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Cited by 45Cholesterol and Lipid MetabolismLipid metabolism and disordersLipoproteins and Cardiovascular Health

The DOI matches its Crossref record; 44 of the 45 references it cites were found in a registry or an index; 14 of the 25 sources its claims cite were checked for support.

Hypomorphic ASGR1 modulates lipid homeostasis via INSIG1-mediated SREBP signaling suppression

Open access
Xu, Yingying, Tao, Jiawang, Yu, Xiaorui, Wu, Yuhang, Chen, Yan, You, Kai, Zhang, Jiaye, Getachew, Anteneh, Pan, Tingcai, Zhuang, Yuanqi, Yuan, Fang, Yang, Fan, Lin, Xianhua, Li, Yin-xiong·2021·JCI Insight
DOI 10.1172/jci.insight.147038PMC PMC8525641
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Abstract

A population genetic study identified that the asialoglycoprotein receptor 1 ( ASGR1 ) mutation carriers had substantially lower non–HDL-cholesterol (non–HDL-c) levels and reduced risks of cardiovascular diseases. However, the mechanism behind this phenomenon remained unclear. Here, we established Asgr1 -knockout mice that represented a plasma lipid profile with significantly lower non–HDL-c and triglyceride (TG) caused by decreased secretion and increased uptake of VLDL/LDL. These 2 phenotypes were linked with the decreased expression of microsomal triglyceride transfer protein and proprotein convertase subtilisin/kexin type 9, 2 key targeted genes of sterol regulatory element–binding proteins (SREBPs). Furthermore, there were fewer nuclear SREBPs (nSREBPs) on account of more SREBPs being trapped in endoplasmic reticulum, which was caused by an increased expression of insulin-induced gene 1 (INSIG1), an anchor of SREBPs. Overexpression and gene knockdown interventions, in different models, were conducted to rescue the ASGR1-deficient phenotypes, and we found that INSIG1 knockdown independently reversed the ASGR1-mutated phenotypes with increased serum total cholesterol, LDL-c, TG, and liver cholesterol content accompanied by restored SREBP signaling. ASGR1 rescue experiments reduced INSIG1 and restored the SREBP network defect as manifested by improved apolipoprotein B secretion and reduced LDL uptake. Our observation demonstrated that increased INSIG1 is a critical factor responsible for ASGR1 deficiency–associated lipid profile changes and nSREBP suppression. This finding of an ASGR1/INSIG1/SREBP axis regulating lipid hemostasis may provide multiple potential targets for lipid-lowering drug development.

Overview

AI snapshot

Written from the paper’s full text.

TL;DR

ASGR1 deficiency lowers plasma non-HDL cholesterol and triglycerides in mice by reducing VLDL/LDL secretion and increasing LDL uptake, through increased INSIG1 and suppressed nuclear SREBP signaling. Knockdown of INSIG1 or restoration of ASGR1 reverses these lipid and SREBP phenotypes, defining an ASGR1/INSIG1/SREBP axis.

Key findings

  • In male Asgr1-knockout mice, serum TC was 3.8 ± 0.50, 2.9 ± 0.43, and 2.7 ± 0.37 mM in WT, Asgr1+/−, and Asgr1−/− mice; non–HDL-c was 2.3 ± 0.45, 1.7 ± 0.3, and 1.63 ± 0.19 mM, respectively. Female ASGR1-deficient mice also had lower TC, LDL-c, HDL-c, and non–HDL-c, with the exception of TG.
  • VLDL/LDL secretion was reduced: after tyloxapol, serum TG at 3 hours decreased by 19% in Asgr1+/− and 21% in Asgr1−/− mice; serum APOB100 and APOB48 decreased by 40% and 60% in Asgr1-null mice, with reduced MTTP and APOB at mRNA and protein levels.
  • LDL uptake increased by 20% and 90% in Asgr1+/− and Asgr1−/− primary hepatocytes, by 45% in ASGR1−/− HepG2 cells, and by 50% in hu-HLC-ASGR1−/− cells; PCSK9 was downregulated and LDLR protein increased.
  • Mechanistically, nSREBP1/2 were lower and INSIG1 was higher; INSIG1 knockdown reversed serum TC, LDL-c, TG, and liver cholesterol and restored nSREBP1/2, MTTP, LDLR, and PCSK9, while ASGR1 restoration increased nSREBP1/2, decreased INSIG1, increased APOB secretion by 50%, and normalized LDL uptake.

Methods

  • Generated Asgr1-knockout mice by CRISPR/Cas9 on a C57BL/6 background; used male mice about 10–13 weeks old, primary hepatocytes, HepG2 cells, and H1-derived hepatocyte-like cells; performed RNA-Seq on WT and Asgr1−/− livers (GSE178370).
  • Measured serum and hepatic lipids, FPLC, oral lipid tolerance, tyloxapol secretion, LDL-dil uptake, APOB ELISA, qPCR, Western blot, immunofluorescence, and nuclear/ER fractionation; used siRNA knockdown and ASGR1/PCSK9 overexpression rescue experiments.
  • Statistics used unpaired 2-tailed t tests for 2 groups and 1-way ANOVA for 3 or more groups; data are means ± SEM, with significance at *P < 0.05, P < 0.01, and *P < 0.001.

