CoChat
Sign Up
Cited by 86Cholesterol and Lipid MetabolismLipid metabolism and disordersCellular transport and secretion

Apolipoprotein E Is Resistant to Intracellular Degradation in Vitro and in Vivo

Open access
Rensen, Patrick C.N., Jong, Miek C., van Vark, Leonie C., van der Boom, Hans, Hendriks, Wendy L., van Berkel, Theo J.C., Biessen, Erik A.L., Havekes, Louis M.·2000·Journal of Biological Chemistry
DOI 10.1074/jbc.275.12.8564
Open this paper in CoChat — free accountOpen PDF

Abstract

Apolipoprotein E (apoE) is an important determinant for the uptake of triglyceride-rich lipoproteins and emulsions by the liver, but the intracellular pathway of apoE following particle internalization is poorly defined. In the present study, we investigated whether retroendocytosis is a unique feature of apoE as compared with apoB by studying the intracellular fate of very low density lipoprotein-sized apoE-containing triglyceride-rich emulsion particles and LDL after LDLr-mediated uptake. Incubation of HepG2 cells with [(3)H]cholesteryl oleate-labeled particles at 37 degrees C led to a rapid release of [(3)H]cholesterol within 30 min for both LDL and emulsion particles. In contrast, emulsion-derived (125)I-apoE was more resistant to degradation (>/=120 min) than LDL-derived (125)I-apoB (30 min). Incubation at 18 degrees C, which allows endosomal uptake but prevents lysosomal degradation, with subsequent incubation at 37 degrees C resulted in a time-dependent release of intact apoE from the cells (up to 14% of the endocytosed apoE at 4 h). The release of apoE was accelerated by the presence of protein-free emulsion (20%) or high density lipoprotein (26%). Retroendocytosis of intact particles could be excluded since little intact [(3)H]cholesteryl oleate was released (<3%). In contrast, the degradation of LDL was complete with virtually no secretion of intact apoB into the medium. The intracellular stability of apoE was also demonstrated after hepatic uptake in C57Bl/6 mice. Intravenous injection of (125)I-apoE and [(3)H]cholesteryl oleate-labeled emulsions resulted in efficient LDLr-mediated uptake of both components by the liver (45-50% of the injected dose after 20 min). At 1 h after injection, only 15-20% of the hepatic (125)I-apoE was degraded, whereas 75% of the [(3)H]cholesteryl oleate was hydrolyzed. From these data we conclude that following LDLr-mediated internalization by liver cells, apoE can escape degradation and can be resecreted. This sequence of events may allow apoE to participate in its hypothesized intracellular functions such as mediator of the post-lysosomal trafficking of lipids and very low density lipoprotein assembly.

AI snapshot

Written from the paper’s full text.

TL;DR

Apolipoprotein E (apoE) delivered to hepatocytes via LDL receptor-mediated uptake of VLDL-sized triglyceride-rich emulsions is relatively resistant to intracellular degradation compared with apoB or the emulsion cholesteryl oleate core. In vitro, intact apoE can be resecreted by retroendocytosis, and in mice hepatic apoE remains largely undegraded at 1 h after uptake.

Key findings

  • In HepG2 cells, emulsion-derived apoE degradation began slowly (≥120 min) and reached only 19% at 4 h, whereas cholesteryl oleate hydrolysis began within 30 min and reached 47% at 4 h; LDL-derived apoB degradation began within 30 min.
  • After 18 °C pulse labeling and 37 °C chase, intact apoE was released from HepG2 cells up to 14% of endocytosed apoE at 4 h, increasing to 20% with protein-free emulsion and 26% with HDL; intact particle retroendocytosis was excluded because <3% of [³H]cholesteryl oleate was released intact.
  • In C57Bl/6KH mice, apoE-enriched emulsions showed efficient LDLr-mediated liver uptake (45–50% of injected dose at 20 min); at 1 h only 15–20% of hepatic ¹²⁵I-apoE was degraded, whereas 75% of [³H]cholesteryl oleate was hydrolyzed.
  • Hepatic uptake of lipid-associated apoE was 43.7 ± 2.3% of dose at 20 min, with 36.3 ± 2.1% still present in liver at 60 min and only 8.8 ± 0.2% degradation products at 60 min.

