Journal: The Journal of Biological Chemistry

Article Title: Apolipoprotein E (APOE) regulates the transport of monosialotetrahexosylganglioside (GM1)

doi: 10.1016/j.jbc.2025.110778

Figure Lengend Snippet: Determination of the interaction affinity between GM1 and APOE. A , schematic illustration of determining the binding affinity between lipid structure and APOE using MST. B , the binding affinity (K D ) between APOE3 or APOE4 and lipid structures containing GM1 or cholesterol (n = 3). C , the binding affinity between APOE3 or APOE4 and lipid structures with different GM1 concentrations. (n = 3). D - H , negative staining images of lipid structures with varying GM1 concentrations. p value: ns (0.05 < p ≤ 1), ∗ (0.01 < p ≤ 0.05, ∗∗ (0.001 < p ≤ 0.01, ∗∗∗ (0.0001 < p ≤ 0.001, ∗∗∗∗ ( p ≤ 0.0001). ( A ) is created with BioRender.com . APOE, apolipoprotein E; MST, microscale thermophoresis.

Article Snippet: We then blocked the cells with 2% bovine serum albumin for 30 min. Next, we added primary APOE recombinant rabbit monoclonal antibody (diluted 1:1000 in 2% bovine serum albumin, Invitrogen) and incubated it for 2 h at room temperature.

Techniques: Binding Assay, Negative Staining, Microscale Thermophoresis

Journal: The Journal of Biological Chemistry

Article Title: Apolipoprotein E (APOE) regulates the transport of monosialotetrahexosylganglioside (GM1)

doi: 10.1016/j.jbc.2025.110778

Figure Lengend Snippet: Analysis of GM1 localization and levels following the cellular uptake of lipid structures. A , schematic illustration of the method used to determine GM1 following the cellular uptake. B , GM1 levels in differentiated PC-12 cells following cellular uptake, determined using the SpectraMax i3 or by quantifying confocal microscopy images. (n ≥ 3) ( C ) representative images showing GM1 and APOE localization in differentiated PC-12 cells following cellular uptake, as determined by confocal microscopy. (n ≥ 6) ( D ) GM1 levels in HEK-293 cells following cellular uptake, determined using the SpectraMax i3 or by quantifying confocal microscopy images. (n ≥ 6) ( E ) (1) cellular uptake of DiD-labeled cholesterol and GM1 lipid structures in U-87 MG cells, and (2) changes in cholesterol levels in U-87 MG cells after cellular uptake of APOE3 and APOE4-enriched cholesterol lipoproteins, as well as a mixture of APOE3 and APOE4-enriched cholesterol and GM1 lipoproteins. (n ≥ 6). p value: ns (0.05 < p ≤ 1), ∗ (0.01 < p ≤ 0.05, ∗∗ (0.001 < p ≤ 0.01, ∗∗∗ (0.0001 < p ≤ 0.001, ∗∗∗∗ ( p ≤ 0.0001). ( A ) is created with BioRender.com . Created in BioRender. Dokholyan, N. (2025) https://BioRender.com/g17z389 . APOE, apolipoprotein E; HEK, human embryonic kidney; PC, l -α-phosphatidylcholine.

Article Snippet: We then blocked the cells with 2% bovine serum albumin for 30 min. Next, we added primary APOE recombinant rabbit monoclonal antibody (diluted 1:1000 in 2% bovine serum albumin, Invitrogen) and incubated it for 2 h at room temperature.

Techniques: Confocal Microscopy, Labeling

Journal: The Journal of Biological Chemistry

Article Title: Apolipoprotein E (APOE) regulates the transport of monosialotetrahexosylganglioside (GM1)

doi: 10.1016/j.jbc.2025.110778

Figure Lengend Snippet: Western blot results of APOE receptors: LDLR, VLDLR, LRP1, and ApoER2 on differentiated PC-12 cells, U-87 MG, bEnd.3, and HEK-293 cells. A , the expression of LDLR on differentiated PC-12 cells, U-87 MG, bEnd.3, and HEK-293 cells (n = 4). B , the expression of VLDLR on differentiated PC-12 cells, U-87 MG, bEnd.3, and HEK-293 cells (n = 4). C , the expression of LRP1 on differentiated PC-12 cells, U-87 MG, bEnd.3, and HEK-293 cells (n = 3). D , the expression of ApoER2 on differentiated PC-12 cells, U-87 MG, bEnd.3, and HEK-293 cells (n = 3). E , schematic illustration of determining the changes of APOE secondary structures. F , the CD results of APOE 3 under the effect of different GM1 content on the lipid structures (n ≥ 3). G , the CD results of APOE 4 under the effect of different GM1 content on the lipid structures (n = 3). p value: ns (0.05 < p ≤ 1), ∗ (0.01 < p ≤ 0.05, ∗∗ (0.001 < p ≤ 0.01, ∗∗∗ (0.0001 < p ≤ 0.001, ∗∗∗∗ ( p ≤ 0.0001). ( E ) is created with BioRender.com . APOE, apolipoprotein E; ApoER2, APOE receptor 2; HEK, human embryonic kidney; LDLR, low-density lipoprotein receptor; LRP1, LRPR-related protein 1; PC, l -α-phosphatidylcholine; VLDLR, very low-density lipoprotein receptor.

Article Snippet: We then blocked the cells with 2% bovine serum albumin for 30 min. Next, we added primary APOE recombinant rabbit monoclonal antibody (diluted 1:1000 in 2% bovine serum albumin, Invitrogen) and incubated it for 2 h at room temperature.

