Igarashi Masato, Kamiya Naoko, Hasegawa Mitsuharu, Kasuya Tomohiro, Takahashi Tomihisa, Takagi Minoru
Department of Periodontology, Nihon University School of Dentistry, 1-8-13 Kanda-Surugadai, Chiyoda-ku, Tokyo 101-8310, Japan.
J Mol Histol. 2004 Jan;35(1):3-10. doi: 10.1023/b:hijo.0000020883.33256.fe.
Runx2/core binding factor alpha 1 (Cbfa1) and Osterix (Osx) are osteoblast-specific transcription factors essential for the development of a mature osteoblast phenotype and are thought to activate osteoblast marker genes in vivo to produce a bone-specific matrix. Dexamethasone (Dex) is known to be a potent stimulator of osteoblastic differentiation in vitro, however, the exact role is still unclear. To investigate the mechanisms of the stimulation of osteoblastic differentiation by Dex, we evaluated the effects of Dex on proliferation and mineralization as well as on mRNA expression of Cbfa1, Osx and osteoblast marker genes, osteocalcin (OC) and bone sialoprotein (BSP) mRNAs in differentiating foetal rat calvarial cells (FRCC), which were cultured for 35 days in the presence or absence of 10(-7) M Dex. Treatment of FRCC with Dex resulted in the stimulation of cell proliferation and increased the number of cells, which are able to produce bone-like nodules with a mineralized matrix when compared to untreated controls. Northern blot analysis revealed that, in the absence of Dex, Cbfa1 mRNA expressed at day 8, while Osx mRNA expressed at day 15. Subsequently expression of these mRNAs increased up to day 21, followed by constant expression during the culture period. The expression of OC and BSP mRNAs appeared to be synchronous with that of Osx mRNA and was detectable at day 15 with an increase thereafter. The presence of Dex resulted in an induction in Cbfa1 and Osx mRNA expression. The former appeared at day 5 and the latter appeared at day 11. Subsequently expression of Cbfa1 and Osx mRNAs increased up to day 15 with a decrease thereafter. Expression of OC and BSP mRNAs appeared to be coincident with that of Osx mRNA and was detectable at day 11 and reached a maximum at day 15 followed by constant expression. These observations indicate that induction of Cbfal and Osx mRNAs by Dex may be followed by activation of osteoblast marker genes such as OC and BSP mRNAs to produce a bone-specific matrix that subsequently becomes mineralized. Thus, it is likely that Dex may promote osteoblastic differentiation and mineralization of FRCC by inducing the expression of Cbfa1 and Osx genes in vitro.
Runx2/核心结合因子α1（Cbfa1）和osterix（Osx）是成骨细胞特异性转录因子，对于成熟成骨细胞表型的发育至关重要，并且被认为在体内激活成骨细胞标记基因以产生骨特异性基质。已知地塞米松（Dex）是体外成骨细胞分化的有效刺激剂，然而，其确切作用仍不清楚。为了研究Dex刺激成骨细胞分化的机制，我们评估了Dex对增殖、矿化以及Cbfa1、Osx和成骨细胞标记基因、骨钙素（OC）和骨涎蛋白（BSP）mRNA在分化的胎鼠颅骨细胞（FRCC）中的mRNA表达的影响，这些细胞在存在或不存在10^(-7) M Dex的情况下培养35天。用Dex处理FRCC导致细胞增殖受到刺激，并且细胞数量增加，与未处理的对照相比，这些细胞能够产生具有矿化基质的骨样结节。Northern印迹分析显示，在没有Dex的情况下，Cbfa1 mRNA在第8天表达，而Osx mRNA在第15天表达。随后这些mRNA的表达在第21天之前增加，并在培养期间持续表达。OC和BSP mRNA的表达似乎与Osx mRNA同步，在第15天可检测到，此后增加。Dex的存在导致Cbfa1和Osx mRNA表达的诱导。前者在第5天出现，后者在第11天出现。随后Cbfa1和Osx mRNA的表达在第15天之前增加，此后下降。OC和BSP mRNA的表达似乎与Osx mRNA一致，在第11天可检测到，并在第15天达到最大值，随后持续表达。这些观察结果表明，Dex对Cbfal和Osx mRNA的诱导可能随后激活成骨细胞标记基因如OC和BSP mRNA以产生随后矿化的骨特异性基质。因此，Dex可能通过在体外诱导Cbfa1和Osx基因的表达来促进FRCC的成骨细胞分化和矿化。
