Figure 5

IL-17A induces the production of pro-inflammatory chemokines and MMPs by triggering distinct signaling pathways. (A) Detection of signaling proteins following SDS-PAGE and immunoblotting in lysates of dermal fibroblasts from one representative HD of three cultured in the presence of IL-17A for the indicated time-points. (B) Densitometric analysis of the signaling proteins normalized to β-tubulin levels. Data are expressed as the ratio of the normalized values relative to time = 0 minutes. One representative experiment of three is shown. (C) Protein levels were assessed in 48-hour culture supernatants by ELISA. Bars represent the mean ± SEM from six distinct HD performed in triplicate relative to the control condition (vehicle). IL-17A (30 ng/ml) was added in all conditions. When used, SP-600125 (JNK inhibitor, 10 μM), SB203580 (p38 inhibitor, 20 μM), U0126 (MEK1/2 inhibitor, 20 μM), LY294002 (PI3K inhibitor, 10 μM) and TPCK (NF-κB inhibitor, 5 μM) were added one hour before IL-17A. Significant differences versus vehicle were assessed by Student’s t-test: * = P <0.05, ** = P <0.01, *** = P <0.001. Light gray, black and dark gray bars depict pathways involved in MMP-1 signaling, MCP-1/IL-8 signaling and common pathways, respectively.

ELISA: enzyme-linked immunosorbent assay; HD: healthy donors; IL: interleukin; JNK: c-jun N-terminal kinase; MCP: monocyte chemotactic protein; MEK: mitogen-activated protein kinase; MMP: matrix metalloproteinase; NF-kB: nuclear factor kappa B; PI3K: phosphoinositol 3 kinase; SDS-PAGE: Sodium Dodecyl Sulphate - PolyAcrylamide Gel Electrophoresis; SEM: standard error of the mean.