Journal: Diabetes

Article Title: Adiponectin reduces glomerular endothelial glycocalyx disruption and restores glomerular barrier function in a mouse model of type 2 diabetes.

doi: 10.2337/db23-0455

Figure Lengend Snippet: Figure 3—Adiponectin ameliorates TNFa-induced glycocalyx shedding. GEnCs were treated with or without 10 ng/mL TNF-a for 2 h in the presence or absence of 2.5 mg/mL gAd. A) qPCR analysis of SDC4 mRNA levels, normalized to GAPDH, are shown. Data are plotted as the mean 2-(DCT) (gene of interest/housekeeping gene) of each triplicate with the mean. B) SDC4 protein expression was quantified by ELISA in the medium of treated cells. C) Sulfated GAGs were quantified in the medium of treated cells by Alcian blue colorimetric assay. MMP2 (D) and MMP9 (E) mRNA levels in CiGEnCs treated with TNF-a (10 ng/mL) and/or gAd (2.5 mg/mL). A–E) One-way ANOVA (n = 5); *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001 by post hoc analysis (Bonferroni). MMP2 was knocked down using three shRNA dif- ferent sequences in GEnCs (v33, v47, v49) or not, using scrambled controls. F) qPCR data analysis highlighting the decreased expression of MMP2 mRNA in CiGEnCs by all three different shRNA sequences. Data are plotted as the mean comparative cycle threshold of each triplicate. GAPDH used as the housekeeping gene control, n = 4; one-way ANOVA, ****P < 0.0001 post hoc analysis (Bonferroni). Gi) Representative Western Blot demonstrating the protein knockdown of MMP2 by shRNA clone v47. Gii) Densitometry showed significant knockdown of MMP2 protein expression by clone v47. Data are normalized to the b-actin loading control. Dots represent mean ± SEM; one-way ANOVA, ***P < 0.001 by post hoc analysis (Bonferroni). H) SDC4 mRNA levels in MMP2 knockdown CiGEnCs treated with TNF-a and/or gAd. Data are plotted as the mean comparative cycle threshold of each triplicate. GAPDH was used as the housekeeping gene control, n = 5; one-way ANOVA, ***P < 0.001 post hoc analysis (Bonferroni).

Article Snippet: Cellular levels of SDC4 were quantified using a SDC4 ELISA (R&D Systems; catalog DY2918) per the manufacturer’s instructions.

Techniques: Expressing, Enzyme-linked Immunosorbent Assay, Colorimetric Assay, shRNA, Control, Western Blot, Knockdown

Journal: Diabetes

Article Title: Adiponectin reduces glomerular endothelial glycocalyx disruption and restores glomerular barrier function in a mouse model of type 2 diabetes.

doi: 10.2337/db23-0455

Figure Lengend Snippet: Figure 4—Adiponectin signals to glomeruli directly and prevents the mRNA upregulation of glycocalyx-related genes in db/db glomeruli. Hu- man (A) and mouse (B) glomeruli were isolated and treated ex vivo with 2.5 mg/mL gAd. Representative Western Blots demonstrating gAd ef- fects at 30 min in human (Ai) and mouse (Bi) ex vivo glomeruli on AMPK phosphorylation (p-). Aii and Bii) Densitometry confirmed significant phosphorylation of AMPK in response to gAd after 30 min in human and mouse glomeruli. Densitometry was performed on blots from inde- pendent repeats (n = 4) showing levels of protein of interest normalized to the b-actin loading control. Dots represent mean ± SEM; one-way ANOVA; *P < 0.05, by post hoc analysis (Bonferroni). C–F) qPCR analysis graph representing the mRNA expression of TNF (C), SDC4 (D), MMP2 (E), and MMP9 (F) in ex vivo (wild type or db/db) sieved glomeruli treated with or without adiponectin. Data are plotted as the mean comparative cycle threshold of each triplicate, n = 5; one-way ANOVA; *P < 0.05, **P < 0.01, ****P < 0.0001 by post hoc analysis (Bonferroni).

Article Snippet: Cellular levels of SDC4 were quantified using a SDC4 ELISA (R&D Systems; catalog DY2918) per the manufacturer’s instructions.

