Characterization of Transverse Aortic Constriction in Mice Based on the Specific Recruitment of Leukocytes to the Hypertrophic Myocardium and the Aorta Ascendens

Figure 1

Methodological procedure and definition of the immune cell subsets. (a) We sacrificed the mice 3, 6, and 21 days after TAC or sham surgery. (b) Heart-weight/body-weight-index in mg/g was calculated 3, 6, and 21 days after TAC or sham surgery. (c) In addition to the LV tissue, we analyzed the tissue of the aorta ascendens, aorta descendens, spleen, blood, and heart draining lymph nodes. (d) The gating strategy for flow cytometry analyses was performed as illustrated for the tissue of the aorta ascendens. All cell-like events (a) were isolated by using the forward scatter area versus sideward scatter area. Doublets were excluded using forward scatter area versus sideward scatter width (b). We defined the living immune cells (c), using CD45 staining versus Hoechst staining. The CD45⁺ immune cells were further classified by staining for the surface markers Ly6G and F4/80 to determine neutrophils (d) and macrophages (e). Next, we checked for the Ly6C surface expression of the macrophage population as a function of CD11b to differentiate between the Ly6C^high (f) and the Ly6C^low macrophages (g). We analyzed the DC compartment based on the population of the CD45⁺ cells (c) and stained for MHC-II to identify MHC-II⁺ immune cells (h). We used CD11c as a function of the autofluorescence channel AF430 for the definition of classical dendritic cells (i). The cDCs were further discriminated according to the CD103 surface expression (j).

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