Fig. 6.

Regulation of fat cell differentiation markers by rhCCN2 or rhTGF-β1 each in the presence of differentiation mix and anti-TGF-β neutralising antibody. (a) Representative images of Oil red O stained cells at day 0 in A, or 10 days post differentiation in B to F. Cells were treated with differentiation mix, in some cases with rhCCN2 (500 ng/ml), active rhTGF-β1 (2 ng/ml) and/or anti- TGF-βantibody (10 μg/ml) at day 0 as indicated, and were then cultured as described in the Methods; at day 10 cells were fixed with 10 % formalin and stained with Oil red O, then photographed. Each size-bar in (a) indicates 400 μM. In (b) Oil red O quantitative data investigating the effect of rhCCN2 (500 ng/ml), active rhTGF-β1 (2 ng/ml) and and/or anti- TGF-βantibody on adipocyte differentation are shown (b). IgG (10 μg/ml), was used as a loading control. Data are expressed as mean ± SD *p < 0.05 each vs. non-differentiated; #P < 0.05 vs the respective rhCCN2 or rhTGF-β1 treatment with differentiation mix (by ANOVA). Adiponectin, Resistin and Pref-1 mRNA levels were determined at day 10 as in (c). Data shown in (c) are generated from 3 independent experiments conducted in triplicate wells and are expressed as mean ± SD; *p < 0.05 each vs differentiation mix alone; #p < 0.05 vs added rhCCN2 or rhTGF-β1 each with differentiation mix (by ANOVA)