Journal: Cell

Article Title: The Bone Marrow Protects and Optimizes Immunological Memory during Dietary Restriction

doi: 10.1016/j.cell.2019.07.049

Figure Lengend Snippet: KEY RESOURCES TABLE

Article Snippet: N/A Mouse: Cxcr4 fl/fl x UBC-CRE ERT2 Laboratory of S.R.S N/A Mouse: S1pr1 fl/fl x UBC-CRE ERT2 Laboratory of S.R.S N/A Mouse: B6Cg.Thy1a/Cy Tg(TcraTcrb)8Rest/J Jackson laboratories Mouse strain: Jax, 005023 Oligonucleotides Hprt Thermofisher 4331182 (Mm03024075_m1) Fabp4 Thermofisher 4331182 (Mm00445878_m1) Software and Algorithms Flowjo software Treestar RRID: SCR_008520 Imaris software Bitplane RRID: SCR_007370 Prism software Graphpad RRID: SCR_002798 ImageJ https://imagej.net/ RRID: SCR_003070 STAR aligner Dobin et al., 2013 RRID: SCR_015899 HOMER http://homer.ucsd.edu/ RRID: SCR_010881 DESeq2 https://bioconductor.org/packages/release/bioc/html/DESeq2.html RRID: SCR_015687 Metascape http://metascape.org/gp/index.html#/main/step1 RRID: SCR_016620 GSEA http://www.broadinstitute.org/gsea/ RRID: SCR_003199 FastQC Babraham Bioinformatics RRID: SCR_014583 R https://www.r-project.org/ N/A Open in a separate window KEY RESOURCES TABLE Dietary restriction promotes memory T cell accumulation in BM BM trophic factors and adipocytes promote memory T cell accumulation in BM Memory T cells display enhanced protective function during dietary restriction

Techniques: Purification, Virus, Recombinant, Expressing, Electron Microscopy, Control, Isolation, Staining, Enzyme-linked Immunosorbent Assay, Software

Journal: bioRxiv

Article Title: Endothelial FABP4 constitutes the majority of basal circulating hormone levels and regulates lipolysis-driven insulin secretion

doi: 10.1101/2022.10.13.511807

Figure Lengend Snippet: ( A ) Left panel: Representative immunoblot of FABP4 expression in perirenal adipose tissue of WT, Adipo-KO, Endo-KO, and Total-KO mice. Right panel: Quantification of FABP4 relative to β-tubulin signal.***p<0.001, **p<0.01. ( B ) FABP4 immunostaining in perigonadal adipose tissue from WT, Adipo-KO, Endo-KO, and Total-KO mice. 40X magnification. ( C ) Left panel: Immunoblot of FABP4 expression in isolated PGWAT adipocytes of WT, Adipo-KO, Endo-KO, and Total-KO mice. Right panel: Quantification of FABP4 relative to β-tubulin signal.*p<0.05, **p<0.01. n=1 for Total-KO. Data were analyzed by one-way ANOVA followed by Tukey’s multiple comparison test and are presented as mean ± SEM. ND = No signal detected.

Article Snippet: For in-house FABP4 ELISA we used anti-FABP4 antibodies produced for the Hotamışlıgil lab by the Dana Farber Antibody Core (clone 351.4.6H7.G3.G9 for capture, HRP-tagged clone 351.4.5E1.H3 for detection) and recombinant human FABP4 as a standard (R&D Systems, cat# DY3150-05).

Techniques: Western Blot, Expressing, Immunostaining, Isolation, Comparison

Journal: bioRxiv

Article Title: Endothelial FABP4 constitutes the majority of basal circulating hormone levels and regulates lipolysis-driven insulin secretion

doi: 10.1101/2022.10.13.511807

Figure Lengend Snippet: ( A ) Left panel: Immunoblot of FABP4 expression in isolated liver endothelial cells of WT, Adipo-KO, and Endo-KO mice. Right panel: Quantification of FABP4 relative to β-tubulin signal. ( B ) FABP4 immunostaining in liver from WT, Adipo-KO, Endo-KO, and Myeloid-KO mice. 20X magnification. ND = No signal detected.

Article Snippet: For in-house FABP4 ELISA we used anti-FABP4 antibodies produced for the Hotamışlıgil lab by the Dana Farber Antibody Core (clone 351.4.6H7.G3.G9 for capture, HRP-tagged clone 351.4.5E1.H3 for detection) and recombinant human FABP4 as a standard (R&D Systems, cat# DY3150-05).

Techniques: Western Blot, Expressing, Isolation, Immunostaining

Journal: bioRxiv

Article Title: Endothelial FABP4 constitutes the majority of basal circulating hormone levels and regulates lipolysis-driven insulin secretion

doi: 10.1101/2022.10.13.511807

Figure Lengend Snippet: ( A) Top panel: Plasma FABP4 levels in WT, Adipo-KO, Endo-KO, and Total-KO lean male mice, after 6h daytime food withdrawal. Data are pooled from samples from 9 experiments. ****p<0.0001, **p<0.01. Bottom panel: Immunoblots of perirenal and mesenteric adipose FABP4 protein expression (same images as in and S2A), for comparison with plasma levels. ( B ) Plasma FABP4 levels in 6h WT vs. Myeloid-KO mice. Data are pooled from samples from 2 experiments. ( C ) Plasma FABP5 levels in WT, Adipo-KO, Endo-KO and Total-KO mice. *p<0.05. Data in 3A, 3C, and 3D were analyzed by one-way ANOVA, followed by Tukey’s multiple comparison test. Data in 3B were analyzed by unpaired t-test. Data are presented as mean ± SEM. ND: No signal detected.

Article Snippet: For in-house FABP4 ELISA we used anti-FABP4 antibodies produced for the Hotamışlıgil lab by the Dana Farber Antibody Core (clone 351.4.6H7.G3.G9 for capture, HRP-tagged clone 351.4.5E1.H3 for detection) and recombinant human FABP4 as a standard (R&D Systems, cat# DY3150-05).

Techniques: Clinical Proteomics, Western Blot, Expressing, Comparison

Journal: bioRxiv

Article Title: Endothelial FABP4 constitutes the majority of basal circulating hormone levels and regulates lipolysis-driven insulin secretion

doi: 10.1101/2022.10.13.511807

Figure Lengend Snippet: ( A ) Plasma FABP4, ( B ) Non-esterified fatty acid (NEFA), and ( C ) glycerol responses to 10mg/kg isoproterenol-induced lipolysis in WT, Adipo-KO, Endo-KO, and Total-KO male mice. FABP4 responses are presented as induction over baseline. Data are pooled from 6 separate experiments. WT n=52, Adipo-KO n=39, Endo-KO n=35, Total-KO n=8 for , n=15 for , . ****p<0.0001, ***p<0.001, **p<0.01, *p<0.05 vs. WT; °°p<0.01 vs. Adipo-KO, °p<0.05 vs. Adipo-KO. ( D ) NEFA, ( E ) glycerol, and ( F ) FABP4 responses to forskolin (FSK)-induced lipolysis in perigonadal adipose explants from male WT, Adipo-KO, and Endo-KO mice. Data are normalized to amount of adipose tissue per culture well. n=4 mice/group, 3 replicates per mouse. ****p<0.0001, ***p<0.001 vs. Adipo-KO. Data were analyzed by repeated measures two-way ANOVA followed by Tukey’s multiple comparison test and are presented as mean ± SEM. ND: No signal detected.

Article Snippet: For in-house FABP4 ELISA we used anti-FABP4 antibodies produced for the Hotamışlıgil lab by the Dana Farber Antibody Core (clone 351.4.6H7.G3.G9 for capture, HRP-tagged clone 351.4.5E1.H3 for detection) and recombinant human FABP4 as a standard (R&D Systems, cat# DY3150-05).

