Figure 4.
RvD2 in vivo actions were enhanced by overexpression and reduced by knockdown of GPR18. (A–C) Naive peritoneal MΦ were collected and transfected ex vivo (106 cells) with either GPR18 (5 µg) or mock plasmids for 48 h. Zymosan (1 mg) was injected into peritoneum to initiate peritonitis. 12 h later, transfected MΦ (1.5 × 105/mouse) and/or RvD2 (10 ng) was injected i.p. (A) Timeline. (B) PMN numbers (Ly6G+ CD11b+) and (C) efferocytosis (Ly6G+ F4/80+) were determined using flow cytometry. Results are expressed as mean ± SEM from 2 independent experiments and 6 mice/group. *, P < 0.05; **, P < 0.01; ***, P < 0.001, versus zymosan (zym) alone. #, P < 0.05, ##, P < 0.01, versus zym+veh+RvD2 (one-way ANOVA with Dunnett’s multiple comparison test). §, P < 0.05, versus zym+MΦ-mock+RvD2 (unpaired Student’s t test). (D–F) Naive peritoneal MΦ (106 cells) were transfected ex vivo with either GPR18 shRNA (5 µg) or control-scrambled shRNA. Zymosan (1 mg) was injected to initiate peritonitis. 12 h later, transfected MΦ (2 × 105/mouse) and/or RvD2 (20 ng) was injected i.p. Inflammatory exudates were collected at 24 h. (D) Timeline. (E) PMN numbers (Ly6G+CD11b+) and (F) efferocytosis (Ly6G+F4/80+) were determined using flow cytometry. Results are expressed as mean ± SEM from 2 independent experiments and 6 mice/group. *, P < 0.05; **, P < 0.01, obtained with unpaired Student’s t test for vehicle versus RvD2 in MΦ + control shRNA group.