Fig. 1
Transgene expression by MSCs during 21 days of aggregate culture following adenoviral gene transfer with GFP, SOX9, TGFB1 or IGF1 alone or in combination. Primary MSCs were infected with Ad.GFP, Ad.SOX9, Ad.TGFB1, Ad.IGF1 alone or in combinations at 5 × 102 vp/cell, seeded into aggregates and analysed for respective transgene expression during a 3 week time course. (a) GFP transgene expression was detected by fluorescence microscopy, panels are reproduced at low magnification (50x; bar = 200 μm) as indicated (b) Production of SOX9 protein was analysed by Western blot of cell lysates. Representative gels of experiments from three independent marrow preparations are shown; lysates from 10 aggregates per time point represent one band of the gel. (c) TGFB1 transgene expression was evaluated by measurement of TGF-β1 protein concentration in the conditioned media of the respective aggregate cultures over a 24-h period at days 3, 7, 14 and 21 compared to Ad.GFP controls. The data represent mean values ± SD from measurements of supernatants of n = 3 aggregates per condition and time point; n = 3 marrow preparations. Asterisks indicate values that are statistically different (p < 0.05) from marker gene–transduced control cultures or between samples as indicated. (d) IGF1 transgene expression was evaluated by quantification of IGF-1 protein concentration in the conditioned media of the respective aggregate cultures over a 24-h period at days 3, 7, 14 and 21 compared to Ad.GFP controls. The data represent mean values ± SD from measurements of supernatants of n = 3 aggregates per condition and time point; n = 3 marrow preparations. Asterisks indicate values that are statistically different (p < 0.05) from marker gene–transduced control cultures