Techniques: Clinical Proteomics, Comparison

Journal: bioRxiv

Article Title: Endothelial FABP4 constitutes the majority of basal circulating hormone levels and regulates lipolysis-driven insulin secretion

doi: 10.1101/2022.10.13.511807

Figure Lengend Snippet: ( A ) Plasma FABP4 responses to 10mg/kg isoproterenol-induced lipolysis in WT and Myeloid FABP4-KO mice, n=8/group, and ( B ) in WT vs. mice with deletion of FABP4 in both adipocytes and endothelial cells (Adipo Endo-KO). n=6/group. ****p<0.0001, **p<0.01. Data were analyzed by repeated measures two-way ANOVA followed by Tukey’s multiple comparison test and are presented as mean ± SEM.

Article Snippet: For in-house FABP4 ELISA we used anti-FABP4 antibodies produced for the Hotamışlıgil lab by the Dana Farber Antibody Core (clone 351.4.6H7.G3.G9 for capture, HRP-tagged clone 351.4.5E1.H3 for detection) and recombinant human FABP4 as a standard (R&D Systems, cat# DY3150-05).

Techniques: Clinical Proteomics, Comparison

Journal: bioRxiv

Article Title: Endothelial FABP4 constitutes the majority of basal circulating hormone levels and regulates lipolysis-driven insulin secretion

doi: 10.1101/2022.10.13.511807

Figure Lengend Snippet: ( A ) 4-hour conditioned media FABP4 levels from CD31-isolated liver, heart, and lung endothelial cells of WT, Adipo--KO, and Endo-KO mice. FABP4 levels are normalized to total cellular protein levels. *p<0.05. ( B ) FABP5 levels in CD31-isolated liver endothelial cell 24-hour conditioned media and (C ) lysates of WT and Endo-KO mice. Data in 6A were analyzed by one-way ANOVA, followed by Tukey’s multiple comparison test and are presented as mean ± SEM. ND: No signal detected.

Article Snippet: For in-house FABP4 ELISA we used anti-FABP4 antibodies produced for the Hotamışlıgil lab by the Dana Farber Antibody Core (clone 351.4.6H7.G3.G9 for capture, HRP-tagged clone 351.4.5E1.H3 for detection) and recombinant human FABP4 as a standard (R&D Systems, cat# DY3150-05).

Techniques: Isolation, Comparison

Journal: bioRxiv

Article Title: Endothelial FABP4 constitutes the majority of basal circulating hormone levels and regulates lipolysis-driven insulin secretion

doi: 10.1101/2022.10.13.511807

Figure Lengend Snippet: ( A ) FABP4 in HUVEC lysates normalized to total cellular protein ( B ) and in 5-hour conditioned media normalized to total cellular protein at days 3 through 14 post-seeding. Pool of 2 experiments. *p<0.05, **p<0.01. ( C ) Time-course of cumulative FABP4 levels in media of day 7 HUVECs. n=4/time point. Inset: Media lactate dehydrogenase (LDH) levels during the same time course. n=4. ( D ) Effects of increasing doses of the ER-Golgi pathway inhibitor, brefeldin A (BFA) on FABP4 and endothelin-1 (ET-1) secretion from day 11 HUVECs. n=3 per BFA dose. *p<0.05. ( E ) Effects of increasing doses of forskolin (FSK) on FABP4 secretion in HUVECs vs. 3T3-L1 adipocytes. n=3 per FSK dose. *p<0.05, **p<0.01 vs. HUVEC. ( F ) Effects of increasing doses of FSK on FABP4 and Von Willibrand Factor (vWF) secretion from day 11 HUVECs. n=3 per FSK dose. *p<0.05, **p<0.01. 7A-C were analyzed by one-way ANOVA followed by Tukey’s multiple comparison test. 7D-F were analyzed by repeated measures 2-way ANOVA with Geisser-Greenhouse correction, followed by Dunnett’s (D,F) or Sidak’s (E) multiple comparison test. Data are presented as mean ± SEM.

Article Snippet: For in-house FABP4 ELISA we used anti-FABP4 antibodies produced for the Hotamışlıgil lab by the Dana Farber Antibody Core (clone 351.4.6H7.G3.G9 for capture, HRP-tagged clone 351.4.5E1.H3 for detection) and recombinant human FABP4 as a standard (R&D Systems, cat# DY3150-05).

Techniques: Comparison

Journal: bioRxiv

Article Title: Endothelial FABP4 constitutes the majority of basal circulating hormone levels and regulates lipolysis-driven insulin secretion

doi: 10.1101/2022.10.13.511807

Figure Lengend Snippet: ( A ) Plasma insulin responses to 10mg/kg isoproterenol-induced lipolysis in WT, Adipo-KO, Endo-KO, and Total-KO mice. Data are pooled from 6 experiments. ****p<0.0001, ***p<0.001, **p<0.01, *p<0.05 vs. WT. °°°°p<0.0001, °°°p<0.001, °°p<0.01, °p<0.05 vs. Adipo-KO. WT, n=51; Adipo-KO, n=39; Endo-KO, n=34; Total-KO, n=15. ( B ) Plasma insulin responses to 10mg/kg isoproterenol-induced lipolysis in WT vs. Myeloid-KO mice. n=8/group. ( C ) Plasma FAPB4 and ( D ) insulin responses in WT and FABP4-KO mice injected with PBS or 7ug of FABP4 prior to induction of lipolysis with 10mg/kg isoproterenol. n=8/group. ***p<0.0005, **p<0.001, *p<0.05 vs. WT. Data were analyzed by repeated measures 2-way ANOVA with Geisser-Greenhouse correction, followed by Sidak’s (C) or Tukey’s (A,D) multiple comparison test. Data are presented as mean ± SEM. ND: No signal detected.

Article Snippet: For in-house FABP4 ELISA we used anti-FABP4 antibodies produced for the Hotamışlıgil lab by the Dana Farber Antibody Core (clone 351.4.6H7.G3.G9 for capture, HRP-tagged clone 351.4.5E1.H3 for detection) and recombinant human FABP4 as a standard (R&D Systems, cat# DY3150-05).

Techniques: Clinical Proteomics, Injection, Comparison

Journal: bioRxiv

Article Title: Endothelial FABP4 constitutes the majority of basal circulating hormone levels and regulates lipolysis-driven insulin secretion

doi: 10.1101/2022.10.13.511807

Figure Lengend Snippet: ( A ) Upper panel: FABP4 immunostaining of pancreas of WT, Adipo-KO, Endo-KO and Myeloid-KO mice. Islets are encircled by black dotted lines. 40X magnification. Lower panel: 2X enlargement of boxed area in upper panel. ( B ) Insulin immunostaining of pancreas from WT, Adipo-KO, Endo-KO, and Total-KO mice, 40X magnification, and ( C ) quantification of insulin-positive area. ( D ) Insulin secretion from isolated islets of WT, Adipo-KO, Endo-KO and Total-KO mice in response to low glucose (2.8mM), high glucose (16.7mM), high glucose + KCl (30mM). *p<0.05. ( E ) Fold-increase in insulin secretion induced by HG + FSK (10uM) over HG. Data in D were analyzed by unpaired t-test. Data in C and E were analyzed by one-way ANOVA followed by Tukey’s multiple comparison test. Data are presented as mean ± SEM.

Article Snippet: For in-house FABP4 ELISA we used anti-FABP4 antibodies produced for the Hotamışlıgil lab by the Dana Farber Antibody Core (clone 351.4.6H7.G3.G9 for capture, HRP-tagged clone 351.4.5E1.H3 for detection) and recombinant human FABP4 as a standard (R&D Systems, cat# DY3150-05).

