Journal: Frontiers in Immunology

Article Title: APOE -associated lipid-handling macrophages in hepatocellular carcinoma: ligand–receptor communication and host Apoe -linked myeloid remodeling

doi: 10.3389/fimmu.2026.1902143

Figure Lengend Snippet: Apoe deficiency increases neutral lipid accumulation in lipid-loaded BMDMs (A) Schematic overview of BMDM isolation and BODIPY 493/503 staining. Bone marrow cells were isolated from WT and Apoe −/− mice and differentiated into BMDMs with M-CSF for 7 days. (B–D) Representative flow cytometry histograms and quantification of BODIPY 493/503 signal in WT and Apoe −/− BMDMs under untreated conditions (B) , after OA treatment (200 μM, 24 h) (C) , and after oxLDL treatment (50 μg/mL, 24 h) (D) . BODIPY 493/503 fluorescence was quantified as background-subtracted median fluorescence intensity (ΔMedian MFI). Data are shown as mean ± SEM. n = 3 biological replicates per group. P values were calculated using unpaired two-tailed Welch’s t-tests. ns, not significant; ** P < 0.01 ; *** P < 0.001 . OA, oleic acid; oxLDL, oxidized low-density lipoprotein; M-CSF, macrophage colony-stimulating factor; BMDMs, bone marrow-derived macrophages.

Article Snippet: BMDMs were left untreated or treated with oleic acid (OA; 200 μM; Xi’an Kunchuang Technology Development Co., Ltd., KC006) or human oxLDL (50 μg/mL; Yeasen Biotechnology, 20605ES) for 24 h. Cells were stained with BODIPY 493/503 (2 μM; MedChemExpress, HY-D1614) for 15 min at 37 °C, washed with PBS, and analyzed by flow cytometry.

Techniques: Isolation, Staining, Flow Cytometry, Fluorescence, Two Tailed Test, Derivative Assay

Journal: Current Biology

Article Title: C. elegans Eats Its Own Intestine to Make Yolk Leading to Multiple Senescent Pathologies

doi: 10.1016/j.cub.2018.06.035

Figure Lengend Snippet: Pathophysiology of Yolk Steatosis in Aging C. elegans (A) Inhibition of yolk uptake by rme-2 RNAi aggravates PLP accumulation; N = 2 trials. (B) Absence of yolk synthesis prevents PLP accumulation in aging males; N = 2 trials. (C) PLPs contain neutral lipids and yolk proteins YP115/YP88 as revealed by co-localization (far right) of Bodipy (center-left) and VIT-6::mCherry (center-right). (D) Estimation of lipid content from measuring intracellular yolk particle (Yp), lipid droplet (LD), and extracellular PLP areas in transmission electron micrographs (TEMs) of mid-body sections reveals a 3.75-fold increase between day 1 and 7 (left), mostly explained by a 15-fold rise in combined PLP area (center) but also by a 3-fold increase in intracellular lipids (right). (E) 8-fold increase in TAG content between day 1 and 7 of adulthood. (F) Lifetime increase in vitellogenins, from SDS-PAGE analysis of wild-type worm lysates. (G) Inhibition of vitellogenin synthesis reduces PLP accumulation. (H) Inhibition of lipid synthesis by fat-6/7 RNAi reduces PLP accumulation. Worms were raised on E. coli B OP50, except for (A), (G), and (H), where E. coli K12 HT115 was used. ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001. See also and and and .

Article Snippet: BODIPY 493/503 (4,4-difluoro-1,3,5,7,8-pentamethyl-4-bora-3a,4a-diaza-s-indacene) , Fisher Scientific, Loughborough, United Kingdom , #D3922.

Techniques: Inhibition, Transmission Assay, SDS Page

Journal: Current Biology

Article Title: C. elegans Eats Its Own Intestine to Make Yolk Leading to Multiple Senescent Pathologies

