PDGF-AA promotes MSC osteogenic differentiation.

<p>A. PDGF-AA promotes ALP expression in MSC, which requires BMP-Smad1/5/8 signaling. Primary mouse MSC cells were cultured with or without PDGF-AA for 7 or 14 days. To test the effect of BMPRI activation, LDN-193189 was included in the culture medium. ALP was stained 7 or 14 days after culture. B. Quantitation data for ALP activities that were normalized to the total protein levels of each culture. C. PDGF-AA promotes bone mineralization in MSC, which requires BMP-Smad1/5/8 signaling. Primary mouse MSC cells were cultured in differentiation medium with or without PDGF-AA (changed every 3 days) for 28 days. To test the effect of BMPRI activation, LDN-193189 was included in the culture medium. Mineralization was stained with Von Kossa and Alizarin red method. D. PDGF-AA up-regulated the mRNA levels of Runx2, Atf4, and Osx via the BMP-Smad1/5/8 signaling, without affecting the mRNA levels of Sox9, C/EBPα, or PPARγ. Primary MSC cells were cultured with or without PDGF-AA for 7 days. To test the effect of BMPRI activation, LDN-193189 was included in the culture medium. Total RNA was isolated from the cells and realtime PCR was carried out to determine the mRNA levels of the six transcription factors. Data are means ±s.e.m. (n = 3 for all panels). * p<0.05, when compared to empty vectors or untreated cells. E. PDGF-AA up-regulated the protein levels of Runx2, Atf4, and Osx via the BMP-Smad1/5/8 signaling. The experiments were carried out as described in Fig. 2D and the protein levels of these proteins were determined by western blot.