Figure 6.

Gating strategy for flow cytometry and quantification of immune cells from epididymal SVF cells. Total SVF cells were first gated on a forward scatter/side scatter plot and then gated for the selection of singlets. Singlets were further gated for live cells followed by CD45 gating for total immune cells. These were then gated for myeloid cells (CD45+/CD11b+) and macrophages (CD45+/Cd11b+/Ly6C+). Fluorescence Minus One (FMO) controls were used for each antibody marker to set the limit for the background signal of the excluded marker and to gate for a positive stained cell population. (A), (B) Representative gating strategy for GR Flox and Adipo GRKO mice. (C) Quantitative analysis of SVF cells prior to cell staining, immune cells, myeloid cells, and macrophages. Epididymal adipose tissues were pooled from 2 mice per genotype for each experiment. n = 4 independent experiments. All values represent mean ± SD. Data were analyzed by Student's t-test. * P < .05.

Abbreviations: Adipo GRKO, adipocyte-specific GR knockout; GR, glucocorticoid receptors; GR Flox, C57BL/6J GR flox/flox; SVF, stromal vascular fraction.