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. 2011 Oct 5;6(10):e25163. doi: 10.1371/journal.pone.0025163

Figure 5. Osteoblast differentiation is arrested by inhibition of dynamin-dependent endocytosis.

Figure 5

(A,B) Confluent C2C12 cells were serum-starved for 2 h prior to stimulation with the indicated concentrations of BMP-2. After 72 h of stimulation, cells were lysed and ALP activity was measured at 405 nm by conversion of para-nitrophenylphosphate. A schematic representation of the treatment is depicted above the respective histogram. The histograms show mean ± s.d. of triplicate measurements representative of three independent experiments. (A) During starvation and initial 4 h of stimulation, 40 µM dynasore or 0.05% DSMO were added. (P-values in relation to control treated samples). (B) After 4 h of stimulation, BMP-2-containing medium was replaced by medium without BMP-2. (** P-values in relation to unstimulated samples; 30 nM: P = 0.008, 60 nM: P = 0.0017) (C–G) Confluent C2C12 cells were treated as described in (A) with 30 nM BMP-2. Total RNA was isolated, cDNA was synthesized and mRNA expression was analyzed by qPCR using specific mouse primers (Table S1). Histograms show mean normalized expression (MNE) with standard error of duplicate measurements relative to the housekeeping gene GAPDH. The results are representative of three independent experiments.