Mechanisms of Synovial Joint Formation

描述(申请人提供)：滑膜关节对于骨骼功能和生活质量是必不可少的，人们对其解剖组织、不同的组织以及对包括年龄相关性骨关节炎在内的常见病理的易感性都有很多了解。相比之下，人们对它们的发育生物学知之甚少。如果有这样的信息，它可以直接使用或与生物工程工具结合使用，以创造新的关节修复和再生策略，例如具有特定关节组织形成能力的工程干细胞。这个项目是五年前开始的，目的是填补信息方面的这种明显空白。在四肢，滑膜关节的形成始于在每个预期的关节部位出现间充质细胞(统称为间带)，两侧是软骨长骨陷窝。然而，长期以来，人们一直不知道带间细胞是如何获得其基本的间充质特征的，它们是否作为一个关键的但短暂的关节分界点，和/或它们是否随着时间的推移实际上产生了关节组织。为了解决这些基本问题，我们对小鼠出生前和出生后的区域间细胞进行了遗传学追踪。值得注意的是，我们发现这些细胞根本不是暂时性的，而是产生关节组织，并且只产生关节组织，因此，构成了一个具有关节形成能力的多能祖细胞的专门队列。我们发现，在早期阶段，细胞表现出很强的Wnt/？-catenin信号。随着关节发育的进展，关节软骨表层的信号逐渐减弱，但仍较强(对关节功能至关重要，具有独特的纤维-软骨表型)。事实上，有条件的？-连环蛋白消融导致了该区域的几乎丧失。更多的初步数据表明，Wnt/？-catenin途径不是单独作用的，而是与维甲酸途径和核维甲酸受体(RARs)合作维持和调节关节的形成。这些数据导致了这种竞争性更新应用的中心假设，即维甲酸和Wnt/？-catenin信号通路建立了早期带间细胞的多潜能间充质特征，然后受到地形的调节，以允许形成不同的关节组织。信号活性的维持将允许带间细胞产生纤维和纤维软骨关节组织，而信号的下降将允许关节软骨的形成。我们的目标是：(1)进一步研究维甲酸信号在区域间功能和关节形成中的作用；(2)确定两条信号通路相互作用调节区域间细胞功能和关节形成的分子机制；(3)确定区域间细胞的可塑性和分化潜能以及维甲酸和Wnt/β-catenin信号的调节作用。这项继续提案中提出的工作结合了关于基本调控机制的实验线和测试区间细胞的可塑性、可移植性和发展潜力的线。因此，它将继续产生对基本生物学知识具有基本价值的信息，并将对翻译医学和创造未来新的关节修复和再生疗法具有重要意义。

DESCRIPTION (provided by applicant): Synovial joints are essential for skeletal function and quality of life and much is known about their anatomical organization, distinct tissues and susceptibility to common pathologies including age-dependent osteoarthritis. In contrast, very little is known about their developmental biology. Were such information available, it could be used directly or in combination with bioengineering tools to create new joint repair and regenerative strategies such as engineered stem cells with specific joint tissue-formation capacity. This project started five years ago to fill such glaring gaps in information. In the limbs, synovial joint formation initiates with the appearance of mesenchymal cells (collectively called the interzone) at each prospective joint site flanked by cartilaginous long bone anlaga. However, it had long remained unknown how the interzone cells acquire their essential mesenchymal character, whether they serve as a critical but transient joint demarcation point, and/or whether they actually produce joint tissues over time. To address these fundamental questions, we genetically traced and tracked the interzone cells prenatally and postnatally in mice. Strikingly, we found that the cells are not at all transient but produce joint tissues and only joint tissues and thus, constitute a specialized cohort of multipotent progenitor cells endowed with joint formation capacity. We showed that at early stages the cells exhibit strong Wnt/?-catenin signaling. As joint development progresses, signaling dwindled but remained strong in the superficial zone of articular cartilage (critical for joint function and characterized by a unique fibro-cartilaginous phenotype). Indeed, conditional ?-catenin ablation caused a near loss of that zone. Additional Preliminary Data now indicate that the Wnt/?-catenin pathway does not act alone, but cooperates with the retinoid pathway and nuclear retinoic acid receptors (RARs) to sustain and regulate joint formation. The data lead to the central hypothesis for this competitive renewal application that the retinoid and Wnt/?- catenin signaling pathways establish the multipotent mesenchymal character of early interzone cells and are then topographically modulated to permit formation of distinct joint tissues. Maintenance of signaling activity would allow interzone cells to produce fibrous and fibro-cartilaginous joint tissues, while a drop in signaling would allow formation of articular cartilage. Our Aims are: (1) to further characterize the roles of retinoid signaling in interzone function and joint formation; (2) to determine molecular mechanisms by which the two signaling pathways interact to regulate interzone cell function and joint formation; and (3) to determine interzone cells' plasticity and differentiation potentials and regulation by retinoid and Wnt/?-catenin signaling. The work proposed in this continuation proposal combines experimental lines on basic regulatory mechanisms with lines testing plasticity, transplantability and developmental potentials of interzone cells. It will thus continue to produce information of fundamental value to basic biologic knowledge and will also significant relevance for translation medicine and creation of future novel joint repair and regeneration therapies.