The Study On Signal Transduction Pathways That Regulate Osteoblast Differentiation

Multiple signal pathways regulate osteoblast differentiation including those initiated by Bone Morphogenetic Proteins. In this study, the regulatory mechanism of MAPK pathway was investigated in culture of mouse MC3T3-E1 pre-osteoblast. Firstly, a reliable and repeatable culture system for osteoblast differentiation in vitro was established, which was based on the combined application of 50 u g/ml L-ascorbate and 10 mM 3 -glycerophosphate. After confirming the involvement of MAPK pathway in regulating osteoblast differentiation by detecting the activation ERK1/2 via western-blot experiment, the investigation of the underlying mechanism was performed according to the following two strategies. On one hand, the activation of MEK/ERK and p38 MAPK pathway was inhibited with the specific inhibitor PD98059 and SB203580, respectively. On the other hand, MC3T3-E1 cells were stably transfected with wild type or constitutively activated form of MEKl gene to enhance the activation of MEK/ERK branch pathway. Besides, the stimulatory effect of BMP-2 on those stable transfectants was also observed. Meanwhile, the regulatory mechanism of PI3K on osteoblast differentiation was studied preliminarily.Our results showed that the inductive and evaluation system for osteoblast differentiation in vitro is stable, reliable and repeatable as well. The MEK/ERK branch of MAPK pathway plays an inhibitory role in regulating osteoblast differentiation, while p38 MAPK and PI3K pathway are necessary for osteoblast differentiation. BMP-2, the most potent regulator of osteoblast differentiation, did not directly depend on the activation of MAPK pathway to induce the differentiation of osteoblast differentiation.