Influence of TRAF2 on CD40-mediated cytokine mRNA expression in macrophages. Macrophages were obtained by exposure of human blood monocytes for 6 days to M-CSF (100 ng/mL) and then transfected with an empty vector (Co) or a TRAF2-expressing vector (TRAF2) or cIAP1 siRNA sequence 1 and 2 and treated with CD40L 24 hours later. (A-D) Real-time PCR analyses of mRNA expression of MCP-1 and IL-8 (A) in Co and TRAF2-transfected macrophages treated for 24 hours with 500 ng/mL CD40L (each line represent an independent sample). (B) Control (open symbols) and TRAF2-transfected (closed symbols) macrophages treated for the indicated times with 500 ng/mL CD40L. (C) Control and c-IAP1 targeting siRNAs macrophages treated for 24 hours with 500 ng/mL CD40L (mean ± SD of 3 independent experiments normalized to controls). (D) Macrophages treated by 500 ng/mL CD40L for 24 hours in the absence (Co) or presence of 10 μM SP600123 (SP) (mean ± SD of 3 independent experiments normalized to controls). (E) Immunoblot analysis of JNK and phosphorylated JNK (p-JNK) in control (Co) and TRAF2-overexpressing macrophages exposed to CD40L for the indicated times. One representative of 3 independent experiments is shown.