Fig 1: MK2/3 are required for neutrophil recruitment following intraperitoneal injection of IL‐33. (a) Wild type, MK2/3 or MSK1/2 were injected with either PBS, 5 μg kg−1 IL‐33 or 2 mg kg−1 LPS. After 3 h, mice were sacrificed and the peritoneum washed to obtain cells. The % of neutrophils (CD11b+/Gr1+) in the peritoneal wash was determined by staining for CD11b and Gr1 and analyzed by flow cytometry. Graphs show average and standard deviation with crosses representing data for an individual mouse. Two‐way ANOVA testing between wild‐type and MK2/3 knockout mice indicated a significant interaction between genotype and treatment (F = 19.34, P < 0.001). For post hoc analysis on the effect of genotype within treatment groups, a P < 0.01 is indicated ** and I < 0.001 by ***. The difference between MSK1/2 knockout and wild‐type animals treated with IL‐33 was not significant (P < 0.05, Student's t‐test). (b) Transwell assays were used to determine the migration of wild‐type neutrophils in response to wild‐type or MK2/3 knockout peritoneal mast cells (4 biological replicates per genotype) that were either unstimulated or treated with IL‐33, as described in the methods. Blank indicated wells with neutrophils but no mast cells. Two‐way ANOVA testing between wild‐type and MK2/3 knockout mast cells indicated a significant interaction between genotype and treatment (F = 12.404, P < 0.004). For post hoc analysis on the effect of genotype within treatment groups, a P < 0.01 is indicated by ** and P < 0.001 by ***. (c) Peritoneal mast cells were stimulated with 10 ng mL−1 IL‐33 for the indicated times and the levels of CCL3, CCL4 and CXCL2 in the media determined. Data show mean and standard deviation of four biological replicates per genotype. Two‐way ANOVA testing between wild‐type and MK2/3 knockout mast cells indicated a significant effect of genotype (F = 160.7, P < 0.001 for CCL3; F = 5.26, P = 0.034 for CCL2 and F = 151.7, P < 0.001 for CXCL2). For post hoc analysis on the effect of genotype at specific times, a P < 0.05 in indicated by *, <0.01 by ** and <0.001 by ***.