Journal: Scientific Reports

Article Title: Chronic exposure to TNF reprograms cell signaling pathways in fibroblast-like synoviocytes by establishing long-term inflammatory memory

doi: 10.1038/s41598-020-77380-9

Figure Lengend Snippet: SAGs regulate pro-inflammatory signaling networks in mouse and human FLS. ( A ) Venn diagram showing the overlap between genes assigned to ATAC-seq and DNase-seq peaks in short-term (16 h) and chronic-TNF conditions with that of mSAGs identified from Fig. A. Overlapping SAGs are shown in blue. ( B ) Venn-diagram showing the overlap between gene upregulated by > twofold in human RA-FLS treated with 10 ng/mL recombinant TNF for 8 h and 24 h with that of 141 mSAGs identified in Fig. 3A. 52 overlapping human SAGs are shown in blue. ( C ) Heatmap of hSAGs. Each box represents one gene and is color-coded based on normalized expression levels. ( D ) Functional protein–protein interaction between hSAGs by STRING analysis. Proteins interacted with RELB are highlighted. ( E ) Top 10 transcriptional factors predicted to regulate hSAGs expression identified by ChEA3 online tool.

Article Snippet: Mice fibroblast cells (FLS) treated with 5 ng/mL recombinant human TNF (R&D Systems) for 16 h were harvested at 95% confluence.

Techniques: Recombinant, Expressing, Functional Assay

Journal: Scientific Reports

Article Title: Chronic exposure to TNF reprograms cell signaling pathways in fibroblast-like synoviocytes by establishing long-term inflammatory memory

doi: 10.1038/s41598-020-77380-9

Figure Lengend Snippet: SRGs regulate BMP signaling in mouse and human FLS. ( A ) Venn diagram showing the overlap between genes assigned to ATAC-seq and DNase-seq peaks in short-term (16 h) and chronic-TNF conditions with that of mSRGs identified from Fig. B. Overlapping SRGs are shown in red. ( B ) Venn-diagram showing the overlap between genes downregulated by > twofold in human RA-FLS treated with 10 ng/mL recombinant TNF for 8 h and 24 h with that of 34 mSAGs identified in Fig. 5A. 7 overlapping human SRGs are shown in red. ( C ) Heatmap of hSRGs. Each box represents one gene and is color-coded based on normalized expression levels. ( D ) Functional protein–protein interaction between hSRGs by STRING analysis. ( E ) Top 5 transcriptional factors predicted to regulate hSAGs expression identified by ChEA3 online tool.

Article Snippet: Mice fibroblast cells (FLS) treated with 5 ng/mL recombinant human TNF (R&D Systems) for 16 h were harvested at 95% confluence.

Techniques: Recombinant, Expressing, Functional Assay

Journal: PLoS ONE

Article Title: The Interaction between Circulating Complement Proteins and Cutaneous Microvascular Endothelial Cells in the Development of Childhood Henoch-Schönlein Purpura

doi: 10.1371/journal.pone.0120411

Figure Lengend Snippet: HMVEC-d were pre-incubated with plasma of patients (N = 30) with acute HSP, plasma of healthy controls (N = 30), or culture medium alone for 48 hr, and then the cells were harvested and analyzed for the expression of C3aR (A) and CD88 (B) by flow cytometry. The expression levels were presented as mean fluorescence intensity (MFI).

Article Snippet: Cells were then harvested by trypsin, washed by PBS, and labeled by PE-conjugated mouse IgG anti-human C3aR, IgG anti-human CD88, or IgG isotype control (BD Biosciences) at 4°C for 30 min. Stained cells were assayed by a FACSCalibur using the CellQuest Software (BD Biosciences).

Techniques: Incubation, Expressing, Flow Cytometry, Fluorescence

Journal:

Article Title: The immunosuppressive surface ligand CD200 augments the metastatic capacity of squamous cell carcinoma

doi: 10.1158/0008-5472.CAN-09-4380

Figure Lengend Snippet: CD200R1+ stromal cells are CD11b+/Gr-1+ MDSCs. A, CD200R1 immunofluorescence in WD SCC, LN Met and Lung Met (white arrows). DAPI counterstain was conducted to visualize nuclei. Dashed line demarcates SCC keratinocytes in LN Met. B, For CD200R1 FACS analysis of CD200R1, CD11b, and Gr-1 or MHC II expression. The total population of CD200R1+ cells were gated and subsequently analyzed for expression of CD11b and Gr-1 or MHC class II. The percentage of the total CD200R1 pool for a single experiment is shown. Far right panel: murine SCC were stained with antibodies against CD200R1 (red) and CD11b (green) and counterstained with DAPI (blue). Arrows point to CD200R1+/CD11b+ (yellow) stromal MDSCs. C, Left: Bar graph showing the densitometric units representing GM-CSF and G-CSF levels in pRS-NS Lung Met/CD200R1+ co-cultures versus pRS-CD200 Lung Met/CD200R1+ co-cultures. *- statistically significant difference (GM-CSF, p = 0.049; G-CSF, p = 0.002). Right: H&E staining (top left) and G-CSF (red) and CD200 (green) immunofluorescence in murine PD SCC. Nuclei were delineated with DAPI (blue). Scale bars mark 50µm.

Article Snippet: Antibodies Antibodies were used against human CD200, mouse CD200, CD3ε, CD86, α6 integrin-FITC (BD Biosciences); NK1.1-488, c-kit, CD123, MHC II, CD11b-FITC, CD11b-PE, Gr-1-FITC, Gr-1-PE, CD11c (BioLegend); Langerin, Foxp3 (eBioscience); Keratin 14 (Covance), CD200R1 (R&D Systems), CD4-APC (Abcam), G-CSF (Santa Cruz), anti-rabbit HRP (Jackson Immunoresearch); species-specific Alexa Fluor-488, -594, -647, and -680 (Invitrogen); and human α6 integrin (kindly provided by Fiona Watt).

Techniques: Immunofluorescence, Expressing, Staining