Techniques: Isolation, Ex Vivo, Western Blot, Phospho-proteomics, Control, Expressing

Journal: International Journal of Molecular Sciences

Article Title: Syndecan-3 as a Novel Biomarker in Alzheimer’s Disease

doi: 10.3390/ijms23063407

Figure Lengend Snippet: SDC3 expression of TNF-α-treated SH-SY5Y and hCMEC/D3 cells. Cells were incubated with or without (i.e., controls) 5 ng/mL TNF-α for 7 days. After incubation, the cells were treated with APC-labeled SDC3 antibody, and SDC3 was measured with imaging flow cytometry. ( A ) Representative flow cytometry histograms showing the SDC3 expression of SH-SY5Y and hCMEC/D3 cells. ( B ) Brightfield (BF) and fluorescent cellular images of SH-SY5Y and hCMEC/D3 cells treated with APC-labeled SDC3 antibody. Scale bar = 20 μm. ( C ) Detected fluorescence intensities were normalized control cells untreated with TNF-α. The bars represent the mean + SEM of ten independent experiments. Statistical significance vs. controls was assessed with ANOVA. *** p < 0.001.

Article Snippet: SDC3 expression of SH-SY5Y and hCMEC/D3 cells incubated with or without 5 ng/mL of recombinant TNF-α (cat. no. 210-TA-100/CF, RnD Systems, Minneapolis, MN, USA) for 7 days was measured with an AMNIS FlowSight imaging flow cytometer (Luminex Corporation, Austin, TX, USA) by using APC-labeled anti-human SDC3 antibody (polyclonal goat IgG, cat. no. FAB3539A, RnD Systems) as described previously [ , ].

Techniques: Expressing, Incubation, Labeling, Imaging, Flow Cytometry, Fluorescence, Control

Journal: International Journal of Molecular Sciences

Article Title: Syndecan-3 as a Novel Biomarker in Alzheimer’s Disease

doi: 10.3390/ijms23063407

Figure Lengend Snippet: APPSWE-Tau mice exhibits increased TNF-α concentrations in the brain and the blood. ( A , B ) TNF-α concentrations of brain extracts ( A ) and whole blood ( B ) of APPSWE-Tau mice, along with representative WT controls, were measured with a mouse SDC3 ELISA kit. Each group contained 8 animals. The bars represent the mean + SEM. Statistical significance vs. WT was assessed with ANOVA. ** p < 0.01. ( C ) Linear regression between the TNF-α content of blood and brain.

Article Snippet: SDC3 expression of SH-SY5Y and hCMEC/D3 cells incubated with or without 5 ng/mL of recombinant TNF-α (cat. no. 210-TA-100/CF, RnD Systems, Minneapolis, MN, USA) for 7 days was measured with an AMNIS FlowSight imaging flow cytometer (Luminex Corporation, Austin, TX, USA) by using APC-labeled anti-human SDC3 antibody (polyclonal goat IgG, cat. no. FAB3539A, RnD Systems) as described previously [ , ].

Techniques: Enzyme-linked Immunosorbent Assay

Journal: International Journal of Molecular Sciences

Article Title: Syndecan-3 as a Novel Biomarker in Alzheimer’s Disease

doi: 10.3390/ijms23063407

Figure Lengend Snippet: APPSWE-Tau mice exhibits increased SDC3 concentrations in the brain and the liver. ( A , B ) SDC3 concentrations of the brain ( A ) and liver ( C ) extracts of APPSWE-Tau mice and representative WT controls were measured with ELISA. Each group contained 8 animals. The bars represent the mean + SEM. Statistical significance vs. WT was assessed with ANOVA. * p < 0.05. ( C , D ) Linear regression between the TNF-α and SDC3 concentrations in the brain ( C ) and liver ( D ).

Article Snippet: SDC3 expression of SH-SY5Y and hCMEC/D3 cells incubated with or without 5 ng/mL of recombinant TNF-α (cat. no. 210-TA-100/CF, RnD Systems, Minneapolis, MN, USA) for 7 days was measured with an AMNIS FlowSight imaging flow cytometer (Luminex Corporation, Austin, TX, USA) by using APC-labeled anti-human SDC3 antibody (polyclonal goat IgG, cat. no. FAB3539A, RnD Systems) as described previously [ , ].

Techniques: Enzyme-linked Immunosorbent Assay

Journal: International Journal of Molecular Sciences

Article Title: Syndecan-3 as a Novel Biomarker in Alzheimer’s Disease

doi: 10.3390/ijms23063407

Figure Lengend Snippet: PBECs of APPSWE-Tau mice exhibits increased SDC3 expression. Isolated PBECs were treated with PECAM-1 and SDC3 antibodies, and SDC3 expression of PECAM-1 positive cells was analyzed with imaging flow cytometry. ( A ) Representative histogram showing the SDC3 expression of PBECs isolated from APPSWE-Tau and WT mice. ( B ) BF and fluorescent cellular images of PECAM-1 and SDC3 antibody-treated PBECs, isolated from APPSWE-Tau and WT mice. Each group contained eight animals. SDC3 expression of each sample was measured twice. ( C ) Detected fluorescence intensities were normalized to WT. The bars represent the mean + SEM. Statistical significance vs. WT was assessed with ANOVA. * p < 0.05. ( D ) Linear regression between in PBECs’ relative SDC3 expression and blood TNF-α concentrations.

