Single Cell RNA Sequencing Analysis of WT and CD169-DTR Adipose Tissue Vascular Fraction

Using t-distributed stochastic neighbor embedding (t-SNE) analysis, we identified 23 transcriptionally distinct cell clusters which are present in the eWAT when the four experimental groups were combined (ND and HFD with AMTs or without).Our analysis identifies six main cell clusters of CD45- cells: two adipocyte stem cell fractions (ASC1 and ASC2), stromal cells, two fractions of vascular endothelial cells (VEC1 and VEC2) as well as a population showing a gene signature for of mesothelial cells, among the CD45+ cells, seventeen lymphoid and myeloid cell subpopulations are detectable in the eWAT ranging from monocytes, monocyte-derived macrophages, dendritic cells, eosinophils, mast cells, B cells, CD4+ Th2, CD4+ Th17, Tregs and CD8+ T cells, NK cells, NKT cells In obese mice, one week absence of ATM in the eWAT affected mostly the non-haematopoetic cell compartment, while the total number of cell clusters and percentage of haematopoetic CD45+ cells were maintained roughtly the same between the four experimental groups, whereas ablation of ATMs in lean mice did not perturb dramatically the cluster distribution of CD45- cells. To investigate the effects of ablation tissue resident macrophages on immune and non-immune cells in adipose tissue under both lean and obese conditions, WT and CD169-DTR mice were fed with Normal Diet or High Fat Diet for 16 weeks, followed by Diphtheria Toxin (DT) treatment for 7 days to ablate tissue resident macrophages. Epidimal adipose tissue were processed to get the stromal vascular fraction cells. CD45+ and CD45- cells (exclude the high proportion of F4/80hi cells as well as Ly6Chi monocytes) were sorted separately and combined with a raton of 1:1 for sequencing analysis.