Figure 2.
Figure 2. eWAT monocytes (eMon) rapidly adapt to the adipose tissue environment. (A) Comparative analysis of eWAT monocyte gene expression compared with blood Ly6C+ and Ly6C− monocytes. eMon RNAseq data were compared with publically available RNAseq data for blood Ly6C+ and Ly6C− monocytes (Mildner et al., 2017). Each column represents consolidated data from at least three biological replicates per cell type. The z-score of the gene expression profiles gives a scale to measure the differential expression. (B) PCA of the transcriptional signature obtained by RNAseq data comparing eWAT monocytes to blood Ly6C+ and Ly6C− monocytes depicted in A. (C and D) Volcano plots depicting genes differentially expressed by eWAT monocytes in relation to blood Ly6C+ (C) and Ly6C− (D) monocytes. Genes differentially expressed with high expression difference and/or high adjusted P values are labeled in red. Consolidated data from at least three biological replicates. (E) Heat map displaying selected genes that are differentially expressed between eWAT monocytes and Ly6C+ and Ly6C− blood monocytes. Consolidated data from at least three biological replicates. (F) Turnover kinetics of Ly6C+ monocytes. WT animals were injected once i.p. with 1 mg of BrdU, and the decay of BrdU+ labeling was analyzed 12, 24, 48, 72, and 96 h after the BrdU injection in the indicated organs (n = 3 per group, representative of two independent experiments). (G) Fate mapping of monocytes and VAM populations. Tamoxifen was administered to Cx3cr1Cre-ERT2-YFP × Rosa26LSL-DsRed mice twice 24 h apart by gavage, before starting the time course. Animals were analyzed 1, 5, 10, 20, and 40 d after the last injection of tamoxifen. Microglia are shown as a well-established internal positive control for DsRed labeling (Parkhurst et al., 2013; n = 3 per group, representative of two independent experiments). (H) Most myeloid eWAT subpopulations are derived from blood progenitors. CD45 congenic animals were precisely irradiated in the hind limbs using an X-RAD 320 (PXI) irradiator, preserving the rest of their bodies from irradiation. Congenic bone marrow (BM) was transferred i.v., and 8 wk later cell chimerism was assessed in the adipose tissue (indicated as percentage of CD45.2 cells among the total population); n = 4 per group, representative of two independent experiments. Graphs are shown as mean ± SEM.

eWAT monocytes (eMon) rapidly adapt to the adipose tissue environment. (A) Comparative analysis of eWAT monocyte gene expression compared with blood Ly6C+ and Ly6C− monocytes. eMon RNAseq data were compared with publically available RNAseq data for blood Ly6C+ and Ly6C− monocytes (Mildner et al., 2017). Each column represents consolidated data from at least three biological replicates per cell type. The z-score of the gene expression profiles gives a scale to measure the differential expression. (B) PCA of the transcriptional signature obtained by RNAseq data comparing eWAT monocytes to blood Ly6C+ and Ly6C− monocytes depicted in A. (C and D) Volcano plots depicting genes differentially expressed by eWAT monocytes in relation to blood Ly6C+ (C) and Ly6C− (D) monocytes. Genes differentially expressed with high expression difference and/or high adjusted P values are labeled in red. Consolidated data from at least three biological replicates. (E) Heat map displaying selected genes that are differentially expressed between eWAT monocytes and Ly6C+ and Ly6C− blood monocytes. Consolidated data from at least three biological replicates. (F) Turnover kinetics of Ly6C+ monocytes. WT animals were injected once i.p. with 1 mg of BrdU, and the decay of BrdU+ labeling was analyzed 12, 24, 48, 72, and 96 h after the BrdU injection in the indicated organs (n = 3 per group, representative of two independent experiments). (G) Fate mapping of monocytes and VAM populations. Tamoxifen was administered to Cx3cr1Cre-ERT2-YFP × Rosa26LSL-DsRed mice twice 24 h apart by gavage, before starting the time course. Animals were analyzed 1, 5, 10, 20, and 40 d after the last injection of tamoxifen. Microglia are shown as a well-established internal positive control for DsRed labeling (Parkhurst et al., 2013; n = 3 per group, representative of two independent experiments). (H) Most myeloid eWAT subpopulations are derived from blood progenitors. CD45 congenic animals were precisely irradiated in the hind limbs using an X-RAD 320 (PXI) irradiator, preserving the rest of their bodies from irradiation. Congenic bone marrow (BM) was transferred i.v., and 8 wk later cell chimerism was assessed in the adipose tissue (indicated as percentage of CD45.2 cells among the total population); n = 4 per group, representative of two independent experiments. Graphs are shown as mean ± SEM.

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