Article Snippet: SDC3 expression of SH-SY5Y and hCMEC/D3 cells incubated with or without 5 ng/mL of recombinant TNF-α (cat. no. 210-TA-100/CF, RnD Systems, Minneapolis, MN, USA) for 7 days was measured with an AMNIS FlowSight imaging flow cytometer (Luminex Corporation, Austin, TX, USA) by using APC-labeled anti-human SDC3 antibody (polyclonal goat IgG, cat. no. FAB3539A, RnD Systems) as described previously [ , ].

Techniques: Expressing, Isolation, Imaging, Flow Cytometry, Fluorescence

Journal: International Journal of Molecular Sciences

Article Title: Syndecan-3 as a Novel Biomarker in Alzheimer’s Disease

doi: 10.3390/ijms23063407

Figure Lengend Snippet: Monocytes isolated from APPSWE-Tau mice exhibits increased SDC3 expression. Isolated monocytes were treated with CD11b and SDC3 antibodies, and SDC3 expression of CD11b positive cells was analyzed with imaging flow cytometry. ( A ) Representative histogram showing the SDC3 expression of monocytes isolated from APPSWE-Tau and WT mice. ( B ) BF and fluorescent cellular images of CD11b and SDC3 antibody-treated monocytes, isolated from APPSWE-Tau and WT mice. Each group contained eight animals. SDC3 expression of each sample was measured twice. ( C ) Detected fluorescence intensities were normalized to WT. The bars represent the mean + SEM. Statistical significance vs. WT was assessed with ANOVA. *** p < 0.001. ( D ) Linear regression between the SDC3 expression of monocytes and Aβ plaque loads.

Article Snippet: SDC3 expression of SH-SY5Y and hCMEC/D3 cells incubated with or without 5 ng/mL of recombinant TNF-α (cat. no. 210-TA-100/CF, RnD Systems, Minneapolis, MN, USA) for 7 days was measured with an AMNIS FlowSight imaging flow cytometer (Luminex Corporation, Austin, TX, USA) by using APC-labeled anti-human SDC3 antibody (polyclonal goat IgG, cat. no. FAB3539A, RnD Systems) as described previously [ , ].

Techniques: Isolation, Expressing, Imaging, Flow Cytometry, Fluorescence

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

Article Title: Targeted Phage Display-based Pulmonary Vaccination in Mice and Non-human Primates

doi: 10.1016/j.medj.2020.10.005

Figure Lengend Snippet: KEY RESOURCES TABLE

Article Snippet: ​ REAGENT or RESOURCE SOURCE IDENTIFIER Antibodies Anti-fd bacteriophage antibody Sigma-Aldrich Cat# B7786 Anti-mouse integrin α3/CD49c antibody Novus Biologicals Cat# NBP1-19724 Anti-human integrin α3 (ASC-1) blocking antibody Millipore Sigma Cat# MAB2056Z anti-proSPC antibody Millipore Cat# AB3786 anti-podoplanin antibody Thermo-Invitrogen Cat# eBio8.1.1 anti-CCSP antibody Millipore-Merck Cat# 07-623 anti-mouse CD16/CD32 antibody BD Pharmigen Cat# 553142 rat anti-mouse EPCAM brilliant violet 421 BioLegend Cat# 118225 anti-mouse CD45 Alexa fluor 700 eBioscience Cat# 56-0451-80 anti-mouse podoplanin monoclonal antibody PE-Cyanine 7 eBioscience Cat# 25-5382-80 anti-CD31 FITC BD Pharmigen Cat# 553372 anti-mouse F4/80 PE-conjugated BD Pharmigen Cat# 565410 Anti-rabbit Horseradish peroxidase (HRP)-conjugated secondary antibody Jackson ImmunoResearch Cat# 111-035-008 Cy3-conjugated goat anti-human secondary antibody Jackson ImmunoResearch Cat# 109-165-008 Anti-monkey IgG KPL Cat# 074-11-021 Anti-monkey IgA KPL Cat# 074-11-011 Bacterial and Virus Strains fUSE5 bacteriophage Laboratory of Dr. George L. Smith N/A K91 E. coli Laboratory of Dr. George L. Smith N/A DH5α electrocompetent Invitrogen Cat# 11319019 pGEX4T-1 plasmid Amersham/ Addgene Cat# 27458001 Biological Samples Chemicals, Peptides, and Recombinant Proteins α3β1 R&D Systems Cat# 2840-A3-050 α6β1 R&D Systems Cat# 7809-A6-050 α6β4 R&D Systems Cat# 5497-A6-050 NRP-1 R&D Systems Cat# 3870-N1-025 SDC-1 R&D Systems Cat# 2780-SD-050 CAKSMGDIVC peptide Biomatik Cat# 293308 Lot# P120420-SL293308 CGRRAGGSC peptide PolyPeptide N/A Triton X-100 Sigma-Aldrich Cat# 9002-93-1 Triton X-114 Sigma-Aldrich Cat# 9036-19-5 Dextran Invitrogen Cat# D1822 Evans Blue Sigma-Aldrich Cat# 314-13-6 LPS - Klebsiella pneumoniae Sigma-Aldrich Cat# L4268 Trypan Blue ThermoFisher Scientific Cat# 15250061 Protein G agarose resin Roche Cat# 11-243-233-001 Jacalin ThermoScientific Cat# 20395 Histochoice Sigma-Aldrich Cat# H2904 Dako Target Retrieval Solution Agilent Dako Cat# S169984-2 T4 DNA ligase Roche Cat# 10799009001 Critical Commercial Assays BCA Protein Assay ThermoFisher Scientific Cat# 23227 QIAGEN Plasmid Midi Kit Qiagen Cat# 12145 Deposited Data Experimental Models: Cell Lines A549 ATCC Cat# CCL-185 Experimental Models: Organisms/Strains Mice/ BALB/cJ The Jackson Laboratory Stock # 000651 Rhesus monkeys Michale E. Keeling Center for Comparative Medicine and Research – MDACC, Bastrop, TX, USA.