Techniques: Immunostaining, Isolation, Comparison

Journal: bioRxiv

Article Title: Endothelial FABP4 constitutes the majority of basal circulating hormone levels and regulates lipolysis-driven insulin secretion

doi: 10.1101/2022.10.13.511807

Figure Lengend Snippet: ( A) The mouse FABP4 gene was first converted to the the human FABP4 open reading frame by substituting the 11 amino acids differing between mouse and human FABP4 located in exons 2 and 3. ( B ) Exon 2 was flanked at the 3’ end by a FRT-neomycin-FRT-loxP cassette and by a single lox P site at the 5’ end. This distal lox P site was positioned upstream of exon 2 within the intron 1 sequences. The model was generated by homologous recombination in embryonic stem cells. The FRT-flanked selection cassette was removed in vivo by crossing with Flp-recombinase-expressing mice. Hatched rectangles represent FABP4 coding sequences. Gray rectangles indicate non-coding exon portions. Solid lines represent chromosome sequences. The 11 murine/human amino acid substitutions are represented by stars.

Article Snippet: For in-house FABP4 ELISA we used anti-FABP4 antibodies produced for the Hotamışlıgil lab by the Dana Farber Antibody Core (clone 351.4.6H7.G3.G9 for capture, HRP-tagged clone 351.4.5E1.H3 for detection) and recombinant human FABP4 as a standard (R&D Systems, cat# DY3150-05).

Techniques: Generated, Homologous Recombination, Selection, In Vivo, Expressing

Journal: Bone Research

Article Title: FABP4 secreted by M1-polarized macrophages promotes synovitis and angiogenesis to exacerbate rheumatoid arthritis

doi: 10.1038/s41413-022-00211-2

Figure Lengend Snippet: The expression of FABP4 in the RA mouse model, BMDMs and RA patients. a , b FABP4 concentrations in the synovial fluid ( a ) and serum ( b ) of controls and RA patients ( n = 8 per group) were assessed by ELISA. C Representative images and quantification of HE staining and FABP4 immunohistochemical staining in the synovium of control and C57BL/6 J mice at 4 and 8 weeks after AIA modeling ( n = 10 per group). Scale bars: 50 μm, 100 μm and 200 μm. d Representative images and quantitative analysis of coimmunostaining of FABP4 with F4/80 or NOS2 in the synovium of control and C57BL/6 J mice at 4 and 8 weeks after AIA modeling ( n = 10 per group). Scale bars: 12.5 μm. e Representative images and quantification of safranin O and fast green staining and FABP4 immunohistochemical staining in knee cartilage from controls and C57BL/6 J mice at 4 and 8 weeks after AIA modeling ( n = 10 per group). Scale bars: 50 μm and 100 μm. f Western blot showing FABP4 and NOS2 in BMDMs stimulated with 50, 200, or 500 ng·mL −1 LPS for 12 h or 24 h. g Quantitative PCR analysis of NOS2, Arg-1, and FABP4 mRNA expression in BMDMs ( n = 3 per group). h FABP4 concentrations in the supernatant of BMDMs stimulated with 50, 200, or 500 ng·mL −1 LPS for 12 h or 24 h ( n = 3 per group) were assessed by ELISA. Student’s t -test or one-way analysis of variance (ANOVA) and Tukey’s multiple comparison test. * P < 0.05, ** P < 0.01; ns, no significance. The data are shown as the mean ± SEM

Article Snippet: We used human and mouse FABP4 ELISA kits (Elabscience Biotechnology, Bethesda, MD, USA: #E-EL-H0285c and #E-EL-M2404c) to analyze the level of FABP4 in the supernatant of macrophages stimulated with lipopolysaccharide (LPS), the serum of C57BL/6 J and TSC1KO mice, and human serum and synovial fluid.

Techniques: Expressing, Enzyme-linked Immunosorbent Assay, Staining, Immunohistochemical staining, Control, Western Blot, Real-time Polymerase Chain Reaction, Comparison

Journal: Bone Research

Article Title: FABP4 secreted by M1-polarized macrophages promotes synovitis and angiogenesis to exacerbate rheumatoid arthritis

doi: 10.1038/s41413-022-00211-2

Figure Lengend Snippet: The effect of FABP4 and M1-polarized macrophage supernatant on HUVECs, FLSs, and chondrocytes in vitro. a , b Tube formation assay and quantification of HUVECs cultured with vehicle (C), rhFABP4 (P), rhFABP4 + BMS309403 (PB), M1-polarized macrophage supernatant (S), or M1-polarized macrophage supernatant+BMS309403 (SB) ( n = 3 per group). Scale bar: 100 µm. c Immunoblot analysis of FABP4 and VEGFα in HUVECs cultured with C, P, PB, S, or SB for 24 h. d , g Representative images and quantification of BrdU (green) immunofluorescence ( d ) and Transwell assays ( g ) in HUVECs and FLSs treated with C, P, PB, S, or SB for 24 h ( n = 3 per group). Scale bar: 25 µm, 100 µm. e , f Immunoblot analysis of ERK1/2, p-ERK1/2, P65, and p-P65 in HUVECs ( e ) and FLSs ( f ) treated with C, P, PB, S, or SB for 1 h. h Toluidine blue staining of ATDC5 cells treated with C, P, PB, S, or SB for 24 h ( n = 3 per group). i Immunoblot analysis of FABP4, MMP13, Sox9, and Col2a1 in primary chondrocytes treated with C, P, PB, S, or SB for 24 h. j Immunoblot analysis of P65 and p-P65 in primary chondrocytes treated with C, P, PB, S, or SB for 1 h. One-way ANOVA and Tukey’s multiple comparison test. * P < 0.05, ** P < 0.01; # P < 0.05, ## P < 0.01 compared to the control. The data are shown as the mean ± SEM

Article Snippet: We used human and mouse FABP4 ELISA kits (Elabscience Biotechnology, Bethesda, MD, USA: #E-EL-H0285c and #E-EL-M2404c) to analyze the level of FABP4 in the supernatant of macrophages stimulated with lipopolysaccharide (LPS), the serum of C57BL/6 J and TSC1KO mice, and human serum and synovial fluid.

Techniques: In Vitro, Tube Formation Assay, Cell Culture, Western Blot, Immunofluorescence, Staining, Comparison, Control

Journal: Bone Research

Article Title: FABP4 secreted by M1-polarized macrophages promotes synovitis and angiogenesis to exacerbate rheumatoid arthritis

doi: 10.1038/s41413-022-00211-2

Figure Lengend Snippet: Recombinant FABP4 exacerbates the development of RA in C57BL/6 J mice. a Representative images and quantitative analysis of FABP4 were assessed by immunohistochemical staining and coimmunostaining of FABP4 with F4/80 or NOS2 in the synovium of control and RA mice treated with vehicle or rmFABP4 for 4 and 8 weeks ( n = 10 per group). Scale bars: 12.5 µm and 100 µm. b – i Representative images and quantification of Vimentin and MMP3 coimmunostaining ( b , f ), CD31 and EMCN coimmunostaining ( c , g ), Col2a1 immunofluorescence staining ( d , h ), and MMP13 immunohistochemical staining ( e , i ) in the knee joints of control and RA mice treated with vehicle or rmFABP4 for 4 and 8 weeks ( n = 10 per group). Scale bars: 12.5 µm, 25 µm, and 100 µm. One-way ANOVA and Tukey’s multiple comparison test. ** P < 0.01. The data are shown as the mean ± SEM

Article Snippet: We used human and mouse FABP4 ELISA kits (Elabscience Biotechnology, Bethesda, MD, USA: #E-EL-H0285c and #E-EL-M2404c) to analyze the level of FABP4 in the supernatant of macrophages stimulated with lipopolysaccharide (LPS), the serum of C57BL/6 J and TSC1KO mice, and human serum and synovial fluid.

