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

描述(由申请人提供)：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，需要开发新的治疗方法来治疗关节疾病。保持关节动态平衡是为了保护关节细胞沿着分化途径向下进行，从而导致软骨细胞肥大和骨替代，这种情况发生在生长板软骨中。如果骨性关节炎是祖细胞未能建立正确的微环境的结果，探索关节如何形成可以引导我们了解软骨是如何退化的，并开发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性和再生性关节基因，以操纵它们的表达，以达到治疗的目的。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骨关节炎中是转化生长因子-α(A)II型受体(TaRII)信号的关键下游介体。我们的中心假设是，MCP-5在带间的表达是受控的，以便在发育过程中形成适当的关节，在成年期也是维持关节完整性和防止骨关节炎退变的关键。提出这项研究的基本原理是，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的目标：1)确定是否需要TaRII信号通过下调MCP-5的表达来维持关节的完整性；2)确定MCP-5/TaRII轴在创伤后骨关节炎中的作用。在第一个目标中，我们将通过他莫昔芬诱导的Cre转基因产生Prx1CreERT2/Tgfbr2lox/lox小鼠，以获得时间和肢体特异性的TaRII表达控制，其中我们将评估存在或不存在MCP-5信号的情况下OA的发展(通过化学和受体操作)。在第二个目标中，我们将：a)通过对TGFBR2-a-Gal-GFP-BAC小鼠内侧半月板进行失稳处理来诱导创伤后骨性关节炎(TGFBR2-a-Gal-GFP-BAC小鼠内侧半月板失稳)，并分析MCP-5在骨性关节炎发育过程中伴随着TaRII的表达；b)对分离的TaRII表达细胞诱导机械负荷，并评估细胞变形是否导致逐渐失去对转化生长因子-α的反应性，从而导致MCP-5表达增加。这一建议是创新的，因为它为评估OA过程提供了一个新的视角：OA不仅被视为系统性细胞因子对被动靶点(关节)的损害的结果，而且被视为ACTVE细胞联合群体未能维持受控的细胞因子环境。我们的研究将为关节发育、动态平衡和骨性关节炎的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.