Methods

  • In vitro: HepG2 cells were incubated with VLDL-sized apoE-containing triglyceride-rich emulsions (mean diameter 44.3 ± 2.6 nm) or LDL; binding was measured at 4 °C, uptake/degradation at 37 °C, and retroendocytosis was assessed by 18 °C pulse-chase followed by 37 °C incubation with or without HDL or protein-free emulsion.
  • Radioactivity was analyzed by TCA precipitation for protein degradation, thin-layer chromatography for cholesteryl oleate hydrolysis, density gradient ultracentrifugation, and SDS-PAGE.
  • In vivo: 10–12-week-old male C57Bl/6KH mice received intravenous ¹²⁵I-apoE- and [³H]cholesteryl oleate-labeled apoE emulsions; serum decay and liver uptake/degradation were measured over time.

Limitations

  • The paper states that in vivo retroendocytosis is hard to establish conclusively under the present experimental conditions; the in vivo work shows hepatic stability but does not directly demonstrate resecretion.
  • The mechanism of apoE retroendocytosis remains unresolved; the authors note it may differ from transferrin recycling and that alternative explanations such as pH-dependent aggregation require further investigation.
  • Evident from the text: the in vivo data are from two experiments (“means ± variation of two experiments”), and the study does not test extrahepatic cells such as macrophages, which the authors mention would be needed to extend implications to reverse cholesterol transport.

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

Integrity signals

Registry

Crossref · 10.1074/jbc.275.12.8564

Cites

63 of 63 references verified0 citation errors · 0 not checkable

Statistics

Nothing to recompute.

Claims the paper makes

0 of 69 cited sources checked · 0 supported · 0 flagged

  • Findingown data
    § Apolipoprotein E Is Resistant to Intracellular Degradation in Vitro and in Vivo EVIDENCE FOR RETROENDOCYTOSIS* (

    Incubation of HepG2 cells with [3H]cholesteryl oleate-labeled particles at 37 °C led to a rapid release of [3H]cholesterol within 30 min for both LDL and emulsion particles.

    “Incubation of HepG2 cells with [ 3 H]cholesteryl oleate-labeled particles at 37 °C led to a rapid release of [ 3 H]cholesterol within 30 min for both LDL and emulsion particles.”
    Quote in the paperNot a substantial claim
  • Findingown data
    § Apolipoprotein E Is Resistant to Intracellular Degradation in Vitro and in Vivo EVIDENCE FOR RETROENDOCYTOSIS* (

    Intravenous injection of 125I-apoE and [3H]cholesteryl oleate-labeled emulsions resulted in efficient LDLr-mediated uptake of both components by the liver (45-50% of the injected dose after 20 min).

    “Intravenous injection of 125 I-apoE and [ 3 H]cholesteryl oleate-labeled emulsions resulted in efficient LDLr-mediated uptake of both components by the liver (45-50% of the injected dose after 20 min).”
    Quote in the paperSubstantial claim
  • Findingown data
    § Apolipoprotein E Is Resistant to Intracellular Degradation in Vitro and in Vivo EVIDENCE FOR RETROENDOCYTOSIS* (

    The intracellular stability of apoE was also demonstrated after hepatic uptake in C57Bl/6 mice.

    “The intracellular stability of apoE was also demonstrated after hepatic uptake in C57Bl/6 mice.”
    Quote in the paperSubstantial claim
  • Findingown data
    § Apolipoprotein E Is Resistant to Intracellular Degradation in Vitro and in Vivo EVIDENCE FOR RETROENDOCYTOSIS* (

    Incubation at 18 °C, which allows endosomal uptake but prevents lysosomal degradation, with subsequent incubation at 37 °C resulted in a time-dependent release of intact apoE from the cells (up to 14% of the endocytosed apoE at 4 h).

    “Incubation at 18 °C, which allows endosomal uptake but prevents lysosomal degradation, with subsequent incubation at 37 °C resulted in a time-dependent release of intact apoE from the cells (up to 14% of the endocytosed apoE at 4 h).”
    Quote in the paperSubstantial claim
  • Findingown data
    § Apolipoprotein E Is Resistant to Intracellular Degradation in Vitro and in Vivo EVIDENCE FOR RETROENDOCYTOSIS* (

    Emulsion-derived 125I-apoE was more resistant to degradation (>120 min) than LDL-derived 125I-apoB (30 min).

    “In contrast, emulsion-derived 125 I-apoE was more resistant to degradation (>120 min) than LDL-derived 125 I-apoB (30 min).”
    Quote in the paperSubstantial claim
  • May combine two claims. 35
  • Classified as probably not a substantial claim. 19

  • May combine two claims.

