Mechanisms of TMJ development and long-term function

描述（由申请人提供）：颞下颌关节（TMJ）与其他滑膜关节不同，它包含一个关节盘，将关节间隙分为两个滑膜腔。这些独特的特征使得在咀嚼和说话期间发生的TMJ的复杂解剖学重排成为可能。这些运动也会产生剪切和摩擦载荷，以及关节的近脱位。为了保护关节完整性免受破坏性机械摩擦，表面和椎间盘细胞产生边界润滑剂，包括透明质酸（HA）和润滑素。因此，随着年龄的增长或其他损害，关节润滑的恶化可能是椎间盘粘连和退行性TMJ疾病的基础。我们建议研究目前知之甚少的TMJ生物学的发展方面，是至关重要的TMJ功能和长期健康。我们的初步数据表明，一个分泌的信号因子，印度刺猬（Ihh），在TMJ发育中起着关键作用。整体Ihh消融导致椎间盘和滑膜腔形成失败，TGF-β1、PTHrP和润滑素表达丧失。出生后消融IHH导致润滑素表达和椎间盘粘连明显减少。这些发现导致了我们的中心假设，即Ihh信号是TMJ形成和出生后功能所必需的。在目标1中，Ihh信号在TMJ发育中的作用将被进一步定义。首先，我们将涉及时空模式的Ihh信号（Gli 1激活）的刺猬受体的表达，信号转导和表型定义基因在TMJ的发展。然后，我们将使用功能的丧失和获得方法来删除TMJ细胞中的Smoothened或Patched 1。在目标2中，我们将通过询问TGF-β 1和/或PTHrP是否足以挽救Ihh缺失的TMJ表型以及TMJ中这些基因的条件性缺失是否产生与Ihh缺失小鼠中所见相似的缺陷来检验TGF-β 1和PTHrP介导Ihh对其TMJ中靶细胞的作用的假设。我们将进一步阐明Ihh-TGF-β和Ihh-PTHrP信号通路及其在调节关节润滑剂产生和相关细胞类型发育中的潜在相互作用。目的3：研究关节润滑剂在Ihh依赖的出生后TMJ维持中的作用. HA和润滑素在TMJ保护中的各自作用将通过分析润滑素（Prg 4）-无效和条件性HA合酶2（HAS 2）-缺陷小鼠来确定。我们还将测试是否可以通过全身或局部注射hedgehog激动剂、PTHrP/PTH、TGF-β1或局部递送润滑素和/或HAS 2表达载体来挽救Ihh缺失小鼠中的TMJ缺陷。总之，该项目将提供新的，重要的和深远的信息刺猬信号在TMJ的形成和维护。我们的抢救实验应该提供一个原则的证明，颞下颌关节的缺陷，包括椎间盘粘连，是经得起治疗干预。

DESCRIPTION (provided by applicant): The temporomandibular joint (TMJ) is different from other synovial joints, in that it contains an articular disc that separates the joint space into tw synovial cavities. These distinctive features enable complex anatomical rearrangements of TMJ that take place during mastication and speech. These movements also impart shearing and frictional loads and a near dislocation of the joint. To protect joint integrity from destructive mechanical friction, superficial and disc cells produce boundary lubricants including hyaluronic acid (HA) and lubricin. Thus, deterioration of joint lubrication with age or by other insults may underlie disc adhesion and degenerative TMJ disorders. We propose to study currently poorly understood developmental aspects of TMJ biology that are critically important for TMJ functions and its long-term health. Our preliminary data showed that a secreted signaling factor, Indian hedgehog (Ihh), plays pivotal roles in TMJ development. Global Ihh ablation led to a failure of disc and synovial cavity formation, and loss of TGF-β1, PTHrP and lubricin expression. Postnatal ablation of Ihh caused markedly reduced lubricin expression and disc adhesions. These findings led to our central hypothesis that Ihh signaling is required for both TMJ formation and postnatal function. In Aim 1, the roles of Ihh signaling in TMJ development will be further defined. Initially, we will relate the spatio-temporal patterns of Ihh signaling (Gli1 activation) o the expression of hedgehog receptors, signal transducers and phenotype defining genes during TMJ development. We will then use loss and gain of function approaches to delete Smoothened or Patched1 in TMJ cells at critical developmental stages. In Aim 2, we will test the hypothesis that TGF-β1 and PTHrP mediate Ihh action on its target cells in TMJ by asking whether TGF-β1 and/or PTHrP are sufficient to rescue Ihh-null TMJ phenotype and whether conditional deletion of these genes in TMJ produce defects similar to those seen in Ihh-null mice. We will further delineate Ihh-TGF-β and Ihh- PTHrP signaling pathways and their potential interactions in the regulation of joint lubricant production and development of relevant cell types. In Aim 3, we will study the roles of joint lubricants in the Ihh- dependent postnatal TMJ maintenance. Respective roles of HA and lubricin in the protection of the TMJ will be determined by analyzing lubricin (Prg4)-null and conditional HA synthase 2 (HAS2)-deficient mice. We will also test whether TMJ defects in Ihh-null mice can be rescued by systemic or local injection of a hedgehog agonist, PTHrP/PTH, TGF-β1 or local delivery of lubricin- and/or HAS2-expression vectors. In sum, this project will provide novel, important and far-reaching information on hedgehog signaling in TMJ formation and maintenance. Our rescue experiments should provide a proof-of-principle that TMJ defects, including disc adhesions, are amenable to therapeutic intervention.