Fig. 6.

Adenoviral expression of FSP27 protects against TNF-α-mediated lipolysis and lipid droplet size diminution. 3T3-L1 adipocytes were infected with control virus (Ad-GFP) or HA-FSP27-expressing virus (Ad-HA-FSP27) on the fourth day of differentiation. On the fifth day, cells were serum starved overnight, treated with 10 ng/ml TNF-α for 24 h, and analyzed for protein, glycerol release, and lipid droplet morphology. A: Western blot analysis using FSP27, HA, GFP, perilipin (PLIN), actin, and Vti1a antibodies show maintenance of HA-tagged FSP27 protein levels even after treatment of TNF-α. Actin is the loading control for HA, GFP, and PLIN, and Vti1a is the loading control for FSP27. B: Glycerol released into the medium in 24 h with or without TNF-α treatment. A. U., arbitrary units. C: Confocal microscopy image showing lipid droplets stained with Oil Red O (red), nuclei stained with DAPI (blue), and GFP expression (green) in 3T3-L1 adipocytes expressing Ad-GFP or Ad-HA-FSP27 in the presence or absence of TNF-α. Data are expressed as means ± SEM for five individual experiments. P values were calculated using ANOVA; *, P < 0.05; **, P < 0.0005.