Abstract 3940: Perihematomal Leukocytes Peak 3 Days After Intracerebral Hemorrhage
Background: Intracerebral hemorrhage (ICH) is followed by a robust inflammatory response in the brain. Therapies inhibiting leukocyte migration to the brain have been developed, but their potential application to ICH relies on detailed understanding of the invading inflammatory cells after injury. Our objective was to quantify the various leukocyte populations between 1 and 7 days after ICH using a mouse model.
Methods: Using the autologous blood injection model in WT (C3H/HeOuJ) mice, we injected 15μ l of blood into the basal ganglia 2.5mm right of bregma. At 24h, 72h, and 7 days following surgery, we isolated the brain and dissociated each hemisphere into a single cell suspension using collagenase, dispase, and mechanical digestions prior to staining with CD45, CD3, CD11b, Ly6G, CD11c, Gr-1, and NK1.1 antibodies. Immune cell populations were quantified by flow cytometry (see figure for gating) and the numbers of excess ipsilateral cells were calculated. Additionally, at 72 hours after ICH or sham surgery, we isolated the ipsilateral hemisphere for quantification of chemokine production by ELISA.
Results: There was a significant increase in blood-derived leukocytes (CD45hi) between 24hr and 72hr, mostly accounted for by inflammatory monocytes and dendritic cells (Figure). We then saw a significant decrease between 72 hours and 7 days in the inflammatory monocyte population as well as a trend towards a decrease in the dendritic cell population. Natural killer cells accounted for less than 0.3% of all leukocytes detected. The monocyte-recruiting chemokine CCL2 (MCP-1) was increased in the brains at 72 hours after ICH (ICH 260.8 ± 31.1 pg/g brain vs sham 148.8 ± 12.4, p<0.05) consistent with the peak of inflammatory monocyte recruitment.
Conclusion: Leukocytes peak in the brain at 72 hours after ICH and are almost completely absent by 7 days. Inflammatory monocytes and dendritic cells account for the greatest numbers of leukocytes at 72 hours. CCL2 levels are elevated when inflammatory monocyte populations peak. Therapies inhibiting the CCL2-dependent migration of inflammatory monocytes across the blood-brain barrier should be investigated to determine if these cells contribute to injury.
- © 2012 by American Heart Association, Inc.