Journal: bioRxiv

Article Title: Activated fatty acid synthesis pathway in macrophages propagates pathogenic fibroblast expansion after myocardial infarction

doi: 10.1101/2025.10.10.681697

Figure Lengend Snippet: A. The schematic diagram of the experimental design to quantify fatty acids in cardiac macrophages using MALDI is shown. B-D. The molecular images ( B ), representative single spectra ( C ), and heatmaps (D) of monoisotopic and D 2 O-incorporated palmitic acid (PA), linoleic acid (LA), oleic acid (OA), stearic acid (SA), and arachidonic acid (AA) acquired from macrophage clusters in the heart sections of C57BL/6 mice with sham and MI surgeries are provided. (n=3/group). E. Quantification of macrophage ATP citrate lyase (ACLY), acetyl-CoA carboxylase (ACC), and fatty acid synthase (FASN) mean fluorescence intensities (MFI) in control and MI patients and mice (day 28) are calculated by confocal microscopy (n=3-5/group). Each datapoint represents a tissue section. F-G. Left ventricular ejection fraction (LV-EF), left ventricle mass (LV mass), fractional shortening (FS), and stroke volume (SV) were calculated by echocardiography 28 days after MI in LysM +/+ Acly fl/fl and LysM cre/+ Acly fl/fl mice (n=5-13/group) ( F ) and in LysM +/+ Fasn fl /fl / LysM cre/+ Fasn fl/fl mice (n=6-10/group) ( G ). H. Representative four-chamber view by cine magnetic resonance images 28 days post MI is shown. I. Various cardiac parameters were assessed by cardiac magnetic resonance imaging (MRI) on 28 days after MI (n=5-7/group). J-K. High resolution diffusion tensor imaging (DTI) tractography in short axis view ( J ) and various DTI parameters ( K ) including quantitative anisotropy (QA), fractional anisotropy (FA), mean diffusivity (MD), and radial diffusivity (RD) on 28 days after MI are provided (n=5-7/group). The data are expressed as mean ± SEM and from 3–5 independent experiments. One way ANOVA for 1F and The Mann–Whitney U test for the other figures were used. * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001.

Article Snippet: Deletion of Acly (The Jackson lab, #030772) and Fasn (Taconic Biosciences, #8034) in myeloid lineages were achieved by crossing these floxed transgenic mice with Lysm Cre/Cre mice (The Jackson lab, #004781).

Techniques: Fluorescence, Control, Confocal Microscopy, Magnetic Resonance Imaging, Diffusion-based Assay, Imaging, MANN-WHITNEY

Journal: bioRxiv

Article Title: Activated fatty acid synthesis pathway in macrophages propagates pathogenic fibroblast expansion after myocardial infarction

doi: 10.1101/2025.10.10.681697

Figure Lengend Snippet: A. MALDI and confocal microscopy were performed on the same cardiac sections in C57BL/6 mice provided with D 2 O in water 28 days after MI. Single spectra of palmitic acid in macrophage clusters were analyzed by mMass (n=3/group). B. ACLY, ACC, and FASN staining in macrophages (CD68+) is quantified by confocal microscopy in the heart of patients and mice with MI (n=3-5/group). C-D. qPCR quantification of Acly in LyzM +/+ Acly fl/fl and LyzM cre/+ Acly fl/fl (n=10-12/group) ( C ) and Fasn in LyzM +/+ Fasn fl/fl and LyzM cre/+ Acly fl/fl ( D ) cardiac macrophages (n=10-12/group). E-F . Representative echocardiogram M-mode images. The green and red lines indicate LVID;s = left ventricular internal diameter in systole and LVID;d = left ventricular internal diameter in diastole respectively ( E ) and quantification of heart function ( F ) in LysM +/+ Acly fl/fl and LysM cre/+ Acly fl/fl mice on 28 days after MI are provided (n=5-13/group). G . Echocardiography quantification in LysM +/+ Acly fl/fl and LysM cre/+ Acly fl/fl mice in the steady state-(without an MI) (n=5/group). H . The images depict representative echocardiograms of the LV in M-mode in SAX view in LyzM +/+ Fasn fl/fl and LyzM cre/+ Fasn fl/fl mice on day 28 after MI (n=6-10/group). The green and red lines indicate LVID;s = left ventricular internal diameter in systole and LVID;d = left ventricular internal diameter in diastole respectively I. Representative long axis four-chambered (LA-4C), long axis two-chambered (LA-2C), and short axis (SA) views of cine magnetic resonance imaging of LysM +/+ Acly fl/fl and LysM cre/+ Acly fl/fl mice on day 28 after MI (n=5/group) are shown. The data are expressed as mean ± SEM and pooled from 3–5 independent experiments. The Mann-Whitney U test was used for S1C-D and One way ANOVA was performed for Fig S1F ** p < 0.01, *** p < 0.001, **** p < 0.0001.

