loading.”

May combine two claims.

The C-terminal domain is necessary for binding of APOE to LDs, but inefficient for proper targeting as observed with the full-length protein.

“We conclude that the C-terminal domain is necessary for binding of APOE to LDs, but inefficient for proper targeting as observed with the full-length protein.”

• AssumptionRests on an assumption: Deleting the signal peptide prevents ER translocation and causes APOE to be present only in the cytoplasm.

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

Western blotting confirmed the expression of each truncation construct at the correct molecular weight.

“Western blotting confirmed the expression of each truncation construct at the correct molecular weight (Fig. 4 B).”

May combine two claims.

Nascent LDs that form in response to OA treatment remain attached to the ER by membrane bridges immediately following biogenesis, which would allow APOE molecules on the ER to exchange with the LD pool.

“Nascent LDs that form in response to OA treatment remain attached to the ER by membrane bridges immediately following biogenesis, which would allow APOE molecules on the ER to exchange with the LD pool.”

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

siRNA-mediated knockdown demonstrated the specificity of the APOE antibody.

“siRNA-mediated knockdown demonstrated the specificity of the APOE antibody (Fig. S1, A–C).”

May combine two claims.

If OA is washed out in a pulse-chase experiment, nascent LDs mature and detach from the ER.

“However, if OA is washed out in a pulse-chase experiment, then these nascent LDs mature and detach from the ER.”

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

iPSC-derived APOE4 astrocytes accumulate lipid droplets rich in highly unsaturated triglycerides.

“Another study demonstrated that iPSC-derived APOE4 astrocytes accumulate LDs rich in highly unsaturated TGs.”

May combine two claims.

APOE targets the cytoplasmic side of the ER before moving onto lipid droplets at membrane bridges between the ER and lipid droplets.

“APOE is trafficked to LDs by avoiding translocation into the ER, targeting the cytoplasmic side of the ER before moving onto LDs at membrane bridges between the ER and LDs.”

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

The N-terminal domain of APOE forms a soluble four-helix bundle and contains a patch of basic residues required for APOE binding to lipoprotein receptors.

“Its N-terminal domain forms a soluble four-helix bundle and contains a patch of basic residues required for APOE binding to lipoprotein receptors; the C-terminal domain is an amphipathic alpha-helix that mediates binding of APOE to the surface of lipoproteins (Westerlund and Weisgraber, 1993).”

• cites Westerlund and Weisgraber, 1993 sourceNot checked — no full text of the source

May combine two claims.

Oleic acid stimulates global APOE upregulation, with the subset of cells that route APOE to lipid droplets accounting for increased intracellular APOE, while the subset lacking lipid-droplet-associated APOE accounts for the increase in APOE secretion.

“Our interpretation of these data is that OA stimulates global APOE upregulation, with the subset of cells that route APOE to LDs accounting for increased intracellular APOE, while the subset of cells which lack LD-associated APOE account for the increase in APOE secretion.”

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

APOE on lipoproteins binds to cell-surface lipoprotein receptors, including LDLR and LRP1, triggering lipoprotein uptake via receptor-mediated endocytosis.

“APOE on lipoproteins binds to cell-surface lipoprotein receptors, including LDLR and LRP1, triggering lipoprotein uptake via receptor-mediated endocytosis (Bu, 2009).”

• cites Bu, 2009Not matched

  Couldn’t be matched to a paper in Crossref, so it can’t be checked.

May combine two claims.

Lipid homeostatic defects in APOE4 sensitize astrocytes to stress, which could promote a reactive, proinflammatory state that contributes to Alzheimer's pathogenesis.

“Our results contribute to growing evidence that lipid homeostatic defects in E4 sensitize astrocytes to stress, which could promote a reactive, proinflammatory state that contributes to Alzheimer’s pathogenesis.”

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

Astrocytes coordinate many aspects of brain lipid homeostasis.

“Astrocytes coordinate many aspects of brain lipid homeostasis (Pfrieger and Ungerer, 2011).”

• cites Pfrieger and Ungerer, 2011 sourceNot checked — no full text of the source

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

The APOE antibody used for immunofluorescence and immunogold experiments is the same one validated by siRNA knockdown.

“The antibody used to probe for endogenous APOE is the same one used for both the immunofluorescence and immunogold experiments.”

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

Nascent APOE lipoproteins are assembled in the lumen of the ER prior to secretion.

“Nascent APOE lipoproteins are assembled in the lumen of endoplasmic reticulum (ER) prior to secretion, although the mechanism for this stage of biogenesis is unknown (Windham and Cohen, 2023).”

