Tracing the origins of craniofacial growth plates

追踪颅面生长板的起源

• 批准号: 10413816
• 负责人: Lindsey Anne Barske
• 金额: $ 15.9万
• 依托单位: CINCINNATI CHILDRENS HOSP MED CTR
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-07-01 至 2024-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10413816
• 关键词: ATAC-seq; Adult; Affect; Birth; Cartilage; Cell Line; Cell Polarity; Cells; Characteristics; Chick; Chondrocytes; Craniofacial Abnormalities; Cre lox recombination system; Data; Development; Dorsal; Embryo; Enhancers; Epiphysial cartilage; Face; Future; Gene Expression; Genes; Growth; Head; Human; Image; Joint structure of suture of skull; Joints; Label; Location; Logic; Maintenance; Mammals; Maps; Methodology; Methods; Modeling; Molecular; Mus; National Institute of Dental and Craniofacial Research; Neural Crest; Pathway interactions; Pattern; Physiologic Ossification; Pilot Projects; Population; Positioning Attribute; Prevalence; Process; Property; RNA; Reagent; Reporter; Research; Research Design; Shapes; Site; Skeletal system; Skeleton; Specific qualifier value; Specificity; Surgical sutures; Testing; Tissues; Transgenes; Transgenic Organisms; Work; Zebrafish; apoAI regulatory protein-1; bone; cartilaginous; clinically relevant; cohort; craniofacial; craniofacial bone; craniofacial tissue; cranium; emx2 protein; experimental study; innovation; intein; limb bone; long bone; novel; osteoprogenitor cell; postnatal; prevent; promoter; reconstitution; single-cell RNA sequencing; skeletal; skeletal dysplasia; stem cells; transcription factor

PROJECT SUMMARY Growth of the skeleton occurs at specialized sites called growth plates and sutures. Where these growth sites are located and how long they persist postnatally determine the final shape and size of the skeleton. The prevalence of skeletal dysplasias affecting bone or cartilage growth in humans (estimated to impact approximately one in 4,000-5,000 births) has motivated a vast body of research into the pathways regulating growth plate and suture development and function. However, the field still lacks a basic understanding of the mechanisms that determine where these critical sites form within the skeleton. Growth plates in particular have been typically assumed to form passively between centers of endochondral ossification. However, a scattering of cell polarity data from zebrafish, mice, and chick indicate that growth plates may be prefigured in the cartilage template well before ossification begins, suggesting an active patterning process. This application presents a novel hypothesis for growth plate origins, to be tested in zebrafish using an innovative fate-mapping strategy newly applied to the skeletal system. Growth plates are present in endochondral bones of the craniofacial skeleton as well as the long bones of the limbs. Prior to the onset of bone or cartilage differentiation in the head, skeletal progenitors for the face express distinct cohorts of genes depending on their position on the dorsal-ventral (DV) axis. These domains are not entirely mutually exclusive: a line of cells co-expressing markers characteristic of the neighboring domains can typically be detected at each boundary. Importantly, the positions of these boundaries appear to correlate with the eventual distribution of growth plates and cartilaginous joints in the adult skull. This observation prompted the hypothesis that cartilaginous growth zones in the skull are destined to form at these molecular boundaries. Testing this hypothesis requires following the fate of embryonic boundary cells to adulthood. To this end, an underutilized but powerful intersectional split-Intein-Cre lineage-tracing method will be used to specifically label cells co-expressing markers of adjacent DV domains. The pilot study outlined here will provide the first test of this original hypothesis, validate the methodology, and create reagents for future extensions of this work, all prerequisites for future R01-level support. This proposal directly aligns with the NIDCR's call to investigate mechanisms of development, maintenance, and remodeling of craniofacial tissues. Though the scope is currently limited to the skull, the principles under study may also apply to many other parts of the endochondral skeleton and possibly also to cranial sutures. Results from this work could stimulate a major advance in understanding the logic governing skeletal patterning, with clear relevance for human craniofacial malformations and other skeletal dysplasias.

项目摘要 骨骼的生长发生在称为生长板和缝合线的专门部位。这些增长 位置的确定以及它们在出生后持续的时间决定了骨骼的最终形状和大小。 影响人类骨或软骨生长的骨骼发育不良的患病率（估计影响 大约每4000 - 5000名新生儿中就有一名）已经激发了大量的研究，研究调节 生长板和缝的发育和功能。然而，该领域仍然缺乏对以下方面的基本了解： 决定这些关键部位在骨骼中形成的机制。尤其是增长板块 通常被认为是在软骨内骨化中心之间被动形成的。但 从斑马鱼、小鼠和小鸡身上获得的细胞极性数据的分散表明，生长板可能是预先形成的， 在骨化开始之前的软骨模板中，这表明了一个活跃的图案化过程。这 应用程序提出了一个新的假设生长板的起源，将在斑马鱼进行测试，使用创新的 新应用于骨骼系统的命运映射策略。 生长板存在于颅面骨骼的软骨内骨以及 四肢在头部的骨或软骨分化开始之前， 根据它们在背腹（DV）轴上的位置，表达不同的基因组。These domains 并不完全相互排斥：一系列细胞共表达邻近细胞的特征性标记物， 通常可以在每个边界处检测域。重要的是，这些边界的位置 与成人头骨中生长板和软骨关节的最终分布相关。这 这一观察提示了一种假设，即颅骨中的软骨生长区注定要在 这些分子边界。验证这一假设需要跟踪胚胎边界细胞的命运 到成年为此，一种未充分利用但功能强大的交叉分裂-内含肽-Cre谱系追踪方法 将用于特异性标记共表达相邻DV结构域标记的细胞。试点研究 这里概述的将提供对这个原始假设的第一次检验，验证方法，并创建 本工作未来扩展的试剂，未来R 01级支持的所有先决条件。 这一建议直接符合NIDCR的呼吁，调查机制的发展，维护， 以及颅面组织的重塑虽然目前的范围仅限于头骨，但根据 这项研究也适用于软骨内骨骼的许多其他部分，也可能适用于颅缝。 这项工作的结果可能会促进对骨骼肌控制逻辑的理解 模式，与人类颅面畸形和其他骨骼发育不良有明确的相关性。