doi: 10.1016/j.cub.2018.06.035

Figure Lengend Snippet: Evidence of Conversion of Intestinal Biomass into Yolk (A) Total protein content of wild-type worms across the course of a lifetime, peaking at day 6. (B) The proportion of YP to total proteins increases 4-fold between day 1 and 7 (gel densitometric analysis). (C) Inhibition of yolk synthesis by vit-5/6 RNAi (left) or lrp-2 or ceh-60 mutations (right) rescues intestinal atrophy; N = 2 trials. All mutants are significantly different from wild-type (p < 0.05) at 2 or more time points. (D) Wild-type males show no gut atrophy (left), whereas yolk-producing mab-3(mu15) males exhibit PLPs and gut atrophy (right). (E) Quantitation of PLP accumulation (top) and gut atrophy (bottom) in mab-3 males; N = 2 trials. Asterisks indicate statistical comparison between mab-3; him-5 males and him-5 or wild-type males. (F) TEM of mid-body sections reveals a redistribution of intestinal lipids to other tissues between day 1 and 7. (G) Lipid redistribution confirmed by analysis of neutral lipid staining by Bodipy. (H) Numbers of DHS-3::GFP-marked lipid droplets decrease with age. (I) Inhibition of yolk uptake by rme-2 RNAi, which aggravates PLP accumulation, does not affect intestinal atrophy. Trials were conducted at 20°C, no FUDR (A–E) or 25°C with FUDR (F–I). ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001. See also Figure S4 .

Article Snippet: BODIPY 493/503 (4,4-difluoro-1,3,5,7,8-pentamethyl-4-bora-3a,4a-diaza-s-indacene) , Fisher Scientific, Loughborough, United Kingdom , #D3922.

Techniques: Inhibition, Quantitation Assay, Comparison, Staining

Journal: Current Biology

Article Title: C. elegans Eats Its Own Intestine to Make Yolk Leading to Multiple Senescent Pathologies

doi: 10.1016/j.cub.2018.06.035

Figure Lengend Snippet: Inhibition of atg-13 Reduces Age-Associated Lipid Redistribution (A) Adult-limited atg-13 RNAi delays the age-associated decrease in DHS-3::GFP-labeled intestinal lipid droplets. Images showing labeling of intestinal lipid droplets using DHS-3::GFP in control and atg-13 RNAi animals (left). Quantification of DHS-3::GFP-labeled intestinal lipid droplets (right). (B) Age-associated lipid redistribution is inhibited by atg-13(bp414) . Bodipy staining of neutral lipids in control and atg-13 RNAi animals reveals preservation of Bodipy-labeled intestinal lipid content (cf. Figure 3 G). (C) Quantification of lipidic organelles using TEM. Age-related increase in lipidic area is suppressed by atg-13(bp414) (top). Relative intestinal lipidic organelle content is preserved during aging in atg-13(bp414) compared to wild-type (middle and bottom). (D) TEM images at 3,000× (top) and 10,000× (bottom) of wild-type (left and center) and atg-13(bp414) (right) worm sections on day 7 of adulthood. atg-13(bp414) worms have reduced PLPs and increased intestinal lipid droplets compared to wild-type. ∗ p < 0.05, ∗∗∗ p < 0.001. See also .

Article Snippet: BODIPY 493/503 (4,4-difluoro-1,3,5,7,8-pentamethyl-4-bora-3a,4a-diaza-s-indacene) , Fisher Scientific, Loughborough, United Kingdom , #D3922.

Techniques: Inhibition, Labeling, Control, Staining, Preserving

Journal: Current Biology

Article Title: C. elegans Eats Its Own Intestine to Make Yolk Leading to Multiple Senescent Pathologies

doi: 10.1016/j.cub.2018.06.035

Figure Lengend Snippet: Reduced Insulin/IGF-1 Signaling Inhibits Gut-to-Yolk Biomass Conversion (A) PLP accumulation (left) and intestinal atrophy (center) are suppressed modestly by daf-2(e1368) and strongly by daf-2(e1370) , which also has a larger effect on lifespan (right), confirming findings in a recent study . (B) YP levels and YP as proportion of total protein in daf-2(e1368) (left) and daf-2(e1370) (right) (cf. F and B). (C) PLP accumulation (top) and intestinal atrophy (bottom) in daf-2(e1370) require daf-16 ; N = 2 trials. (D) Age-associated neutral lipid redistribution (Bodipy) is suppressed in daf-2(e1370) mutants, and this requires daf-16 . ∗ p < 0.05, ∗∗∗ p < 0.001, &&& p < 0.001, compared to age-matched daf-16 , daf-16; daf-2 , or wild-type (cf. Figure 3 G). (E) RNAi of atg-2 , atg-13 , and atg-18 all suppress intestinal atrophy (top), whereas atg-2 and atg-13 also increase lifespan (bottom; p < 0.01 all trials; N = 2 and 5, respectively) in a daf-16(mgDf50) mutant (25°C, FUDR). ∗∗∗ p < 0.001. (F) Proposed mechanism of antagonistically pleiotropic action of IIS promoting reproduction at the expense of senescent pathologies, through gut-to-yolk biomass conversion. See also Figure S7 .