Techniques: Blocking Assay, Virus, Plasmid Preparation, Recombinant, Bicinchoninic Acid Protein Assay, Sequencing, shRNA, Software

Journal: bioRxiv

Article Title: Mucin-type O -glycans regulate proteoglycan stability and chondrocyte maturation

doi: 10.64898/2025.12.11.693745

Figure Lengend Snippet: Venn diagrams comparing significantly downregulated (A) and upregulated (B) genes (Fold change ≥ 2, FDR p ≤ 0.01) from RNA-seq datasets for COSMC -/- and C1GALT1 -/- knockout cells. Gene Ontology enrichment analysis of overlapping 128 downregulated (C) and 219 upregulated (D) genes shared in COSMC -/- and C1GALT1 -/- cells. (E) Differential expression analysis of proteoglycan and GAG biosynthetic genes (FDR p ≤ 0.01) compared to wild-type control cells. Flow cytometry analysis of (F) syndecan-1 (SDC1) and (G) syndecan-2 levels in TC28a2 wild-type, COSMC -/- , and C1GALT1 -/- cells. Data points are shown as mean ± SD (n ≥ 3 independent experiments); p-values were determined by one-way ANOVA with Tukey post-test with **** p < 0.0001, *** p < 0.001, ** p < 0.01, * p < 0.05.

Article Snippet: For the detection of other proteins and ligands, lifted cells were incubated in suspension for 30 min at 4 °C with 0.5 μg/mL mAb 10E4 (AMSBio #370255-1, Clone F58-10E4, 1:5000), 1 μg/mL mAb 3G10 (AMSBio, #370260-S, clone F69-3G10, 1:1000), 1 μg/mL mAb 2B6 (AMSBio # 270432-CS, Clone 2B6, 1:200), 0.1 μg/mL pan-CD44 mAb (Thermo #14-0441-82, Clone IM7, 1:1000), 0.5 μg/mL anti-CD44v3 mAb (R&D Systems #BBA11, Clone 3G5, 1:1000), 0.2 μg/mL anti-SDC1 mAb (Novus Biologicals #NB100-64980, Clone B-A38, 1:500), 0.5 μg/mL anti-SDC2 mAb (R&D Systems #MAB2965, Clone 305515, 1:1000), 80 nM biotin-FGF1, or 2.5 nM biotin-FGF2, respectively.

Techniques: RNA Sequencing, Knock-Out, Quantitative Proteomics, Control, Flow Cytometry

