Fig. 4

Isolation of NCPs coexisting with TGF-β in the G2 extract. A Heparin affinity chromatogram showing absorbance at 280 nm for the G2 extract obtained from bone powder of rat tibiae and femora (50 mg). Downward-pointing arrows are the starting point of the step gradient with 0.05, 0.1, 0.2 and 1 M NaCl. B SDS–PAGE (5–20% gradient gel) stained with Bio-Safe Coomassie Brilliant Blue (Coomassie Brilliant Blue, left) and Stains-all (right) showing fractions a, b, c, d, and e on a heparin affinity chromatogram. C ALP-inducing activity of HPDL cells exposed to fractions a, b, c, d, and e (5 μg/mL each) and CF-hTGF-β1 (1 ng/mL) without (−) or with (+) SB431542. Data in box indicate the 25-75% tiles of IQR, and the end of whiskers indicates maximum and minimum values, respectively. The median is represented as line located in the middle of box (*p < 0.05, Mann–Whitney U test). Control: ALP-inducing activity of HPDL cells cultured in the absence of fractions a, b, c, d, and e or CF-hTGF-β1