Differential activation of infiltrating monocyte-derived cells after mild and severe traumatic brain injury

Microglia are the resident innate immune cells of the brain. Although embryologically and functionally distinct, they are morphologically similar to peripheral monocyteYderived cells, resulting in a poor ability to discriminate between the two cell types. The purpose of this study was to develop a rapid and reliable method to simultaneously characterize, quantify, and discriminate between whole populations of myeloid cells from the brain in a murine model of traumatic brain injury. Male C57BL/6 mice underwent traumatic brain injury (n = 16) or sham injury (n = 14). Brains were harvested at 24 h after injury. Multiparameter flow cytometry and sequential gating analysis were performed, allowing for discrimination between microglia and infiltrating leukocytes as well as for the characterization and quantification of individual subtypes within the infiltrating population. The proportion of infiltrating leukocytes within the brain increased with the severity of injury, and the predominant cell types within the infiltrating population were monocyte derived (P = 0.01). In addition, the severity of injury altered the overall makeup of the infiltrating monocyte-derived cells. In conclusion, we describe a flow cytometryYbased technique for gross discrimination between infiltrating leukocytes and microglia as well as the ability to simultaneously characterize and quantify individual myeloid subtypes and their maturation states within these populations.