Techniques: Recombinant, Immunohistochemical staining, Staining, Control, Immunofluorescence, Comparison

Journal: Bone Research

Article Title: FABP4 secreted by M1-polarized macrophages promotes synovitis and angiogenesis to exacerbate rheumatoid arthritis

doi: 10.1038/s41413-022-00211-2

Figure Lengend Snippet: Activation of the mTORC1 pathway enhances the secretion of FABP4 by M1-polarized macrophages to exacerbate RA progression. a , b Representative images and quantification of F4/80 and pS6 coimmunostaining in the mouse ( n = 10 per group) ( a ) and human synovium ( n = 16 per group) ( b ) of the control and RA groups. Scale bar: 12.5 µm. c Western blot showing S6 and pS6 in human synovial tissue. d Western blot showing FABP4, NOS2, S6 and pS6 in BMDMs treated with LPS or rapamycin. e Representative immunofluorescence staining images and quantitative analysis of FABP4 in BMDMs stimulated with LPS or rapamycin ( n = 3 per group). Scale bar: 100 µm. f Western blot showing FABP4, NOS2, S6 and pS6 in BMDMs treated with MHY1485 or rapamycin. g Representative immunofluorescence staining images and quantitative analysis of FABP4 in BMDMs treated with MHY1485 or rapamycin ( n = 3 per group). Scale bar: 100 µm. h Western blot showing FABP4, NOS2, S6 and pS6 in BMDMs from control and TSC1KO mice. i Representative images and quantification of F4/80 and pS6 coimmunostaining in the synovium of wild-type and TSC1KO mice at 4 and 8 weeks after AIA modeling ( n = 10 per group). Scale bar: 12.5 µm. j Representative images and quantification of FABP4 immunohistochemical staining and coimmunostaining of FABP4 with F4/80 or NOS2 in the synovium of control and TSC1KO mice at 4 and 8 weeks after AIA modeling ( n = 10 per group). Scale bars: 12.5 µm and 100 µm. f–j Representative images ( f ) and quantification of Vimentin and MMP3 coimmunostaining ( g ), CD31 and EMCN coimmunostaining ( h ), Col2a1 immunofluorescence staining ( i ), and MMP13 immunohistochemical staining ( j ) in the knee joints of controls and TSC1KO mice at 4 and 8 weeks after AIA modeling ( n = 10 per group). Scale bar: 12.5 µm, 25 µm, and 100 µm. Student’s t -test or one-way ANOVA and Tukey’s multiple comparison test. ** P < 0.01. The data are shown as the mean ± SEM

Article Snippet: We used human and mouse FABP4 ELISA kits (Elabscience Biotechnology, Bethesda, MD, USA: #E-EL-H0285c and #E-EL-M2404c) to analyze the level of FABP4 in the supernatant of macrophages stimulated with lipopolysaccharide (LPS), the serum of C57BL/6 J and TSC1KO mice, and human serum and synovial fluid.

Techniques: Activation Assay, Control, Western Blot, Immunofluorescence, Staining, Immunohistochemical staining, Comparison

Journal: Bone Research

Article Title: FABP4 secreted by M1-polarized macrophages promotes synovitis and angiogenesis to exacerbate rheumatoid arthritis

doi: 10.1038/s41413-022-00211-2

Figure Lengend Snippet: Inhibition of the mTORC1 pathway reduces FABP4 secretion by M1-polarized macrophages to attenuate RA progression. a Representative images and quantification of F4/80 and pS6 coimmunostaining in the synovium of controls and Rheb1KO mice at 4 and 8 weeks after AIA modeling ( n = 10 per group). Scale bar: 12.5 µm. b – e Representative images ( b ) and quantitative analysis of FABP4 immunohistochemical staining ( c ) and coimmunostaining of FABP4 with F4/80 ( d ) or NOS2 ( e ) in the synovium of controls and Rheb1KO mice at 4 and 8 weeks after AIA modeling ( n = 10 per group). Scale bars: 12.5 µm and 100 µm. f – j Representative images ( f ) and quantitative analysis of Vimentin and MMP3 coimmunostaining ( g ), CD31 and EMCN coimmunostaining ( h ), Col2a1 immunofluorescence staining ( i ), and MMP13 immunohistochemical staining ( j ) in the knee joints of controls and Rheb1KO mice at 4 and 8 weeks after AIA modeling ( n = 10 per group). Scale bar: 12.5 µm, 25 µm, and 100 µm. Student’s t -test. ** P < 0.01. The data are shown as the mean ± SEM

Article Snippet: We used human and mouse FABP4 ELISA kits (Elabscience Biotechnology, Bethesda, MD, USA: #E-EL-H0285c and #E-EL-M2404c) to analyze the level of FABP4 in the supernatant of macrophages stimulated with lipopolysaccharide (LPS), the serum of C57BL/6 J and TSC1KO mice, and human serum and synovial fluid.

Techniques: Inhibition, Immunohistochemical staining, Staining, Immunofluorescence

Journal: Bone Research

Article Title: FABP4 secreted by M1-polarized macrophages promotes synovitis and angiogenesis to exacerbate rheumatoid arthritis

doi: 10.1038/s41413-022-00211-2

Figure Lengend Snippet: BMS309403 and anagliptin reduce FABP4 expression in murine synovial macrophages. a – c Representative images and quantification of FABP4 immunohistochemical staining and coimmunostaining of FABP4 with F4/80 or NOS2 in the synovium of TSC1KO mice treated with vehicle, BMS309403 or anagliptin for 4 and 8 weeks after AIA modeling ( n = 10 per group). Scale bar: 12.5 µm and 100 µm. d – f Representative images and quantification of FABP4 immunohistochemical staining and coimmunostaining of FABP4 and F4/80 or NOS2 in the synovium of RA mice treated with vehicle or BMS309403 for 4 and 8 weeks ( n = 10 per group). Scale bar: 12.5 µm and 100 µm. Student’s t -test or one-way ANOVA and Tukey’s multiple comparison test. * P < 0.05, ** P < 0.01. The data are shown as the mean ± SEM

Article Snippet: We used human and mouse FABP4 ELISA kits (Elabscience Biotechnology, Bethesda, MD, USA: #E-EL-H0285c and #E-EL-M2404c) to analyze the level of FABP4 in the supernatant of macrophages stimulated with lipopolysaccharide (LPS), the serum of C57BL/6 J and TSC1KO mice, and human serum and synovial fluid.

Techniques: Expressing, Immunohistochemical staining, Staining, Comparison

Journal: Bone Research

Article Title: FABP4 secreted by M1-polarized macrophages promotes synovitis and angiogenesis to exacerbate rheumatoid arthritis

doi: 10.1038/s41413-022-00211-2

Figure Lengend Snippet: Inhibiting FABP4 prevents RA development. a – d Representative images of Vimentin and MMP3 coimmunostaining ( a ), CD31 and EMCN coimmunostaining ( b ), Col2a1 immunofluorescence staining ( c ), and immunohistochemical staining of MMP13 ( D ) in the knee joints of TSC1KO mice treated with vehicle, BMS309403 or anagliptin for 4 and 8 weeks after AIA modeling. Scale bar: 12.5 µm, 25 µm, and 100 µm. e – h Representative images of Vimentin and MMP3 coimmunostaining ( e ), CD31 and EMCN coimmunostaining ( f ), Col2a1 immunofluorescence staining ( g ), and immunohistochemical staining of MMP13 ( h ) in the knee joints of RA mice treated with vehicle or BMS309403 for 4 and 8 weeks. Scale bar: 12.5 µm, 25 µm, and 100 µm. i – l Quantification of Vimentin-MMP3 ( i ), CD31-EMCN ( j ), Col2a1 ( k ), and MMP13 ( l ) in TSC1KO mice treated with vehicle, BMS309403, or anagliptin for 4 and 8 weeks after AIA surgery ( n = 10 per group). m – p Quantification of Vimentin-MMP3 ( m ), CD31-EMCN ( n ), Col2a1 ( o ), and MMP13 ( p ) in RA mice treated with vehicle or BMS309403 for 4 and 8 weeks ( n = 10 per group). Student’s t -test or one-way ANOVA and Tukey’s multiple comparison test were used. ** P < 0.01. The data are shown as the mean ± SEM

Article Snippet: We used human and mouse FABP4 ELISA kits (Elabscience Biotechnology, Bethesda, MD, USA: #E-EL-H0285c and #E-EL-M2404c) to analyze the level of FABP4 in the supernatant of macrophages stimulated with lipopolysaccharide (LPS), the serum of C57BL/6 J and TSC1KO mice, and human serum and synovial fluid.