Limitations

  • Stated: the deep mechanistic roots of the ASGR1/INSIG1 linkage remain to be clarified, and the authors suggest further investigation into whether ASGR1 mediates insulin receptor degradation and regulates Insig1.
  • Stated/evident: the findings come from mouse models and cell lines (HepG2, H1-derived hepatocytes, primary mouse hepatocytes), not from human ASGR1 mutation carriers or human intervention studies.
  • Evident: sex differences were observed—female ASGR1-deficient mice did not show the TG decrease seen in males—and subsequent experiments used male mice to avoid hormonal effects, so sex-specific mechanisms are not fully resolved.

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

Integrity signals

Registry

Crossref · 10.1172/jci.insight.147038

Cites

44 of 44 references verified0 citation errors · 0 not checkable

Statistics

Nothing to recompute.

Verification

Claims the paper makes

14 of 25 cited sources checked · 5 supported · 1 flagged · accuracy not yet measured

  • Premisecitation

    The gene expression of PCSK9 is negatively regulated by SREBPs.

    “The gene expression of PCSK9 is negatively regulated by SREBPs, and PCSK9 protein is a key mediator for LDLR internalization and degradation by ubiquitination.”
    Quote in the paperSubstantial claim1 of 1 sources checked
    • cites [34] source

      The section “Regulation of the promoter activity of PCSK9 by overexpression of SREBPs” reports that nuclear SREBPs dramatically increased PCSK9 promoter transcription. The paper describes SREBP-1 and SREBP-2 as activators of PCSK9 expression, contrary to the claimed negative regulation. (LLM, conf=0.94)

      Checked by a reasoning model — may be wrong.

      Experimental — accuracy not yet measured
  • Premisecitation

    LDLR and PCSK9 are downstream genes of SREBPs.

    “LDLR and PCSK9 are downstream genes of SREBPs.”
    Quote in the paperSubstantial claim2 of 2 sources checked
    • cites [33] source

      The Discussion concludes that SREBP-1 is involved in LDL receptor gene activation, supporting the LDLR part of the claim. Our copy does not address PCSK9, so it does not support the claim that PCSK9 is also downstream of SREBPs (searched: PCSK9, proprotein convertase subtilisin/kexin 9). (LLM, conf=0.89)

      Experimental — accuracy not yet measured
    • cites [34] source
  • Premisecitation

    ASGR1 mutation carriers have reduced risks of cardiovascular diseases.

    “A human population genetic study provided evidence that variant asialoglycoprotein receptor 1 (ASGR1) is associated with lower non–HDL-c content and reduced risk of coronary artery disease (CAD), indicating a potential target for CVD prevention and treatment (3).”
    Quote in the paperSubstantial claim1 of 1 sources checked
  • Premisecitation

    PCSK9 protein is a key mediator for LDLR degradation by ubiquitination.

    “The PCSK9 protein is a key mediator for LDLR degradation by ubiquitination, and its inhibitors have been applied to clinical trials for hypocholesterolemia (33, 34).”
    • Also stated in: § Discussion · plain
      “The gene expression of PCSK9 is negatively regulated by SREBPs, and PCSK9 protein is a key mediator for LDLR internalization and degradation by ubiquitination.”
    Quote in the paperSubstantial claim2 of 2 sources checked
    • cites [33] source

      The introduction and Discussion describe SREBP-1 regulation of LDL receptor gene transcription, not degradation of the receptor protein. Searches for PCSK9, ubiquitination, and PCSK9 inhibitors found no passage addressing the attributed claim; page numbers were unavailable in our copy. (LLM, conf=0.78)

      Experimental — accuracy not yet measured
    • cites [34] source

      The Abstract says PCSK9 promotes LDLR internalization and degradation, and describes prevention of LDLR recycling or targeting it for lysosomal degradation. Neither that passage nor the Discussion identifies ubiquitination as the mechanism, so the mechanistic part of the claim is unsupported. (LLM, conf=0.88)

      Experimental — accuracy not yet measured
  • Premisecitation

    Previously established Asgr1-knockout mice by replacing exons 2-3 were unable to clear asialofetuin and asialo-orosomucoid.

    “Previously, Asgr1-knockout mice were established by replacing exons 2–3, which contain the ATG translational initiation codon and transmembrane domain (27). These ASGR1-knockout mice were unable to clear asialofetuin and asialo-orosomucoid.”
    Quote in the paperNot a substantial claim1 of 1 sources checked
    • cites [27] source

      The Experimental Procedures describe replacing the fragment containing exons 2–3 to generate MHL-1 knockout mice. The Results report severely impaired clearance of both asialofetuin and asialo-orosomucoid, but not an inability to clear them: at 30 minutes, 40% and 65% of the respective doses remained in knockout mice. (LLM, conf=0.91)

      Experimental — accuracy not yet measured
  • Probably not a substantial claim. 16
  • It may state two separate claims as one. 44

  • Probably not a substantial claim. The paper’s conclusions may not depend on it. It may state two separate claims as one.

    Premisecitation

    LDLR recycling plays a critical role in the endocytosis of LDL, and hepatic uptake of serum LDL-c facilitates serum lipid homeostasis.