    Findingown data
    § Apolipoprotein E Is Resistant to Intracellular Degradation in Vitro and in Vivo EVIDENCE FOR RETROENDOCYTOSIS* (

    The degradation of LDL was complete with virtually no secretion of intact apoB into the medium.

    “In contrast, the degradation of LDL was complete with virtually no secretion of intact apoB into the medium.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data
    § Apolipoprotein E Is Resistant to Intracellular Degradation in Vitro and in Vivo EVIDENCE FOR RETROENDOCYTOSIS* (

    At 1 h after injection, only 15-20% of the hepatic 125I-apoE was degraded, whereas 75% of the [3H]cholesteryl oleate was hydrolyzed.

    “At 1 h after injection, only 15-20% of the hepatic 125 I-apoE was degraded, whereas 75% of the [ 3 H]cholesteryl oleate was hydrolyzed.”
    Quote in the paperSubstantial claim

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

    Findingown data

    The apoE contents of the reisolated emulsions were 4.6 ± 1.1 g/mg TG (50:0.3) and 34.2 ± 2.6 g/mg TG (50:3.0), corresponding to 4 ± 1 and 27 ± 2 molecules of apoE per particle.

    “The apoE contents of the reisolated emulsions were 4.6 Ϯ 1.1 g/mg TG (50:0.3)and 34.2 Ϯ 2.6 g/mg TG (50:3.0),which corresponds to 4 Ϯ 1 and 27 Ϯ 2 molecules of apoE per particle.”
    Quote in the paperNot a substantial claim

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

    Findingcitation

    The emulsions were homogeneous with respect to size (low polydispersity of 0.23-0.31) and the mean particle diameter was 44.3 ± 2.6 nm (mean ± S.D.; n = 6) as determined by photon correlation spectroscopy.

    “The emulsions were homogeneous with respect to size (low polydispersity of 0.23-0.31)and the mean particle diameter was 44.3 Ϯ 2.6 nm (mean Ϯ S.D.; n ϭ 6) as determined by photon correlation spectroscopy using a Malvern 4700 C system (Malvern Instruments, Malvern, UK) (18).”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [18]

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

  • May combine two claims.

    Findingown data

    Incubation of HepG2 cells with [3H]CO or 125I-apoB-labeled LDL (20 g of protein/ml), which resulted in a similar rate of particle uptake as compared with apoE emulsions (~10^12 particles per mg of cell protein at 4 h), led to the rapid onset of both [3H]CO hydrolysis and 125I-apoB degradation (both within 30 min).

    “Incubation of HepG2 cells with [ 3 H]CO or 125 I-apoB-labeled LDL (20 g of protein/ ml), which resulted in a similar rate of particle uptake as compared with apoE emulsions (ϳ10 12 particles per mg of cell protein at 4 h), led to the rapid onset of both [ 3 H]CO hydrolysis and 125 I-apoB degradation (both within 30 min) (not shown).”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    The hydrolysis of CO started within 30 min after incubation and was very effective (47% of the total uptake at 4 h).

    “However, whereas the hydrolysis of CO started within 30 min after incubation and was very effective (47% of the total uptake at 4 h)”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    At 60 min, the cell association was reduced to 45 ± 1% (p < 0.05) and 38 ± 3% (p < 0.01) in the presence of the protein-free emulsion or HDL, respectively.

    “At this time point, the cell association was reduced to 45 Ϯ 1% (Student's t test; p Ͻ 0.05) and 38 Ϯ 3% (p Ͻ 0.01) in the presence of the protein-free emulsion or HDL, respectively.”
    Quote in the paperSubstantial claim

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

    Findingcitation

    Emulsion-bound 125I-apoE was only monomeric.

    “In contrast, emulsion-bound 125 I-apoE was only monomeric, which confirms earlier observations that the interaction of apoE homodimers with lipidic particles does not withstand ultracentrifugation conditions (48).”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [48]

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

  • May combine two claims.

    Findingown data

    The presence of protein-free emulsion or HDL had no effect on the degradation rate of 125I-apoE but did result in a significantly increased secretion rate of trichloroacetic acid-precipitable radioactivity (representing intact protein) as compared with the absence of acceptor.