Techniques: Immunofluorescence, Staining, Immunohistochemical staining, Comparison

Journal: Scientific Reports

Article Title: Circulating FABP4 is eliminated by the kidney via glomerular filtration followed by megalin-mediated reabsorption

doi: 10.1038/s41598-018-34902-w

Figure Lengend Snippet: Circulating FABP4 is taken up by kidney. Blood, kidney, liver and heart were collected at indicated time points after intravenous injection of 125 I-labeled recombinant FABP4 ( 125 I-FABP4) to assess FABP4 uptake by each tissue. The data were expressed as the percent injected dose per gram tissue (%dose/g). n = 5. ***p < 0.001.

Article Snippet: Recombinant human FABP4 (ab133145, abcam, MA) was labeled with Na 125 I (NEZ033A, Perkin Elmer, MA) by Chloramine T method .

Techniques: Injection, Labeling, Recombinant

Journal: Scientific Reports

Article Title: Circulating FABP4 is eliminated by the kidney via glomerular filtration followed by megalin-mediated reabsorption

doi: 10.1038/s41598-018-34902-w

Figure Lengend Snippet: Circulating FABP4 is reabsorbed by PTECs. Kidney was isolated 10 min after intravenous injection of AF647-FABP4 (red) for subsequent immunofluorescence. Saline iv served as a negative control. LTL, Lotus Tetragonolobus Lectin, a marker for PTECs (green); DAPI, 4′,6-diamidino-2-phenylindole, a marker for nuclei (blue).

Article Snippet: Recombinant human FABP4 (ab133145, abcam, MA) was labeled with Na 125 I (NEZ033A, Perkin Elmer, MA) by Chloramine T method .

Techniques: Isolation, Injection, Immunofluorescence, Negative Control, Marker

Journal: Scientific Reports

Article Title: Circulating FABP4 is eliminated by the kidney via glomerular filtration followed by megalin-mediated reabsorption

doi: 10.1038/s41598-018-34902-w

Figure Lengend Snippet: Circulating FABP4 levels were elevated by BLN. Blood was collected from retro-orbital plexus at indicated time points after surgery to measure serum levels of FABP4, creatinine and glycerol (right panels). FABP4 KO mice were used as negative control for serum levels of FABP4. Blood was also collected from non-treated mice as a control after a 6 h fast (left panels). n = 5. **p < 0.01 and ***p < 0.001. BLN, bilateral nephrectomy; ULN, unilateral nephrectomy.

Article Snippet: Recombinant human FABP4 (ab133145, abcam, MA) was labeled with Na 125 I (NEZ033A, Perkin Elmer, MA) by Chloramine T method .

Techniques: Negative Control

Journal: Scientific Reports

Article Title: Circulating FABP4 is eliminated by the kidney via glomerular filtration followed by megalin-mediated reabsorption

doi: 10.1038/s41598-018-34902-w

Figure Lengend Snippet: Circulating FABP4 levels were more elevated in mice model with severe renal dysfunction after intraperitoneal injection of CL316,243 (1 mg/kg). Blood was collected from retro-orbital plexus before, 10 and 20 minutes after injection to measure serum levels of creatinine, glycerol and FABP4. n = 5. * < 0.05. ULN, unilateral nephrectomy; IRI, ischemia reperfusion injury.

Article Snippet: Recombinant human FABP4 (ab133145, abcam, MA) was labeled with Na 125 I (NEZ033A, Perkin Elmer, MA) by Chloramine T method .

Techniques: Injection

Journal: Scientific Reports

Article Title: Circulating FABP4 is eliminated by the kidney via glomerular filtration followed by megalin-mediated reabsorption

doi: 10.1038/s41598-018-34902-w

Figure Lengend Snippet: FABP4 is excreted to urine in kidney-specific megalin-KO mice (NDRG1 cre ERT2, megalin flox/flox ). ( A ) Levels of FABP4 and albumin in urine. Note that FABP4 levels were markedly elevated in urine in megalin KO mice while it was hardly detected in urine in control (megalin flox/flox ). ( B ) Levels of FABP4 and albumin in serum. n = 5. *p < 0.05 and ***p < 0.001.

Article Snippet: Recombinant human FABP4 (ab133145, abcam, MA) was labeled with Na 125 I (NEZ033A, Perkin Elmer, MA) by Chloramine T method .

Techniques:

Journal: Scientific Reports

Article Title: Circulating FABP4 is eliminated by the kidney via glomerular filtration followed by megalin-mediated reabsorption

doi: 10.1038/s41598-018-34902-w

Figure Lengend Snippet: QCM analysis for the binding of FABP4 to megalin. FABP4 (1 mg/ml) was injected at the time point indicated by the arrow into the QCM chamber in which megalin or FA-free BSA had been immobilized on the sensor chips. The binding of FABP4 to the chip-immobilized protein is indicated by a change in the frequency, ΔF. All the experiments were repeated at least three times and the representative result is shown. Note that FABP4 bound to megalin, but not to albumin.

Article Snippet: Recombinant human FABP4 (ab133145, abcam, MA) was labeled with Na 125 I (NEZ033A, Perkin Elmer, MA) by Chloramine T method .

Techniques: Binding Assay, Injection

Journal: Neoplasia (New York, N.Y.)

Article Title: FABP4 facilitates epithelial-mesenchymal transition via elevating CD36 expression in glioma cells

doi: 10.1016/j.neo.2024.101050

Figure Lengend Snippet: FABP4 is abundantly expressed in GBM samples. (a) Representative MRI, H&E and FABP4 IHC staining images of specimens from patients with GBM and lower grade glioma. Fluorescence staining for FABP4 of GBM and lower grade glioma patients. (b) IHC images of high and low grade glioma from Human Protein Atlas datasets. Scale bar= 50/20 μm. (c) IHC score of FABP4 in human glioma specimens of different grades. (d) Correlation of FABP4 expression with gender, age, tumor size and tumor grade. (e) Protein level of FABP4 in primary glioma cells and cell lines. (f) Fluorescence staining of FABP4 in glioma cells. (g) Secretion level of FABP4 in glioma cells. Data are expressed as Mean ± SD.** p < 0.01, *** p < 0.001. H&E, hematoxylin and eosin. IHC, Immunohistochemistry.

Article Snippet: Human FABP4 ELISA kit (R&D Systems) was used to determine the protein concentration of FABP4 from culture media according to manufacturer's instruction. supernatants of medium from glioma cells cultured for 24 h were collected and analyzed by Elisa kit.

Techniques: Immunohistochemistry, Fluorescence, Staining, Expressing

Journal: Neoplasia (New York, N.Y.)

Article Title: FABP4 facilitates epithelial-mesenchymal transition via elevating CD36 expression in glioma cells

doi: 10.1016/j.neo.2024.101050

Figure Lengend Snippet: FABP4 is closely associated with mesenchymal subtype in GBM. (a) Fluorescence staining of FABP4 and vimentin in GBM samples. (b) Fluorescence staining of FABP4 and α-SMA in GBM samples. (c) FABP4 mRNA expression in different molecular subtypes of GBM samples from TCGA, CGGA and Ivy database. (d) Correlation between the expression of FABP4 and EMT-related markers (ACTA2 and S100A4) in TCGA GBM dataset was evaluated. Results are represented as Mean ± SD of biologically triplicate assays. * p < 0.05, ** p < 0.01, *** p < 0.001.