Journal: bioRxiv

Article Title: Mucin-type O -glycans regulate proteoglycan stability and chondrocyte maturation

doi: 10.64898/2025.12.11.693745

Figure Lengend Snippet: (A) Time course quantification of cell surface SDC1 levels via flow cytometry prior to and after trypsinization (0-24 hours). Data is normalized to untreated wild-type cells (dashed line). (B) Quantification of SDC1 recovery to the cell surface. Data were normalized to the respective untreated control for each genotype and fitted using linear regression. Table inset includes slopes ± SD for each. Statistical comparisons were performed using an F-test to compare the slopes of knockout lines versus wild-type cells (* p < 0.05). (C) Quantification of total SDC1 protein levels in wild-type, COSMC -/- , and C1GALT1 -/- cells treated with bafilomycin A1 (0.25 µM) or DMSO vehicle for 16 hours. (D) Representative histograms from flow cytometry analysis of total SDC1 levels in cells treated with bafilomycin A1. (E) Flow cytometry analysis of cell surface SDC1 levels after treatment with bafilomycin A1 or DMSO vehicle. Data points are shown as mean ± SD (n ≥ 3 independent experiments); p-values were determined by one-way ANOVA with Tukey post-test (A,C), and student’s T-test (E) with **** p < 0.0001, *** p < 0.001, ** p < 0.01, * p < 0.05.

Article Snippet: For the detection of other proteins and ligands, lifted cells were incubated in suspension for 30 min at 4 °C with 0.5 μg/mL mAb 10E4 (AMSBio #370255-1, Clone F58-10E4, 1:5000), 1 μg/mL mAb 3G10 (AMSBio, #370260-S, clone F69-3G10, 1:1000), 1 μg/mL mAb 2B6 (AMSBio # 270432-CS, Clone 2B6, 1:200), 0.1 μg/mL pan-CD44 mAb (Thermo #14-0441-82, Clone IM7, 1:1000), 0.5 μg/mL anti-CD44v3 mAb (R&D Systems #BBA11, Clone 3G5, 1:1000), 0.2 μg/mL anti-SDC1 mAb (Novus Biologicals #NB100-64980, Clone B-A38, 1:500), 0.5 μg/mL anti-SDC2 mAb (R&D Systems #MAB2965, Clone 305515, 1:1000), 80 nM biotin-FGF1, or 2.5 nM biotin-FGF2, respectively.

Techniques: Flow Cytometry, Control, Knock-Out

Journal: bioRxiv

Article Title: Mucin-type O -glycans regulate proteoglycan stability and chondrocyte maturation

doi: 10.64898/2025.12.11.693745

Figure Lengend Snippet: (A) Representative confocal images of SDC1 (red) and nuclei (DAPI; blue) in DMSO-treated cells. The scale bar represents 20 nm. (B) Quantification of SDC1 fluorescence intensity using ImageJ. (C) Representative confocal images of SDC1 (red) and nuclei (DAPI; blue) in cells after treatment with bafilomycin A1 (250 nM) for 16 hours. The scale bar represents 20 nm. (B) Quantification of SDC1 fluorescence intensity in bafilomycin A1-treated cells versus wild-type controls. Data points are shown as mean ± SD (n = 25-35 cells from three independent biological replicates); p-values were determined by one-way ANOVA with Tukey post-test with **** p < 0.0001.

Article Snippet: For the detection of other proteins and ligands, lifted cells were incubated in suspension for 30 min at 4 °C with 0.5 μg/mL mAb 10E4 (AMSBio #370255-1, Clone F58-10E4, 1:5000), 1 μg/mL mAb 3G10 (AMSBio, #370260-S, clone F69-3G10, 1:1000), 1 μg/mL mAb 2B6 (AMSBio # 270432-CS, Clone 2B6, 1:200), 0.1 μg/mL pan-CD44 mAb (Thermo #14-0441-82, Clone IM7, 1:1000), 0.5 μg/mL anti-CD44v3 mAb (R&D Systems #BBA11, Clone 3G5, 1:1000), 0.2 μg/mL anti-SDC1 mAb (Novus Biologicals #NB100-64980, Clone B-A38, 1:500), 0.5 μg/mL anti-SDC2 mAb (R&D Systems #MAB2965, Clone 305515, 1:1000), 80 nM biotin-FGF1, or 2.5 nM biotin-FGF2, respectively.

Techniques: Fluorescence

Journal: Scientific Reports

Article Title: Glycoprotein non-metastatic melanoma protein B is a potential biomarker for arthroplasty aseptic loosening

doi: 10.1038/s41598-025-13922-3

Figure Lengend Snippet: Transcriptome analysis of eight sex- and age-balanced BM-MSC preparations by RNA-seq. ( A ) Volcano plot emphasizing differentially expressed genes with DESeq2-based p adj -value cut-off 0.05. These genes were included in the gene set enrichment analysis in C , D . Arrows highlight GPNMB and SDC4 . ( B ) Heatmap showing differentially expressed genes with DESeq2-based p adj -value cut-off 0.05. Qlucore Omics Explorer was used for the heatmap visualisation and normalization of the data (each column has mean of 0 and variance of 1) of four primary hip surgeries vs. four revisions (2x hip, 2x knee). Arrows highlight GPNMB and SDC4 . ( C ) KEGG analyses (KEGG: https://www.kegg.jp/ ; of downregulated genes (upper panel) as identified using DESeq2. Gene ontologies for Biological Processes (middle panel) and Molecular Functions (lower panel) are included. ( D ) KEGG analyses of upregulated genes (upper panel) as identified using DESeq2. Gene ontologies for Biological Processes (middle panel) and Molecular Functions (lower panel) are included.