    “LDLR recycling plays a critical role in the endocytosis of LDL, and hepatic uptake of serum LDL-c facilitates serum lipid homeostasis (32).”
    Quote in the paperNot a substantial claim1 of 1 sources checked

  • It may state two separate claims as one.

    Premisecitation

    Heterozygous ASGR1 mutation carriers have lower non-HDL-c and TG, and ASGR1 haploinsufficiency is associated with lower risk of CAD.

    “The heterozygous mutation carriers had a lower level of non–HDL-c and TG, and the ASGR1 haploinsufficiency was associated with a lower risk of CAD (3).”
    Quote in the paperSubstantial claim1 of 1 sources checked

  • Probably not a substantial claim. It may state two separate claims as one.

    Premisecitation

    MTTP is transcriptionally regulated by SREBPs, and lack of MTTP results in APOB cotranslational degradation.

    “MTTP is transcriptionally regulated by SREBPs, and the lack of MTTP results in APOB cotranslational degradation (29–31).”
    Quote in the paperSubstantial claim3 of 3 sources checked
    • cites [29] source

      The Introduction says that adding lipids to apo B as it is translated is thought to help it avoid degradation, and the results section reports substantially reduced apo B-100 secretion from Mttp-deficient hepatocytes. Neither passage establishes that the loss causes cotranslational degradation; the paper also reports little change in apo B-48 secretion. The passages do not support the claim that SREBPs transcriptionally regulate MTTP (section labels available; page numbers not provided). (LLM, conf=0.84)

      Experimental — accuracy not yet measured
    • cites [30] source

      The section on SREBP1a binding states that SREBP negatively regulates MTTP expression, supporting the transcriptional-regulation claim. The Introduction says MTTP stabilizes newly synthesized apoB, but does not state that lack of MTTP causes its cotranslational degradation. Page numbers are unavailable in our copy. (LLM, conf=0.86)

      Experimental — accuracy not yet measured
    • cites [31] source

      The section on cotranslational production of ApoB48-like protein proposes that insufficient MTP activity can interrupt apoB translocation and leave apoB susceptible to degradation during translation. This supports the degradation part, though the mechanism is proposed from experiments in transfected COS-7 cells. No passage found supports the separate claim that SREBPs transcriptionally regulate MTTP (searched for SREBP and sterol regulatory element-binding proteins). (LLM, conf=0.88)

      Experimental — accuracy not yet measured

  • Probably not a substantial claim. The paper’s conclusions may not depend on it. It may state two separate claims as one.

    Premisecitation

    INSIG1 is ubiquitinated and degraded by GP78 when oxysterols are depleted, while oxysterols stabilize it by binding within its transmembrane domains.

    “INSIG1 is ubiquitinated and degraded by GP78 when oxysterols are depleted, while oxysterols stabilize it by binding within its transmembrane domains (37, 38).”
    Quote in the paperNot a substantial claim1 of 2 sources checked
    • cites [37] source
    • cites [38] source

      The Abstract reports that oxysterols bind Insigs and that the third and fourth transmembrane helices are important for binding. The Discussion says the binding experiments used Insig-2, though the authors believe the conclusions also apply to Insig-1. Our copy does not say that oxysterol binding stabilizes Insig-1 or that GP78 ubiquitinates and degrades it upon oxysterol depletion; its Discussion instead describes ubiquitination and degradation of HMGR. (LLM, conf=0.88)

      Experimental — accuracy not yet measured

  • Probably not a substantial claim. It may state two separate claims as one.

    Premisecitation

    INSIG1 is a protein promoting ER retention of SREBPs, and it is the targeted gene that exerts negative feedback control of SREBPs.

    “INSIG1 is a protein promoting ER retention of SREBPs, while it is the targeted gene that exerts negative feedback control of SREBPs (35, 36).”
    Quote in the paperSubstantial claim1 of 2 sources checked
    • cites [35] source
    • cites [36] source

  • It may state two separate claims as one.

    Findingown data

    mRNA expression of SREBP target genes in lipogenesis (ACC, FASN, SCD1) and cholesterol synthesis (HMGCS, HMGCR) is downregulated in mouse livers and HepG2 cells, indicating lipogenesis is inhibited in the absence of ASGR1.

    “All of those inspected genes were transcriptionally downregulated, indicating that lipogenesis was inhibited in the absence of ASGR1 (Figure 5, E and F); in addition, the expression of MTTP and PCSK9 related to LDL assembling and endocytosis was also significantly decreased (Figure 2, C and F; and Figure 3, B and E).”
    Quote in the paperSubstantial claim

  • It may state two separate claims as one.

    Findingown data

    In Asgr1-null mice, serum contents of APOB100 and APOB48 are decreased by 40% and 60%, respectively.

    “Particularly, in the null Asgr1-nullmice, the serum contents of APOB100 and APOB48 were decreased by 40% and 60% (Figure 2, D and E).”
    Quote in the paperSubstantial claim

  • It may state two separate claims as one.

    Findingown data

    Upon overexpression of PCSK9 in HepG2-ASGR1-/- cells and WT HepG2 cells, LDLR protein content decreases in both cell lines, leading to no substantial difference of LDLR between HepG2-ASGR1-/- and WT cells.

