Figure 4.

Osteogenic BMPs effectively induce early osteogenic genes but fail to induce late osteogenic markers. (a) Osteogenic BMPs upregulate Id expression in the four OS lines. Subconfluent OS lines were cultured in 1% FCS and infected with AdBMP2, AdBMP6, AdBMP9, and AdGFP. At 30 h after infection, total RNA was isolated from the infected cells and subjected to reverse transcription and qPCR analysis using primers specific for human Id1, Id2, and Id3. (b) Osteogenic BMPs upregulate Id expression in the patient-derived primary OS lines. Subconfluent cells were cultured in 1% FCS and infected with AdBMP2, AdBMP9, and AdGFP for 30 h. Total RNA and qPCR analysis were carried out essentially the same as that described in panel (a). (c) Osteogenic BMPs exert no significant effect on the expression of late osteogenic marker osteocalcin and osteopontin in human OS lines. Subconfluent OS lines were infected with AdBMP2, AdBMP6, AdBMP9, and AdGFP. At 7 days after infection, total RNA was isolated from the infected cells and subjected to reverse transcription and qPCR analysis using primers specific for human osteocalcin and osteopontin (listed in Supplementary Table 1). Each assay condition was done in triplicate. Fold of changes was calculated by dividing the relative expression level from BMP-treated samples with that of GFP-treated samples. The dotted lines indicate that the fold of change is 1.0 (ie, BMP-induced expression is equal to that of GFP control-treated cells). See text for detail.