Tissue-specific murine neutrophil activation states in health and inflammation

Neutrophils are quickly recruited to tissues in response to proinflammatory cues; however, little is known about tissue neutrophil phenotypes in health. We employ a multicolor flow cytometric approach to assess surface markers of activation on neutrophils from the bone marrow, blood, peritoneum, spleen, liver, fat, colon, and oral cavity of healthy mice. Cell preparations were promptly fixed to preserve native surface marker expression levels. Peritoneal, colonic, and oral neutrophils were also assessed in the setting of pHrodo-induced peritonitis, dextran sodium sulfate-induced colitis, and ligature-induced periodontal disease, respectively. Our results demonstrate consistent detectable neutrophil populations in various sterile and nonsterile tissues of healthy mice, and these cells had tissue-specific neutrophil immunophenotypes. Neutrophils derived from biofilm-associated mucosal tissues had 2- to 3-fold higher expression of surface markers of activation, including CD66a, CD11b, and CD62L, compared to neutrophils derived from both sterile healthy tissues as well as tissues in animals treated with broad-spectrum antibiotics. Furthermore, the unique cluster of differentiation (CD) marker activation signatures of tissue-specific neutrophils from the peritoneum, colon, and oral cavity were altered to a proinflammatory immunophenotype with the presence of an inflammatory stimulus. Based on our results, we propose a model whereby a hierarchy of tissue neutrophil immunophenotypes, based on the differential expression of CD markers of activation, correlates with sterile, healthy commensal biofilm-associated and inflamed tissue states.