    “The presence of these acceptors had no effect on the degradation rate of 125 I-apoE but did result in a significantly increased secretion rate of trichloroacetic acid-precipitable radioactivity (representing intact protein) as compared with the absence of acceptor (repeated measures analysis of variance, p Ͻ 0.01 and p Ͻ 0.001, respectively).”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    ApoE released from cells by retroendocytosis can recombine with both protein-deficient emulsion and HDL.

    “We observed that apoE, which is released from cells by retroendocytosis, can recombine with both the protein-deficient emulsion and HDL.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    Both the presence of emulsion and HDL in the medium stimulated retroendocytosis of apoE, without an effect on apoE degradation.

    “Both the presence of the emulsion and HDL in the medium stimulated retroendocytosis of apoE, without an effect on apoE degradation.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    Radioiodinated apoE contained some disulfide-linked apoE homodimers, but analysis of emulsion-bound 125I-apoE confirmed that radioactivity was associated with 34-kDa apoE only.

    “Whereas radioiodinated apoE did contain some disulfide-linked apoE homodimers, analysis of emulsion-bound 125 I-apoE confirmed that the radioactivity was associated with 34-kDa apoE only.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    The degradation of the apoE moiety started slowly (≥120 min) and was to a much lower extent (19% at 4 h).

    “the degradation of the apoE moiety started slowly (Ն120 min) and was to a much lower extent (19% at 4 h).”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    Hepatic uptake of lipid-associated apoE (43.7 ± 2.3% of dose at 20 min) is not coupled to rapid degradation, with 36.3 ± 2.1% still present in liver at 60 min after injection in mice.

    “The hepatic uptake of lipid-associated apoE (43.7 Ϯ 2.3% of the dose at 20 min after injection) was not coupled to rapid degradation, as 36.3Ϯ 2.1% was still present within the liver at 60 min after injection.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    Injection of an equal dose of lipid-free 125I-apoE into mice leads to rapid elimination of 80% of the injected dose from serum within 2 min and high liver uptake (70% at 5 min).

    “In contrast, injection of an equal dose of 125 I-apoE in a lipid-free state led to the rapid elimination of 80% of the injected dose from the serum within 2 min, with a high uptake by the liver (70% at 5 min after injection).”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    Preassociation of apoE with [3H]CO-labeled emulsion accelerates serum clearance (t1/2 <10 min) and liver uptake (~50% at 20 min) of the emulsion in mice.

    “As shown in Fig. 7, preassociation of apoE with the [ 3 H]CO-labeled emulsion accelerated the serum clearance (t1 ⁄2 Ͻ 10 min) and liver uptake (ϳ50% at 20 min) of the emulsion.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    In HepG2 cells loaded with [3H]CO-labeled apoE emulsions at 18°C, more than 90% of the cell-associated radiolabel is hydrolyzed to [3H]cholesterol after 4 h at 37°C, while less than 3% of the initially cell-associated radiolabel remains as intact [3H]CO in the medium.

    “Loading the cells with [ 3 H]CO-labeled apoE emulsions at 18 °C led to the hydrolysis of the vast majority of radiolabel into [ 3 H]cholesterol (Ͼ90%) after 4 h of incubation at 37 °C, whereas hardly any intact [ 3 H]CO could be detected in the medium (Ͻ3% of the initially cell-associated radiolabel) (not shown).”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingcitation

    Most LDL-derived apoB is recovered in the medium in a degraded state (>95%), confirming previous observations.

    “By contrast, most LDL-derived apoB was recovered in the medium in a degraded state (Ͼ95%), which confirms previous observations (28,62).”
    Quote in the paperSubstantial claim0 of 2 sources checked
    • cites [28]

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

    • cites [62]

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

  • May combine two claims.

    Findingown data

    An apoE-deficient emulsion shows low cellular association and degradation, which is not substantially increased by addition of 4 ± 1 molecules of apoE per emulsion particle at a TG:apoE = 50:0.3 weight ratio.

    “The apoE-deficient emulsion showed a low cellular association and degradation, which was not substantially increased by the addition of 4 Ϯ 1 molecules of apoE per emulsion particle, obtained at a TG:apoE ϭ 50:0.3 weight ratio (not shown).”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    Emulsion-derived apoE is much more resistant to intracellular degradation than LDL-derived apoB, as evident from a later onset of protein degradation (120 versus 30 min) and a much lower degradation rate.