Article Snippet: Deletion of Acly (The Jackson lab, #030772) and Fasn (Taconic Biosciences, #8034) in myeloid lineages were achieved by crossing these floxed transgenic mice with Lysm Cre/Cre mice (The Jackson lab, #004781).

Techniques: Confocal Microscopy, Staining, Magnetic Resonance Imaging, MANN-WHITNEY

Journal: The Journal of Biological Chemistry

Article Title: Fatty acid synthase inhibits the O- GlcNAcase during oxidative stress

doi: 10.1074/jbc.M116.760785

Figure Lengend Snippet: Oxidative stress induces the association of OGA with FAS, FLNA, HSC70, and OGT. U2OS cells were treated with Vehicle (V) or H2O2 (2.5 mm, 1–3 h). n = 3. A, anti-OGA antibody (IP: OGA; top panel) or a rabbit isotype control immunoglobulin (IP: IgG; middle panel) was used to enrich endogenous OGA from NETN cell lysates (500 μg), of which 1.5–2% (input) and 30–40% (immunoprecipitate) were analyzed by SDS-PAGE. OGA, FAS, FLNA, HSC70, OGT (positive control), and actin (loading/negative control) were detected by Western blotting. B, anti-FAS antibody (IP: FAS; top panel) or a rabbit isotype control immunoglobulin (IP: IgG; middle panel) was used to enrich endogenous FAS from NETN cell lysates (250 μg), of which 3% (input) and 60% (immunoprecipitate) were analyzed by SDS-PAGE. FAS, OGA, HSC70, OGT, and actin (loading/negative control) were detected by Western blotting. C, U2OS cells were transfected with pcDNA3.1 (control) or pCMV-SPORT6 V5-FAS (test). An anti-V5 antibody was used to enrich V5-FAS from control and test NETN cell lysates (300 μg), of which 1.7% (input) and 33.3% (immunoprecipitate) were analyzed by SDS-PAGE. V5, OGA, HSC70, OGT, and actin (loading/negative control) were detected by Western blotting. A–C, FAS (CST) and FAS (NB) represent anti-FAS antibody from Cell Signaling Technology and Novus Biologicals, respectively. To ensure that images were in the linear range, Western blot exposures from the input and immunoprecipitated fractions are often different. The exposure lengths for the test and control isotype antibody immunoprecipitates are always identical. The migration of molecular mass (MW) markers is indicated.

Article Snippet: The following antibodies were used for Western blotting analysis: anti-OGA (345), anti-O-GlcNAc (CTD110.6) (gift from The Johns Hopkins University School of Medicine Core C4); anti-FLNA (A301-135A; Bethyl Laboratories); anti-HA (H6908), anti-actin (A5060), anti-OGT (DM17, O6264), anti-mouse IgM-HRP (A8786; Sigma); NeutrAvidin-HRP (31030), anti-V5 (461157; Thermo Fisher Scientific); anti-Myc (9E10, CRL-1729; American Type Culture Collection (ATCC), Manassas, VA); anti-FAS (3180; Cell Signaling Technology, Danvers, MA); anti-FAS (H00002194-M01; Novus Biologicals, Littleton, CO); anti-HSC70 (sc-7298), anti-chicken IgY-HRP (sc-2428; Santa Cruz Biotechnology, Dallas, TX); anti-rabbit IgG-HRP (NA934V) and anti-mouse IgG-HRP (NA931V; GE Healthcare).

Techniques: SDS Page, Positive Control, Negative Control, Western Blot, Transfection, Immunoprecipitation, Migration