Influence of Genetic Background on Bone Anabolic Response to Mechanical Loading

摘要 骨质疏松症是一种骨量低、骨折风险增加的遗传性疾病。药物可用于 治疗低骨量，但它们有风险，适应症有限。因此，仍然需要新的 防治骨质疏松症的途径。机械负荷可以建立和维持骨量。 对人类来说是一种强效的骨合成刺激物质。对小鼠骨骼负荷的研究提供了一种 在基因、细胞和组织水平上研究骨合成代谢的强大平台。然而，到目前为止的研究一直是 受缺乏遗传多样性的限制-大多数小鼠骨负荷研究都是利用近交系进行的 C57BL/6(B6)小鼠，以及少数几个使用多个品系的小鼠使用的是遗传多样性有限的小鼠。因此， 关于遗传背景对骨骼机械合成代谢反应的贡献，我们知之甚少。 正在装车。我们的目标是检验总体假设，即骨骼对机械载荷的反应 可遗传性(即取决于遗传背景)。我们将使用来自八个不同基因的小鼠来测试这一点 近交系小鼠品系，包括5个常见的实验室品系和3个野生来源品系。这些菌株 解释了小鼠基因组变异的%。在目标1中，我们将应用应变匹配的胫骨负荷 对8个近交系的幼年-成年小鼠，并测定皮质骨形成反应。我们会 包括雌鼠和雄鼠，以评估基因和性别主效应，以及性别基因 互动。然后，我们将对被鉴定为低反应者和高反应者的小鼠品系进行rna-seq分析。 以探索可能驱动差异加载反应的转录差异。骨骼 负荷驱动骨细胞陷窝管网络(LCN)中的液体流动，人们普遍认为 骨细胞转导这种液体流动刺激，调节对负荷的合成代谢反应。对 LCN随着骨质疏松和衰老而发生，并可能改变骨细胞的微机械环境，在 将合成代谢反应转为负荷。在目标2中，我们建议表征骨骼中LCN的形态 8个近交系菌株，并测试这些特性是否在不同品系之间存在差异。我们将使用亚微米 分辨率X射线显微镜和共聚焦成像，以表征骨细胞LCN。然后我们将探索 LCN形态的菌株间变异是否与合成代谢反应的变异有关 菌株之间的加载(来自目标1)。这项建议适合作为R21探索性/开发性 格兰特。目标1是发育阶段的；它将确定骨骼对负荷的合成代谢反应是否可遗传。如果 因此，这将为未来利用遗传多样性的小鼠发现基因的大规模研究提供理论基础 这会影响骨骼对负荷的反应。目的2将评估骨细胞LCN形态的遗传力， 这可能同样会激励未来的研究，以确定因果基因。它还将探索一个基本的 骨力学生物学中的问题--骨对负荷的反应是否取决于骨细胞LCN 形态学。

SUMMARY Osteoporosis is a heritable disease of low bone mass and increased fracture risk. Drugs are available to treat low bone mass, yet they carry risks and have limited indications. Thus, there remains a need for new approaches to prevent and treat osteoporosis. Mechanical loading can build and maintain bone mass in humans, and is a potent bone anabolic stimulus in animals. Studies of skeletal loading in mice provide a powerful platform to study bone anabolism at the gene, cell and tissue levels. Yet studies to date have been limited by a lack of genetic diversity – most murine bone loading studies have been done using inbred C57Bl/6 (B6) mice, and the few done with multiple strains used mice with limited genetic diversity. Thus, little is known about the contribution of genetic background to bone's anabolic response to mechanical loading. Our goal is to test the overall hypothesis that the response of bone to mechanical loading is heritable (i.e., depends on genetic background). We will test this using mice from eight genetically diverse inbred mouse strains, including five common laboratory strains and three wild-derived strains. These strains account for 89% of the variation in the mouse genome. In Aim 1, we will apply strain-matched tibial loading to young-adult mice of the eight inbred strains, and determine cortical bone formation responses. We will include female and male mice, to allow assessment of genotype and sex main effects, and sex-genotype interactions. We will then perform RNA-seq analysis on mouse strains identified as low and high responders in order to explore the transcriptomic differences that may drive the differential loading responses. Skeletal loading drives fluid flow in the osteocyte lacunocanalicular network (LCN), and it is widely believed that osteocytes transduce this fluid flow stimulus and regulate the anabolic response to loading. Changes to the LCN occur with osteoporosis and aging, and may alter the osteocyte micromechanical environment, and in turn the anabolic response to loading. In Aim 2, we propose to characterize LCN morphology in bones from the eight inbred strains, and test whether these traits vary between strains. We will use sub-micron resolution X-ray microscopy and confocal imaging to characterize the osteocyte LCN. We will then explore whether inter-strain variations in LCN morphology are associated with variations in anabolic response to loading between strains (from Aim 1). This proposal is appropriate as an R21 Exploratory/ Developmental grant. Aim 1 is developmental; it will establish whether bone's anabolic response to loading is heritable. If so, it will provide rationale for large-scale future studies using genetically diverse mice to discover genes that influence bone's response to loading. Aim 2 will assess the heritability of osteocyte LCN morphology, which may likewise motivate future studies to identify causal genes. It will also explore a fundamental question in bone mechanobiology – whether bone's response to loading depends on osteocyte LCN morphology.