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. 2012 Apr 23;7(4):e35258. doi: 10.1371/journal.pone.0035258

Figure 1. Ets-1 is essential for TGF-β1 induction of CCN2 protein in osteoblasts.

Figure 1

(A) Primary osteoblasts were cultured until they were 80% confluent, serum deprived for 24 hrs and then treated with TGF-β1 (5 ng/ml) (+) or mock treated with TGF-β1 diluent (−). At 24 hrs post treatment, cell lysates were harvested and assessed for CTGF and Ets-1 expression by Western blot analysis. (B) Primary osteoblasts were transfected with Ets-1-Myc (+) or an empty vector (−) and then treated with TGF-β1 (5 ng/ml) (+) or mock treated with TGF-β1 diluent (−). At 24 hrs post treatment, cell lysates were harvested and assessed for CCN2, Ets-1 and Myc expression by Western Blot analysis. (C) Primary osteoblasts were transfected with 100 nM of Ets-1 siRNA (Ets-1) or control siRNA (C) for 48 hrs. Following transfection, the cells were serum starved for 24 hrs and then treated with 5 ng/ml of TGF-β1 (+) or TGF-β1 diluent (−) for 24 hrs. RNA was harvested and assessed for Ets-1 expression by RT-PCR. Cell lysates were harvested and assessed for Ets-1 and CCN2 expression by Western Blot analysis. Each experiment is representative of at least three independent experiments.