Rosiglitazone enhances osteoblastic differentiation of murine preosteoblast cells

Abstract

Peroxisome proliferator-activated receptor (PPAR) γ belongs to the nuclear receptor family of transcription factors. In mouse, there are at least two PPARγ isoforms, PPARγ1 and PPARγ2, that are generated by alternative splicing and promoter usage. The 2 isoforms show distinctive expression patterns. While the more ubiquitously expressed PPARγ1 can be detected in many cell types including adipocytes and osteoblasts, PPARγ2 is expressed primarily in adipose tissues. As PPARγ is demonstrated to play central role in adipogenesis, the reciprocal relationship between osteoblastogenesis and adipogenesis has led to the hypothesis that PPARγ is an inhibitor of osteoblastogenesis. Although there are increasing evidence for the inhibitory role of PPARγ2 isoform, little is known about the roles of PPARγ1 in osteoblast differentiation. This study examined how PPARγ1 was involved in osteoblast differentiation. Preosteoblast MC3T3-E1 (MC) cells express only PPARγ1 but not PPARγ2. Thus, they serve as a good in vitro model system for the study. We treated MC cells with rosiglitazone (ROS), a specific agonist of PPARγ that exerts most of its biological effects via PPARγ, at different concentrations to examine its effect on osteoblastic differentiation. Treatment with ROS at concentrations of 1-10μM resulted in a significant increase in alkaline phosphatase (ALP) activity but their effect on the cellular proliferation of MC cells remained insignificant. Similar upregulation of ALP activities was observed using other full and partial PPARγ agonists such as pioglitazone and MCC-555 respectively but not PPARγ antagonists including GW9662. Using real-time RT-PCR, we further demonstrated that ROS treatment upregulated the expression of osteoblast-specific transcription factors, including Runx2 and osterix. Reporter assays using Runx2 and osteocalcin promoters also showed that ROS enhanced Runx2 transcription as well as Runx2-dependent transcription in MC cells. It is noteworthy that the osteoblastogenic effects of ROS were inhibited by overexpressing PPARγ2 in MC cells. Taken together, our findings suggested that ROS induce osteoblastic differentiation of preosteoblasts in the absence of PPARγ2 and that PPARγ1 play an important role in osteoblastic differentiation.