    “Upon overexpression of PCSK9 in HepG2-ASGR1–/– cells and WT HepG2 cells, respectively, the LDLR protein content decreased in both cell lines, leading to no substantial difference of the LDLR between HepG2-ASGR1–/– and WT cells (Supplemental Figure 4D).”
    Quote in the paperSubstantial claim

  • It may state two separate claims as one.

    Findingown data

    LDLR mRNA expression declines in ASGR1-deficient livers, and a slight increase is observed in HepG2 cells.

    “The LDLR mRNA expression declined in ASGR1-deficient livers, and a slight increase was observed in HepG2 cells.”
    Quote in the paperNot a substantial claim

  • It may state two separate claims as one.

    Findingown data

    INSIG1 knockdown reverses reduction of nSREBP1/2 proteins in Asgr1-/- mice; scramble siRNA groups have no marked effects on lipid profile or nSREBP levels.

    “while the protein level of INSIG1 in Asgr1–/– mice was reduced to the level similar to the one in WT mice, the reduction of nSREBP1/2 proteins was also reversed, and the scramble siRNA groups (WT-NC and Asgr1–/–-NC) had no marked effects on lipid profile or on nSREBP levels (Figure 7I).”
    Quote in the paperSubstantial claim

  • It may state two separate claims as one.

    Findingown data

    The majority of transcriptomes of Asgr1-/- mouse liver are similar to those of WT counterparts, and the significantly altered genes are very limited.

    “The majority of transcriptomes of Asgr1–/– mouse liver were similar to those of WT counterparts, and the significantly altered genes were very limited (Figure 4A).”
    Quote in the paperNot a substantial claim

  • It may state two separate claims as one.

    Findingown data

    PCSK9 overexpression reduced LDLR levels and increased PCSK9 protein levels in HepG2 and HepG2-ASGR1-/- cells.

    “we conducted a rescue experiment through overexpression of PCSK9 in HepG2 and HepG2-ASGR1–/– cells and found out that the LDLR levels were reduced while the PCSK9 protein levels were increased either in HepG2 or in HepG2-ASGR1–/– cells.”
    Quote in the paperSubstantial claim

  • It may state two separate claims as one.

    Findingown data

    After tyloxapol administration, serum TG levels of ASGR1-deficient mice are less than those of WT mice, with the most significant difference at 3 hours (Asgr1+/− decreased by 19%; Asgr1−/− decreased by 21%).

    “we measured serum lipid level after administration of lipoprotein lipase inhibitor tyloxapol and found serum TG levels of ASGR1-deficient mice were comparatively less than those of WT mice. The most significant difference was observed at 3 hours after intravenous injection (Asgr1+/−, decreased by 19%; Asgr1−/−, decreased by 21%; Figure 2, A and B).”
    Quote in the paperSubstantial claim

  • It may state two separate claims as one.

    Findingown data

    Bile acid content in liver and feces, as well as expressions of bile acid synthesis-related genes in liver, have no significant difference among mice with or without ASGR.

    “In addition, bile acid content in the liver and feces, as well as the expressions of bile acid synthesis-related genes in liver, had no significant difference among mice with or without ASGR (Supplemental Figure 3, D–F).”
    Quote in the paperNot a substantial claim

  • It may state two separate claims as one.

    Findingown data

    Among altered genes, 12 are associated with lipid metabolism, of which 11 are downregulated and 1 is upregulated.

    “Among those genes with altered expression, 12 of them were associated with lipid metabolism, in which 11 genes were downregulated and 1 gene was upregulated, as shown in the heatmap.”
    Quote in the paperSubstantial claim

  • It may state two separate claims as one.

    Findingown data

    In HepG2 cells, INSIG1 knockdown reverses phenotypic features with 40% increase of APOB secretion and LDL uptake down to WT level.

    “Then phenotypic features were reversed with 40% increase of APOB secretion and LDL uptake down to the level of WT (Figure 7, L and M).”
    Quote in the paperSubstantial claim

  • It may state two separate claims as one.

    Findingown data

    181 genes are significantly changed (fold change > 1.5, q value < 0.05), among which 104 genes are downregulated and 77 genes are upregulated.

    “Further analysis of the differentially expressed genes showed that 181 genes were significantly changed (fold change > 1.5, q value < 0.05), among which 104 genes were downregulated and 77 genes were upregulated (Figure 4C).”
    Quote in the paperSubstantial claim

  • It may state two separate claims as one.

    Findingown data

    Body weight and hepatic histology show no significant difference among mice with different Asgr1 genotypes.

    “The body weight of mice and hepatic histology showed no significant difference among mice with different genotypes (Supplemental Figure 1, C and E).”
    Quote in the paperNot a substantial claim

  • It may state two separate claims as one.

    Findingown data

    LDL uptake rates are increased by 20% and 90% in Asgr1+/- and Asgr1-/- primary hepatocytes, respectively, compared with WT counterparts.

    “In detail, the uptake rates increased by 20% and 90% in Asgr1+/– and Asgr1–/– primary hepatocytes, respectively, compared with the WT counterparts.”
    Quote in the paperSubstantial claim

  • It may state two separate claims as one.

    Findingown data

    ASGR1 is highly expressed in liver tissues of WT mice but is substantially decreased or undetectable in Asgr1+/− or Asgr1−/− mice, respectively.

    “ASGR1 was highly expressed in liver tissues of WT mice but was substantially decreased or undetectable in Asgr1+/− or Asgr1−/− mice, respectively (Figure 1B).”
    Quote in the paperSubstantial claim

  • It may state two separate claims as one.

