In vitro and in vivo synergistic interactions between the Runx2/Cbfa1 transcription factor and bone morphogenetic protein-2 in stimulating osteoblast differentiation
In vitro and in vivo synergistic interactions between the Runx2/Cbfa1 transcription factor and bone morphogenetic protein-2 in stimulating osteoblast differentiation
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DOI：
10.1359/jbmr.2003.18.4.705
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发表时间：
2003-04-01
影响因子：
6.2
通讯作者：
Franceschi, RT
中科院分区：
文献类型：
--
作者：
Yang, SY;Wei, DY;Franceschi, RT
骨再生需要许多因素之间的相互作用，包括骨形态发生蛋白（BMP），生长因子和转录调节因子，如Runx 2/Cbfa 1（Runx 2）。因为每种成分对整体成骨反应都有独特的贡献，我们假设骨形成可以通过互补因子的组合来增强。作为该概念的初始测试，在多能C3 H10 T1/2细胞系中使用基于腺病毒的表达载体（AdCMV-Runx 2、AdCMV-BMP 2）检查BMP 2和Runx 2之间的相互作用。AdCMV-Runx 2转导的细胞强烈表达成骨细胞标志物，如碱性磷酸酶和骨钙素，但在体外仅形成弱矿化的细胞外基质，而AdCMV-BMP 2转导的细胞表现出较高的矿化水平，但仅表达低水平的Runx 2和骨钙素mRNA。值得注意的是，当细胞用两种病毒的最佳滴度转导时，成骨细胞分化被刺激到比单独使用AdCMV-Runx 2或AdCMV-BMP 2所观察到的水平高10倍的水平。为了测量体内成骨活性，将病毒转导的细胞皮下植入免疫缺陷小鼠中。用对照病毒转导的细胞仅产生纤维组织，而用AdCMV-Runx 2转导的细胞产生有限量的软骨和骨。相反，单独用AdCMV-BMP 2或AdCMV-BMP 2加AdCMV-Cbfa 1转导的细胞产生含有软骨、骨和骨髓腔的大听小骨。然而，AdCMV-BMP 2 + AdCMV-Cbfa 1组的骨化更广泛，因为矿物质含量和骨面积分数均大于AdCMV-BMP 2组。因此，在体外用组合腺病毒处理观察到的成骨细胞分化的增加也通过体内骨形成的增加来证明。这些结果表明，Runx 2和BMP 2在骨生成中具有不同但互补的作用，并且它们的联合作用可能是最佳骨形成所必需的。
Bone regeneration requires interactions between a number of factors including bone morphogenetic proteins (BMPs), growth factors, and transcriptional regulators such as Runx2/Cbfa1 (Runx2). Because each component may provide a unique contribution to the overall osteogenic response, we hypothesized that bone formation may be enhanced by using combinations of complimentary factors. As an initial test of this concept, interactions between BMP2 and Runx2 were examined using adenovirus-based expression vectors (AdCMV-Runx2, AdCMV-BMP2) in the pluripotent C3H10T1/2 cell line. Cells transduced with AdCMV-Runx2 strongly expressed osteoblast markers, such as alkaline phosphatase and osteocalcin, but formed only a weakly mineralized extracellular matrix in vitro, whereas cells transduced with AdCMV-BMP2 exhibited higher levels of mineralization, but only expressed low levels of Runx2 and osteocalcin mRNA. Significantly, when cells were transduced with optimal titers of both viruses, osteoblast differentiation was stimulated to levels that were 10-fold greater than those seen with either AdCMV-Runx2 or AdCMV-BMP2 alone. To measure in vivo osteogenic activity, virally transduced cells were subcutaneously implanted into immunodeficient mice. Cells transduced with control virus produced only fibrous tissue while those with AdCMV-Runx2 produced limited amounts of both cartilage and bone. In contrast, cells transduced with either AdCMV-BMP2 alone or AdCMV-BMP2 plus AdCMV-Cbfa1 generated large ossicles containing cartilage, bone, and a marrow cavity. However, ossification in the AdCMV-BMP2 plus AdCMV-Cbfa1 group was more extensive in that both mineral content and fractional bone area were greater than that seen in the AdCMV-BMP2 group. Thus, the increased osteoblast differentiation observed with combined adenovirus treatment in vitro is also manifested by increased bone formation in vivo. These results suggest that Runx2 and BMP2 have distinct, but complementary, roles in osteogenesis and that their combined actions may be necessary for optimal bone formation.