    “Emulsion-derived apoE is much more resistant to intracellular degradation as compared with LDL-derived apoB, as evident from a later onset of protein degradation (120 versus 30 min) and a much lower degradation rate (Fig. 4).”
    Quote in the paperSubstantial claim

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

    Findingown data

    After radiolabeling of apoE, 96% of the 125I-activity could be recovered at the position of monomeric apoE (34 kDa).

    “After radiolabeling of apoE, 96% of the 125 I-activity could be recovered at the position of monomeric apoE (34 kDa).”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Findingown data

    After simultaneous hepatic uptake of both the particle core and preassociated apoE (45-50% of injected dose at 20 min), only 10-20% of apoE is degraded in contrast to as much as 75% of the CO moiety at 1 h after injection in mice.

    “Indeed, it appeared that after simultaneous hepatic uptake of both the particle core (reflected by the CO moiety) and the preassociated apoE (45-50% of the injected dose at 20 min after injection), only 10 -20% of the apoE is degraded in contrast to as much as 75% of the CO moiety at 1 h after injection (Figs. 7 and 8).”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Interpretationown data
    § Apolipoprotein E Is Resistant to Intracellular Degradation in Vitro and in Vivo EVIDENCE FOR RETROENDOCYTOSIS* (

    Following LDLr-mediated internalization by liver cells, apoE can escape degradation and can be resecreted.

    “From these data we conclude that following LDLr-mediated internalization by liver cells, apoE can escape degradation and can be resecreted.”
    Quote in the paperSubstantial claim

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

    Premisecitation
    § Apolipoprotein E Is Resistant to Intracellular Degradation in Vitro and in Vivo EVIDENCE FOR RETROENDOCYTOSIS* (

    ApoE plays a key role in the hepatic metabolism of triglyceride-rich lipoproteins such as chylomicrons and very low density lipoproteins and of triglyceride-rich emulsions.

    “Apolipoprotein E (apoE) 1 plays a key role in the hepatic metabolism of triglyceride (TG)-rich lipoproteins such as chylomicrons and very low density lipoproteins (VLDL) (1-3) and TG-rich emulsions (4 -5).”
    Quote in the paperNot a substantial claim0 of 5 sources checked
    • cites [1]

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

    • cites [2]

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

    • cites [3]

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

    • cites [4]

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

    • cites [5]

      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).

    Interpretationreasoning
    § Apolipoprotein E Is Resistant to Intracellular Degradation in Vitro and in Vivo EVIDENCE FOR RETROENDOCYTOSIS* (

    This sequence of events may allow apoE to participate in its hypothesized intracellular functions such as mediator of the post-lysosomal trafficking of lipids and very low density lipoprotein assembly.

    “This sequence of events may allow apoE to participate in its hypothesized intracellular functions such as mediator of the post-lysosomal trafficking of lipids and very low density lipoprotein assembly.”
    Quote in the paperNot a substantial claim

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

    Interpretationreasoning
    § Apolipoprotein E Is Resistant to Intracellular Degradation in Vitro and in Vivo EVIDENCE FOR RETROENDOCYTOSIS* (

    Since these emulsions mimic TG-rich lipoproteins, apoE delivered to the hepatocyte as a constituent of chylomicron remnants can possibly be re-used for intracellular lipid trafficking, such as the assembly and secretion of VLDL, or may participate in lipoprotein remnant internalization via the secretion-recapture pathway.

    “Since these emulsions mimic TG-rich lipoproteins, we anticipate that apoE, which has been delivered to the hepatocyte as a constituent of chylomicron remnants, can possibly be re-used for intracellular lipid trafficking, such as the assembly and secretion of VLDL, or may participate in lipoprotein remnant internalization via the “secretion-recapture” pathway (40,41).”
    Quote in the paperSubstantial claim0 of 2 sources checked
    • cites [40]

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

    • cites [41]

      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).

    Premisecitation
    § Apolipoprotein E Is Resistant to Intracellular Degradation in Vitro and in Vivo EVIDENCE FOR RETROENDOCYTOSIS* (

    The affinity of TG-rich lipoproteins and emulsions for the LDLr pathway and non-LDLr pathway is governed by their particle size.

    “It has been shown that the affinity of TG-rich lipoproteins and emulsions for the LDLr pathway and non-LDLr pathway is governed by their particle size.”
    Quote in the paperNot a substantial claim0 of 2 sources checked
    • cites [17]

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

    • cites [18]

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

  • May combine two claims.