    Findingown data

    Nuclear and cytoplasmic protein separation confirms that nSREBP1/2 level is reduced in nuclei of ASGR1-/- cells, while more full-length SREBP1/2 is accumulated in cytoplasm.

    “Next, we separated nuclear and cytoplasmic proteins from HepG2 cells and confirmed that the level of nSREBP1/2 was reduced in the nuclei of ASGR1–/– cells, while more full-length SREBP1/2 was accumulated in the cytoplasm (Figure 5C and Supplemental Figure 5A).”
    Quote in the paperSubstantial claim

  • It may state two separate claims as one.

    Findingown data

    nSREBP1 and nSREBP2 protein content is lower in ASGR1-deficient mouse livers and HepG2 cells, while full-length SREBP proteins are increased in mice, indicating that SREBP activation is inhibited.

    “By detecting SREBP1 and SREBP2, we found that the protein content of nSREBP1 and nSREBP2 was lower in ASGR1-deficient mouse livers and HepG2 cells, while the content of full-length proteins was increased in mice, indicating that the activation was inhibited (Figure 5, A and B).”
    Quote in the paperSubstantial claim

  • It may state two separate claims as one.

    Findingown data

    INSIG1 knockdown or ASGR1 restoration reversed the ASGR1 deficiency-associated lipid profile and SREBP suppression in ASGR1-deficient mice and cells.

    “we performed knockdown or restored expression of key factors in the ASGR1-deficient mice and cells and found that INSIG1 knockdown or ASGR1 restoration reversed the ASGR1 deficiency–associated lipid profile and SREBP suppression.”
    Quote in the paperSubstantial claim

  • It may state two separate claims as one.

    Findingown data

    GSEAs show improved metabolic signaling and reduced cholesterol-related signaling in Asgr1-/- mouse livers.

    “Gene set enrichment analyses (GSEAs) showed improved metabolic signaling and reduced cholesterol-related signaling in Asgr1–/– mouse livers (Figure 4B).”
    Quote in the paperSubstantial claim

  • It may state two separate claims as one.

    Findingown data

    In female mice, serum TC, LDL-c, HDL-c, and non-HDL-c levels are decreased in ASGR1-deficient mice, with the exception of TG.

    “Consistent with male mice, the serum TC, LDL-c, HDL-c, and non–HDL-c levels were decreased in ASGR1-deficient female mice, with the exception of TG (Supplemental Figure 2, A–E).”
    Quote in the paperSubstantial claim

  • It may state two separate claims as one.

    Findingown data

    LDL-c, HDL-c, and TG levels are reduced in ASGR1-deficient male mice in similar trends to total cholesterol.

    “In detail, the serum TC level showed significant changes among groups, which were 3.8 (± 0.50) mM, 2.9 (± 0.43) mM, and 2.7 (± 0.37) mM in WT, Asgr1+/−, and Asgr1−/− mice, respectively, followed by the reduced level of LDL-c, HDL-c, and TG in similar trends.”
    Quote in the paperSubstantial claim

  • It may state two separate claims as one.

    Findingreasoning

    MTTP is reduced along with lower levels of APOB when ASGR1 is deficient, resulting in decreased VLDL/LDL secretion.

    “We observed that the MTTP was reduced along with lower levels of APOB, when ASGR1 was deficient, resulting in decreased VLDL/LDL secretion.”
    Quote in the paperSubstantial claim

  • It may state two separate claims as one.

    Findingown data

    Restoring ASGR1 expression in ASGR1−/− HepG2 cells increases nSREBP1/2 and decreases INSIG1.

    “Compared with the control cells (ASGR1−/−-EGFP-tg), nSREBP1/2 were increased, while INSIG1 was decreased (Figure 8A).”
    Quote in the paperSubstantial claim

  • It may state two separate claims as one.

    Findingown data

    INSIG1 knockdown in Asgr1-/- mice causes no obvious changes in serum HDL-c and liver TG content.

    “the phenotypic features were reversed in Asgr1–/– mice as manifested by decreased serum TC, LDL-c, and TG and liver cholesterol content (Figure 7, C, D, F, and G), while there were no obvious changes in serum HDL-c and liver TG content (Figure 7, E and H).”
    Quote in the paperNot a substantial claim

  • It may state two separate claims as one.

    Findingreasoning

    The mRNA of both Ldlr and Pcsk9 declined in ASGR1-deficient conditions.

    “While nSREBPs were decreased, the mRNA of both genes declined.”
    Quote in the paperSubstantial claim

  • It may state two separate claims as one.

    Findingown data

    Immunofluorescence staining shows INSIG1 level is markedly increased and predominantly located on the ER of ASGR1−/− HepG2 cells.

    “Immunofluorescence staining of INSIG1 in HepG2 cells showed that the level of INSIG1 was markedly increased and predominantly located on the ER of ASGR1−/−-HepG2 cells (Figure 6C).”
    Quote in the paperSubstantial claim

  • It may state two separate claims as one.

    Findingown data

    In ASGR1−/−-ASGR1-tg cells, APOB secretion increases by 50% and LDL uptake returns to WT level.

