Figure 2

Adipogenic differentiation. (a) FAPs and ASCs were grown to confluence in adjacent culture wells and then treated with differentiation-inducing medium. Cells were fixed and assayed for intracellular lipid droplets with oil red O staining from day 0 (start of differentiation induction) to day 30, as indicated. Cell cultures were counterstained with crystal violet. Pictures were visualized by light microscopy with × 200 magnification. Fields representative of whole cell culture wells are shown and were obtained with ASC biopsy 4 and FAP biopsy 6. Scale bar=50 μm. (b) Adipocyte and lipid droplet sizes (μm²) were analyzed using ImageJ software in cultures after 2 weeks of adipogenic differentiation. Lipid droplet quantity was estimated by the intracellular droplet number counted per adipocyte. ASC-As are represented in gray bars (n=3; ASCs biopsies 1, 3, and 4) and FAP-As in black bars (n=3; FAP biopsies 4, 6, and 7). Results are mean±S.E. of the mean for three independent measurements. *P<0.05, **P<0.01. (c) FAP-As mRNA content of classical early (PPARγ and CEBPβ) and late (FABP4, CD36, adipsin, adiponectin, leptin, and LPL) adipogenic markers was quantified by quantitative RT-PCR. The results are expressed as ratios (%) to respective ASC-As mRNA contents, which are represented by the horizontal line. FAP-As (n=3; FAP biopsies 4, 6, and 7) and ASC-As (n=3; ASC biopsies 1, 2, and 4) were strictly maintained under the same culture conditions. Results are mean±S.E. of the mean for three independent ratio measurements *P<0.05. (d) UCP1 expression was measured with quantitative RT-PCR in FAP-As, which were prepared from a panel of biopsies (n=10; FAP biopsies 1, 4, 6, 7, 8, 9, 11, 12, 14, and 15) in comparison with ASCs-As from biopsies 1, 3, and 4. *P<0.05, **P<0.01