Fig 1: Single-cell scRNA-seq reveals that subsets of lung IMs have unique iCCR expression patterns and that iCCR expression is not random. (A) UMAP visualization of CD11b+ Ly6G− SiglecF− lung cells. (B) UMAP plot of data subsetted from panel A depicting cells positive for monocyte or macrophage markers. Nonclassical monocytes were also removed from the dataset. Colors represent unique clusters termed “classical monocytes,” “Retnla+ macrophages,” “MHCII hi macrophages,” and “MHCII lo macrophages.” (C) (i) UMAP displaying hashtagged populations and how they correspond to the monocyte and macrophage clusters presented in panel B. Blue = CCR1− CCR5−; red = CCR1+ CCR5+; green = CCR1− CCR5+; purple = CCR1+ CCR5−. UMAP plots in panels B and C have the same coordinates. (ii) Bar graph presenting the proportion of hashtagged cells corresponding to each macrophage population. (D) (i) Violin plot showing the relative expression of Ccr2 by hashtagged populations across the 4 clusters presented in panel B. (ii) Bar graph presenting combinatorial iCCR expression across the 3 macrophage populations presented in panel B. Cells were considered positive for Ccr2 if relative expression was >1. Feature plots depicting the expression of Cd2 and Cd226 within the scRNA-seq dataset. UMAP coordinates are identical to the plot shown in panel B. (i) Antibodies against CD2 and CD226 were used to identify Retnla+ macrophages by flow cytometry and assess MHCII and iCCR expression in the CD2+ CD226+ population. DC, dendritic cell; NK, natural killer.