LPS challenge induces greater neutrophil infiltration and reduced iNOS expression in male mice compared with female mice. (A) Schematic of the experimental design. Male and female mice were injected IP with 5 mg/kg LPS, and peritoneal lavage cells were collected 1 day later. Control male and female mice were left untreated to assess baseline immune cell populations. Flow cytometry was used to analyze immune cell composition, as well as iNOS and arginase expression, in lavage cells from control and 1-day LPS-challenged male (n = 6) and female (n = 6) mice. (B) Total peritoneal lavage cell counts from control and 1-day LPS-treated male and female mice. (C) Gating strategy for flow cytometry analysis of peritoneal lavage cells, identifying distinct myeloid cell subsets: (A) CD11b^mod cells; (B) CD11b^hiLy6G^– cells (peritoneal macrophages); (C) CD11b⁺Ly6G⁺ cells (neutrophils). (D) Representative flow cytometry plots of CD11b^mod, CD11b^hiLy6G^– and CD11b⁺Ly6G⁺ cells in peritoneal lavage fluid from control and 1-day LPS-treated male and female mice. (E) Quantification of the frequency of CD11b^mod, CD11b^hiLy6G^– and CD11b⁺Ly6G⁺ cells in peritoneal lavage fluid from control and 1-day LPS-treated male and female mice (n = 6 per group). (F) Representative flow cytometry plots showing iNOS and arginase expression in CD11b^hiLy6G^– macrophages from peritoneal lavage fluid of control and 1-day LPS-treated male and female mice. (G) Quantification of the frequency of iNOS-positive, arginase-positive and arginase-positive/iNOS-positive populations within the CD11b^hiLy6G^– macrophages in peritoneal lavage fluid from control and 1-day LPS-treated male and female mice. Statistical significance was determined using 2-tailed t tests (Mann-Whitney U tests) for 2-group comparisons. P values < .05 were considered to indicate statistical significance. ∗P < .05; ∗∗P < .01; ∗∗∗∗P < .0001. ns, not significant.