• cites Windham and Cohen, 2023 sourceNot checked — no full text of the source

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

The C-terminal domain of APOE is an amphipathic alpha-helix that mediates binding of APOE to the surface of lipoproteins.

“Its N-terminal domain forms a soluble four-helix bundle and contains a patch of basic residues required for APOE binding to lipoprotein receptors; the C-terminal domain is an amphipathic alpha-helix that mediates binding of APOE to the surface of lipoproteins (Westerlund and Weisgraber, 1993).”

• cites Westerlund and Weisgraber, 1993 sourceNot checked — no full text of the source

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

APOE has two domains separated by a short hinge region.

“APOE has two domains separated by a short hinge region.”

• cites Westerlund and Weisgraber, 1993 sourceNot checked — no full text of the source

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

Astrocyte-derived lipoproteins supply key lipids, predominantly cholesterol and phospholipids, to neurons.

“Astrocyte-derived lipoproteins supply key lipids, predominately cholesterol and phospholipids, to neurons for building their high surface area membranes and cholesterol-rich lipid nanodomains at synapses (van Deijk et al., 2017).”

• cites van Deijk et al., 2017 sourceNot checked — no full text of the source

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

APOE is a key protein mediator of brain lipid homeostasis, expressed primarily by astrocytes and microglia as well as neurons under stress.

“A key protein mediator of brain lipid homeostasis is apolipoprotein E (APOE), a 34-kD secreted protein expressed primarily by astrocytes and microglia as well as neurons under stress (Xu et al., 2006).”

• cites Xu et al., 2006Not matched

  Couldn’t be matched to a paper in Crossref, so it can’t be checked.

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

Individuals possessing the APOE4 variant are significantly more likely to develop late-onset Alzheimer's disease than those homozygous for the APOE3 variant in a dose-dependent manner.

“The APOE4 variant is a cysteine to arginine substitution at residue 112, with individuals possessing the APOE4 variant significantly more likely to develop late-onset Alzheimer’s disease than those homozygous for the APOE3 variant in a dose-dependent manner (Corder et al., 1993).”

• cites Corder et al., 1993 sourceNot checked — no full text of the source

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

For lipidomics experiments, two independent biological replicates were performed, and within each biological replicate, three separate plates were collected for each condition and considered as technical replicates.

“For lipidomics experiments, two independent biological replicates were performed. Within each biological replicate, three separate plates were collected for each condition and considered as technical replicates.”

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

Ischemia is correlated with both increased lipid peroxidation and neutral lipid accumulation in the brain.

“Ischemia is correlated with both increased lipid peroxidation and neutral lipid accumulation in the brain (Yoshida et al., 1982; Gasparovic et al., 2001; Ioannou et al., 2019).”

• cites Yoshida et al., 1982 sourceNot checked — no full text of the source
• cites Gasparovic et al., 2001 sourceNot checked — no full text of the source
• cites Ioannou et al., 2019 sourceNot checked — no full text of the source

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

Cells were confirmed free of mycoplasma and monitored at regular intervals using a Mycoplasma PCR Detection Kit.

“Cells were confirmed free of mycoplasma and monitored at regular intervals using a Mycoplasma PCR Detection Kit (G238; ABM).”

May combine two claims.

The APOE4 phenotype is in the same direction as APOE knockdown, yet more severe.

“With respect to LDs, the APOE4 phenotype is in the same direction as APOE knockdown, yet, more severe.”

May combine two claims.

Western blot confirmed the depletion of endogenous APOE and expression of both HA-tagged rescue constructs.

“Western blot confirmed the depletion of endogenous APOE and expression of both HA-tagged rescue constructs (Fig. 6 B).”

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

The innate immune protein cathelicidin reroutes to lipid droplets in response to OA loading or lipopolysaccharide treatment, retaining its N-terminal signal peptide.

“One study demonstrated that the innate immune protein cathelicidin reroutes to LDs in response to OA loading or lipopolysaccharide treatment, retaining its N-terminal signal peptide (Bosch et al., 2020).”

• cites Bosch et al., 2020 sourceNot checked — no full text of the source

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

Three to five wells per condition were performed per independent experiment.

“Three to five wells per condition were performed per independent experiment”

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

Parts of APOB's nascent polypeptide chain become exposed to the solution during in vitro translocation.

“Another apolipoprotein, APOB, possesses pause transfer sequences that cause it to stall during in vitro translocation, and parts of its nascent polypeptide chain become exposed to the solution (Chuck et al., 1990; Kivlen et al., 1997).”

• cites Chuck et al., 1990 sourceNot checked — no full text of the source
• cites Kivlen et al., 1997 sourceNot checked — no full text of the source

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

Expression of astrocyte markers ALDH1L1 and GFAP was tested at day 14 after induction.

