Fig. 3. Macrophages show transcriptional heterogeneity after ischemia and shift from a pro-inflammatory to a wound healing transcriptional profile.

a tSNE plot indicating transcriptomic similarities across macrophages obtained from all conditions. Different colors, symbols and numbers highlight the clusters as determined by k-medoids clustering of 1 −  Pearson’s correlation coefficient. Stars highlight a cell cluster formed by macrophages from all conditions; triangles highlight clusters containing mainly 1 dp IR macrophages and circles depict clusters containing mainly 3 dp IR macrophages. b Bar graph showing the proportion of macrophages originating from the different conditions per cluster. c Heatmap showing expression of all genes significantly enriched in at least one cluster. Examples of genes enriched per cluster are depicted on the right side of the heatmap. d Bubble plot of top GO terms in gene ontology analysis on genes significantly enriched within either cluster one, in at least one of the 1 dp IR enriched clusters or in at least one of the 3 dp IR clusters. e, f Cell trajectory analysis of macrophages, showing pseudotime on a color-coded scale (e) or highlighting the k-medoids cluster of each macrophage in the trajectory plot (f). The branches in the trajectory plot that contain mostly cells from cluster 1, from the 1 dp IR clusters or from the 3 dp IR clusters are highlighted by number or tekst. g RT-qPCR analysis on the infarcted cardiac tissue at different time points post IR for Arg1 and Gpnmb. Per time point post IR, expression levels are relative to the corresponding time point post sham surgery (n = 6 animals). Data are presented as mean ± SEM. *P < 0.05, **P < 0.01, Two-sample t-test vs corresponding time point post sham, with Holm–Sidak adjustment for multiple comparisons. n.d. not determined.