Corosolic Acid Stimulates Osteoblast Differentiation by Activating Transcription Factors and MAP Kinases

Recently, the use of anabolic agents to enhance bone mass has been a source of interest. Previous work by us suggested that corosolic acid (2a-hydroxyursolic acid), an active component of banaba leaves (Lagerstroemia speciosa L.), potentially stimulates the differentiation of mouse osteoblasts. Therefore, the present study investigated whether corosolic acid stimulates osteoblast differentiation, and its possible mechanisms of action. At low concentrations (up to 5 mM), corosolic acid significantly stimulated osteoblast differentiation and mineralization without cytotoxicity. Corosolic acid induced NF-kB and MAP kinase activity at an early stage of osteoblast differentiation and increased the activity of the transcription factor AP-1 during late-stage osteoblast differentiation. These results suggest that the anabolic effects of corosolic acid upon osteoblast differentiation could result from its activation of transcription factors and MAP kinases.

In this study, it was found that corosolic acid had a stimulatory effect on osteoblast differentiation. Corosolic acid induced mRNA expression and the activity of ALP, along with activation of MAP kinases (especially ERK and p38) and NF-kB during early-stage osteoblast differentiation. During later stages, it induced mineralization and activation of AP-1 family members.

MAP kinases and NF-kB signaling play an important role in osteoblast differentiation. ERK regulates ALP activity and the deposition of bone matrix proteins (Lai et al., 2001). Activation of p38 is critical for ALP expression during differentiation of MC3T3-E1 cells (Suzuki et al., 2002). Furthermore, NF-kB regulates the expression of genes specific to osteoblast differentiation (Yamamoto et al., 1995). In several studies, corosolic acid triggered the induction of gene expression by activating MAP kinase and NF-kB (Ikeda et al., 2005; You et al., 2001). Thus, osteoblast differentiation induced by corosolic acid could be due to the ability of corosolic acid to induce the activation of MAP kinases and NFkB, which would consequently trigger the expression of osteoblast-specific genes such as ALP.

AP-1 activity and NFAT signaling is also critical for osteoblast differentiation (Eferl et al., 2004; Wagner, 2002; Koga et al., 2005; Winslow et al., 2006). In fact, ERK and p38 activation activates AP-1 in osteoblast differentiation (Hipskind and Bilbe; 1998). AP-1 is a dimeric transcription factor complex formed by Jun proteins (c-Jun, JunB and JunD) and Fos proteins (c-Fos, FosB, Fra-1, Fra-2 and DFosB). AP-1 family members affect the maturation of osteoblasts and the production of bone matrix, which is essential to bone formation and remodeling (Eferl et al., 2004; Wagner, 2002). Interestingly, osteoblast-specific genes such as osteocalcin have AP-1 binding sites within their promoter regions (Akhouayri and St-Arnaud, 2007; Lu et al., 2006). The dephosphorylation of NFAT proteins in the cytoplasm allows them to enter the nucleus and activate gene expression in concert with binding partners such as members of the AP-1 family (Boise et al., 1993). In the present study, it was shown that corosolic acid increased the mRNA levels and nuclear translocation of AP-1 family members during late-stage differentiation; this supports a role for AP-1 in the mineralization stage of corosolic acid-induced osteoblast differentiation.

Recently, corosolic acid was shown to inhibit the enzymatic activities of protein tyrosine phosphatases (PTPs) such as PTP1B in vitro (Shi et al., 2008). Interestingly, PTP1B inhibitors increased ALP activity and collagen synthesis in normal human bone cells (Yoon et al., 2000). These data suggest that the inhibitory effect of corosolic acid on the activity of PTP1B could result in the stimulation of osteoblast differentiation.

In conclusion, it is suggested that corosolic could induce osteoblast differentiation in at least two ways: (1) by activating MAP kinases and transcription factors that consequently increase the expression of osteogenic factors, and (2) by inhibiting PTP1B. Further studies are required to investigate the biological efficacy of corosolic acid in an in vivo model.

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