Article Snippet: Afterwards, sandwich ELISA was performed to analyse the secretion of GPNMB (Human Osteoactivin/GPNMB DuoSet ELISA; DY2550, R&D Systems) or SDC4 (Human Syndecan-4 DuoSet ELISA; DY2918, R&D Systems) according to the manufacturer’s protocols.

Techniques: RNA Sequencing

Journal: Scientific Reports

Article Title: Glycoprotein non-metastatic melanoma protein B is a potential biomarker for arthroplasty aseptic loosening

doi: 10.1038/s41598-025-13922-3

Figure Lengend Snippet: Transcriptome analysis of all primary vs. all revision arthroplasties at both implantation sites (28 samples). ( A ) KEGG analyses (KEGG: https://www.kegg.jp/ ; of downregulated genes (upper panel). Gene ontologies for Biological Processes (middle panel) and Molecular Functions (lower panel) are included. ( B ) KEGG analyses of upregulated genes (upper panel). Gene ontologies for Biological Processes (middle panel) and Molecular Functions (lower panel) are included. ( C ) RNA-seq data for GPNMB in BM-MSCs (normalised counts). For the statistical comparisons between primary and revision the parametric unpaired Welch’s t-test was used. Experimental data are shown as individual values. ( D , E ) GPNMB relative expression levels in BM-MSCs based on RPS29 as housekeeping gene. E indicates the levels for individual samples. ( F ) GPNMB relative expression levels in MNCs from blood (PB) and synovial fluid (SF). ( G ) RNA-seq data for SDC4 in BM-MSCs (normalised counts). For the statistical comparisons between primary and revision the parametric unpaired Welch’s t-test was used. Experimental data are shown as individual values. (H , I) SDC4 relative expression levels in BM-MSCs based on RPS29 as housekeeping gene. I indicates the levels for individual samples. ( J ) SDC4 relative expression levels in MNCs from blood (PB) and synovial fluid (SF). The black bar in C, D, G, H represents the median value. * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001.

Article Snippet: Afterwards, sandwich ELISA was performed to analyse the secretion of GPNMB (Human Osteoactivin/GPNMB DuoSet ELISA; DY2550, R&D Systems) or SDC4 (Human Syndecan-4 DuoSet ELISA; DY2918, R&D Systems) according to the manufacturer’s protocols.

Techniques: RNA Sequencing, Expressing

Journal: Cellular and Molecular Life Sciences: CMLS

Article Title: Syndecan-3 positively regulates the pro-inflammatory function of macrophages

doi: 10.1007/s00018-025-05649-1

Figure Lengend Snippet: SDC3, but not the other syndecans, is upregulated in response to IFNγ in human macrophages. ( a ) SDC1 , SDC2 , SDC3 and SDC4 mRNA levels in control and IFNγ-treated THP-1 macrophages, as assessed by RT-qPCR ( n = 4–6). ( b ) Representative flow histograms (upper panel) and pooled data (lower panel) showing the protein expression of SDC1, SDC2, SDC3 and SDC4 in control and IFNγ-treated THP-1 macrophages ( n = 4–6). ( c ) Representative confocal images ( n = 3) of control, IFNγ- or IL-4/IL-13-treated THP-1 macrophages immunostained for DAPI and SDC3 (scale bar 5 μm). ( d ) SDC3 mRNA levels in control, IFNγ- and IL-4/IL-13-treated THP-1 macrophages, as assessed by RT-qPCR ( n = 3). Means ± SEM. Statistically significant difference from controls or between indicated groups is shown by ns: not significant, ** p < 0.01, *** p < 0.001 and **** p < 0.0001

Article Snippet: SDC3 primary antibody (R&D Systems Cat. #: AF3539), was incubated overnight at 4 °C in blocking buffer.