    Premisecitation
    § Apolipoprotein E Is Resistant to Intracellular Degradation in Vitro and in Vivo EVIDENCE FOR RETROENDOCYTOSIS* (

    Small apoE-containing TG-rich emulsions mimic the metabolic fate of TG-rich lipoproteins in the blood and are exclusively taken up via the LDLr in mice.

    “For this purpose, we utilized small apoE-containing TGrich emulsions that mimic the metabolic fate of TG-rich lipoproteins in the blood (4,39) and are exclusively taken up via the LDLr in mice (18).”
    Quote in the paperSubstantial claim0 of 3 sources checked
    • cites [4]

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

    • cites [39]

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

    • cites [18]

      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).

    Study design and conductown data

    Using this method, 99.6% of the emulsion-associated 3H radioactivity appeared as [3H]CO.

    “Using this method, 99.6% of the emulsion-associated 3 H radioactivity appeared as [ 3 H]CO.”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Premisecitation
    § Apolipoprotein E Is Resistant to Intracellular Degradation in Vitro and in Vivo EVIDENCE FOR RETROENDOCYTOSIS* (

    Preliminary data on radioiodinated apoE emulsions and LDL suggest that, in contrast to the efficient degradation of apoB, the degradation of apoE may be retarded, whereas fluorescently labeled lipids in TG-rich particles and LDL followed a similar intracellular route toward lysosomes.

    “Preliminary data on radioiodinated apoE emulsions and LDL suggest that, in contrast to the efficient degradation of apoB, the degradation of apoE may be retarded, whereas fluorescently labeled lipids in TG-rich particles and LDL followed a similar intracellular route toward lysosomes (29,30).”
    Quote in the paperSubstantial claim0 of 2 sources checked
    • cites [29]

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

    • cites [30]

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

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

    Premisecitation
    § Apolipoprotein E Is Resistant to Intracellular Degradation in Vitro and in Vivo EVIDENCE FOR RETROENDOCYTOSIS* (

    LDL's protein and lipid components are completely degraded within perinuclear lysosomes while the receptor recycles back to the cell surface.

    “of which both the protein and lipid components are completely degraded within perinuclear lysosomes while the receptor recycles back to the cell surface (19,20).”
    Quote in the paperSubstantial claim0 of 2 sources checked
    • cites [19]

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

    • cites [20]

      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).

    Premisecitation
    § Apolipoprotein E Is Resistant to Intracellular Degradation in Vitro and in Vivo EVIDENCE FOR RETROENDOCYTOSIS* (

    ApoE appeared to have a physiological role in the assembly and secretion of VLDL.

    “In addition, apoE appeared to have a physiological role in the assembly and secretion of VLDL (34 -36).”
    Quote in the paperNot a substantial claim0 of 3 sources checked
    • cites [34]

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

    • cites [35]

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

    • cites [36]

      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).

    Premisecitation
    § Apolipoprotein E Is Resistant to Intracellular Degradation in Vitro and in Vivo EVIDENCE FOR RETROENDOCYTOSIS* (

    Large particles are mainly processed via the liver-specific non-LDLr recognition site.

    “Whereas large particles are mainly processed via the liver-specific non-LDLr recognition site”
    Quote in the paperNot a substantial claim0 of 2 sources checked
    • cites [17]

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

    • cites [18]

      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).

    Premisecitation
    § Apolipoprotein E Is Resistant to Intracellular Degradation in Vitro and in Vivo EVIDENCE FOR RETROENDOCYTOSIS* (

    Initial data suggested that apoE affects the transport and metabolism of free fatty acids and free cholesterol released from the lysosomes.

    “Initial data suggested that apoE affects the transport and metabolism of free fatty acids and free cholesterol released from the lysosomes (33).”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [33]

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

  • May combine two claims.

    Premisecitation

    Incubation at 18 °C does not impair binding and endocytosis, but blocks the fusion of endosomes with lysosomes.

    “Cells were preincubated in the presence of 125 I-apoE-containing emulsions (60 g of TG/ ml) for 3 h at 18 °C, which does not impair binding and endocytosis, but blocks the fusion of endosomes with lysosomes.”
    Quote in the paperSubstantial claim0 of 2 sources checked
    • cites [51]

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

    • cites [52]

      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.

    Study design and conductown data

    More than 97, 98.5, and 99.5% of the radiolabels in LDL, apoE, and bLf, respectively, were 10% trichloroacetic acid-precipitable.