Techniques: Western Blot, Expressing

Journal: The Journal of Biological Chemistry

Article Title: Apolipoprotein E (APOE) regulates the transport of monosialotetrahexosylganglioside (GM1)

doi: 10.1016/j.jbc.2025.110778

Figure Lengend Snippet: The binding affinity of APOE-enriched lipoprotein and APOE receptor LDLR using MST. A and B , the binding affinity between APOE3-enriched lipoprotein with varying GM1 concentration to LDLR. (n = 3) ( C and D ) the binding affinity between APOE4-enriched lipoprotein varying GM1 concentration to LDLR (n = 3). p value: ns (0.05 < p ≤ 1), ∗ (0.01 < p ≤ 0.05, ∗∗ (0.001 < p ≤ 0.01, ∗∗∗ (0.0001 < p ≤ 0.001, ∗∗∗∗ ( p ≤ 0.0001). APOE, apolipoprotein E; LDLR, low-density lipoprotein receptor; MST, microscale thermophoresis.

Article Snippet: We then blocked the cells with 2% bovine serum albumin for 30 min. Next, we added primary APOE recombinant rabbit monoclonal antibody (diluted 1:1000 in 2% bovine serum albumin, Invitrogen) and incubated it for 2 h at room temperature.

Techniques: Binding Assay, Concentration Assay, Microscale Thermophoresis

Journal: Journal of the American Heart Association

Article Title: Hepatic Abnormal Secretion of Apolipoprotein C3 Promotes Inflammation in Aortic Dissection

doi: 10.1161/jaha.124.037172

Figure Lengend Snippet: Figure 1. Apo C3 is abundant in the plasma of patients with AD and deposits in the aorta. A, Heat map results of plasma exosome sequencing in patients with AD (n=20) and normal controls (n=20). B, ELISA results of apo C3 in the plasma of patients with AD (n=20) and NCs (n=20). C, Representative western blot of apo C3 protein in the aortas of patients with AD (n=6) and NCs (n=6). D, Representative immunohistochemistry of apo C3 in the aortas of patients with AD (n=8) and normal controls (n=8). E, Representative immunofluorescence of apo C3 in the aortas of patients with AD (n=9) and normal controls (n=9). F through H, Quantitative protein levels of apo C3 in western blot (C), immunohistochemistry (D), and immunofluorescence (E). Data are expressed as mean±SEM. Bars represent the means, and caps represent the SEM. Statistical comparisons were made using an unpaired t test (B, F through H). A value of P<0.05 was considered significant. The layers of the aorta are denoted as I for intima, M for media, and A for adventitia. AD indicates aortic dissection; apo C3 apolipoprotein C3; α-SMA, α-smooth muscle actin; CD: cluster of differentiation; ENO, enolase; LPL, lipoprotein lipase; MMP, matrix metalloproteinase; NC, normal control; and RBP, retinol-binding protein.

Article Snippet: Primary antibodies against apo C3 (1:100; Affinity; DF8054), TLR2 (1:100; Proteintech; 17 236- 1- AP), NLRP3 (1:100; Invitrogen; MA5- 23919), CD86 (1:100, Proteintech; 13 395- 1- AP), INOS (1:100; Invitrogen; PA1- 036), MMP2 (1:100; Proteintech; 10 373- 2- AP), MMP9 (1:100; Proteintech; 10 375- 2- AP), CD68 (1:100; Invitrogen; 14- 0688- 82), F4/80 (1:100 Invitrogen; 14–4801- 82), CD31(1:100; Abcam; ab9498) and α- smooth muscle actin (1:100; Boster; BM4172), were incubated overnight at 4°C.

Techniques: Clinical Proteomics, Sequencing, Enzyme-linked Immunosorbent Assay, Western Blot, Immunohistochemistry, Immunofluorescence, Dissection, Control, Binding Assay

Journal: Journal of the American Heart Association

Article Title: Hepatic Abnormal Secretion of Apolipoprotein C3 Promotes Inflammation in Aortic Dissection

doi: 10.1161/jaha.124.037172

Figure Lengend Snippet: Figure 3. Transcriptome sequencing reveals inhibition of TLR2/NLRP3 pathway activation, M1 macrophage polarization, and MMP release in the aortas of mice after hepatic apo C3 interference. A through E, Results of transcriptome sequencing in the aortas of mice in the BAPN group (n=5) and BAPN+Sh-apo C3 group (n=5). A, Volcano plot. B, Heat map. C, Protein–protein interaction network of apo C3 with differentially expressed genes. D, GO analysis results. E, KEGG analysis results. apo C3 indicates apolipoprotein C3; BAPN, β-aminopropionitrile; GO, gene ontology; KEGG, Kyoto Encyclopedia of Genes and Genomes; MMP, matrix metalloproteinase; NLRP3, NOD-like receptor pyrin domain containing 3; and TLR2, Toll-like receptor 2.

Article Snippet: Primary antibodies against apo C3 (1:100; Affinity; DF8054), TLR2 (1:100; Proteintech; 17 236- 1- AP), NLRP3 (1:100; Invitrogen; MA5- 23919), CD86 (1:100, Proteintech; 13 395- 1- AP), INOS (1:100; Invitrogen; PA1- 036), MMP2 (1:100; Proteintech; 10 373- 2- AP), MMP9 (1:100; Proteintech; 10 375- 2- AP), CD68 (1:100; Invitrogen; 14- 0688- 82), F4/80 (1:100 Invitrogen; 14–4801- 82), CD31(1:100; Abcam; ab9498) and α- smooth muscle actin (1:100; Boster; BM4172), were incubated overnight at 4°C.

Techniques: Sequencing, Inhibition, Activation Assay