Enamel matrix derivative: protein components and osteoinductive properties.
Enamel matrix derivative: protein components and osteoinductive properties.
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DOI：
10.1902/jop.2013.130264
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发表时间：
2014-02
影响因子：
4.3
通讯作者：
Bradshaw M Stout;Brian J. Alent;Peter M. Pedalino;Ryan Holbrook;J. Gluhak‐Heinrich;Yong Cui;M. Harris;A. Gemperli;D. Cochran;D. E. Deas;S. Harris
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文献类型：
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作者：
Bradshaw M Stout;Brian J. Alent;Peter M. Pedalino;Ryan Holbrook;J. Gluhak‐Heinrich;Yong Cui;M. Harris;A. Gemperli;D. Cochran;D. E. Deas;S. Harris
背景技术虽然牙釉质基质衍生物(EMD)已被证明能够在体外和体内促进血管生成和骨生成，但EMD化合物中负责这些作用的具体元素仍然未知。方法 根据分子量从商业生产的 EMD 中收集九种不同的蛋白质库。其中六个池以及完整的 EMD 未分级化合物以及阳性和阴性对照，在颅骨诱导测定中测试了它们诱导骨形成的能力。在选定的时间点对磷酸化 SMAD1/5/8（磷酸化 SMAD）、osterix 和血管内皮生长因子 A (VEGF-A) 进行免疫细胞化学分析。最后，完成蛋白质组分析以确定各种骨诱导库的特定蛋白质-肽含量。结果 所测试的较低分子量池之一（池 7）显示骨诱导反应显着高于其他池和完整 EMD 化合物，并且呈浓度依赖性。动态骨形成率分析表明，7 号池在 5 至 10 μg 浓度下具有最佳活性。结果表明，EMD 和池 7 诱导磷酸化 SMAD、osterix 和 VEGF-A，这表明骨形态发生蛋白 (BMP) 信号传导增加。蛋白质组组成分析表明，池 7 具有最高浓度的生物活性牙釉蛋白-富含亮氨酸的牙釉蛋白肽和成釉细胞蛋白 17-kDa 肽。结论 这些研究表明，EMD 中的低分子量蛋白库（7 至 17 kDa）比市售的完整 EMD 化合物具有更大的骨诱导潜力，并且其作用机制部分是通过增加 BMP 信号传导以及增加 osterix 和 VEGF-A 来实现的。有了这些信息，现在就可以配制 EMD 的选定成分，以实现最佳的骨生成和血管生成。
BACKGROUND Although enamel matrix derivative (EMD) has demonstrated the ability to promote angiogenesis and osteogenesis both in vitro and in vivo, the specific elements within the EMD compound responsible for these effects remain unknown. METHODS Nine different protein pools from a commercially produced EMD were collected based on molecular weight. Six of these pools, along with the complete EMD unfractionated compound and positive and negative controls, were tested for their ability to induce bone formation in a calvarial induction assay. Immunocytochemistry of phosphorylated SMAD1/5/8 (phospho-SMAD), osterix, and vascular endothelial growth factor A (VEGF-A) was carried out at selected time points. Finally, proteomic analysis was completed to determine the specific protein-peptide content of the various osteoinductive pools. RESULTS One of the lower-molecular-weight pools tested, pool 7, showed bone induction responses significantly greater than those of the other pools and the complete EMD compound and was concentration dependent. Dynamic bone formation rate analysis demonstrated that pool 7 was optimally active at the 5- to 10-μg concentration. It was demonstrated that EMD and pool 7 induced phospho-SMAD, osterix, and VEGF-A, which is indicative of increased bone morphogenetic protein (BMP) signaling. Proteomic composition analysis demonstrated that pool 7 had the highest concentration of the biologically active amelogenin-leucine-rich amelogenin peptide and ameloblastin 17-kDa peptides. CONCLUSIONS These studies demonstrate that the low-molecular-weight protein pools (7 to 17 kDa) within EMD have greater osteoinductive potential than the commercially available complete EMD compound and that the mechanism of action, in part, is through increased BMP signaling and increased osterix and VEGF-A. With this information, selected components of EMD can now be formulated for optimal osteo- and angio-genesis.