Rspondin-Lgr Axis in Bone Regeneration

骨再生中的 Rspondin-Lgr 轴

• 批准号: 10261766
• 负责人: Kurt David Hankenson
• 金额: $ 40.01万
• 依托单位: UNIVERSITY OF CONNECTICUT SCH OF MED/DNT
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-09-18 至 2022-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10261766
• 关键词: Address; Adult; Attenuated; Binding; Bone Injury; Bone Regeneration; Bone callus; Calvaria; Cell Compartmentation; Cell Lineage; Cells; Chondrocytes; Clinical; Data; Development; Embryo; Family; Family member; Femoral Fractures; Fracture; Fracture Healing; Funding; GPR4 gene; Genes; Goals; Grant; Hand; Histology; In Vitro; Incidence; Injury; Knockout Mice; Lead; Leucine-Rich Repeat; Ligands; Limb Development; Limb structure; LoxP-flanked allele; Marrow; Mechanics; Mesenchymal; Mesenchymal Stem Cells; Mesenchyme; Minerals; Modeling; Molecular; Mus; Natural regeneration; Osteoblasts; Osteogenesis; Pathway interactions; Periosteal Cell; Periosteum; Play; Positioning Attribute; Process; Property; Proteins; Public Health; Publishing; Receptor Signaling; Regulation; Research; Role; Signal Transduction; Skeletal bone; Source; Testing; Therapeutic Uses; Time; WNT Signaling Pathway; base; bone; bone healing; cranium; experimental study; healing; in vivo; injured; insight; long bone; mRNA Expression; mouse model; novel; novel strategies; osteoblast differentiation; osteochondral tissue; osteogenic; osteoprogenitor cell; parent grant; prevent; progenitor; receptor; regenerative therapy; repaired; response; response to injury; skeletal; stem cell proliferation; stem cells; tissue regeneration

Our funded grant, R01DE030716, ”Rspondin-Lgr Axis in Bone Regeneration”, is focused on examining the role of Rspo2-Lgr6 signaling in calvarial regeneration. While there are some similarities in healing processes that exist between long bones and bones of the skull, there are also important differences. As an example, the calvaria heals exclusively through an intramembranous process while long bones heal through both intramembranous and endochondral bone formation. An additional important difference that exists between calvarial and long bone healing is the source and type of progenitor cell that contributes to bone healing. In long bones, a critical aspect of the bone healing process begins with the expansion of mesenchymal progenitors from the marrow and periosteum, which occurs immediately after injury. These cells then become bone forming osteoblasts and chondrocytes. However, mechanisms that control long-bone skeletal progenitor/stem cell activation, expansion, and differentiation in response to injury are poorly described. This supplement will study the role of the R-spondin (ligand) – Lgr (receptor) signaling axis in regulating long bone progenitors and bone regeneration. R-spondins (roof plate specific spondin) are a family of four secreted matricellular proteins (Rspo1-4) that bind to Leucine-rich repeat-containing G-protein coupled receptors 4/5/6 (Lgrs). Rspo-Lgr interaction potentiate canonical Wnt pathway by preventing the turnover of Wnt Frizzled receptors, and hence determines canonical Wnt signaling levels. While canonical Wnt signaling is known to play an important role in bone regeneration, very little research has explored positive modulators of Wnt signaling. In particular, the requirement of Rspo-Lgr in the context of long-bone fracture healing has never been examined, due to the lack of appropriate models. Our primary goal is to define the requirement of Rspo2/3 and Lgr6 in mesenchymal progenitors in response to femoral fracture. We have defined two specific aims to address this goal. In Aim1, we will use single and compound Rspo2 and Rspo3 floxed mice crossed with an alphaSMACreERT2 mouse to disrupt the Rspo2/3 genes in long-bone mesenchymal progenitors at the time of fracture. We will assess in vivo proliferation, Wnt signaling, and osteogenesis of progenitors, and analyze healing by using µCT, histology, and mechanical testing. In Aim 2, Lgr6 knockout mice will be investigated for their bone healing properties using parameters similarly to Aim 1. Completion of this project will identify the requirement of Rspo2/3-Lgr6 interaction in long-bone fracture healing and will provide new mechanistic insights into the action of cWnt signaling in bone healing that can be directly compared to similar experiments conducted with calvarial healing in the parent grant.

我们的资助基金，R 01 DE 030716，“骨再生中的Rspondin-Lgr轴”，专注于研究 Rspo 2-Lgr 6信号在颅骨再生中的作用。虽然在愈合过程中有一些相似之处， 除了头骨和长骨之间存在的差异外，还有重要的区别。为例 颅骨完全通过膜内过程愈合，而长骨通过两者愈合 膜内和软骨内骨形成。另一个重要的区别是， 颅骨和长骨愈合是有助于骨愈合的祖细胞的来源和类型。在 长骨，骨愈合过程的一个关键方面始于间充质细胞的扩张， 骨髓和骨膜的祖细胞，这在损伤后立即发生。然后这些细胞变成 形成骨的成骨细胞和软骨细胞。然而，控制长骨骨骼的机制 祖细胞/干细胞响应损伤的活化、扩增和分化的描述很少。这 补充剂将研究R-spondin（配体）- Lgr（受体）信号传导轴在调节长骨中的作用 祖细胞和骨再生。R-spondins（顶板特异性spondin）是一个由四个分泌的家族， 与含有富含亮氨酸重复序列的G蛋白偶联受体4/5/6结合的基质细胞蛋白（Rsp 1 -4） （Lgrs）。Rspo-Lgr相互作用通过阻止Wnt Frizzled的周转来加强经典Wnt途径 受体，并因此决定典型的Wnt信号传导水平。虽然已知典型的Wnt信号传导 在骨再生中发挥重要作用，但很少有研究探索Wnt的正性调节剂 发信号。特别是，在长骨骨折愈合的背景下，Rspo-Lgr的需求从未被证实。 由于缺乏合适的模型，因此进行了研究。我们的主要目标是定义 间充质祖细胞中的Rspo 2/3和Lgr 6对股骨骨折的反应我们定义了两个具体的 旨在实现这一目标。在Aim 1中，我们将使用单个和复合Rspo 2和Rspo 3 floxed小鼠杂交 用alphaSMACreERT 2小鼠破坏长骨间充质祖细胞中的Rspo 2/3基因， 骨折的时间我们将评估祖细胞的体内增殖、Wnt信号传导和成骨作用，并 使用µCT、组织学和机械测试分析愈合情况。在目标2中，Lgr 6敲除小鼠将被 使用与目标1类似的参数研究其骨愈合特性。该项目的完成将 确定Rspo 2/3-Lgr 6相互作用在长骨骨折愈合中的需求，并将提供新的 对cWnt信号传导在骨愈合中的作用的机械见解，可以直接与类似的 在父母补助金中进行的颅骨愈合实验。