“Expression of astrocyte markers was tested at day 14 after induction, ALDH1L1 (1:500, ab190298; Abcam) and GFAP (1:500, MA5-12023; Thermo Fisher Scientific).”

May combine two claims.

APOE knockdown causes defects in the formation and growth of LDs.

“The distribution of lipids in fewer, larger LDs after an OA pulse suggests APOE knockdown causes defects in the formation and growth of LDs.”

Classified as probably not a substantial claim.

The Rosner-Glynn-Lee variation of the Wilcoxon rank-sum test for clustered data was used to assess statistical significance between groups from data collected from multiple independent experiments performed at different times, treating each independent experiment as a cluster and accounting for variability both within and between independent experimental trials.

“The Rosner–Glynn–Lee variation of the Wilcoxon rank-sum test for clustered data was applied to assess statistical significance between groups from data collected from multiple independent experiments performed at different times. This test treats data from each independent experiment as a “cluster” and accounts for variability both within and between independent experimental trials (Rosner et al.,”

• cites Rosner et al., 2006 sourceNot checked — no full text of the source

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

P values were calculated via the Wilcoxon rank-sum test and Bonferonni-corrected for multiple comparisons.

“P values were calculated via the Wilcoxon rank-sum test and Bonferonni-corrected for multiple comparisons.”

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

Three independent biological replicates were conducted for the ELISA.

“Three independent biological replicates were conducted.”

May combine two claims.

The bright BODIPY C11 signal of lipid droplets was easily segmented from the rest of the cell, allowing measurement of peroxidation specifically in lipid droplets.

“The bright BODIPY C11 signal of LDs was easily segmented from the rest of the cell, allowing for the measurement of peroxidation specifically in LDs.”

Classified as probably not a substantial claim.

Optimization experiments determined 30 μM digitonin was the lowest concentration that showed full loss of cytoplasmic mEmerald signal in most cells after 1 min.

“Optimization experiments determined 30 µM digitonin was the lowest concentration that showed full loss of cytoplasmic mEmerald signal in most cells after 1 min, and this concentration was used in subsequent experiments.”

Probably not a substantial claim: may not pass S3 (load-bearing). The quote only partly states this claim.

Data for peroxidation quantification were collected and pooled from three independent experiments.

“Data were collected and pooled from three independent experiments.”

May combine two claims.

While APOE depletion does not significantly alter the total amount of cellular TG, it does result in the selective enrichment of polyunsaturated TG species.

“Thus, while APOE depletion does not significantly alter the total amount of cellular TG, it does result in the selective enrichment of polyunsaturated TG species.”

• AssumptionRests on an assumption: One mechanism that affects LD size is their composition.

May combine two claims.

Although the same total amount of neutral lipids is present in APOE knockdown cells, LD biogenesis is altered, causing neutral lipids to concentrate in a smaller number of larger LDs.

“This suggests that, although the same total amount of neutral lipids is present in these cells, LD biogenesis is altered, causing neutral lipids to concentrate in a smaller number of larger LDs.”

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

APOB possesses pause transfer sequences that cause it to stall during in vitro translocation.

“Another apolipoprotein, APOB, possesses pause transfer sequences that cause it to stall during in vitro translocation, and parts of its nascent polypeptide chain become exposed to the solution (Chuck et al., 1990; Kivlen et al., 1997).”

• cites Chuck et al., 1990 sourceNot checked — no full text of the source
• cites Kivlen et al., 1997 sourceNot checked — no full text of the source

May combine two claims.

Like APOE-depleted cells, E4 cells form a smaller number of large LDs that contain more unsaturated triglycerides.

“Like APOE-depleted cells, E4 cells form a smaller number of large LDs that contain more unsaturated triglycerides.”

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

BODIPY C11 contains an 11-carbon fatty acid moiety that allows it to incorporate into lipid membranes.

“BODIPY C11 contains an 11-carbon fatty acid moiety that allows it to incorporate into lipid membranes (Drummen et al., 2002).”

• cites Drummen et al., 2002 sourceNot checked — no full text of the source

May combine two claims.

During LD biogenesis, neutral lipids are partitioned into a smaller number of larger LDs in E4 cells, similar to the phenotype observed upon APOE knockdown.

“This suggests that during LD biogenesis, neutral lipids are partitioned into a smaller number of larger LDs in E4, similar to the phenotype we observed upon APOE knockdown.”

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

Three to five wells per condition were performed per independent experiment, and data were collected and pooled from three independent experiments.

“Three to five wells per condition were performed per independent experiment, and data were collected and pooled from three independent experiments.”