Journal: Stem Cell Research & Therapy

Article Title: uPAR deficiency triggers TGFβ1-mediated fibrotic remodeling in a cardiac perivascular-like microenvironment

doi: 10.1186/s13287-026-04923-8

Figure Lengend Snippet: uPAR deficiency promotes TGFβ1 activation, uPA nuclear accumulation and SNAIL upregulation. A Protein and gene expression levels of transforming growth factor β1 (TGFβ1) in Wt and uPAR-/- mCSs, * p < 0.05, N = 3–4. B Protein levels of single-chain urokinase (sc-uPA) and two-chain urokinase (tc-uPA) in Wt and uPAR-/- mCSs, ** p < 0.01, N = 4. C uPA nuclear accumulation (percent of uPA-positive nuclei) within Wt and uPAR-/- mCSs, * p < 0.05, N = 3. D Confocal images of intracellular uPA staining (green) of Wt and uPAR-/- mCS cryosections. Nuclei were stained with DAPI. White segmented arrows indicate the direction of fluorescence intensity profiling presented on panel “E”. Straight arrows indicate colocalization of uPA and DAPI. E Intensity profile plots of the uPA and nuclei/DAPI fluorescence signal. Arrows indicate overlay of fluorescence signals. F Nuclear uPA expression (mean fluorescence intensity per nuclei) within Wt and uPAR-/- mCSs, * p < 0.05, N = 3. G Protein levels of SNAIL and TWIST1 in Wt and uPAR-/- mCSs, **** p < 0.0001, N = 4. H Representative images of western blot results for panels “A”, “B” and “G”. Full-length blots/gels are presented in Supplementary Figure S7. I Confocal images of double-staining for CD31 (red) and α smooth-muscle actin (SMA, green) in Wt and uPAR-/- mouse cardiospheres (mCSs). Nuclei were stained with DAPI. White arrows indicate colocalization of CD31 and SMA. J Intensity profile plots of the CD31 and SMA fluorescence signal. Arrows indicate overlay of fluorescence signals

Article Snippet: To analyze transforming growth factor beta 1 (TGFβ1)-induced signaling activation, cells were treated with 10 ng/mL mouse TGFβ1 (PKSM041167, Elabscience) in DMEM low glucose (Servicebio) for 30 min. Activation of SMAD2/SMAD3 and Akt pathways was assessed by immunoblotting using phospho/total protein ratios.

Techniques: Activation Assay, Gene Expression, Staining, Fluorescence, Expressing, Western Blot, Double Staining

Journal: Stem Cell Research & Therapy

Article Title: uPAR deficiency triggers TGFβ1-mediated fibrotic remodeling in a cardiac perivascular-like microenvironment

doi: 10.1186/s13287-026-04923-8

Figure Lengend Snippet: Plaur knockout in fibroblasts resulted in enhanced ECM deposition and TGFβ1 activation. A Schematic representation of Plaur knockout in fibroblasts using CRISPR/Cas9 system with three single guide RNAs (sgRNA). The illustration was created with Servier Medical Art, licensed under CC-BY 4.0. B Flow cytometry analysis of Wt, Scrambled (control plasmid) and Plaur knockout ( Plaur KO) fibroblasts stained with anti-uPAR antibodies (red), isotype IgG (green), or unstained (blue). C Plaur gene expression in fibroblasts quantified by qPCR, ** p < 0.01, *** p < 0.001, **** p < 0.0001, N = 3. D Analysis of surface uPAR expression on fibroblasts by flow cytometry. Data are presented as median fluorescence intensity (MFI), *** p < 0.001, **** p < 0.0001, N = 3. E Representative images of western blot results for panels “F” and “G”. Full-length blots/gels are presented in Supplementary Figure S8. F Protein levels of COL I (pro-collagen I and mature collagen I α-chain), FN, and αSMA measured in fibroblasts, * p < 0.05, ** p < 0.01, N = 3. G Protein levels of latent and active (dimer and monomer) forms of TGFβ1 in fibroblasts, * p < 0.05, ** p < 0.01, N = 3. H Tgfb1 gene expression in fibroblasts, * p < 0.05, ** p < 0.01, N = 3

Article Snippet: To analyze transforming growth factor beta 1 (TGFβ1)-induced signaling activation, cells were treated with 10 ng/mL mouse TGFβ1 (PKSM041167, Elabscience) in DMEM low glucose (Servicebio) for 30 min. Activation of SMAD2/SMAD3 and Akt pathways was assessed by immunoblotting using phospho/total protein ratios.