Article Snippet: BODIPY 493/503 (4,4-difluoro-1,3,5,7,8-pentamethyl-4-bora-3a,4a-diaza-s-indacene) , Fisher Scientific, Loughborough, United Kingdom , #D3922.

Techniques: Mutagenesis

Journal: Current Biology

Article Title: C. elegans Eats Its Own Intestine to Make Yolk Leading to Multiple Senescent Pathologies

doi: 10.1016/j.cub.2018.06.035

Figure Lengend Snippet:

Article Snippet: BODIPY 493/503 (4,4-difluoro-1,3,5,7,8-pentamethyl-4-bora-3a,4a-diaza-s-indacene) , Fisher Scientific, Loughborough, United Kingdom , #D3922.

Techniques: Virus

Journal: Nature Communications

Article Title: Grasp55 −/− mice display impaired fat absorption and resistance to high-fat diet-induced obesity

doi: 10.1038/s41467-020-14912-x

Figure Lengend Snippet: a , b Measurements of ATGL in the lipid droplet (LD) fraction of Caco-2 cells after treatment with 400 μM oleic acid for 16 h. Representative immunoblots are shown in ( a ) and a summary of multiple experiments is shown in ( b , n = 3). c Presence of ATGL on the LD surface was examined by co-staining with anti-ATGL antibodies and BODIPY 493/503 (an LD marker) in Caco-2 cells, in which endogenous GRASP55 was depleted ( shGRASP55, shG55 ) and/or exogenously supplemented ( GRASP55-Myc, G55-Myc ). Cells were treated with 400 μM oleic acid for 4 h before immunostaining. Arrows indicate the location of ATGL, which is targeted to the peripheral regions of LDs. Four independent experiments showed similar results. d , e ATGL protein stability was examined in Caco-2 cells, in which GRASP55 was depleted ( shGRASP55 , open arrowhead) and/or exogenously supplemented ( GRASP55-Myc , filled arrowhead). Cytosolic proteins were collected 0, 2, and 4 h after treatment with cycloheximide (0.1 mg ml −1 ), an inhibitor of protein biosynthesis. Aldolase A was used as a cytosolic protein loading control. Representative immunoblot is shown in ( d ), and the results of multiple experiments are summarized in ( e , n = 3). f Immunoblot results of Caco-2 monolayers, in which GRASP55 was depleted ( shGRASP55 ) and exogenous ATGL ( ATGL-Myc ) was supplemented. Levels of endogenous ATGL (open arrowhead) and exogenous ATGL (filled arrowhead) are shown. A high level of ATGL-Myc supplementation restored LD-associated ATGL levels in GRASP55-depleted cells, although the LD transport efficiency was much lower (compare endogenous and exogenous ATGL levels in lanes 3 and 4). g Transepithelial lipid transport of Caco-2 monolayers were determined using 14 C-labeled oleic acid. The basolateral (B) to apical (A) transport of oleic acid (left, B to A) was much smaller than that of apical to basolateral transport (middle, A to B), and it was not affected by GRASP55 depletion or ATGL overexpression. Transepithelial lipid absorption was determined by subtracting values of (B to A) from those of (A to B) in each paired experiment (right, n = 4). ATGL overexpression rescued lipid absorption in GRASP55-deficient Caco-2 Cells. Unprocessed blots are presented in Supplementary Fig. . CPM, counts per min. TM, total membrane. Data are shown as mean ± SEM. Scale bars: 10 μm. n.s.: not significant, * p < 0.05. P values were caluculated by paired ( b ), unpaired ( e ) two-tailed Student’s t tests or ANOVA followed by Tukey’s multiple comparison tests ( g ). Source data are provided as a Source Data file.

Article Snippet: For LD staining, cells were also stained with BODIPY 493/503 (Merck-Millipore).

