Cellular and molecular mechanisms regulating synovial joint development

调节滑膜关节发育的细胞和分子机制

• 批准号: 10454425
• 负责人: Minwook Kim
• 金额: $ 7.32万
• 依托单位: CHILDREN'S HOSP OF PHILADELPHIA
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-08-01 至 2023-05-01
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10454425
• 关键词: Ablation; Adult; Anatomy; Appearance; Back; Biochemical; Biochemistry; Biological Assay; Biological Process; Biology; Biomedical Engineering; CD44 gene; Cartilage; Cell Adhesion Molecules; Cell Line; Cell Separation; Cell surface; Cells; Complement; Cytoskeletal Modeling; Data; Degenerative polyarthritis; Development; Developmental Biology; Digit structure; Disease; Embryo; Embryonic Development; Engineering; Enzymes; Extracellular Matrix; GDF5 gene; HAS2 gene; Homeostasis; Hyaluronan; Hyaluronic Acid; Hyaluronidase; Immobilization; Impairment; In Situ Hybridization; In Vitro; Individual; Inflammatory; Injections; Integral Membrane Protein; Joint by Site; Joints; Knee; Knee joint; Lead; Ligaments; Limb structure; Link; Liquid substance; LoxP-flanked allele; Lubricants; MAP2K1 gene; Maintenance; Mentored Research Scientist Development Award; Mentors; Mesenchymal; Metabolism; Molecular; Molecular Biology; Molecular Genetics; Molecular Weight; Morphology; Motion; Movement; Mus; Natural regeneration; Pathway interactions; Phenotype; Platelet-Derived Growth Factor beta Receptor; Play; Process; Property; Quality of life; Recombinants; Research; Role; Shapes; Shoulder; Signal Pathway; Signal Transduction; Site; Spectroscopy, Fourier Transform Infrared; Stress Fibers; Structure; Surface; Synovial Fluid; Synovial joint; System; Testing; Therapeutic; Time; Tissues; Toll-like receptors; Training; articular cartilage; base; capsule; career; cartilaginous; clinically relevant; fetal; growth differentiation factor 5; infrared spectroscopy; insight; joint formation; joint function; lubricin; mouse model; mutant; novel; novel therapeutics; physical separation; prospective; rational design; receptor; repaired; response; skeletal; skills; stem cell function; sugar; therapy design; tool; trait; transcriptome sequencing

Project Summary and Abstract Synovial joints are essential for body motion and quality of life. Their synovial cavity and lubricant-rich fluid permit unhindered joint motion and function and provide tissue protection and nourishment. While these aspects of synovial joint biology are well understood, little is known about how the cavity and its fluid actually develop during embryogenesis. At early fetal stages, the limb skeletal primordia are composed of continuous cartilaginous structures without joints. Joint development starts with appearance of an “interzone”, a tissue made of mesenchymal cells expressing the growth and differentiation factor 5 gene (Gdf5). We previously showed that Gdf5+ cell progenies produce most joint tissues over time and the synovial cavity forms in the middle of the interzone. Because the interzone cells are initially attached to each other, the cavitation process must involve their physical separation along the prospective articular line to facilitate the creation of a fluid-filled cavity. Previous studies indicated that interzone cells produce hyaluronan (HA) around the cavitation time, and this is accompanied by accumulation of a HA-rich matrix in local tissues. HA is a major component of extracellular matrix and synovial fluid and plays important roles in tissue homeostasis. In my preliminary studies, I found that just before cavitation onset, interzone cells in mouse embryo limbs express hyaluronan synthase 2 (HAS2, ‘the HA synthesizer’) and transmembrane protein 2 (TMEM2), a cell surface hyaluronidase that specifically cleaves high molecular weight HA into intermediate and biologically-active fragments. I also discovered that, morphologically, cavitation initiates with formation of microlumens along the prospective articular line and is completed soon afterwards when the pockets coalesce to generate a single one synovial cavity. This process is extremely rapid in the developing knee but is slower in digits. These and other novel data lead to my central hypothesis that joint cavitation is brought about by convergence of diverse but coordinated biological processes. Accordingly, Aim 1 is to determine the role of HAS2 and TMEM2 in joint cavitation using genetically modified mouse models. I will conditionally delete Has2 and/or Tmem2 in interzone cells (using Gdf5Cre mice) and subject resulting mutant embryos to detailed analysis. Aim 2 is to determine cellular and molecular mechanisms of cavitation. I will investigate downstream signaling pathways in response to changes in HA sizes and resulting interactions with cell surface CD44 receptor, regulating HA metabolism in synovial joint development and long-term maintenance. The project will provide novel insights into mechanisms underlying joint development and cavitation. In line with the K01 mechanism, the project will allow me to acquire new expertise in skeletal developmental and molecular biology and to integrate it with my previous training in bioengineering. This unique combination of expertise in two distinct but interrelated fields will allow me to establish an independent career distinct from my mentors and create novel therapeutic tools to repair and regenerate cartilage for the treatment of joint conditions such as osteoarthritis (OA).