    “Correspondingly, the related phenotype of APOB secretion increased by 50%, and LDL uptake returned to the WT level in ASGR1−/−-ASGR1-tg cells (Figure 8, C and D).”
    Quote in the paperSubstantial claim

  • It may state two separate claims as one.

    Interpretationown data

    ASGR1 deficiency leads to a defect in LDL secretion that might be caused by less MTTP and APOB for the construction of the LDL complex and its secretion from hepatocytes.

    “In summary, ASGR1 deficiency led to a defect in LDL secretion that might be caused by less MTTP and APOB for the construction of the LDL complex and its secretion from hepatocytes.”
    Quote in the paperSubstantial claim

  • It may state two separate claims as one.

    Premisecitation

    The first report of ASGR1 mutation found a 12 bp deletion in ASGR1, leading to loss of function.

    “The first report of ASGR1 mutation was based on a population genetic study and found a 12 bp deletion in ASGR1, which led to a loss of function.”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [3] source

  • It may state two separate claims as one.

    Interpretationreasoning

    Both nSREBP1 and nSREBP2 are decreased in ASGR1 deficiencies, which led to the physiological benefits of the plasma lipid profile.

    “Both nSREBP1 and nSREBP2 were decreased in ASGR1 deficiencies, which led to the physiological benefits of the plasma lipid profile.”
    Quote in the paperSubstantial claim

  • Probably not a substantial claim.

    Interpretationreasoning

    SREBP suppression is responsible for the higher LDLR level in ASGR1 deficiency.

    “It seems plausible that the SREBP suppression is responsible for the higher LDLR level in ASGR1 deficiency.”
    Quote in the paperSubstantial claim

  • Probably not a substantial claim. It may be too vague to check against evidence. The paper’s conclusions may not depend on it.

    Interpretationreasoning

    Other mechanisms might play a major role in the increased INSIG1 in ASGR1 deficiency.

    “Thus, it seems that other mechanisms might play a major role in the increased INSIG1 in ASGR1 deficiency.”
    Quote in the paperNot a substantial claim

  • It may state two separate claims as one.

    Interpretationreasoning

    The hypomorphic effect of ASGR1 results in a healthy and favorable lipid profile, primarily generated from differential turnover of plasma lipoproteins.

    “The hypomorphic effect of ASGR1 results in a healthy and favorable lipid profile, which is primarily generated from the differential turnover of the plasma lipoproteins.”
    Quote in the paperSubstantial claim

  • It may state two separate claims as one.

    Interpretationreasoning

    The failure to observe significant changes of lipid profile in previous Asgr1-knockout mice may be due to particular knockout mouse lines, limited analyzed group samples, and sex-influenced variation.

    “The failure to observe significant changes of lipid profile may be due to conditions including those particular knockout mouse lines, limited analyzed group samples, and the sex-influenced variation.”
    Quote in the paperSubstantial claim

  • Probably not a substantial claim. It may state two separate claims as one.

    Premisecitation

    VLDL/LDL particles mediate lipid secretion from the liver, and two steps are generally involved in VLDL/LDL assembly: MTTP transports lipids onto APOB to form precursor particles, then precursor particles and TG droplets combine to form mature VLDL/LDL.

    “VLDL/LDL particles mediated lipid secretion from the liver, and there are 2 steps generally involved in VLDL/LDL assembly. First, the MTTP transports the lipids onto APOB to form precursor particles. Then, the precursor particles and TG droplets combine to form mature VLDL/LDL (28).”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [28] source

  • Probably not a substantial claim. The paper’s conclusions may not depend on it.

    Premisecitation

    In 104 patients, ASGR1 expression in peripheral blood monocytes is positively correlated with serum TC and LDL-c levels at ages younger than 60.

    “Moreover, a recent study on 104 patients revealed that the expression level of ASGR1 in peripheral blood monocytes was positively correlated with serum TC and LDL-c levels at ages younger than 60 (26).”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [26] Zhao, 2020, Asialoglycoprotein receptor 1 gene expression in peripheral blood monocytes associates with serum total cholesterol and low-density lipoprotein cholesterol levels, Chin Med J (Engl) source

      Checked before — its earlier verdict is held back until a new check runs.

  • It may state two separate claims as one.

    Interpretationreasoning

    Suppression of SREBP accounted for low PCSK9 and less degradation of LDLR, which may result in higher LDLR level and LDL endocytosis.

    “Collectively, the suppression of SREBP accounted for low PCSK9 and less degradation of LDLR that may result in higher LDLR level and LDL endocytosis.”
    Quote in the paperSubstantial claim

  • It may state two separate claims as one.

    Interpretationreasoning

    In a dose-dependent manner, hypomorphic manipulations lead to more LDLR on cell membrane and less assembled VLDL in hepatocytes, resulting in more LDL uptake and less VLDL secretion in cell and mouse models, which represents substantially lower serum lipid content.

    “In a dose-dependent manner, hypomorphic manipulations led to more LDLR on cell membrane and less assembled VLDL in hepatocytes, resulting in more LDL uptake and less VLDL secretion in cell and mouse models, which represented a substantially lower serum lipid content.”
    Quote in the paperSubstantial claim

  • It may state two separate claims as one.

    Interpretationown data

    There is a signaling axis, ASGR1/INSIG1/SREBP, in which INSIG1 is the primary change factor and independent of SREBP under ASGR1 deficiency.

