Elucidating Mechanisms of Mechanosensitivity During Secondary Chondrogenesis

描述（由申请人提供）：在生命的所有阶段，骨骼都经过优化，用于检测和适应生物力学力。当维持骨骼健康的微妙平衡被疾病或伤害破坏时，个人的生活质量会迅速恶化。这个项目的目标是确定机制，使骨骼感觉和响应机械力。次级软骨是一种高度适应于检测机械力的骨骼组织。次级软骨最初在颌骨骨骼中的骨区域上发育，以响应胚胎运动期间产生的力。在没有适当的机械力的情况下，继发性软骨不能形成，并且也可以在任何时候退化，如在颞下颌关节紊乱病（TMD）和颌骨固定的患者中。虽然继发性软骨对颌骨功能至关重要，但对诱导和维持继发性软骨的分子机制知之甚少。为了解决这个问题，目前的建议采用了一个鸟类模型系统，利用物种特异性差异的方式，二级软骨已经演变，以支持专门的喂养模式。鸭饲料使用他们的下巴舀和过滤通过潮湿的沉积物。即使在孵化前，次级软骨出现在鸭下颌内收肌起点，插入横向，从而大大延长冠状突。这在下颌内收肌肌腱和下颌骨之间形成了一个坚固的界面，并传递了抬起下颌所需的强大收缩。与此相反，鸡饲料主要是在种子啄食，和他们的下颌内收肌插入背沿着的冠状突的下颌骨没有任何次级软骨。这些关键的区别在鸭和鸡的胚胎中是明显的，即使在胚胎的下颌运动性上没有显著差异。这表明，物种特异性颌骨结构产生的机械力，是存在于鸭，但不是鸡，导致在发展过程中的机械敏感信号通路的差异激活。基于已发表的和初步的数据，我们假设成纤维细胞生长因子（FGF）和钙（Ca 2+）信号在使下颌内收肌附着点检测生物力学力和产生次级软骨中发挥作用。目的1涉及的实验，将确定是否FGF和Ca 2+信号是必要的二次软骨形成。浸泡在FGF和Ca 2+信号传导的小分子抑制剂中的珠将被植入覆盖在假定的鸭冠状突上的上皮下。目的2中的实验将揭示FGF和Ca 2+信号传导是否足以通过使用鸡中的FGF和Ca 2+信号传导激动剂来促进继发性软骨形成。目标3的实验将采用鸡-鸭嵌合体来确定鸡细胞在鸭环境中是否有能力形成次级软骨。了解调节继发性软骨形成的机制将导致再生治疗涉及继发性软骨损失的条件，如TMD和那些发生在创伤后。

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.