Molecular and cellular mechanisms of heterotopic ossification

描述(申请人提供)：异位骨化(HO)，在骨骼肌和软组织中形成的异位软骨内骨，是关节不能活动和疼痛的重要原因。HO的确切机制尚不清楚；然而，它与手术后和创伤后环境的关联表明了一个混乱的损伤修复过程。从先天性HO综合征-进行性骨化性纤维发育不良(FOP)可以获得对HO原因的进一步了解。FOP是由骨形态发生蛋白(BMP)I型受体ALK2的“结构性激活”突变引起的，这种突变会导致由轻微创伤或炎症引发的进行性和广泛性关节骨化。FOP和获得性HO都缺乏有效的治疗方法。事实上，有重要证据表明，FOP和HO都是由BMP信号通路的不适当激活引起的。目前尚不清楚增强的BMP信号如何偏离损伤修复程序，也不知道哪些细胞群介导了增强的BMP信号的影响。为了解决这些问题，我们开发了一种FOP的小鼠模型，在该模型中，ALK2的成分活性突变形式(CaALK2)被诱导表达。与受影响的人类相似，该基因的表达不会自发地引发HO，但在炎症和肌肉损伤的额外刺激下，会发生剧烈的骨化和关节融合。我们随后对这个模型的研究表明，这些caALK2蛋白可能不像以前认为的那样具有结构性活性，但可能会使细胞对传统的配体介导的BMP信号敏感。在这项资助的目标1中，我们将使用这个模型来识别caALK2使细胞对BMP信号敏感的机制，测试caALK2是否需要配体介导的信号或独立发挥作用。为了确定介导BMP信号增强效应的细胞前体，以及哪些细胞前体参与了异位骨损伤，在目标2中，我们将突变的caALK2的表达靶向几个候选的祖细胞谱系。利用基于细胞的、基因靶向和过继转移技术的补充，我们将系统地确定caALK2在具有已知成骨潜力的隔室中表达的影响，包括骨骼肌卫星细胞、血管周细胞以及骨髓来源的谱系。在目标3中，我们在这个模型中研究了先天免疫信号在异位骨发育中的作用。了解caALK2突变如何改变BMP信号转导的结果，可以突出管理HO的新的分子或细胞靶点。这项建议试图阐明增强的BMP信号如何影响间充质祖细胞，并在损伤、炎症和再生的界面上调控适应性和非适应性成骨。 公共卫生相关性：这项建议提出了骨形态发生蛋白信号，它通常调节参与肌肉、血管、结缔组织和骨骼修复的前体细胞的活动，但在人类异位骨化和进行性骨纤维发育不良的疾病中，如何可能变得失调而导致不适当的骨形成。除了提供对这些鲜为人知的过程的机制的见解外，这些研究还可能为它们的管理确定亟需的新方法。这些机制与更广泛的条件有关，在这些条件下，炎症和损伤似乎导致自身免疫性、心脏和血管疾病中的异常骨化。

DESCRIPTION (provided by applicant): Heterotopic ossification (HO), the formation of ectopic endochondral bone in skeletal muscle and soft tissues, is a significant cause of morbidity from joint immobility and pain. The precise mechanisms responsible for HO are not known; however, its association with postsurgical and posttraumatic contexts suggests a process of disordered injury repair. Further insights into the causes of HO may be gained from a congenital HO syndrome, fibrodysplasia ossificans progressiva (FOP). FOP is caused by "constitutively-activating" mutations in the bone morphogenetic protein (BMP) type I receptor ALK2, which result in progressive and widespread joint ossifications triggered by minimal trauma or inflammation. Both FOP and acquired forms of HO lack effective therapies. In fact, there is significant evidence that both FOP and HO are caused by inappropriate activation of the BMP signaling pathway. It is not known how enhanced BMP signaling deviates the injury repair program, or which populations of cells mediate the effects of enhanced BMP signaling. To address these questions we have developed a mouse model of FOP in which a constitutively-active mutant form of ALK2 (caALK2) is inducibly expressed. Similar to affected humans, expression of this gene does not spontaneously induce HO, but vigorous ossification and joint fusion occur with additional stimuli of inflammation and muscle injury. Our subsequent studies with this model suggest that these caALK2 proteins may not be constitutively-active, as previously thought, but may sensitize cells to traditional ligand-mediated BMP signals. In Aim 1 of this grant, we will employ this model to discern the mechanisms by which caALK2 sensitizes cells to BMP signals, testing whether caALK2 requires ligand-mediated signaling or functions independently. To identify the cellular progenitors which mediate the effects of enhanced BMP signaling, and which contribute to the ectopic bone lesions, in Aim 2 we have targeted the expression of mutant caALK2 to several candidate progenitor lineages. Using a complement of cell-based, genetic targeting, and adoptive transfer techniques, we will systematically determine the impact of expressing caALK2 in compartments with known osteogenic potential, including skeletal muscle satellite cells, vascular pericytes, as well as bone-marrow derived lineages. In Aim 3, we examine the role of innate immune signaling in the development of ectopic bone in this model. Understanding how caALK2 mutations alter the consequences of BMP signaling could highlight novel molecular or cellular targets for management of HO. This proposal seeks to elucidate how enhanced BMP signaling impacts mesenchymal progenitors and governs adaptive and maladaptive osteogenesis at the interface of injury, inflammation, and regeneration. PUBLIC HEALTH RELEVANCE: This proposal asks how bone morphogenetic protein signals, which normally regulate the activity of progenitor cells involved in the repair of muscle, blood vessels, connective tissues, and bone, may become dysregulated to cause inappropriate bone formation in the human diseases of heterotopic ossification and fibrodysplasia ossificans progressiva. In addition to providing insights into the mechanism of these poorly understood processes, these studies may identify much needed novel approaches for their management. These mechanisms have relevance to a broader set of conditions in which inflammation and injury appear to lead to abnormal ossification in autoimmune, cardiac, and vascular disease.