Techniques: Control, Quantitative RT-PCR, Expressing

Journal: Cellular and Molecular Life Sciences: CMLS

Article Title: Syndecan-3 positively regulates the pro-inflammatory function of macrophages

doi: 10.1007/s00018-025-05649-1

Figure Lengend Snippet: SDC3 deficient macrophages exhibit aberrant proliferation, adhesion and expression of cell surface markers. ( a ) Representative immunoblots ( n = 3) of WT or SDC3 KO THP-1 macrophages stimulated with either 100 ng/ml of IFNγ or 20 ng/ml of IL-4/IL-13 cocktail. ( b ) SDC3 mRNA levels in control, IFNγ- and IL-4/IL-13-treated WT or SDC3 KO THP-1 macrophages, as assessed by RT-qPCR ( n = 3) ( c ) Representative brightfield images of WT or SDC3 KO THP-1 macrophages (scale bar 20 μm, n = 3). ( d ) Proliferation of WT or SDC3 KO THP-1 cells up to 5 days ( n = 3). ( e ) Adhesion activity of WT or SDC3 KO THP-1 macrophages, as quantified by Crystal Violet ( n = 2–3). ( f ) Flow cytometry data showing the expression of CD40, CD86 and CD163 in WT or SDC3 KO THP-1 macrophages stimulated with 100 ng/ml of IFNγ ( n = 7). Means ± SEM. Statistically significant difference from controls or between indicated groups is shown by ns: not significant, * p < 0.05, ** p < 0.01, *** p < 0.001, and **** p < 0.0001

Article Snippet: SDC3 primary antibody (R&D Systems Cat. #: AF3539), was incubated overnight at 4 °C in blocking buffer.

Techniques: Expressing, Western Blot, Control, Quantitative RT-PCR, Activity Assay, Flow Cytometry

Journal: Cellular and Molecular Life Sciences: CMLS

Article Title: Syndecan-3 positively regulates the pro-inflammatory function of macrophages

doi: 10.1007/s00018-025-05649-1

Figure Lengend Snippet: SDC3 -defective macrophages show distinctive gene expression patterns. ( a ) Heatmap of differentially expressed genes in IFNγ-treated THP-1 WT and SDC3 KO macrophages ( n = 3, purple represents upregulation and green represents downregulation). ( b ) Differentially expressed pathways in IFNγ-treated THP-1 WT and SDC3 KO macrophages where blue represents pathways upregulated in SDC3 KO macrophages, and orange represents downregulated pathways in SDC3 KO macrophages ( n = 3). ( c ) TNF , IL10 , iNOS , PD-L1 , CD86 , and VEGFA mRNA levels in IFNγ-treated THP-1 WT and SDC3 KO macrophages, as assessed by RT-qPCR ( n = 3). Means ± SEM. Statistically significant difference from controls or between indicated groups is shown by * p < 0.05, ** p < 0.01, *** p < 0.001 and **** p < 0.0001

Article Snippet: SDC3 primary antibody (R&D Systems Cat. #: AF3539), was incubated overnight at 4 °C in blocking buffer.

Techniques: Gene Expression, Quantitative RT-PCR

Journal: Cellular and Molecular Life Sciences: CMLS

Article Title: Syndecan-3 positively regulates the pro-inflammatory function of macrophages

doi: 10.1007/s00018-025-05649-1

Figure Lengend Snippet: SDC3 promotes macrophage phagocytic capacity and inhibits tumour-spheroid formation. ( a ) Left: Representative images of IFNγ-stimulated WT or SDC3 KO THP-1 macrophages co-cultured with MDA-MB-231-GFP breast cancer cells. Images were taken with a Leica SP8 Lightning confocal microscope (scale bar 20.5 μm, n = 3). Right: Quantification of phagocytosis rate by flow cytometry normalized to WT, using the inhibitor of actin polymerization Cytochalasin D as control ( n = 3). ( b ) Left: Representative images of IFNγ-treated WT or SDC3 KO THP-1 macrophages forming spheroids with MDA-MB-231-GFP cells. Images were taken using Nikon Eclipse TD 100 microscope at the indicated times (scale bar 100 μm, white colour represents GFP + tumour cells). Right: Proliferation of MDA-MB-231-GFP + tumour cells in each condition normalized to MDA-MB-231 condition, calculated after acquiring single cell suspensions of spheroids by flow cytometry ( n = 3). Means ± SEM. Statistically significant difference from controls or between indicated groups is shown by * p < 0.05, ** p < 0.01 and **** p < 0.0001

Article Snippet: SDC3 primary antibody (R&D Systems Cat. #: AF3539), was incubated overnight at 4 °C in blocking buffer.

