Defective neutrophil recruitment in CCRL2-deficient mice. (A) Cytofluorimetric profiles of CXCR2 and CCRL2 expression in purified BM neutrophils from WT and CCRL2-deficient mice (CCRL2 KO) stimulated with tumor necrosis factor α (TNF-α; 20 ng/mL), LPS (100 ng/mL), a combination of TNF-α, LPS, and 50 ng/mL interferon gamma (MIX), or medium for 18 hours. Cells were stained with a rat anti-mouse CCRL2 moAb followed by an anti-rat phycoerythrin (PE) moAb and with a rat anti-mouse CXCR2-Alexa Fluor 647 moAb. Representative plots from 3 independent experiments are shown in the left panel; right panels show summarized results of single CXCR2 and CCRL2 staining. (B-C) Peritoneal recruited cells from WT and CCRL2 KO mice injected intraperitoneally with (B) LPS (15 ng per mouse) for 2 hours or (C) CXCL8 (300 ng per mouse) for 4 hours. Control mice received sterile phosphate-buffered saline (saline). The number of CD11b⁺Ly6G⁺ neutrophils per mouse was evaluated by fluorescence-activated cell sorter analysis. The results are expressed as mean ± standard error of the mean (SEM) of 3 independent experiments for a total of 10 mice per group. (B) The rightmost graph shows mean fluorescent intensity (MFI) values of CCRL2 expression by neutrophils collected 2 hours after LPS or saline injection. (D) Cells recruited from the synovial cavity at the indicated time points after the injection of methylated bovine serum albumin (metBSA) or saline into the knee joints of metBSA-immunized mice. Results are expressed as mean ± SEM of 3 independent experiments (n = 14). *P < .05; **P < .01 by Student t test. ns, not significant.