Fig. 3.
a Effects of KRO-105714 on chemokine levels in stimulator-induced RAW 264.7 cells. The RAW 264.7 cells were seeded in 96-well plates at a density of 2 × 104 cells/well and pre-treated with various concentrations (1, 3, 10 μM) of KRO-105714 for 30 min and then treated with 10 μM of SPC for 24 h. Expression levels of MCP-1 were measured using ELISA. Each dose of the test compound was assayed in triplicate and the assays were repeated three times. Data are shown as mean ± standard deviation (SD). *P < 0.05, ** P < 0.01 and *** P < 0.001 is the significance level compared to the SPC treated group. b-c Inhibitory effects of KRO-105714 on the expression of pro-inflammatory cytokines and adhesion molecule. The cytokines used were PBMCs isolated from the blood of healthy individual donors. The human PBMCs were seeded in 96-well plates at a density of 5 × 105 cells/well and pre-treated with concentrations (1 and 10 μM) of KRO-105714 for 30 min and then treated with 10 μM of SPC and 5 μg/ml of PHA for 24 h (b) of 72 h (c). IL-4 (b), and IL-5 (c) levels were measured using ELISA. Each dose of the test compound was assayed in triplicate. ICAM-1 expression level of PBMCs (d) was measured using flow cytometry. MFI means “mean fluorescence intensity”. Data are shown as mean ± standard deviation (SD). *P < 0.05, ** P < 0.01 and *** P < 0.001 versus compound-untreated cells (ANOVA multiple comparison)