项目摘要和摘要 滑膜关节对身体运动和生活质量是必不可少的。他们的滑膜腔和富含润滑剂的液体 允许不受阻碍的关节运动和功能，并提供组织保护和营养。而这些 滑膜关节生物学的各个方面都已被很好地了解，但对空洞及其液体实际上是如何形成的知之甚少。 在胚胎发育过程中发育。在胎儿早期，四肢骨骼原基由连续的 无关节的软骨结构。关节发育始于“中间带”的出现，即一种组织 由表达生长分化因子5基因(GDF5)的间充质细胞制成。我们之前 结果显示，随着时间的推移，Gdf5+细胞后代产生的关节组织最多，滑膜腔在 在中间地带。因为区域间细胞最初彼此连接，所以空化过程 必须将它们沿预期的关节线进行物理分离，以便于产生充满液体的 空洞。以前的研究表明，带间细胞在空化时间附近产生透明质酸(HA)，并且 伴随而来的是局部组织中富含HA的基质的积聚。房委会是一个主要组成部分 细胞外基质和滑液，在组织内稳态中发挥重要作用。在我的初赛中 研究发现，就在空化开始之前，小鼠胚胎四肢中的带间细胞表达透明质酸 合酶2(HAS2，HA合成器)和跨膜蛋白2(TMEM2)，一种细胞表面透明质酸酶 这会将高分子量的HA特异性地裂解成具有生物活性的中间片段。我也是 发现，在形态上，空化始于沿预期方向的微腔的形成。 关节线，完成后不久，口袋结合在一起，形成一个单一的滑膜 空洞。这一过程在发育中的膝盖非常迅速，但在手指上则较慢。这些和其他小说 这些数据引出了我的中心假设，即关节空化是由不同但 协调的生物过程。因此，目标1是确定HAS2和TMEM2在关节中的作用。 利用转基因小鼠模型进行空化。我将有条件地删除区域间的Has2和/或Tem2 细胞(使用Gdf5Cre小鼠)，并对产生的突变胚胎进行详细分析。目标2是确定 空化的细胞和分子机制。作为回应，我将研究下游信号通路 与HA大小的变化以及由此产生的与细胞表面CD44受体的相互作用，调节HA的代谢 滑膜关节发育和长期维护。该项目将提供对机制的新见解 潜在的节理发育和空化。根据K01机制，该项目将使我能够 在骨骼发育和分子生物学方面获得新的专业知识，并将其与我以前的 生物工程方面的培训。这两个不同但相互关联的领域的独特专业知识组合将使 我要建立一个独立的职业生涯，不同于我的导师，并创造新的治疗工具来修复 并再生软骨，用于治疗骨关节炎(OA)等关节疾病。