Elucidating Mechanisms of Mechanosensitivity During Secondary Chondrogenesis

描述(申请人提供)：在生命的所有阶段，骨骼都被优化以检测和适应生物力学的力量。当维持骨骼健康的微妙平衡被疾病或伤害破坏时，个人的生活质量可能会迅速恶化。该项目的目标是确定使骨骼能够感知机械力并对其做出反应的机制。次级软骨是一种高度协调的检测机械力的骨骼组织。次级软骨最初在颌骨骨骼区域发育，以响应胚胎运动过程中产生的力。在没有适当的机械力的情况下，次级软骨不能形成，也可以在任何时候退化，就像在颞下颌关节紊乱病(TMD)和颌骨不动的患者中一样。虽然次级软骨是颌骨功能所必需的，但对诱导和维持次级软骨的分子机制知之甚少。为了解决这个问题，目前的提案采用了一个鸟类模型系统，该系统利用物种特定的差异，利用次级软骨进化的方式来支持专门的喂养模式。鸭子通过用它们的下巴铲取和过滤湿沉积物来进食。甚至在孵化之前，鸭的下颌内收肌吻合处就出现了次级软骨，它向外侧插入，从而极大地延长了冠突。这在下颌内收肌的肌腱和下颌之间创造了一个坚固的界面，并传递了抬起下颌所必需的强大的收缩。相比之下，雏鸟主要通过啄食种子的方式进食，它们的下颌内收肌沿着下颌冠突的背侧插入，没有任何次级软骨。这些关键的区别在鸭子和鸡的胚胎中很明显，即使在胚胎的颌骨运动方面没有显著的差异。这表明，物种特有的颌部结构产生了在鸭子中存在的机械力，而不是在鸡中，导致了发育过程中机械敏感信号通路的差异激活。根据已发表的初步数据，我们推测成纤维细胞生长因子(成纤维细胞生长因子)和钙(钙)信号在下颌内收肌诱导过程中发挥作用，使其能够检测生物力并产生次级软骨。目的1通过实验确定成纤维细胞生长因子和钙离子信号在继发性软骨形成中是否必需。浸泡在成纤维细胞生长因子和钙信号的小分子抑制剂中的珠子将被植入覆盖在假定的鸭冠突上皮下。Aim 2中的实验将揭示通过使用成纤维细胞生长因子和钙信号激动剂，成纤维细胞生长因子和钙信号是否足以促进鸡的继发性软骨形成。Aim 3的实验将使用鸡和鸭的嵌合体来确定当在鸭子环境中时，鸡细胞是否有能力形成次级软骨。了解调控继发性软骨形成的机制将导致对继发性软骨丧失的再生治疗，如TMD和创伤后发生的情况。

DESCRIPTION (provided by applicant): Through all stages of life, the skeleton is optimized for detecting and adapting to biomechanical forces. When the delicate balance that maintains skeletal health is disrupted by disease or injury, an individual's quality of life can deteriorate rapidly. The goal of this project is to identify mechanisms that allow the skeleton to sense and respond to mechanical forces. One skeletal tissue that is highly attuned to detecting mechanical force is secondary cartilage. Secondary cartilage initially develops on regions of bone in the jaw skeleton in response to forces arising during embryonic motility. In the absence of proper mechanical forces, secondary cartilage fails to form, and can also degenerate at any point as in temporomandibular disorders (TMD) and in patients with immobilized jaws. Though secondary cartilage is essential for jaw functionality, little is known about the molecular mechanisms that induce and maintain secondary cartilage. To address this issue, the current proposal employs an avian model system that exploits species-specific differences in the way secondary cartilage has evolved to support specialized modes of feeding. Duck feed by using their jaws to scoop and filter through wet sediment. Even before hatching, secondary cartilage arises in the duck mandibular adductor enthesis, which inserts laterally and thus greatly extends the coronoid process. This creates a robust interface between the tendon of the mandibular adductor muscle and the mandible, and transmits the powerful contractions necessary to lift the jaw. In contrast, chick feed primarily by pecking at seed, and their mandibular adductor muscle inserts dorsally along the coronoid process of the mandible without any secondary cartilage. These key distinctions are apparent in duck and chick embryos, even though there are no significant differences in embryonic jaw motility. This suggests that species-specific jaw architecture generates mechanical forces that are present in duck but not chick, leading to the differential activation of mechanosensitive signaling pathways during development. Based on published and preliminary data, we hypothesize that Fibroblast Growth Factor (FGF) and Calcium (Ca2+) signaling play a role in enabling the mandibular adductor enthesis to detect biomechanical forces and produce secondary cartilage. Aim 1 involves experiments that will determine whether FGF and Ca2+ signaling are necessary for secondary chondrogenesis. Beads soaked in small molecule inhibitors of FGF and Ca2+ signaling will be implanted beneath the epithelium overlying the presumptive duck coronoid process. Experiments in Aim 2 will uncover whether FGF and Ca2+ signaling are sufficient to promote secondary chondrogenesis by using FGF and Ca2+ signaling agonists in chick. Experiments of Aim 3 will employ chick-duck chimeras to determine whether chick cells are competent to form secondary cartilage when in a duck environment. Understanding mechanisms that regulate secondary chondrogenesis will lead to regenerative therapies for conditions involving loss of secondary cartilage such as TMD and those that occur following trauma.