Effects of hypoxia on adipogenesis. (A) Hypoxia inhibits TG accumulation. Cells were allowed to differentiate for 15 days with differentiation mix (black bars) under normoxia (20%), gaseous hypoxia (1%) or chemical hypoxia (100 μM DFO). TG accumulation was quantified by colorimetric determination of Red-Oil staining. Data are presented as optical density at 495 nm (OD495 nm). n = 4, ** p < 0.001. (B) Hypoxia modulated adipocyte gene expression. Confluent cells were treated for 24 hours with 100 μM DFO (+) grey bars or were not treated (-) white bars. RNA was extracted and expression of the PPARγ, leptin and VEGF genes was followed by quantitative PCR. n = 2, ** p < 0.01. (C) Localization of HIF-1α and leptin protein in hypoxia-stimulated cells. Cells were treated for 24 hours with 100 μM DFO (+DFO) or were not treated (-DFO). Proteins were detected with mAB against HIF1-α tagged with a goat anti-mouse IgG coupled to FITC (a) or with mAB against leptin tagged with a goat anti-rabbit IgG coupled to cy3 (b), or a co-staining (c).