Integrin Function in Cartilage

描述(申请人提供)：该项目的长期目标是确定通过整合素受体产生的信号调节软骨细胞功能的基本机制。这项工作的总体假设是，软骨细胞外基质(ECM)的变化，包括ECM蛋白片段的产生，可由软骨细胞整合素识别，并启动一系列旨在重塑ECM但在关节炎中导致进一步基质破坏的事件(软骨细胞软骨溶解)。这项建议的重点是确定控制信号的基本细胞机制，这些信号通过α5？1整合素来调节分解代谢介质的产生，包括细胞因子和基质金属蛋白酶(MMPs)。在之前的资助期间，我们已经确定了在纤维连接蛋白片段(Fn-f)刺激α5？1整合素时介导基质金属蛋白酶-13产生的信号通路，并发现活性氧物种(ROS)是必要的第二信使。利用一种新的蛋白质组学方法，我们发现MAP激酶家族成员JNK2在Fn-f刺激的细胞中被氧化，形成半胱氨酸-磺酸(Cys-SOH)中间体。磺酸的形成是ROS调节细胞信号转导的主要机制，但其在软骨细胞信号转导中的作用尚未被研究。在他的竞争性更新中，我们建议确定磺酸形成调节软骨细胞中JNK2活性的机制。我们将通过研究手术诱导的JNK2-/-小鼠骨性关节炎的发生发展来确定JNK2激活在体内骨性关节炎中的作用。最后，我们将确定当软骨细胞受到FN-f刺激时，HB-EGF是否上调并释放，是否促进RAC活性，以激活共同信号通路，从而增加MMP-13的表达和软骨基质的破坏。这些研究将通过确定介导软骨基质破坏的信号网络中的关键枢纽，对该领域产生重大影响。通过发现ROS调节这个信号网络的新机制，这些信息可以用来开发一种独特的方法来改变关节炎中针对特定蛋白质修饰的氧化还原调节的分解代谢信号网络。与一般抑制ROS产生相比，这是一个重大进步，后者在治疗由过量ROS促进的疾病(包括关节炎)方面并未被证明有效。

DESCRIPTION (provided by applicant): The long-term goal of this project is to determine the basic mechanisms by which signals generated through integrin receptors regulate chondrocyte function. The overall hypothesis driving this work is that changes in the cartilage extracellular matrix (ECM), including production of ECM protein fragments, are recognized by chondrocyte integrins and initiate a cascade of events intended to remodel the ECM but which in arthritis result in further matrix destruction ("chondrocytic chondrolysis"). The focus of this proposal is to determine the basic cellular mechanisms that control signals generated through the α5ß1 integrin which regulate production of catabolic mediators including cytokines and matrix metalloproteinases (MMPs). During the previous funding period, we have defined the signaling pathways that mediate MMP-13 production in response to fibronectin fragment (FN-f) stimulation of the α5ß1integrin and discovered reactive oxygen species (ROS) are necessary second messengers. Using an innovative proteomics approach we found that the MAP kinase family member JNK2 is oxidized in FN-f stimulated cells forming a Cys-SOH (sulfenic acid) intermediate. Sulfenic acid formation serves as a major mechanism by which ROS regulate cell signaling but its in chondrocyte signaling has not been investigated. In his competitive renewal, we propose to determine the mechanism by which sulfenic acid formation regulates JNK2 activity in chondrocytes. We will determine the role of JNK2 activation in OA in vivo by studying the development of surgically-induced OA in JNK2-/- mice. Finally, we will determine if HB-EGF, upregulated and released when chondrocytes are stimulated by FN-f, promotes Rac activity to activate a co-signaling pathway that augments MMP-13 expression and cartilage matrix destruction. These studies will have significant impact on the field by defining key hubs in a signaling network that mediates cartilage matrix destruction. By discovering novel mechanisms by which ROS regulate this signaling network, the information can be used to develop a unique approach to altering redox-regulated catabolic signaling networks in arthritis that targets specific protein modifications. This represents a significant advance over the general inhibition of ROS production which not proven successful in treating conditions promoted by excessive ROS, including arthritis.