FIGURE 6.

Ac₂PIM regulates NOD2-β-catenin-mediated COX-2, SOCS-3, and MMP-9 expression. A, peritoneal macrophages were pretreated with a pharmacological inhibitor of JAK kinase (AG490) followed by treatment with MDP or IFN-γ (200 units/ml) to analyze phosphorylation status of STAT1 and STAT3. B and C, inhibitory phosphorylation status of β-catenin and GSK-3β during the following conditions: treatment of macrophages with MDP for the indicated time points (B), pretreatment of macrophages with PI3K-MAPK pathway-specific pharmacological inhibitors followed by MDP treatment for 6 h (C). D, peritoneal macrophages were treated with AG490, β-catenin inhibitor or LiCl (GSK-3β inhibitor) prior to 12 h MDP treatment. Lysates were assessed for COX-2, SOCS-3, and MMP-9 by immunoblotting. E and F, Phosphorylation status of β-catenin and GSK-3β was assessed by immunoblotting under following conditions: Peritoneal macrophages were pretreated with the indicated inhibitors for 1 h followed by Ac₂PIM and MDP treatment for 6 h (E), miR-150- (left panel) or miR-143- (right panel) specific miRNA inhibitor-transfected RAW 264.7 macrophages were treated with Ac₂PIM prior to MDP treatment for 6 h (F). All blots are representative of three independent experiments. The cells were treated with 2 μg/ml Ac₂PIM for 2 h followed by 200 ng/ml MDP. Med, medium; DMSO, dimethyl sulfoxide; NC, negative control; Inhi., inhibitor; LiCl, lithium chloride.