Tracing the origins of craniofacial growth plates

项目总结 骨骼的生长发生在称为生长板和缝合线的专门位置。在那里，这些增长 这些部位的位置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。 影响人类骨骼或软骨生长的骨骼发育不良的流行情况(估计影响 大约每4,000-5,000个新生儿中就有一个)推动了对调节途径的大量研究 生长板和缝合线的发育和功能。然而，该领域仍然缺乏对 决定这些关键部位在骨骼中形成位置的机制。特别是生长板 通常被认为是在软骨内骨化中心之间被动形成的。然而，a 斑马鱼、小白鼠和小鸡的细胞极性数据的散布表明，生长板可能是预先形成的 在骨化开始之前很久就在软骨模板中，这表明有一个活跃的图案化过程。这 应用程序提出了生长板起源的新假说，将在斑马鱼身上使用创新的 命运映射策略新应用于骨骼系统。 生长板存在于头面部骨骼的软骨内骨以及长骨中。 四肢。在头部骨骼或软骨分化开始之前，面部的骨骼祖细胞 根据基因在背腹(DV)轴上的位置表达不同的基因序列。这些域 并不是完全互斥的：一系列共同表达邻近细胞特征的标记的细胞 通常可以在每个边界上检测到域。重要的是，这些边界的位置会出现 与成人颅骨生长板和软骨关节的最终分布相关。这 观察引发了一种假设，即颅骨中的软骨生长带注定是在 这些分子边界。验证这一假设需要追踪胚胎边界细胞的命运。 为成年干杯。为此，一种未得到充分利用但功能强大的交叉性分裂-整合-CRE血统追踪方法 将被用来特异性地标记相邻DV结构域的共表达标记。先导性研究 这里概述的将提供对这一原始假设的第一次测试，验证方法，并创建 试剂为这项工作的未来延伸，都是未来R01级支持的先决条件。 这项建议直接与NIDCR关于调查开发、维护、 以及头面部组织的重塑。虽然目前的范围仅限于头骨，但根据 这项研究也可能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刺激对支配骨骼的逻辑的理解取得重大进展 花纹，与人类颅面畸形和其他骨骼发育不良明显相关。

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.