FACS Analysis of Immune Cells from Adipose Tissue

To test whether TCDD would stimulate infiltration of immune cells into adipose tissue, we isolated adipose tissue cells from epididymal adipose tissue of male wt and AhRR Tg mice and assessed their phenotype by flow cytometry. Fluorescence-activated cell sorting (FACS) analyses indicated a rapid increase of F4/80–CD11b+ cell subset 1 day post TCDD treatment in male wt mice (Figure 6A). This cell subset is composed of two major cell types: CD11c–Ly-6G+MHC-II– neutrophils and CD11c+Ly-6G–MHC-IIhi dendritic cells. Although frequencies of F4/80–CD11b+ cells gradually declined (Figure 6C), their numbers remained significantly higher than those in untreated mice (Figure 6D), as a result of continuous accumulation of immune cells in adipose tissues induced by TCDD (Figure 6B). The F4/80+CD11b+ cell subset was phenotypically more homogenous (CD11c±Ly-6G–MHC-IIint/hi), in contrast to the F4/80–CD11b+ cell subset (Figure 6A, left panel). The phenotype of F4/80+CD11b+ cells resembles inflammatory macrophages and became the dominant cell population 3 days post TCDD treatment (Figure 6C). On day 6, the frequencies of F4/80+CD11b+ cells stayed at the peak level (Figure 6C) and the cell number of this cell subset continued to rise (Figure 6D). The number of F4/80+CD11b+ and F4/80–CD11b+ cells in adipose tissues of wt mice were marginally higher than those of AhRR Tg mice on day 3 and 6 post TCDD treatment (Figure 6E, middle and right panels). The differences were resulting from slight increases of cell accumulation in adipose of wt mice (Figure 6E, left panel). For the frequencies of adipose tissue F4/80+CD11b+ or F4/80-CD11b+ cell subsets, there were no differences between those of wt and AhRR tg mice (data not shown).