Techniques: Knock-Out, Activation Assay, CRISPR, Flow Cytometry, Control, Plasmid Preparation, Staining, Gene Expression, Expressing, Fluorescence, Western Blot

Journal: Stem Cell Research & Therapy

Article Title: uPAR deficiency triggers TGFβ1-mediated fibrotic remodeling in a cardiac perivascular-like microenvironment

doi: 10.1186/s13287-026-04923-8

Figure Lengend Snippet: TGFβ1 further enhances Akt phosphorylation and ECM synthesis in Plaur knockout fibroblasts. A Schematic representation of the experimental design for assessing TGFβ1 effects on Plaur knockout ( Plaur KO) fibroblasts. B Analysis of SMAD2/SMAD3 and Akt phosphorylation in Scrambled and Plaur KO fibroblasts following 30 min TGFβ1 stimulation. Phospho-protein levels were normalized to total proteins levels, * p < 0.05, ** p < 0.01, N = 3. C Representative images of western blot results for panel “B”. Full-length blots/gels are presented in Supplementary Figure S9. D Immunofluorescence images of Wt, Scrambled and Plaur KO fibroblasts after 48-hour TGFβ1 stimulation stained for collagen I (COL I, green) and EDA-fibronectin (EDA-FN, red). Nuclei were stained with DAPI. E Protein levels of COL I (pro-collagen I and mature collagen I α-chain), FN, and αSMA measured in fibroblasts after 48-hour TGFβ1 stimulation, * p < 0.05, N = 3. F Representative images of western blot results for panel “E”. Full-length blots/gels are presented in Supplementary Figure S10. G Schematic summary illustrating that uPAR deficiency in fibroblasts promotes TGFβ1 activation, enhances TGFβ1-dependent signal transduction, and stimulates ECM synthesis. “A” and “G” were created with Servier Medical Art, licensed under CC-BY 4.0

Article Snippet: To analyze transforming growth factor beta 1 (TGFβ1)-induced signaling activation, cells were treated with 10 ng/mL mouse TGFβ1 (PKSM041167, Elabscience) in DMEM low glucose (Servicebio) for 30 min. Activation of SMAD2/SMAD3 and Akt pathways was assessed by immunoblotting using phospho/total protein ratios.

Techniques: Phospho-proteomics, Knock-Out, Western Blot, Immunofluorescence, Staining, Activation Assay, Transduction

Journal: Stem Cell Research & Therapy

Article Title: uPAR deficiency triggers TGFβ1-mediated fibrotic remodeling in a cardiac perivascular-like microenvironment

doi: 10.1186/s13287-026-04923-8

Figure Lengend Snippet: Fibrotic remodeling of the cardiac perivascular microenvironment induced by uPAR deficiency. uPAR: urokinase receptor; uPA: urokinase; EC: endothelial cell; SM: smooth muscle; EndMT: endothelial-to-mesenchymal transition; ECM: extracellular matrix; COL: collagen; TGFβ1: transforming growth factor β1; Created with Servier Medical Art, licensed under CC-BY 4.0

Article Snippet: To analyze transforming growth factor beta 1 (TGFβ1)-induced signaling activation, cells were treated with 10 ng/mL mouse TGFβ1 (PKSM041167, Elabscience) in DMEM low glucose (Servicebio) for 30 min. Activation of SMAD2/SMAD3 and Akt pathways was assessed by immunoblotting using phospho/total protein ratios.

Techniques: