Extended Data Fig. 4: CD38+ resident macrophages accumulate in ageing adipose tissue.
From: Senescent cells promote tissue NAD+ decline during ageing via the activation of CD38+ macrophages
a, LC-MS quantification of NAD in eWAT from WT young male mice (6 month) n=7 mice/group, Cd38 KO young male mice (3 month) n=5 mice/group, WT old male mice (25 month) n=10 mice/group, and Cd38 KO old male mice (26 month) n=5 mice/group. NAD concentrations are shown as pmol/mg of tissue. (same WT data from Fig. 4a). b, mRNA levels of Il-1α and IL-10 in eWAT from 6 and 25 month-old WT male mice. (WT young male mice (6 month) n=7 mice/group, WT old male mice (25 month) n=9 mice/group) c, Western analysis of adipose tissue from young (3 Month) and old (19 month) WT male mice to detect PARP activity (PARylation) and DNA damage (γH2AX). Each lane represents one mouse (young n=4 mice/group, old n=4 mice/group). d, mRNA levels of Cd38 in visceral adipose tissue, the stromal vascular fraction, and adipocyte fraction from young (3 month) and old (19 month) WT male mice. (young n=4 mice/group, old n=4 mice/group). e, Flow cytometry gating strategy to identify CD45+ immune cells from the stromal vascular fraction of eWAT. Cells were first gated on forward scatter (FSCA) vs side scatter (SSCA) to discard cell debris and dead or dying cells. Next FSCH (height) vs FSCA (Area) was used to select single cells. Single cells were then gated for auto-fluorescent using the Empty(E) BV421 vs BV711 channels (not used as antibody fluorophores) to discard cells that showed auto-fluorescence in these channels. Then CD45+ cells were selected and analyzed for CD38 and macrophage markers. Flow cytometry gating strategy to identify resident and non-resident macrophages from the stromal vascular fraction of eWAT, showing representative flow plots and histograms for the indicated ages of mice. f, Flow cytometry quantification of CD38- (low) resident macrophages, CD38- non-resident macrophages, and CD38+ (high) non-macrophage immune cells from eWAT of WT male mice for the ages shown. (2 months n=6 mice/group, 6 months n=5 mice/group, 12 months n=5 mice/group, 18 months n=5 mice/group, 25+ months n=7 mice/group) For in vivo experiments, data from individual mice are shown. Statistical significance indicated as *P<0.05, **P<0.01, and ***P<0.001; two-sided Student’s t-test.
