Recruited lung mononuclear phagocytes are a distinct cell population with a broad spectrum of maturity and immune activation in cystic fibrosis airways. (A) Potential of heat diffusion for affinity-based transition embedding (PHATE) of monocytes and monocyte-derived macrophages colored by pseudotime, all starting from quiescent monocytes toward 1) activated monocytes, 2) mature monocyte-derived macrophages, 3) monocytes expressing a heat-shock response, and 4) monocytes and monocyte-derived macrophages, colored by disease state. All three archetypes are accompanied by three PHATE plots colored by the gene expression of typical genes, ramping up along a specific pseudotime. For the corresponding uniform manifold approximation and projection embedding colored by the gene expressions of the same genes, see Figure E4. For the corresponding PHATE colored by cell type and subjects, see Figure E5. (B) Heatmap of gene expression and regulon activity in monocytes undergoing activation (ordered by pseudotime distances along PHATE manifolds that transition from quiescent monocytes toward an activated monocyte archetype). (C) Heatmap of gene expression and regulon activity in monocytes undergoing maturation (ordered by pseudotime distances along PHATE manifolds that transition from quiescent monocytes toward a control-enriched mature monocyte-derived macrophage archetype). In both heatmaps, annotation bars represent the pseudotime distance, disease status, and subject for each cell; expression values are centered and scaled. (D) Violin plots of pathway activity scores grouped by cell type and separated by disease state. Depicted pathway scores from left to right are as follows: GO:0045087 innate immune response, GO:0006958 complement activation and classical pathway, GO:0019882 antigen processing and presentation, and GO:0006911 phagocytosis and engulfment. *False discovery rate–adjusted P values < 0.05 calculated using the Wilcoxon signed-rank test. alvMΦ = alveolar macrophage; CF = cystic fibrosis; Gene Expr. = gene expression; HC = healthy control; Mo = monocyte; MoMΦ = monocyte-derived macrophage; PMN = polymorphonuclear neutrophil; Regulon Act. = regulon activity.