Article Snippet: Human FABP4 ELISA kit (R&D Systems) was used to determine the protein concentration of FABP4 from culture media according to manufacturer's instruction. supernatants of medium from glioma cells cultured for 24 h were collected and analyzed by Elisa kit.

Techniques: Fluorescence, Staining, Expressing

Journal: Neoplasia (New York, N.Y.)

Article Title: FABP4 facilitates epithelial-mesenchymal transition via elevating CD36 expression in glioma cells

doi: 10.1016/j.neo.2024.101050

Figure Lengend Snippet: FABP4 confers increased invasiveness in glioma cells. (a) primary glioma cells were transfected with empty vector or FABP4-expressing lentiviral vector, fluorescent staining of FABP4 and Phalloidin were assessed. White arrowheads indicate filopodia on glioma cells. (b) Glioma cells were treated with PBS or rhFABP4, FABP4 and Phalloidin expressions were evaluated. (c) primary glioma cells with scrambled shRNA or shFABP4 were stained with FABP4 and Phalloidin. (d) Invasion abilities of glioma cells with FABP4 expression were assessed. (e) Gene set enrichment analysis (GSEA) was performed according to FABP4 expression in TCGA GBM dataset showed enriched pathways associated with cell invasiveness. Results are represented as Mean ± SD of biologically triplicate assays. * p < 0.05, ** p < 0.01, *** p < 0.001.

Article Snippet: Human FABP4 ELISA kit (R&D Systems) was used to determine the protein concentration of FABP4 from culture media according to manufacturer's instruction. supernatants of medium from glioma cells cultured for 24 h were collected and analyzed by Elisa kit.

Techniques: Transfection, Plasmid Preparation, Expressing, Staining, shRNA

Journal: Neoplasia (New York, N.Y.)

Article Title: FABP4 facilitates epithelial-mesenchymal transition via elevating CD36 expression in glioma cells

doi: 10.1016/j.neo.2024.101050

Figure Lengend Snippet: FABP4 induces EMT process in GBM. (a) differential gene expression analysis between high and low FABP4 groups from TCGA GBM dataset. Several key molecules related to EMT are indicated. (b) GSEA analysis reveals a significant correlation between FABP4 expression and EMT process in TCGA GBM dataset. (c) Expression of EMT-related molecules in glioma cells that transfected with FABP4-lentiviral vector or treated with recombinant human FABP4 protein. (d) Immunofluorescence analysis of N-Cadherin and Vimentin in glioma cells with FABP4 over-expression. (e) Immunoblot analysis of N-Cadherin and Vimentin in glioma cells when FABP4 is downregulated. (f) Immunofluorescence analysis of N-Cadherin in glioma cells treated with shRNA targeting FABP4. Scale bar=20 μm. Results are represented as Mean ± SD of biologically triplicate assays. * p < 0.05, ** p < 0.01, *** p < 0.001. ns, not significant.

Article Snippet: Human FABP4 ELISA kit (R&D Systems) was used to determine the protein concentration of FABP4 from culture media according to manufacturer's instruction. supernatants of medium from glioma cells cultured for 24 h were collected and analyzed by Elisa kit.

Techniques: Gene Expression, Expressing, Transfection, Plasmid Preparation, Recombinant, Immunofluorescence, Over Expression, Western Blot, shRNA

Journal: Neoplasia (New York, N.Y.)

Article Title: FABP4 facilitates epithelial-mesenchymal transition via elevating CD36 expression in glioma cells

doi: 10.1016/j.neo.2024.101050

Figure Lengend Snippet: FABP4-induced EMT is mediated in CD36-dependent manner. (a) Protein-protein interaction analysis of FABP4 via using STRING database. (b) CD36 mRNA expression in high or low FABP4 group from TCGA GBM dataset. (c) Immunoblot analysis of CD36 and PPARG in glioma cells with FABP4 upregulation or downregulation. (d) Transcriptional levels of CD36 and PPARG in glioma cells upon FABP4 overexpression. (e) CD36 mRNA expression in glioma samples with different grades. (f) CD36 mRNA expression in three GBM subtypes. (g) Kaplan-Meier survival analysis for CD36 in patients with glioma from TCGA dataset. (h) Correlation between FABP4 and CD36 in TCGA glioma dataset. (i) Immunofluorescence staining of CD36 in glioma cells with FABP4 upregulation. (j) Indicated proteins in glioma cells treated with exogenous FABP4 followed by CD36 knockdown. (k) Immunofluorescence staining of N-Cadherin and Vimentin in FABP4-expressed glioma cells treated with CD36 downregulation. (l) Glioma cells with stable FABP4 expression are treated with CD36 knockdown, F-actin staining is detected with phalloidin. White arrowheads indicate filopodia on glioma cells. Scale bar=50 μm. Results are represented as Mean ± SD of biologically triplicate assays. * p < 0.05, ** p < 0.01, *** p < 0.001. ns, not significant.

Article Snippet: Human FABP4 ELISA kit (R&D Systems) was used to determine the protein concentration of FABP4 from culture media according to manufacturer's instruction. supernatants of medium from glioma cells cultured for 24 h were collected and analyzed by Elisa kit.

Techniques: Expressing, Western Blot, Over Expression, Immunofluorescence, Staining, Knockdown

Journal: Neoplasia (New York, N.Y.)

Article Title: FABP4 facilitates epithelial-mesenchymal transition via elevating CD36 expression in glioma cells

doi: 10.1016/j.neo.2024.101050

Figure Lengend Snippet: CD36 enhances EMT process by activating non-canonical TGFβ pathways. (a) Association between CD36 mRNA level and EMT proteins (PAI.1, fibronectin) in TCPA. (b) Primary glioma cells with empty vector or CD36 transfection were treated with 100 pM TGFβ1 for 6 h, indicated proteins of canonical and non-canonical TGFβ pathways were examined. (c) Immunofluorescence staining of ROCK1 and CD36 in glioma cells with EV or CD36 transfection. (d) PGCs with (without) CD36 knockdown were treated with recombinant human FABP4 proteins at different time point, expressions of indicated proteins were analyzed by immunoblotting. (e) PGCs with CD36 downregulation were stimulated with rhFABP4, immunofluorescence stainings of ROCK1 and p-Smad3 were evaluated. (f) PGC-shScr or PGC-shCD36 was added with rhFABP4, cell invasiveness and filopodia formation were measured. Results are represented as Mean ± SD of biologically triplicate assays. * p < 0.05, ** p < 0.01, *** p < 0.001.

Article Snippet: Human FABP4 ELISA kit (R&D Systems) was used to determine the protein concentration of FABP4 from culture media according to manufacturer's instruction. supernatants of medium from glioma cells cultured for 24 h were collected and analyzed by Elisa kit.

Techniques: Plasmid Preparation, Transfection, Immunofluorescence, Staining, Knockdown, Recombinant, Western Blot

Journal: Neoplasia (New York, N.Y.)

Article Title: FABP4 facilitates epithelial-mesenchymal transition via elevating CD36 expression in glioma cells

doi: 10.1016/j.neo.2024.101050

Figure Lengend Snippet: Targeting FABP4 effectively reduces tumor progression and EMT in a xenograft model. (a) Luminescent imaging of representative nude mice xenografts from mCherry-LUC-labeled shScr ( n = 5) or shFABP4 PGC#2 ( n = 5) at day 3 and 25. (b) Luminescent signal intensity of GBM-bearing mice in two groups were evaluated. (c) Representative immunofluorescence images of FABP4, N-Cadherin, Vimentin and α-SMA. Scale bar=20 μm. (d) Sample sections from control (upper) and shFABP4 tumors (bottom) were immunofluorescently labeled with mCherry. (e) Schematic diagram illustrating the FABP4-CD36 signaling pathway in EMT regulation. Results are represented as Mean ± SD of biologically triplicate assays. * p < 0.05, ** p < 0.01, *** p < 0.001.

