FIG. 2.

Adipose obELVs promote the differentiation and proliferation of BMDMs and impair activation of the insulin signaling pathway in vitro and macrophage infiltration into adipose tissues in vivo. C2C12 cells at 80% confluency were cultured for 24 h in the presence of conditioned medium harvested from 14-day cultures of bone marrow cells that had been pretreated with obELVs, wtELVs, or thymus exosomes (10 μg/ml) at day 0 of the culture and cultured in the presence or absence of GM-CSF. After a 3-h starvation, the C2C12 cells were stimulated with insulin (100 nmol/l) for 20 min and either lysed for Western blot analysis of phosphorylated Akt (A) or used for glucose uptake testing (B). Results presented are representative of a minimum of three experiments (A) or data are the means ± SE of three experiments with two replicates of each (B). *P < 0.05; **P < 0.01. C: B6 mice fed an HFD were intravenously injected with a mixture (1:1) of 4 × 10⁶ of BMDMs^PKH26+ that had been preincubated with obELVs with BMDMs^PKH67+ that had been preincubated with wtELVs. The percentage of injected macrophages infiltrating adipose tissue, liver, spleen, and bone marrow were determined by FACS analysis of PKH67 and PKH26 14 days after the injection. D: The PKH67⁺ or PKH26⁺ cells were gated and analyzed for CD11b⁺F4/80⁺. The proliferation of injected fluorescent dye–labeled CD11b⁺F4/80⁺ macrophages was then determined by FACS analysis of BrdU⁺ cells in the adipose tissue, liver, spleen, and bone marrow. A representative graph of FACS analysis of macrophage infiltration in adipose tissue is shown and the data represent the means ± SE from five mice from each group. **P < 0.01. BM, bone marrow.