Journal: International Journal of Molecular Sciences
Article Title: CD200 Modulates S. aureus-Induced Innate Immune Responses Through Suppressing p38 Signaling
Figure Lengend Snippet: CD200 restrains p38 signaling upon Staphylococcal infection. ( A – L ) Mouse PEMs were transfected with CD200 siRNA (siCD200) or control siRNA (NC) for 48 h, and then stimulated with S. aureus (MOI = 1) for indicated time periods. The protein expression levels of p-p38, p38, p-ERK1/2, ERK1/2, p-JNK, JNK, p-TAK1, TAK1, p-MMK3/6, MMK3/6, p-CREB, CREB, p-MSK1, MSK, p-c-Fos, c-Fos, p-c-Jun, and c-Jun were detected by western blotting. ( M – Q ) Mouse BMDMs were pre-treated with CD200-Fc (2 μg/mL) or IgG1-Fc (2 μg/mL) for 1 h, and then stimulated with S. aureus (MOI = 1) for indicated time periods. The protein expression levels of p-p38, p38, p-c-Fos, c-Fos, p-c-Jun, and c-Jun were detected by western blotting. ( R ) RAW264.7 macrophages were pre-transfected with CD200 siRNA for 48 h and then challenged with S. aureus (MOI = 10) for 2 h. The AP-1 luciferase activity was analyzed by the dual luciferase. Representative blots from three independent experiments are shown ( A , E , J , M , and O ). Changes in the phosphorylation state of different proteins were semi-quantitated using ImageJ software ( B – D , F – I , K – L , N , P – Q ). ND means not detected; * p
Article Snippet: Transfection of siRNA The siRNA molecules targeting CD200 mRNA were purchased from GenePharma Corporation (Shanghai, China), and delivered into macrophages using X-tremeGENE siRNA transfection reagent (Roche, Basel, Switzerland), according to the manufacturer’s instructions.
Techniques: Infection, Transfection, Expressing, Western Blot, Luciferase, Activity Assay, Software