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. 2014 Mar 3;9(3):e89884. doi: 10.1371/journal.pone.0089884

Figure 2. Cyclin G2 is down-regulated in estrogen-mediated osteogenesis in vitro.

Figure 2

(A and B) Semi-quantitative RT-PCR cyclin G2 mRNA expression and Western Blot analysis of cyclin G2 protein level in steroid-free OS-medium cultured C2C12 cells with or without E2 treatment. C2C12 cells were induced to osteogenic differentiation by steroid-free OS-medium treated with E2 (100 nM) or DMSO. Total RNA and protein were extracted after 48 h of induction, followed by semi-quantitative RT-PCR and Western Blot analysis of cyclin G2. The relative integrated density of each band was digitized by Quantity One. Results are shown as mean ± SEM of data at least three separate experiments, each performed with triplicate samples. *P<0.05 and **P<0.01 vs. control groups by one-way ANOVA. (C and D) ALP activity (Results are shown as mean ± SEM of data at least three separate experiments, each performed with triplicate samples. *P<0.01 vs. control groups by one-way ANOVA) and mineralization (ARS staining) confirmed osteogenic differentiation in C2C12 cells induced by steroid-free OS-medium and E2. Cells were treated as in A, and the amount of ALP activities on day 7 and calcium deposition on day 14 was determined as described under Materials and Methods .