Article Snippet: Human FABP4 ELISA kit (R&D Systems) was used to determine the protein concentration of FABP4 from culture media according to manufacturer's instruction. supernatants of medium from glioma cells cultured for 24 h were collected and analyzed by Elisa kit.

Techniques: Imaging, Labeling, Immunofluorescence, Control

Journal: Cell reports

Article Title: Dynamic interplay between IL-1 and WNT pathways in regulating dermal adipocyte lineage cells during skin development and wound regeneration

doi: 10.1016/j.celrep.2023.112647

Figure Lengend Snippet: KEY RESOURCES TABLE

Article Snippet: Rabbit anti-FABP4 , Proteintech , Cat# 15872-1-AP ; RRID: AB_2102440.

Techniques: Recombinant, Staining, SYBR Green Assay, Red Blood Cell Lysis, Protease Inhibitor, cDNA Synthesis, Generated, Transgenic Assay, Software

Journal: American Journal of Cancer Research

Article Title: 25-HC promotes hepatocellular carcinoma metastasis through up-regulation of TLR4 dependent FABP4

doi:

Figure Lengend Snippet: The primer sequence of target genes

Article Snippet: Recombinant human FABP4 was bought from Cloud-Clone Corp. (Houston, TX, USA) and dissolved in PBS.

Techniques: Sequencing

Journal: American Journal of Cancer Research

Article Title: 25-HC promotes hepatocellular carcinoma metastasis through up-regulation of TLR4 dependent FABP4

doi:

Figure Lengend Snippet: Sequences of siRNA

Article Snippet: Recombinant human FABP4 was bought from Cloud-Clone Corp. (Houston, TX, USA) and dissolved in PBS.

Techniques:

Journal: American Journal of Cancer Research

Article Title: 25-HC promotes hepatocellular carcinoma metastasis through up-regulation of TLR4 dependent FABP4

doi:

Figure Lengend Snippet: 25-HC increases FABP4 expression which could directly enhance HepG2 cells migration. A. HepG2 cells were treated with the indicated concentrations of 25-HC for 24 hours before mRNA or protein was extracted to determine the expression of FABP4 by RT-qPCR or Western blotting, respectively. B. HepG2 cells were treated with the indicated concentrations of FABP4 (0, 20, 40 and 100 ng/mL) for 24 hours before mRNA or protein was extracted. The mRNA expressions of MMP2 and MMP9 were examined by RT-qPCR and the protein expressions of MMP1, MMP2, MMP3, MMP9 were detected by Western blotting, respectively. C. Migration assay of HepG2 cells after stimulation of FABP4 for 36 hours was performed. Results were obtained from 3 independent experiments and are expressed as the means ± SEM. Statistical significance was determined by one-way ANOVA followed by Dunnett’s test. ns, not significant, *P<0.05, **P<0.01, ***P<0.001.

Article Snippet: Recombinant human FABP4 was bought from Cloud-Clone Corp. (Houston, TX, USA) and dissolved in PBS.

Techniques: Expressing, Migration, Quantitative RT-PCR, Western Blot

Journal: American Journal of Cancer Research

Article Title: 25-HC promotes hepatocellular carcinoma metastasis through up-regulation of TLR4 dependent FABP4

doi:

Figure Lengend Snippet: Inhibition of FABP4 decreases the promotive effects of 25-HC on HepG2 cells migration. A. HepG2 cells were transfected with siNC or siFABP4 and CCK-8 assay was performed. Results are shown as the cells absorbance at 450 nm. HepG2 cells were transfected with siNC or siFABP4 for 24 hours before cells were treated with 25-HC at 10 μM. 36 hours later, migratory ability of HepG2 cells was determined by Transwell assay. Or 24 hours later, cells were collected for mRNA or protein extraction to determine the expressions of MMPs by RT-qPCR or Western blotting, respectively. B. HepG2 cells were stimulated with 10 μM 25-HC with or without 30 μM BMS-309403. 36 hours later, migration assay was performed. Or 24 hours later, cells were collected for mRNA or protein extraction to determine the expressions of MMPs by RT-qPCR or Western blotting, respectively. C. HepG2 cells were transfected with siNC or siTLR4 for 24 hours before 25-HC treatment. 24 hours later, the mRNA and protein expression of FABP4 was assessed by RT-qPCR and Western blotting, respectively. Results were obtained from 3 independent experiments and are expressed as the means ± SEM. Statistical significance was determined by Student’s t-test. *P<0.05, **P<0.01, ***P<0.001.

Article Snippet: Recombinant human FABP4 was bought from Cloud-Clone Corp. (Houston, TX, USA) and dissolved in PBS.

Techniques: Inhibition, Migration, Transfection, CCK-8 Assay, Transwell Assay, Protein Extraction, Quantitative RT-PCR, Western Blot, Expressing

Journal: American Journal of Cancer Research

Article Title: 25-HC promotes hepatocellular carcinoma metastasis through up-regulation of TLR4 dependent FABP4

doi:

Figure Lengend Snippet: Inhibition of FABP4 decreases the 25-HC-induced HepG2 cells intrahepatic and abdominal metastasis. BALB/C nude mice were intrahepatic inoculation of 5 × 106 HepG2 cells followed by intraperitoneally injection with PBS or 25-HC (10 mg/kg) or 25-HC with BMS-309403 (45 mg/kg). At the end of the experiment, mice were sacrificed, (A) livers were removed and imaged. (B) The number of abdominal metastasis was counted, and body weight was recorded. Results were obtained from 3 independent experiments and are expressed as the means ± SEM. Statistical significance was determined by Student’s t-test, **P<0.01. ns: no significant.

Article Snippet: Recombinant human FABP4 was bought from Cloud-Clone Corp. (Houston, TX, USA) and dissolved in PBS.

Techniques: Inhibition, Injection

Journal: American Journal of Cancer Research

Article Title: 25-HC promotes hepatocellular carcinoma metastasis through up-regulation of TLR4 dependent FABP4

doi:

Figure Lengend Snippet: FABP4 was up-regulated in HCC tissues. (A) The mRNA expression of FABP4 in hepatocellular carcinoma and normal tissues (GEPIA). T, tumor tissues, N, normal tissues. (B) DNA copy number of FABP4 in hepatocellular carcinoma from The Cancer Genome Atlas (TCGA). (C) The expression of FABP4 in hepatocellular carcinoma cell lines (EMBL-EBI). (D) FABP4 transcription in subgroups of patients with hepatocellular carcinoma, stratified based on gender, age, weight, race, tumor grade and cancer stages (UALCAN). (E) Correlation between FABP4 expression and the prognosis of patients with hepatocellular carcinoma (OncoLnc). Data are expressed as the means ± SEM. Statistical significance was determined by Student’s t-test (A and B) or one-way ANOVA followed by Dunnett’s test (D). *P<0.05, **P<0.01, ***P<0.001.

Article Snippet: Recombinant human FABP4 was bought from Cloud-Clone Corp. (Houston, TX, USA) and dissolved in PBS.

Techniques: Expressing

Journal: BMC Cancer

Article Title: FABP4-mediated ERK phosphorylation promotes renal cancer cell migration

doi: 10.1186/s12885-025-13989-1

Figure Lengend Snippet: Serum FABP4 levels of ccRCC patients were higher compared to HD ( A ), whereas no significant differences were observed stratifying patient according to BMI ( B ), diabetes ( C ) and hypertension ( D ). Box plots denote median and 25th to 75th percentiles (boxes) and Tukey whiskers. The p value was evaluated using Mann–Whitney test. Ns indicates a p- value > 0.05 and *Indicates a p value < 0.05

Article Snippet: Recombinant FABP4 was purchased from Elabscience.