    “This suggests that there is a signaling axis, ASGR1/INSIG1/SREBP, in which INSIG1 is the primary change factor and independent of SREBP under ASGR1 deficiency, and hypomorphic ASGR1 modulates lipid homeostasis via INSIG1-mediated SREBP signaling suppression.”
    Quote in the paperSubstantial claim

  • It may state two separate claims as one.

    Premisecitation

    In previously established Asgr1-knockout mice, measurements of LDL and VLDL revealed a decreased trend with large standard deviations without a statistical difference, and serum TC and TG contents had no significant difference.

    “However, the measurements of LDL and VLDL revealed a decreased trend with large standard deviations without a statistical difference, and the serum TC and TG contents also had no significant difference.”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [27] source

      Checked before — its earlier verdict is held back until a new check runs.

  • Probably not a substantial claim. The paper’s conclusions may not depend on it.

    Study design and conductnone given

    Genome-edited F0 Asgr1+/- mice were bred with C57BL/6 mice for 2 rounds to dilute off-target effects.

    “Genome-edited F0 Asgr1+/– mice (Cyagen) were bred with C57BL/6 mice for 2 rounds to dilute the off-target effects.”
    Quote in the paperNot a substantial claim

  • It may state two separate claims as one.

    Interpretationown data

    Increased INSIG1 was a primary change factor and responsible for the ASGR1 deficiency-associated lipid profile changes and nSREBP suppression.

    “Overall, our observation clearly demonstrated that the increased INSIG1 was a primary change factor and responsible for the ASGR1 deficiency–associated lipid profile changes and nSREBP suppression.”
    Quote in the paperSubstantial claim

  • Probably not a substantial claim. The paper’s conclusions may not depend on it. It may state two separate claims as one.

    Study design and conductnone given

    Mice used for experiments were aged about 10-13 weeks, and the age difference of the mice was no more than 1 week for the same experiment.

    “Mice aged about 10–13 weeks were used for experiments, and the age difference of the mice was no more than 1 week for the same experiment.”
    Quote in the paperNot a substantial claim

  • Probably not a substantial claim. The paper’s conclusions may not depend on it.

    Study design and conductnone given

    Experiments were repeated 3 times individually at least.

    “Experiments were repeated 3 times individually at least.”
    Quote in the paperNot a substantial claim

  • Probably not a substantial claim. The paper’s conclusions may not depend on it.

    Study design and conductnone given

    For each sample, no less than 5 representative images were analyzed to quantify fluorescence intensity values and calculate an average.

    “For each sample, no less than 5 representative images were analyzed to quantify the fluorescence intensity values and calculate an average.”
    Quote in the paperNot a substantial claim

  • Probably not a substantial claim. The paper’s conclusions may not depend on it.

    Study design and conductnone given

    All animal studies were performed in accordance with the experimental animal ethics committee of Guangzhou Institutes of Biomedicine and Health.

    “All animal studies, including mouse feeding and operation experiments, were performed in accordance with the experimental animal ethics committee of Guangzhou Institutes of Biomedicine and Health.”
    Quote in the paperNot a substantial claim

  • Probably not a substantial claim.

    Study design and conductnone given

    WT and Asgr1-/- littermates were generated by breeding Asgr1+/- mice.

    “WT and Asgr1–/– littermates were generated by breeding Asgr1+/– mice.”
    Quote in the paperSubstantial claim

  • Probably not a substantial claim. The paper’s conclusions may not depend on it.

    Premisecitation

    Insig1 is remarkably induced by insulin.

    “Moreover, Insig1 is remarkably induced by insulin (39, 40).”
    Quote in the paperNot a substantial claim0 of 2 sources checked
    • cites [39] Mohn, 1991, The immediate-early growth response in regenerating liver and insulin-stimulated H-35 cells: comparison with serum-stimulated 3T3 cells and identification of 41 novel immediate-early genes, Mol Cell Biol source
    • cites [40] source

  • Probably not a substantial claim.

    Premisecitation

    Desialylation of the insulin receptor (IR) induces its activation and insulin signal transduction.

    “Notably, desialylation of the insulin receptor (IR) induces its activation and insulin signal transduction (41, 42).”
    Quote in the paperSubstantial claim0 of 2 sources checked
    • cites [41] source

      Checked before — its earlier verdict is held back until a new check runs.

    • cites [42] source
54 of 54

  • The paper relies on this without support.

    Cholesterol and protein chaperones affect the activation of SREBPs.

    “Cholesterol and protein chaperones are known to affect the activation of SREBPs.”

  • The paper relies on this without support.

    The uptake of LDL-c by the liver is the predominant way to clear it from plasma.

    “The uptake of LDL-c by the liver is the predominant way to clear it from plasma.”

  • The paper relies on this without support.

    SREBPs are transported from the ER to the Golgi apparatus for proteolytic double cleavages and activation.

    “SREBPs were transported from the ER to Golgi apparatus for proteolytic double cleavages and activation.”

  • The paper relies on this without support.

    Functions of SREBPs depend on their activation by double proteolytic cleavage.

    “Functions of SREBPs depend on their activation by double proteolytic cleavage.”

  • The paper relies on this without support.