    “More than 97, 98.5, and 99.5% of the radiolabels in LDL, apoE, and bLf, respectively, were 10% trichloroacetic acid-precipitable.”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Interpretationown data

    Both emulsion particles and HDL can function as acceptors of secreted apoE, with HDL more effective than emulsion particles in stimulating total release of apoE under the given conditions.

    “Apparently, both emulsion particles and HDL can function as acceptors of secreted apoE, albeit that HDL is more effective than emulsion particles in stimulation of the total release of apoE under the given conditions (Fig. 5).”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Premisecitation

    The apoE content of rat VLDL is 29 μg/mg TG, corresponding to 26 molecules per particle.

    “which is similar to the apoE content of rat VLDL (29 g/mg of TG, corresponding to 26 molecules per particle) (61)”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [61]

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

  • Classified as probably not a substantial claim.

    Premisecitation

    After cellular uptake of LDL, both its protein and lipid constituents follow the same lysosomal pathway.

    “After cellular uptake of LDL, both its protein and lipid constituents follow the same lysosomal pathway (19,20)”
    Quote in the paperSubstantial claim0 of 2 sources checked
    • cites [19]

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

    • cites [20]

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

  • May combine two claims.

    Study design and conductcitation

    Heparinase treatment inhibited binding of bovine lactoferrin (bLf) to HepG2 cells by 45%, demonstrating that HSPG had been effectively removed.

    “In accordance with previous observations by Ji and Mahley (59), heparinase treatment inhibited the binding of bLf to HepG2 cells by 45%, which demonstrates that HSPG had been effectively removed.”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [59]

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

  • May combine two claims.

    Interpretationown data

    The present data rule out that secretion of intact particles occurs upon endocytosis and show that apoE can selectively undergo retroendocytosis.

    “Our present data rule out that secretion of intact particles occurs upon endocytosis but rather show that apoE can selectively undergo retroendocytosis.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Interpretationown data

    The finding that HSPG are hardly involved in binding of apoE emulsion particles is in agreement with previous observations that, as opposed to LPL, apoE is not essential for binding of β-VLDL to HSPG.

    “The finding that HSPG are hardly involved in the binding of apoE emulsion particles is in agreement with our previous observations that, as opposed to LPL, apoE is not essential for the binding of ␤-VLDL”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [60]

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

  • May combine two claims.

    Interpretationown data

    ApoE emulsions, after LDLr-mediated uptake, may have a different intracellular fate as compared with LDL, as a result of which apoE may partially escape from lysosomal degradation.

    “These data thus indicate that apoE emulsions, after LDLr-mediated uptake, may have a different intracellular fate as compared with LDL, as a result of which apoE may (partially) escape from lysosomal degradation.”
    Quote in the paperSubstantial claim

  • Classified as probably not a substantial claim.

    Premisecitation

    Cell surface-bound apoE is released by a wash with heparin.

    “cell surface-bound apoE was released by a subsequent wash with heparin (770 units/ml in DMEM ϩ 1% BSA) (53,54).”
    Quote in the paperSubstantial claim0 of 2 sources checked
    • cites [53]

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

    • cites [54]

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

  • Classified as probably not a substantial claim.

    Premisecitation

    Degradation of 125I-protein in the medium can be determined by separating 125I-protein from degradation products by 10% trichloroacetic acid precipitation.

    “Degradation of 125 I-protein in the medium was determined by separating 125 I-protein from degradation products by 10% trichloroacetic acid precipitation as described (19).”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [19]

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

  • May combine two claims.

    Interpretationown data

    The applied apoE-emulsion particles are almost exclusively recognized by the LDLr on HepG2 cells in vitro, whereas HSPG play only a minor role in particle recognition.

    “Taking these data together, it is evident that the applied apoE-emulsion particles are also almost exclusively recognized by the LDLr on HepG2 cells in vitro, whereas HSPG play only a minor role in particle recognition.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Interpretationcitation

    The fact that transferrin recycling in HepG2 cells occurs with a half-time of less than 10 min, whereas apoE is slowly and gradually released after pulse labeling, suggests that mechanisms other than transferrin-like CURL shuttling account for apoE escaping degradation.

    “The fact that the recycling of transferrin in HepG2 cells occurs with a half-time of less than 10 min (64), whereas apoE is slowly and gradually released from these cells after pulse labeling, suggests that other mechanisms should account for escaping degradation.”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [64]

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

  • Classified as probably not a substantial claim.

