Figure 4.

Proteomic membrane analysis of synovial fibroblast supernatants. (a) Comprehensive protein membrane array map. The human protein array simultaneously profiles 30 proteins in duplicate, a set of six positive controls and four negative controls. EGF = epidermal growth factor; GCSF = granulocyte colony stimulating factor; Gro = growth-related oncogene; HGF = hepatocyte growth factor; ICAM-1 = intercellular cell adhesion molecule-1; IFN-γ = interferon-γ; IGF-1 = insulin-like growth factor-1; IGFBP-3, 4 = insulin-like growth factor binding protein-3, 4; IL-1α, 1β, 6, 8 = interleukin-1α, 1β, 6, 8; IL-1ra = interleukin-1 receptor antagonist; MCP-1 = monocyte chemoattractant protein-1; MIP-1α, 1β, 3α = macrophage inflammatory protein-1α, 1β, 3α; MMP-1, 3, 9, 13 = matrix metalloproteinase-1, 3, 9, 13; Neg = negative control; pos = positive control; RANTES = regulated on activation, normal T cell expressed and secreted; SDF-1 = stromal cell derived factor-1; TGF-β3 = transforming growth factor-β3; TNF-α = tumor necrosis factor-α; VCAM-1 = vascular cell adhesion molecule-1; VEGF = vascular endothelial growth factor. The sensitivity of antibodies of the RayBio™ human array for the respective proteins differs; proteins in italics: high sensitivity (1–25 pg/ml), in bold: medium sensitivity (100 – 300 pg/ml), underlined: low sensitivity (1,000 – 10,000 pg/ml), Gro determines Groα (low sensitivity) and Groβ (low sensitivity) and Groγ (high sensitivity). (b) Supernatants of a synovial fibroblast cell line derived from a patient with rheumatoid arthritis (RASF) and a synovial fibroblast cell line derived from a normal donor (NDSF) were examined for cytokine secretion with the use of antibody-based protein arrays. Because cultivation was performed in medium supplemented with serum, the protein content of the cultivation medium was analyzed as a control. Results are shown after exposure of the array membranes to X-ray films for 2 minutes. The cytokines/chemokines IL-6, CXCL8 (IL-8), monocyte chemoattractant protein-1 (MCP-1), and growth-related oncogene (Gro) showed increased secretion from RASF compared with NDSF and with serum control.