Fig. 8: Defective efferocytosis by peritoneal macrophages from Axl^−/− infected mice.

PECs from infected Axl^−/− (open triangles) and infected WT (closed triangles) mice were treated with CFSE-labelled or CFSE-negative T cells or thymocytes for 1 h. a, b Adherent PECs were then analysed by flow cytometry for the presence of a intracellular or b extracellular CFSE⁺ cells. a, c PECs treated with CFSE-labelled T cells were washed and then stained with anti-CD11b and anti-TCRβ mAbs. For evaluation of intracellular CFSE⁺ cells, doublets and TCR⁺ cells were gated out and excluded from analysis. To define CD11b⁺CFSE⁺ macrophages, gates were based on macrophages treated with CFSE-negative T cells, employed as negative controls. c Graphs show the percentages of intracellular T cells in PECs from infected WT (n = 4) and Axl^−/− (n = 5) mice. b, d For analysis of tethering, a more homogenous population of CFSE-labelled apoptotic thymocytes was used in the assays. PECS were washed and stained with anti-F4/80 and anti-CD8. Then, gated F4/80⁺ cells were analysed for extracellular CD8⁺CFSE^hi thymocytes. Gates were based on macrophages treated with CFSE-negative thymocytes. d Graphs depict the percentages of PECs from infected WT (n = 5) and Axl^−/− (n = 4) mice bound to extracellular (CD8⁺) thymocytes. e F4/80⁺ macrophages containing intracellular thymocytes. Selection of singlets followed by gating of F4/80⁺ macrophages was used to identify macrophages that internalized CFSE⁺ cells. Each symbol represents the average of duplicates/triplicate determinations for each mouse. Dotted lines stand for WT PECs treated with CFSE-negative cells. Means and SEM are represented for each mouse group. Significant differences, as analysed by unpaired Student’s t tests, are indicated as for ^*P < 0.05, ^**P < 0.01, and ^***P < 0.001.