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. 2013 Aug 21;9(8):830–841. doi: 10.7150/ijbs.7039

Figure 6.

Figure 6

PTHrP knockdown decreases the tumor-induced inhibition of osteoblast differentiation in vitro. (A-C) MC-4 preosteoblastic cells were cultured in osteoblast differentiation media for 7d (qPCR analysis) (A) or 12d (Alizarin red staining) (B). Cells were treated with and without NC-CM, 995-CM or 996-CM for the last 48h. mRNAs were normalized to Gapdh mRNA. Alizarin red was extracted with 10% cetylpyridinium chloride and the dye concentration in the extracts was determined at OD 562 nm. (C). Bars represent means ± S.D. from three independent experiments. (D) Primary BMSCs were cultured in osteoblast differentiation media for 7d and treated with and without conditioned media (CM) from NC (NC-CM), 995 (995-CM) or 996 cells (996-CM) for the last 48h, followed by ALP (alkaline phosphatase) activity assay. ALP activity was normalized to total protein.