Isolation of primary peritoneal Mϕs

Peritoneal Mϕs were elicited by intraperitoneal (i.p.) injection of 4% thioglycollate (Sigma-Aldrich, T9032). Five d postinjection, Mϕs were harvested via peritoneal lavage using 10 mL of sterile 1X Dulbecco's phosphate-buffered saline (Life Technologies, 14190-144), centrifuged, resuspended, and cultured in RPMI-1640 (Life Technologies, 22400154) supplemented with 10% fetal bovine serum, 1% penicillin/streptomycin and 1% GlutaMAX™ (Life Technologies, 35050061) overnight to allow adherence. Mϕs were then washed twice with RPMI-1640 to remove nonadherent cells.

For macrophage polarization studies, cells were treated with 100 ng/ml LPS (Sigma-Aldrich, L9641) for 6 h.