Techniques: MANN-WHITNEY

Journal: BMC Cancer

Article Title: FABP4-mediated ERK phosphorylation promotes renal cancer cell migration

doi: 10.1186/s12885-025-13989-1

Figure Lengend Snippet: Human recombinant FABP4 does not affect cell proliferation in 786-O and ACHN. 786-O ( A ) and ACHN ( B ) cells were seeded (1 × 10 3 cells/ well) in 96-well. and incubated with medium containing 10% FBS, 0.25% BSA and recombinant FABP4 10000 pg/mL alone or in combination with BMS309403 40 µM for 24 h. Cell proliferation was calculated as percentage of BrdU incorporation compared to control. Data represent the mean ± SD of 5 independent experiments for 786-O and 3 independent experiments for ACHN. The p -value was calculated using the one-way ANOVA test. Ns indicates a p- value > 0.05, * Indicates a p value < 0.05

Article Snippet: Recombinant FABP4 was purchased from Elabscience.

Techniques: Recombinant, Incubation, BrdU Incorporation Assay, Control

Journal: BMC Cancer

Article Title: FABP4-mediated ERK phosphorylation promotes renal cancer cell migration

doi: 10.1186/s12885-025-13989-1

Figure Lengend Snippet: Ad-CM ccRCC enhances migration in 786-O and ACHN cells through FABP4 release. 786-O ( A ) and ACHN ( B ) cells were seeded (1 × 10 5 cells/ well) in 12-well. In vitro “wound” was created by a straight-line scratch across the cell monolayer and cells were incubated with medium containing 10% FBS, 0.25% BSA, and human recombinant FABP4 10000 pg/mL and BMS309403 40 µM. The images were acquired at 0 and 24 h using a camera connected to the microscope. The distance toward the wound edges was photographed and measured (magnification 10x). Cell motility was calculated as percentage of wound healing rate. Data represent the mean ± SD of 6 independent experiments for 786-O and 6 independent experiments for ACHN. The p -value was calculated using the one-way ANOVA test. Ns indicates a p -value > 0.05, *Indicates a p value < 0.05, **indicates a p -value ≤ 0.01, and **** a p -value ≤ 0.0001

Article Snippet: Recombinant FABP4 was purchased from Elabscience.

Techniques: Migration, In Vitro, Incubation, Recombinant, Microscopy

Journal: BMC Cancer

Article Title: FABP4-mediated ERK phosphorylation promotes renal cancer cell migration

doi: 10.1186/s12885-025-13989-1

Figure Lengend Snippet: Ad-CM ccRCC displayed a higher concentration of e-FABP4 ( A ) while no significant differences were observed by stratifying patient according to BMI ( B ), hypertension ( C ) and type 2 diabetes ( D ). Peri-renal adipocytes were seeded (200 µL of adipocytes suspension/well) and incubated with 500 µL of medium containing 0,25% BSA to isolate the Ad-CM. After 16 h the Ad-CMs were collected below 80 °C until the assay. Box plots denote median and 25th to 75th percentiles (boxes) and Tukey whiskers. The p -value was evaluated using Mann-Whitney test. Ns indicates a p- value > 0.05 and **indicates a p -value ≤ 0.01

Article Snippet: Recombinant FABP4 was purchased from Elabscience.

Techniques: Concentration Assay, Suspension, Incubation, MANN-WHITNEY

Journal: BMC Cancer

Article Title: FABP4-mediated ERK phosphorylation promotes renal cancer cell migration

doi: 10.1186/s12885-025-13989-1

Figure Lengend Snippet: Ad-CM affect cell proliferation in ACHN, whereas no effect was observed in 786-O cell line. 786-O ( A ) and ACHN ( B ) cells were seeded (1 × 10 3 cells/ well) in 96-well. and incubated with medium containing 10% FBS, 0.25% BSA and recombinant FABP4 10000 pg/mL alone or in combination with BMS309403 40 µM for 24 h. Cell proliferation was calculated as percentage of BrdU incorporation compared to control. Data represent the mean ± SD of 5 independent experiments for 786-O and 3 independent experiments for ACHN. The p -value was calculated using the one-way ANOVA test. Ns indicates a p- value > 0.05, * Indicates a p value < 0.05

Article Snippet: Recombinant FABP4 was purchased from Elabscience.

Techniques: Incubation, Recombinant, BrdU Incorporation Assay, Control

Journal: BMC Cancer

Article Title: FABP4-mediated ERK phosphorylation promotes renal cancer cell migration

doi: 10.1186/s12885-025-13989-1

Figure Lengend Snippet: Ad-CM ccRCC enhances migration in 786-O and ACHN cells through FABP4 release. 786-O ( A ) and ACHN ( B ) cells were seeded (1 × 10 5 cells/ well) in 12-well. In vitro “wound” was created by a straight-line scratch across the cell monolayer and cells were incubated with medium containing 10% FBS, 0.25% BSA, Ad-CM obtained from mature adipocytes both from HD (Ad-CM) and from ccRCC patients (Ad-CM ccRCC), and BMS309403 40 µM. The images were acquired at 0 and 24 h using a camera connected to the microscope. The distance toward the wound edges was photographed and measured (magnification 10x). Cell motility was calculated as percentage of wound healing rate. Data represent the mean ± SD of 6 independent experiments for 786-O and 6 independent experiments for ACHN. The p -value was calculated using the one-way ANOVA test. Ns indicates a p -value > 0.05, *Indicates a p value < 0.05, **indicates a p -value ≤ 0.01, and **** a p -value ≤ 0.0001, Ns indicates a p- value > 0.05, *Indicates a p value < 0.05, **indicates a p -value ≤ 0.01, ***indicates a p -value < 0.001 and **** a p -value ≤ 0.0001

Article Snippet: Recombinant FABP4 was purchased from Elabscience.

Techniques: Migration, In Vitro, Incubation, Microscopy

Journal: BMC Cancer

Article Title: FABP4-mediated ERK phosphorylation promotes renal cancer cell migration

doi: 10.1186/s12885-025-13989-1

Figure Lengend Snippet: Effect of Ad-CM on the MAPK pathway in ccRCC cancer cells. 786-O and ACHN cells were seeded (1 × 10 5 cells/well) and incubated with medium containing 0.25% BSA, Ad-CM from HD, Ad-CM from ccRCC, alone or in combination with BMS309403 40 µM and human recombinant FABP4 10000 pg/mL. GAPDH was used as loading control. A representative blot for 4 experiments for 786-O and 6 experiment for ACHN is presented in figure ( A ) and ( B ) respectively. Quantification of Western Blot is reported for 786-O ( C ) and ACHN ( D ). The p value was calculated using Kruskal-Wallis test. Ns indicates a p -value > 0.05, *Indicates a p value < 0.05. Full-length blots/gels are presented in Supplementary Fig.

Article Snippet: Recombinant FABP4 was purchased from Elabscience.

Techniques: Incubation, Recombinant, Control, Western Blot

Journal: BMC Cancer

Article Title: FABP4-mediated ERK phosphorylation promotes renal cancer cell migration

doi: 10.1186/s12885-025-13989-1

Figure Lengend Snippet: Schematic representation of the proposed role of PRAT adipocytes in promoting migration in ccRCC cells. e-FABP4 released by mature peri-renal adipocytes surrounding ccRCC stimulates ccRCC cells inducing ERK phosphorylation. Parts of the figure were drawn using pictures from Server Medical Art. Servier Medical Art by Servier is licensed under a Creative Commons Attribution 3.0 Unported License

Article Snippet: Recombinant FABP4 was purchased from Elabscience.

Techniques: Migration, Phospho-proteomics