Figure 3: In Vivo CD47 Blockade Versus CD47 Induction Regulates CM Phagocytosis Efficiency and Cardiac Inflammation Resolution

(A) Flow cytometric measure of Mϕ engulfment of transgenic mCherry CMs. Gating strategy is shown gating CD11b myeloid cells that are mCherry+ and Ly6g low (non-neutrophil phagocytes or Mϕs). Mice were subjected to 45 minutes of ischemia followed by injection of isotype IgG control (ctrl) versus CD47 monoclonal antibodies in phosphate-buffered saline, just before unoccluding the left anterior descending coronary artery. Right image depicts sorted Mϕs with internalized CM mCherry. (B) Dot plots, histogram, and bar graph of flow cytometry analysis of CM mCherry-association with myocardial CD11b+ phagocytes after ischemia and reperfusion of transgenic Myh6-mCherrry mice. (C) Adeno-associated virus CD47 (myocyte Cre) was injected intravenously before surgery and fluorescent intensity (scale bar, 100 μm) is a marker of adeno-associated virus expression and quantified as a function of viral titer. To the right is cardiac phagocytosis enumerated as in (B and C). (D) Inflammation resolution kinetics at indicated day (D) time points. Flow cytometric analysis of (CD45-hi CD11b-hi Ly6g-lo) Ly6c and CD11c cells in the myocardium. (Left) Percentage of CD45-hi CD11b-hi Ly6g-hi neutrophils and polymorphonuclear neutrophils. (Right) (CD45-hi CD11b-hi Ly6g-lo) Ly6C-hi monocytes and Ly6c-lo Mo/Mϕs. *p ≤ 0.05 relative to ctrls. ns, not significant. Abbreviations as in Figures 1 and 2.



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