Evaluation of Prg4 as a New Therapy for TMJ Disc Degeneration

颞下颌关节（temporo-mandibular joint，TMJ）由下颌髁状突、颞下颌关节窝等组成。 颞骨和关节盘，是说话和咀嚼的基本。光盘扮演着重要的角色 在TMJ运动中，通过允许髁状突自由地滑下关节窝的斜坡。颞下颌关节疾病 （TMJ的结构、功能或生理学的破坏）折磨着大约20%的美国人。 人群，高达70%的条件是由于颞下颌关节盘移位和/或损伤。一般 认为，在润滑系统，这主要取决于关节润滑剂，包括 透明质酸、磷脂和蛋白聚糖4（Prg 4）导致TMJ功能障碍。一个关键的差距， 知识是澄清关节润滑剂建立和维持关节结构和功能的程度， 颞下颌关节盘我们发现Prg 4-null（Prg 4-/-）小鼠的TMJ表现出明显更大的退行性变， 与包括膝关节在内的其他滑膜关节相比，我们在Prg 4-/-小鼠中发现：（a）TMJ 椎间盘未能形成其双凹形状，并随着时间的推移而显著增厚;（B）大量 的椎间盘细胞分化为纤维软骨细胞，导致椎间盘矿化;以及（c）随后， 髁突软骨破坏，骨小梁表现出结构变化。此外该 与对照相比，Prg 4-/-盘的机械性能显著改变。这些和其他 新的数据导致我们的中心假设，Prg 4是必要的：1）维持TMJ椎间盘细胞表型， 功能; 2）保护TMJ免受退行性变化。在目标1中，我们将确定 途径，PRG 4通过其调节多个椎间盘细胞的区域和阶段特异性表型 解剖部位，并阐明如何全球损失Prg 4导致异常的椎间盘细胞功能和椎间盘带状 organization.通过在出生后诱导Prg-4表达细胞的细胞死亡，我们还将确定Prg-4在哺乳动物中的作用。 在椎间盘形态发生和体内平衡中表达细胞。在目标2中，我们将测试TMJ椎间盘退变是否是 可治疗的，如果是，还确定是否存在成功干预的治疗窗口。我们将恢复 通过病毒驱动的Prg 4和他莫昔芬或病毒驱动的Cre过表达的出生后Prg 4表达 ROSA 26 CreERT 2; Prg 4GT（Prg 4cGT）小鼠中的重组酶，其在Prg 4中具有可逆基因陷阱（GT） （Prg 4GT），其中Prg 4表达在Cre介导的基因陷阱切除后被激活。表型 分析将包括组织病理学，基因和蛋白质表达，以及生物力学测试。口面疼痛会 通过机械反射测试和啮齿动物鬼脸量表进行评估。我们的项目将提供新颖和远- 获得有关关节润滑剂在颞下颌关节盘维护和疾病中的作用的信息。我们的救援 实验应该提供一个原则的证明，即颞下颌关节盘退变是适合治疗的， 干预虽然我们的实验使用TMJ作为模型系统，但我们相信这种应用的结果 也可直接应用于其他滑膜关节。

The temporo-mandibular joint (TMJ), composed of the mandibular condyle, the glenoid fossa of the temporal bone and the articular disc, is essential for speech and mastication. The disc plays an important role in TMJ movement, by allowing the condyle to freely slide down the slope of the glenoid fossa. TMJ disorders (disruption in the structure, function or physiology of the TMJ) afflict approximately 20% of the U.S. population, and up to 70% of the conditions are due to TMJ disc displacement and/or damage. It is generally believed that aberrations in the lubrication system, which depends primarily on joint lubricants, including hyaluronic acid, phospholipid, and Proteoglycan 4 (Prg4), contribute to TMJ dysfunction. A critical gap in knowledge is to clarify the extent to which joint lubricants establish and maintain the structure and function of the TMJ disc. We found that TMJs of Prg4-null (Prg4-/-) mice exhibited significantly greater degenerative changes compared to other synovial joints, including the knee. We found in Prg4-/- mice that: (a) the TMJ discs failed to develop their biconcave shape and became substantially thickened over time; (b) a large number of the disc cells differentiated into fibrochondrocytes, leading to disc mineralization; and (c) subsequently, the condylar cartilage broke down and the trabecular bone exhibited architectural changes. In addition, the mechanical properties of the Prg4-/- discs were significantly altered compared to controls. These and other novel data lead to our central hypothesis that Prg4 is necessary: 1) to maintain TMJ disc cell phenotype and function, and 2) to protect the TMJ from degenerative changes. In Aim 1, we will determine the signaling pathways, by which PRG4 regulates region- and stage-specific phenotypes of disc cells across multiple anatomical sites, and elucidate how global loss of Prg4 leads to abnormal disc cell function and disc zonal organization. By inducing cell death in Prg-4 expressing cells postnatally, we will also define the roles of Prg4- expressing cells in disc morphogenesis and homeostasis. In Aim 2, we will test if TMJ disc degeneration is treatable, and, if so, also determine if there is a therapeutic window for successful intervention. We will restore Prg4 expression postnatally via virally-driven overexpression of Prg4 and tamoxifen or virally-driven Cre recombinase in ROSA26CreERT2;Prg4GT (Prg4cGT) mice, which have a reversible gene-trap (GT) in Prg4 (Prg4GT), in which Prg4 expression is activated upon Cre-mediated excision of the gene-trap. Phenotypic analyses will include histopathology, gene and protein expression, and biomechanical tests. Orofacial pain will be evaluated by mechanical reflex testing and rodent grimace scale. Our project will provide novel and far- reaching information regarding the role of joint lubricants in TMJ disc maintenance and disease. Our rescue experiments should provide a proof-of-principle that TMJ disc degeneration is amenable to therapeutic intervention. While our experiments use the TMJ as a model system, we believe results from this application will also be directly applicable to other synovial joints.