Figure 3.

Rora^fl/flLysM^Cre/+ mice show decreased Ly6C^hi ATM and fewer CD9⁺ inflammatory ATM in obese animals. Groups of Rora^fl/fl and Rora^fl/flLysM^Cre/+ mice were fed a control diet (CD; n = 5 Rora^fl/fl, n = 3 Rora^fl/flLysM^Cre/+) or a high-fat diet (HFD; n = 7 Rora^fl/fl, n = 6 Rora^fl/flLysM^Cre/+) for 16 weeks and the E-WAT and I-WAT isolated and prepared for flow cytometry. Cells isolated from the E-WAT were gated as Live-dead⁻CD45⁺SiglecF⁻ and assessed for Ly6C expression (A). Ly6Clo cells were further assessed for CD64 and CD9 expression (B). This gating strategy was repeated in cells isolated from I-WAT (C,D, n = 2 Rora^fl/fl CD, n = 1 Rora^fl/flLysM^Cre/+ CD; n = 7 Rora^fl/fl HFD, n = 6 Rora^fl/flLysM^Cre/+ HFD). Rora^Cre and Rora^CreRosa-YFP mice were fed a high-fat diet for 12 weeks and flow cytometry performed on E-WAT. Cells were gated as Live-dead⁻CD45⁺SiglecF⁻CD11b⁺ then gated as described previously based on Ly6C expression, then CD9 expression, as indicated, and assessed for Rora-YFP expression (E, data is representative of 3 Rora^Cre and 3 Rora^CreRosa-YFP mice). (F) Cells isolated from the E-WAT from HFD fed Rora^fl/fl and Rora^fl/flLysM^Cre/+ mice were stained as CAM (CD11b⁺SiglecF⁻F4/80⁺CD206^lo) or AAM (CD11b⁺SiglecF⁻F4/80^hiCD206^hi; n = 6–7). Data is representative of mean ± SEM. Student's t-test: ns, not significant, *P < 0.05, **P < 0.01.