    Transportation of SREBPs from the ER to the Golgi and to the nucleus is the key event involved with many factors for trafficking and cleavage.

    “During the process, the transportation of SREBPs from the ER to the Golgi and to the nucleus is the key event that is involved with many factors for trafficking and cleavage.”
5 of 5

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?

  • Hepatic silencing of Asgr1 in mice proposed that ASGR1 attenuates hepatic SREBP2 signaling and reduces PCSK9 levels.

    “This was similarly observed in a study employing hepatic silencing of Asgr1 in mice [87], which proposed that ASGR1 attenuates hepatic SREBP2 signaling and reduces PCSK9 levels.”

    From The low-density lipoprotein receptor: Emerging post-transcriptional regulatory mechanisms (2025) · cites this paper as [87]

    The Results section on SREBP activation reports that ASGR1-deficient mouse livers had lower nuclear SREBP2, and the LDL-uptake Results section reports lower PCSK9 under ASGR1-deficient conditions. The Results section on restoring ASGR1 expression reports the opposite effect: SREBP1/2 and PCSK9 increased. The cited paper therefore attributes attenuation to loss of ASGR1, not to ASGR1 itself; our copy provides no page numbers for these passages. (LLM, conf=0.90)

    Checked by a reasoning model — may be wrong.

    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.

  • LDL uptake incubation duration differs between Results and Methods

    Major

    The Results say labeled LDL was incubated with hepatic cells for 4 hours, whereas the LDL uptake method specifies 3 hours.

  • Direction of SREBP regulation of PCSK9 is inconsistent

    Minor

    The Discussion calls PCSK9 negatively regulated by SREBPs, but the Results report that PCSK9 decreases when SREBP activation and signaling are suppressed.

  • INSIG1 knockdown reverses lipid changes in opposite directions

    Minor

    The abstract says INSIG1 knockdown reversed the ASGR1-deficient phenotype by increasing serum TC, LDL-c, TG, and liver cholesterol. The Results describe the same reversal as decreases in those measures.

3 of 3

Citations

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1 of the 45 references couldn’t be matched to a known paper.

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Metadata

Authors
Xu, Y., Tao, J., Yu, X., Wu, Y., Chen, Y., You, K., Zhang, J., Getachew, A., Pan, T., Zhuang, Y., Yuan, F., Yang, F., Lin, X., & Li, Y.-X.
Affiliations
  • Chinese Academy of Sciences (CN)
  • University of Science and Technology of China (CN)
  • Center for Health Research, Guangdong Provincial Key Laboratory of Biocomputing, Guangzhou Institutes of Biomedicine and Health, Chinese Academy of Sciences, Guangzhou, China.
ORCID
Venue
JCI Insight
Publisher
American Society for Clinical Investigation
Published
2021-10-08
Volume
6
Issue
19
ISSN
2379-3708 (ISSN-L)
DOI
10.1172/jci.insight.147038
PMC
PMC8525641
OpenAlex
W3203214670
Semantic Scholar
cd41ec78332b3b070560f4aa88b0faa2fc80a9cc
Cited by
45
References
45
Citations by year
2026: 13 · 2025: 8 · 2024: 13 · 2023: 5 · 2022: 6
FWCI
2.42
Type
journal article
Language
en
Open access
Licence
Locations
  • JCI Insight (publishedVersion, cc-by)
  • Europe PMC (PubMed Central) (publishedVersion, cc-by)
  • PubMed Central (publishedVersion, cc-by)
  • PubMed (publishedVersion)
  • DOAJ (DOAJ: Directory of Open Access Journals) (submittedVersion)
Funders
  • the National Key R&D Program of China (2019YFA0111300)
  • the National Natural Science Foundation of China (31871379,31900539)
  • Guangzhou City Science and Technology Plan (202102020223)
  • Guangdong Province Science and Technology Plan (2018A050506070)
  • Frontier Research Program of Guangzhou Regenerative Medicine and Health Guangdong Laboratory (2018GZR110105011)
Topics
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Biochemistry › Cholesterol and Lipid Metabolism
  • Health Sciences › Medicine › Cardiology and Cardiovascular Medicine › Lipid metabolism and disorders
  • Health Sciences › Medicine › Cardiology and Cardiovascular Medicine › Lipoproteins and Cardiovascular Health
Keywords
ASGR1, INSIG1, SREBP signaling, lipid homeostasis, non-HDL cholesterol, triglycerides, LDL cholesterol, VLDL secretion, LDL uptake, microsomal triglyceride transfer protein, PCSK9, apolipoprotein B, endoplasmic reticulum, cardiovascular risk
MeSH terms
Proprotein Convertase 9, Animals, Carrier Proteins, Cell Nucleus, Endoplasmic Reticulum, Homeostasis, Cholesterol, HDL, Cholesterol, LDL, Membrane Proteins, Triglycerides, Cholesterol, VLDL, Signal Transduction, Mice, Knockout, Asialoglycoprotein Receptor, Lipid Metabolism, Mice, Sterol Regulatory Element Binding Proteins

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.

A language model drew the claims from the paper on 2026-11-09, under version 3 of the claim definition (model deepseek-v4.1-flash, prompt extract-v3).

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Cited sources are checked with a language model’s help (gpt-6-sol).

Statistics and arithmetic are recomputed, not judged.

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