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Fig. 3 | Arthritis Research & Therapy

Fig. 3

From: Inflammatory properties of inhibitor of DNA binding 1 secreted by synovial fibroblasts in rheumatoid arthritis

Fig. 3

RA ST fibroblasts secrete Id1 and upregulate Id1 expression upon stimulation with TGFβ. a EPCs, HMVECs, monocytes, NL, RA, and OA fibroblasts were plated and serum starved. Cell culture media was replaced and the supernatants were collected 24 h later. The cell culture supernatants were examined for Id1 expression by ELISA (MyBioSource). EPCs, HMVECs as well as all fibroblasts were collected from human tissues, which were digested in a mix of cell culture media supplemented with FBS, collagenase, and hyaluronidase. HMVECs were isolated from skin biopsies while the fibroblasts were isolated from STs of patients with either RA, OA, or from NL patients. HMVECs were isolated and purified using CD31 microbeads (Miltenyi Biotec). All cell lines were between passages 3 and 6. No cytokine stimulation was used. b NL fibroblasts were plated and serum starved under the same conditions as panel a. Cell culture media with the respective cytokine was exchanged and collected 24 h later. CXCL16 (R&D Systems), IL-17 (R&D Systems), TNF-α (Life Technologies) and TGF-β (R&D Systems) were used at 10 ng/mL and 50 ng/mL concentrations. A not stimulated (NS) well was also used with no added cytokine (n = number of experimental replicates). c Synovial fibroblasts were plated under the same conditions as panel a. Cell culture media with TGF-β was exchanged and collected 24 h later. CXCL16 chemokine (C-X-C motif) ligand 16, EPC endothelial progenitor cell, HMVEC human dermal microvascular endothelial cell, Id1 inhibitor of DNA binding 1, IL-17 interleukin 17, NL normal, NS nonstimulated, OA osteoarthritis, RA rheumatoid arthritis, TGF-β transforming growth factor beta, TNF-α tumor necrosis factor alpha

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