The Mechanistic Control of Bone Matrix Material Properties by TGF-beta and Runx2

描述（由申请人提供）：骨基质是一种有机和矿物质复合物，赋予骨韧性和硬度。这种基质的材料特性可以预测骨折风险，并可以补充目前依赖于骨矿物质密度的临床骨折风险评估。与骨质量的其他方面一样，基质材料的特性是由生物学决定的，在解剖学上是不同的。然而，对它们在健康骨骼中的调节或在疾病中的失调知之甚少。这项研究的长期目标是确定控制基质材料特性的机制，以便最终可以利用它们进行治疗以预防骨骼疾病并改善颅面骨修复。转化生长因子-<$（TGF-<$）最近被确定为第一个调节骨基质材料性质的生长因子。TGF-β通过抑制Runx 2（一种关键的成骨细胞转录因子）的功能，抑制成骨细胞基因表达和分化，以及骨基质材料特性。虽然Runx 2整合了来自许多途径的信号，但只有TGF-β和糖皮质激素被证明可以调节基质材料的性质。因此，该提案旨在研究TGF-β和Runx 2调节基质材料性质的细胞和分子机制。成骨细胞是TGF-β调节基质材料特性所必需的。成骨细胞是否直接改变基质材料的性质，或通过与破骨细胞合作间接改变骨重建活性仍不清楚。使用遗传学和药理学工具来操纵小鼠破骨细胞和成骨细胞的功能，目的1旨在确定两种细胞类型在基质材料特性规范中的相对作用。将通过纳米压痕、断裂韧性测试和X射线断层扫描显微镜评价骨基质材料特性和矿物质浓度。TGF-β可以抑制或激活Runx 2的表达，显然是通过下游信号通路的差异利用。由于Runx 2的功能水平影响基质材料的性质，目的2试图确定TGF-β信号调节Runx 2以控制成骨细胞分化和基质材料性质的机制。将使用新型小鼠模型和体外系统。此外，导致基质材料性质改变的TGF-β和Runx 2作用的靶点仍有待确定。几个TGF-β和Runx 2调节的基因产物有可能影响基质材料特性，包括非胶原性骨基质蛋白、蛋白酶和其他信号分子。目的3旨在确定TGF-β和Runx 2调节基质材料性质的必要下游靶点，使用体内和体外方法的组合。总的来说，拟议的研究试图验证TGF-β利用Smad 3和非Smad 3机制来差异控制Runx 2成骨细胞基因的激活的假设，这反过来又直接和间接地通过调节破骨细胞介导的骨重建来建立基质材料特性。这项研究将阐明骨基质材料特性在骨骼健康和疾病中的作用。公共卫生相关性：对控制骨基质质量的机制的研究将导致预防骨骼疾病和改善骨骼组织修复的策略，特别是颅面骨和牙齿。该项目将研究参与骨基质质量调节的两个特定因子（TGF-β和Runx 2）。这将导致对它们在健康骨骼中的调节或在疾病过程中的误调节进行更仔细的检查。

DESCRIPTION (provided by applicant): Bone matrix is an organic and mineral composite that confers toughness and hardness to bone. The material properties of this matrix may predict bone fracture risk and could complement clinical fracture risk assessment, which currently relies on bone mineral density. Matrix material properties, like other aspects of bone quality, are biologically determined and anatomically distinct. However, little is known about their regulation in healthy bone or their misregulation in disease. The long-term goal of this research is to identify mechanisms that control matrix material properties, so that ultimately, they can be harnessed therapeutically to prevent skeletal disease and improve craniofacial bone repair. Transforming growth factor-¿ (TGF-¿) was recently identified as the first growth factor to regulate bone matrix material properties. TGF-¿ inhibits osteoblast gene expression and differentiation, as well as bone matrix material properties, by repressing the function of Runx2, a key osteoblast transcription factor. Although Runx2 integrates signals from many pathways, only TGF-¿ and glucocorticoids have yet been shown to regulate matrix material properties. Therefore, this proposal aims to investigate cellular and molecular mechanisms by which TGF-¿ and Runx2 regulate matrix material properties. Osteoblasts are required for the regulation of matrix material properties by TGF-¿. Whether osteoblasts modify matrix material properties directly, or indirectly by cooperating with osteoclasts to change bone remodeling activity remains unclear. Using genetic and pharmacologic tools to manipulate osteoclast and osteoblast function in mice, Aim 1 seeks to determine the relative roles of both cell types in the specification of matrix material properties. Bone matrix material properties and mineral concentration will be evaluated by nanoindentation, fracture toughness testing, and X-ray tomographic microscopy. TGF-¿ can repress or activate the expression of Runx2, apparently by differential utilization of downstream signaling pathways. Since the level of Runx2 function affects matrix material properties, Aim 2 seeks to identify mechanisms by which TGF-¿ signaling regulates Runx2 to control osteoblast differentiation and matrix material properties. Novel mouse models and in vitro systems will be used. Furthermore, the targets of TGF-¿ and Runx2 action that result in altered matrix material properties remain to be determined. Several TGF-¿ and Runx2-regulated gene products have the potential to affect matrix material properties, including non-collagenous bone matrix proteins, proteases, and other signaling molecules. Aim 3 seeks to identify essential downstream targets through which TGF-¿ and Runx2 regulate matrix material properties, using a combination of in vivo and in vitro approaches. Overall, the proposed studies seek to test the hypothesis that TGF-¿ utilizes Smad3 and non-Smad3 mechanisms to differentially control Runx2 activation of osteoblast genes, which in turn establish matrix material properties both directly, and indirectly through the regulation of osteoclast-mediated bone remodeling. This research will elucidate the role of bone matrix material properties in skeletal health and disease. PUBLIC HEALTH RELEVANCE: Relevance Research into the mechanisms that control the bone matrix quality will lead to strategies to prevent skeletal disease and improve skeletal tissue repair, specifically craniofacial bones and perhaps teeth. This project will investigate two specific factors (TGF-¿ and Runx2) that participate in the regulation of bone matrix quality. This will lead to a closer examination of their regulation in healthy bone or their misregulation during disease processes.