Techniques: Western Blot, Staining, Marker, Immunostaining, Control, Labeling, Over Expression, Membrane, Two Tailed Test, Comparison

Journal: Nature Communications

Article Title: Grasp55 −/− mice display impaired fat absorption and resistance to high-fat diet-induced obesity

doi: 10.1038/s41467-020-14912-x

Figure Lengend Snippet: The dGrasp -null mutant allele ( dGrasp −/− ), also known as dGrasp 302 , harbors a deletion of the whole dGrasp coding region. a , b Lipid accumulation in enterocytes of posterior midguts of dGrasp −/− flies. Enterocytes of NP1>mCD8::RFP fly guts were stained with antibodies against DsRed (green) and nuclei were counterstained with TO-PRO-3 (blue). Lipids were visualized by staining with BODIPY solution (red). For consistency, pseudocolors were used. Confocal microscopic images of whole guts from control ( NP1>mCD8::RFP ) and dGrasp -mutant flies ( NP1>mCD8::RFP;dGrasp −/− ) are shown in ( a ). Scale bars: 1 mm. Boxed area indicates the posterior midgut where lipids accumulated. Image sections of the boxed area taken from the top (upper, scale bar: 25 μm) and lateral (bottom, scale bar: 10 μm) orientations are shown in ( b ). c , d Confocal microscopic images of lipid accumulation in the fly guts of the indicated genotypes. The fly genes dGrasp and bmm are homologs of mammalian GRASP and ATGL , respectively. Each gene was expressed in the enterocytes of dGrasp −/− flies under the control of NP1-GAL4. Lipids were visualized by staining with Nile Red (red) and nuclei were counterstained with TO-PRO-3 (blue). Representative images of the posterior midguts are shown in ( c ), and quantitative analyses of intestinal lipid accumulation in the indicated genotypes of the flies are presented in ( d ), n = 4–6. Scale bars: 50 μm. e Quantitative analyses of relative whole-body TG contents in the indicated genotypes of the flies ( n = 5). UAS, upstream activation sequence. Data are shown as mean ± SEM. * p < 0.05, ** p < 0.01. All p values were calculated by ANOVA followed by Tukey’s multiple comparison tests. Source data are provided as a Source Data file.

Article Snippet: For LD staining, cells were also stained with BODIPY 493/503 (Merck-Millipore).

Techniques: Mutagenesis, Staining, Control, Activation Assay, Sequencing, Comparison

Journal: Journal of Immunology Research

Article Title: Tuftelin 1 Facilitates Hepatocellular Carcinoma Progression through Regulation of Lipogenesis and Focal Adhesion Maturation

doi: 10.1155/2022/1590717

Figure Lengend Snippet: TUFT1 expression promoted lipid accumulation in HCC cells. (a) GSEA analysis showed that TUFT1 was involved in fatty acid metabolism. (b) Analysis of FFA level in TUFT1 overexpression or knockdown cells. (c) Lipid accumulation detected by oil red O and BODIPY 493/503 dye staining, in TUFT1 overexpression or knockdown cells. Scale bar: 10 μ m.

Article Snippet: For BODIPY 493/503 dye staining, HCC cells were seeded in a 12-well u-Chamber (ibidi, Germany).

Techniques: Expressing, Over Expression, Staining

Journal: Journal of Immunology Research

Article Title: Tuftelin 1 Facilitates Hepatocellular Carcinoma Progression through Regulation of Lipogenesis and Focal Adhesion Maturation

doi: 10.1155/2022/1590717

Figure Lengend Snippet: Inhibition of CREB1 reversed the promotion of cell proliferation by TUFT1 overexpression. (a) Western blot analysis of FASN and SCD in TUFT1-overexpressing or knockdown cells. (b) Western blot analysis of p-CREB1, FASN, and SCD in TUFT1-overexpressing Huh7 cells, treated with KG-501 or vehicle control. (c) Upper panel: immunofluorescence of p-CREB1 (Ser133) in vector, TUFT1 overexpression, and KG-501-treated TUFT1-overexpressing Huh7 cells; down panel: BODIPY 493/503 dye staining in these cells. Scale bar: 10 μ m. (d) Cell proliferation analysis of Huh7 cells, treated with KG-501 or vehicle control. ∗∗ P < 0.01.

Article Snippet: For BODIPY 493/503 dye staining, HCC cells were seeded in a 12-well u-Chamber (ibidi, Germany).

Techniques: Inhibition, Over Expression, Western Blot, Immunofluorescence, Plasmid Preparation, Staining