    Premisecitation

    Newly synthesized apoE that is secreted and bound to cell surface HSPG can be re-endocytosed upon binding of lipoproteins (secretion-recapture pathway).

    “It is already known that newly synthesized apoE that is secreted and bound to cell surface HSPG can be re-endocytosed upon the binding of lipoproteins (so called secretion-recapture pathway) (40,41,57).”
    Quote in the paperSubstantial claim0 of 3 sources checked
    • cites [40]

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

    • cites [41]

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

    • cites [57]

      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).

    Premisecitation

    Newly synthesized apoE that binds to the cell surface may partially be proteolytically degraded upon re-entry into the cell.

    “In addition, newly synthesized apoE that binds to the cell surface may partially be proteolytically degraded upon re-entry into the cell (57).”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [57]

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

49 of 49

  • The paper relies on this without support.

    The intracellular pathway of apoE following particle internalization is poorly defined.

    “but the intracellular pathway of apoE following particle internalization is poorly defined.”

  • The paper relies on this without support.

    The VLDL-sized TG-rich emulsion enriched with radiolabeled apoE is a valid model for the intracellular fate of apoE in native TG-rich lipoproteins.

    “In order to chase the intracellular fate of apoE only, we thus decided to utilize VLDL-sized TG-rich emulsions (44 Ϯ 3 nm) that can be enriched with radiolabeled apoE (4,18).”
2 of 2

Cites

Flagged first, then by citations

Research works printed but not linked: 3

  1. Beisiegel, 1995, Circulation, 92, I
  2. Chen, 1995, Circulation, 92, I
  3. Chen, 1997, Circulation, 96, I

Cited by

Sorted by citations

Related

Authors
Rensen, P. C. N., Jong, M. C., van Vark, L. C., van der Boom, H., Hendriks, W. L., van Berkel, T. J. C., Biessen, E. A. L., & Havekes, L. M.
Affiliations
  • Leiden University (NL)
  • Netherlands Organisation for Applied Scientific Research (NL)
Corresponding author
Rensen, Patrick C.N.
ORCID
Venue
Journal of Biological Chemistry
Publisher
Elsevier BV
Published
2000-03
Volume
275
Issue
12
Pages
8564-8571
ISSN
0021-9258 (ISSN-L)
DOI
10.1074/jbc.275.12.8564
OpenAlex
W1982686316
Semantic Scholar
12bda1313988653c9aba69a2666f7405a7c86220
Cited by
86
References
62
Citations by year
2023: 2 · 2021: 2 · 2019: 3 · 2018: 3 · 2016: 1 · 2015: 2 · 2014: 8 · 2013: 2 · 2012: 3
FWCI
7.08
Type
journal article
Language
en
Open access
Licence
Locations
  • Journal of Biological Chemistry (publishedVersion, cc-by)
  • PubMed (publishedVersion)
  • Leiden Repository (Leiden University) (submittedVersion)
  • TNO Repository (submittedVersion)
  • Repository hosted by TU Delft Library (TU Delft) (submittedVersion)
  • Data Archiving and Networked Services (DANS) (submittedVersion, other-oa)
Topics
  • Health Sciences › Medicine › Surgery › Cholesterol and Lipid Metabolism
  • Health Sciences › Medicine › Cardiology and Cardiovascular Medicine › Lipid metabolism and disorders
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Cell Biology › Cellular transport and secretion
Keywords
apolipoprotein E, intracellular degradation, triglyceride-rich lipoproteins, apolipoprotein B, cholesterol ester hydrolysis, hepatic lipoprotein uptake, HepG2 cells, C57BL/6 mice, post-translational processing
MeSH terms
Animals, Apolipoproteins B, Apolipoproteins E, Biological Transport, Blood, Cells, Cultured, Cholesterol Esters, Emulsions, Endocytosis, Lipoproteins, Lipoproteins, LDL, Lipoproteins, VLDL, Liver, Lysosomes, Male, Mice, Inbred C57BL, Protein Processing, Post-Translational, Mice

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 2026-10-31.

Classified by jev-1.13-20260917 (gold-guideline-v0.2) on 2026-10-31.

Statistics and arithmetic are recomputed, not judged.

Confidence bands are shown only for fields calibrated on our annotated test set. None is calibrated yet, so no band is shown.

What counts as a claim ›How we measure confidence ›How we check this ›

Are you an author, or do you see a problem with this page? Request a review