MCP5 as key mediator of TGFbeta signaling in joint maintenance and osteoarthritis

项目摘要骨关节炎（OA）是最常见的关节炎形式。骨性关节炎患者缺乏有效的药物治疗来再生受损的软骨，因此需要开发新的治疗方法来治疗关节疾病。维持关节内稳态，保持关节细胞沿着分化途径进行，导致软骨细胞肥大和骨置换，这种情况发生在生长板软骨中。如果OA是祖细胞未能建立正确微环境的结果，那么探索关节的形成方式可以让我们了解软骨如何退化，并开发出能够重新激活形成机制的药物。我们的长期目标是阐明关节祖细胞如何表达生殖和再生关节基因，以便操纵其表达以达到OA的治疗目的。总体目标是确定细胞因子单核细胞趋化蛋白-5 （MCP-5）的调节是否在关节维持和创伤后OA中是tgf - α (a) II型受体（TaRII）信号传导的关键下游介质。我们的中心假设是，在发育过程中，需要MCP-5的控制表达，以允许适当的关节形成，并且在成年期保持关节完整性和防止OA变性也至关重要。本研究的基本原理是细胞因子控制的环境是促进发育和维持体内平衡的共同机制。基于我们的初步数据，我们提出了两个具体目标：1)确定是否需要TaRII信号通过下调MCP-5表达来维持关节完整性；2)探讨MCP-5/TaRII轴在创伤后骨性关节炎中的作用。在第一个目标中，我们将使用他莫昔芬诱导的cre转基因产生Prx1CreERT2/Tgfbr2lox/lox小鼠，以获得TaRII表达的时间和肢体特异性控制，其中我们将评估OA在存在或不存在MCP-5信号（通过化学和受体操作）的情况下的发展。在第二个目标中，我们将：A)通过对Tgfbr2-a-Gal-GFP-BAC小鼠（TaRII表达的成像报告者）进行内侧半月板失稳诱导创伤后OA，并分析OA发展过程中MCP-5与TaRII的表达；B)对分离的表达TaRII的细胞诱导机械负荷，并评估细胞变形是否导致对TGF-a的反应性逐渐丧失，从而导致MCP-5表达增加。这一建议具有创新性，因为它为评估OA过程提供了一个新的视角：OA不仅被视为细胞因子对被动靶点（关节）的系统性流入引起的损伤的结果，而且是活性细胞关节群未能维持受控的细胞因子环境的结果。我们的研究将为关节发育稳态和OA病理生理学提供重要的见解。

DESCRIPTION (provided by applicant): PROJECT ABSTRACT Osteoarthritis (OA) is the most common form of arthritis. Patients with OA lack effective medical treatments to regenerate damaged cartilage and there is a need for developing novel therapies to treat joint disorders. Joint homeostasis is maintained preserving the articular cells to proceed down the differentiation pathway that leads to chondrocyte hypertrophy and bone replacement, such it occurs in the growth plate cartilage. If OA is the result of the failure of progenitor cells to establish a corret microenvironment, exploring how joints form can lead us to understand how cartilage degenerates and develop drugs that are able to reactivate the forming mechanism. Our long-term goal is to elucidate how joint progenitor cells express generative and regenerative joint genes in order to manipulate their expression for therapeutic purpose in OA. The overall objective is to determine whether the regulation of the cytokine monocyte chemoattractant protein-5 (MCP-5) is a key down- stream mediator of TGF-alpha (a) type II receptor (TaRII) signaling in joint maintenance and post-traumatic OA. Our central hypothesis is that a controlled expression of MCP-5 is needed in the interzone to allow proper joint formation during development and is also crucial in adulthood to maintain joint integrity and to prevent OA degeneration. The rationale for the proposed research is that a cytokine controlled environment is a common mechanism in joint to promote development and maintain homeostasis. Based on our preliminary data we are proposing two Specific Aims: 1) To determine whether TaRII signaling is needed to maintain joint integrity through down-regulation of MCP-5 expression; 2) To determine the role of MCP-5/TaRII axis in post-traumatic OA. In the first aim, we will generate the Prx1CreERT2/Tgfbr2lox/lox mouse to obtain temporal and limb-specific control of TaRII expression using a tamoxifen-inducible-Cre transgenic in which we will evaluate OA development in the presence or absence of MCP-5 signaling (by chemical and receptor manipulation). In the second aim, we will: A) induce post-traumatic OA by performing a destabilization of medial meniscus in Tgfbr2-a-Gal-GFP-BAC mice (imaging reporters of TaRII expression) and analyze expression of MCP-5 in concomitance with TaRII during OA development; B) induce mechanical loading on isolated TaRII expressing cells and evaluate whether cell deformation leads to progressive loss of responsiveness to TGF-a with consequent increase in MCP-5 expression. This proposal is innovative because provides a new perspective to evaluate the OA process: OA is not only seen as the result of the damage induced by a systemic influx of cytokines against a passive target (the joint), but rather the failure of an actve cell joint population to maintain a controlled cytokine environment. Our studies will provide critical insights on joint development homeostasis and OA pathophysiology.