Techniques: Cell Culture, Microscopy, Flow Cytometry, Control

Journal: Cellular and Molecular Life Sciences: CMLS

Article Title: Syndecan-3 positively regulates the pro-inflammatory function of macrophages

doi: 10.1007/s00018-025-05649-1

Figure Lengend Snippet: SDC3 plays a role in macrophage pro-inflammatory functions. ( a ) Left: Representative confocal microscopy images of WT or SDC3 KO THP-1 macrophages stimulated with 100 ng/ml of IFNγ and incubated with pHrodo™ Green S. aureus bioparticles. Images were taken using a Leica SP8 Lightning confocal microscope (scale bar 10 μm). Right: Flow cytometry quantification of S. aureus phagocytosis normalized to WT, using the inhibitor of actin polymerization Cytochalasin D as control ( n = 3). ( b ) Cytokine quantification in cell supernatants from WT or SDC3 KO THP-1 macrophages stimulated with 100 ng/ml of IFNγ, using the LEGENDplex™ HU Essential Immune Response Panel ( n = 5). ( c ) Quantification of T cell proliferation index using CellTrace CFSE staining ( n = 5) in CD8 + T cells co-cultured with IFNγ-treated WT or SDC3 KO THP-1 macrophages. ( d ) Quantification of T cell activation markers PD-1 and CD95 by flow cytometry in CD8 + T cells co-cultured with IFNγ-treated WT or SDC3 KO THP-1 macrophages ( n = 5). Means ± SEM. Statistically significant difference from controls or between indicated groups is shown by ns: not significant, * p < 0.05, ** p < 0.01, *** p < 0.001 and **** p < 0.0001

Article Snippet: SDC3 primary antibody (R&D Systems Cat. #: AF3539), was incubated overnight at 4 °C in blocking buffer.

Techniques: Confocal Microscopy, Incubation, Microscopy, Flow Cytometry, Control, Staining, Cell Culture, Activation Assay

Journal: Cellular and Molecular Life Sciences: CMLS

Article Title: Syndecan-3 positively regulates the pro-inflammatory function of macrophages

doi: 10.1007/s00018-025-05649-1

Figure Lengend Snippet: SDC3 deficient macrophages regulate EC migration and tube formation. ( a ) HUVEC migration over the indicated times in response to conditioned media (CM) from WT or SDC3 KO THP-1 macrophages stimulated with 100 ng/ml of IFNγ. Serum-free media (SFM) was used as negative control ( n = 4). ( b ) Quantification of HUVEC migration (Cell Index) shown in (a) at 48 h ( n = 4). ( c ) Representative images of Matrigel-grown HUVECs treated with supernatants from WT or SDC3 KO THP-1 macrophages stimulated with 100 ng/ml of IFNγ. Tube formation was imaged using an Olympus IX-83 inverted microscope (scale bar 1 mm, n = 4). ( d ) Quantification of total tube number shown in (c) at 24 h, relative to WT untreated control ( n = 4). ( e ) VEGFA quantification by ELISA in the supernatants of WT or SDC3 KO THP-1 macrophages stimulated with 100 ng/ml of IFNγ ( n = 4). ( f ) Angiogenic factor quantification in cell supernatants from WT or SDC3 KO THP-1 macrophages stimulated with 100 ng/ml of IFNγ, using the LEGENDplex™ HU Angiogenesis Panel 1 ( n = 4). Means ± SEM. Statistically significant difference from controls or between indicated groups is shown by ns: not significant, * p < 0.05 and *** p < 0.001

Article Snippet: SDC3 primary antibody (R&D Systems Cat. #: AF3539), was incubated overnight at 4 °C in blocking buffer.

Techniques: Migration, Negative Control, Inverted Microscopy, Control, Enzyme-linked Immunosorbent Assay

Journal: Cellular and Molecular Life Sciences: CMLS

Article Title: Syndecan-3 positively regulates the pro-inflammatory function of macrophages

doi: 10.1007/s00018-025-05649-1

Figure Lengend Snippet: Macrophage-derived SDC3 plays a role in the regulation of the TME. The plethora of cells present in the TME interact to promote or inhibit tumour progression and growth. SDC3 is expressed by macrophages in response to the pro-inflammatory cytokine IFNγ and regulates many aspects of the TME. Firstly, SDC3 in macrophages reduces tumour proliferation and is necessary for macrophage-mediated tumour-cell phagocytosis. The expression of SDC3 promotes a pro-inflammatory phenotype in macrophages that results in the secretion of cytokines such as MCP-1 and IL-10, while reducing the release of IL-8. Additionally, SDC3 is necessary for the acquisition of an effector phenotype by T cells, shown by an increased expression of PD-1 and CD95 on T cells. Finally, macrophage-derived SDC3 plays a role in the inhibition of endothelial cell migration and proliferation through a reduction in the secretion of angiogenic factors such as VEGFA, PECAM-1 and IL-8

Article Snippet: SDC3 primary antibody (R&D Systems Cat. #: AF3539), was incubated overnight at 4 °C in blocking buffer.

Techniques: Derivative Assay, Expressing, Inhibition, Migration