Journal: Scientific Reports

Article Title: The E3 ubiquitin-protein ligase UHRF1 promotes adipogenesis and limits fibrosis by suppressing GPNMB-mediated TGF-β signaling

doi: 10.1038/s41598-024-62508-y

Figure Lengend Snippet: Overexpression of UHRF1 reverses the phenotype observed in UHRF1-depleted cells. ( A ) Representative fluorescence and brightfield microscopy images of lentivirus-mediated expression of GFP control or UHRF1-GFP in HEK293T cells (Scale bar, 100 μm). GFP-positive cells in the total cell population are shown in the merged image. ( B ) Representative confocal microscopy images of GFP control and UHRF1-GFP in HEK293T cells. Nuclei were stained with Hoechst (blue). Scale bar, 50 μm. ( C – H ) Representative western blot data and quantification of changes in the expression levels of UHRF1-GFP, TGF-β, GPNMB, MMP3 and MMP13 levels in UHRF1-GFP HEK293T cells versus GFP control. Target protein levels were normalized to Actin using ImageJ software. ( I ) Fluorescence and brightfield microscopy images of lentivirus-mediated expression of GFP control and UHRF1-GFP in SW872 cells (Scale bar, 100 μm). GFP-positive cells in the total cell population are shown in the merged image. ( J ) Representative confocal microscopy images of GFP control and UHRF1-GFP in SW872 cells. Nuclei were stained with Hoechst (blue). Scale bar, 50 μm. ( K – N ) Representative western blot data and quantification of changes in the expression levels of UHRF1-GFP, PPAR-γ and TGF-β in UHRF1-GFP SW872 cells versus GFP control. Target protein levels were normalized to Actin using ImageJ software. ( O , P ) Oil Red O staining images of oleic acid (50 µM)-induced differentiation of SW872 cells overexpressing GFP control or UHRF1-GFP on day 4 post-differentiation. Scale bar 100 μm. ( P ) Bar graph showing the total number of lipid droplets present in the entire 40 × field Oil Red O images of GFP control and UHRF1-GFP. Data are mean ± SEM of 3 independent experiments. Statistical analyses were performed using ordinary one-way ANOVA, * P < 0.05.

Article Snippet: Human SW872 preadipocytes (ATCC) were cultured in DMEM/F12 medium supplemented with 8% fetal bovine serum (FBS) (catalog no. AlphaFBS-HI, Alphabioregen, Boston, MA, USA), 1% penicillin–streptomycin and 15 mM Hepes.

Techniques: Over Expression, Fluorescence, Microscopy, Expressing, Control, Confocal Microscopy, Staining, Western Blot, Software