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Effects of Bone Morphogenetic Protein-2 and Transforming Growth Factor β 1 on Gene Expression of Transcription Factors, AJ18 and Runx2 in Cultured Osteoblastic Cells

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Abstract

Osteoblast differentiation is controlled by multiple transcription factors, Runx2, AJ18, Osterix, Dlx5 and Msx2. The mechanisms of regulation of AJ18 mRNA expression by the transforming growth factor β (TGF-β) superfamily remain poorly understood. However, it is known that BMP-2 induces differentiation of C26 cells into more mature osteoblastic cells. The present study, using Northern blot and real-time reverse transcription polymerase chain reaction analyses, investigated the effects of bone morphogenetic protein-2 (BMP-2) and TGF-β 1 on mRNA expression of AJ18 and Runx2 in a clonal osteoblast precursor cell line ROB-C26 (C26) cultured for 3, 6 or 9 days in the presence or absence of BMP-2. Although mRNA expression of Osterix and bone sialoprotein (BSP) was undetectable in the C26 culture, BMP-2 induced Osterix expression on days 3–9, but not BSP expression. BMP-2 also stimulated significantly Dlx5 expression on days 3–9, Msx2 and matrix Gla protein expressions on days 3 and 6, Runx2, alkaline phosphatase and osteocalcin expressions on days 6 and 9 in the culture. Furthermore, BMP-2 increased significantly Smad5 mRNA in the culture on day 3, indicating BMP-2 involvement in the regulation of Smad5 mRNA expression. In contrast, the inhibitory effects of BMP-2 on AJ18 mRNA expression were significant on days 3–9, indicating that a decrease in AJ18 mRNA expression is essential for the increased osteoblastic differentiation. Furthermore, TGF-β 1 (0, 0.1, 1.0 and 5.0 ng/ml) treatment of C26 cells cultured for 6 days in the presence or absence of BMP-2 for 24 h stimulated mRNA levels of AJ18 and Runx2, maximal stimulation occurring principally at 1.0 ng/ml. These observations indicate that the expression of AJ18 and Runx2 mRNAs in C26 cells is under the control of BMP-2 and TGF-β 1, which exert different effects on AJ18 mRNA expression, but are potent stimulators of Runx2 mRNA expression during osteoblast differentiation.

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Authors and Affiliations

  1. Department of Anatomy, Nihon University School of Dentistry, Tokyo, 101-8310, Japan

    Minoru Takagi, Naoko Kamiya, Tomihisa Takahashi & Shinsuke Ito

  2. Division of Functional Morphology of Dental Research Center, Nihon University School of Dentistry, Tokyo, 101-8310, Japan

    Minoru Takagi, Naoko Kamiya, Tomihisa Takahashi & Naoto Suzuki

  3. Department of Oral and Maxillofacial Surgery, Nihon University School of Dentistry, Tokyo, 101-8310, Japan

    Mitsuharu Hasegawa & Koji Nakanishi

  4. Department of Biochemistry, Nihon University School of Dentistry, Tokyo, 101-8310, Japan

    Naoto Suzuki

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  1. Minoru Takagi
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Takagi, M., Kamiya, N., Takahashi, T. et al. Effects of Bone Morphogenetic Protein-2 and Transforming Growth Factor β 1 on Gene Expression of Transcription Factors, AJ18 and Runx2 in Cultured Osteoblastic Cells. Histochem J 35, 81–90 (2004). https://doi.org/10.1023/B:HIJO.0000021151.31118.e3

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