THE INSULIN-LIKE GROWTH FACTOR AXIS IN OSTEOARTHRITIS

描述(申请人提供)：胰岛素样生长因子(IGFS)是关节软骨中基质稳态的关键调节因子，有研究认为骨关节炎(OA)时代谢失调是由于IGF不敏感所致。胰岛素样生长因子结合蛋白(IGFBPs)可以促进或抑制IGF活性，也可以独立于IGF发挥作用。申请人的初步研究表明，IGFBP-3在骨关节炎期间增加，直接在未培养的骨关节炎软骨提取液中进行评估。工作假说认为IGFBP-2定位在细胞膜上，并起到维持基质内环境平衡的作用。软骨中IGFBP-3的含量在骨性关节炎期间增加2-3倍，而IGFBP-2不增加。IGFBP-3可以与软骨细胞上自身特异的、高亲和力的蛋白结合，也可以与丰富的细胞外锚定位点结合。后者包括纤维连接蛋白(FN)基质中的位点，其中锚定的IGFBP-3作用于隔离IGFS。IGFBP-3的总体作用是限制IGF的作用和软骨的修复。其具体目标是：(1)IGFBP-3、IGFBP-2、IGF-I、IGF-II和IGF受体(IGF-IR)在人软骨骨关节炎进展过程中的定位(免疫组织化学研究)以及IGF与BP-3和FN可能的共定位(通过免疫电子显微镜)；在体外研究IGFBP-3和IGF的选择性相互作用；以及检测分离的软骨细胞是否存在高亲和力的IGFBP-3结合蛋白(可能的受体)；(2)骨关节炎期间IGF轴各成分(IGFBPs、IGFS、IGFIR)以及IGFBP-3锚定纤维连接蛋白水平和合成的变化；(3)IGFBPs的功能，通过它们对基质合成和细胞分裂的影响来评估。在骨关节炎期间，将通过培养人软骨切片，并通过添加IGFS和各种IGF类似物来检测IGFBPs的作用，从而确定常驻IGFBPs的整体功能。IGFPs和各种IGF类似物已经发生突变，选择性地失去了它们与IGFPs结合的能力，或者只失去了它们与IGF受体结合的能力。此外，反义寡核苷酸将被用来阻断IGFBP-3，以及在软骨基质通透后阻断抗体。长期目标是设计合理的方案，使IGF轴转向修复模式，从而确定用于人类骨关节炎治疗的候选治疗方案。

DESCRIPTION (provided by applicant): The Insulin-like growth factors (IGFs) are key regulators of matrix homeostasis in articular cartilage, and it has been proposed that dysregulation of metabolism during osteoarthritis (OA) is due to IGF insensitivity. The IGF binding proteins (IGFBPs) can either promote or inhibit IGF activity or may act independently of IGFs. Preliminary studies by the applicant indicate that IGFBP-3 is increased during OA, as assessed directly in extracts of uncultured OA cartilage. The working hypothesis states that IGFBP-2 is Iocated on the cell membrane and acts to maintain matrix homeostasis. IGFBP-3 content in cartilage, but not IGFBP-2, increases 2-3 fold during OA. IGFBP-3 can bind to its own specific, high affinity proteins on the chondrocyte and can also bind to abundant extracellular anchoring sites. The latter include sites in the fibronectin (FN) matrix, where anchored IGFBP-3 acts to sequester IGFs. The overall effect of IGFBP-3 is to restrict IGF action and repair of cartilage. The Specific Aims are: (1) the localization of IGFBP-3, IGFBP-2, IGF-I and II and the IGF receptor (IGF-IR) during OA progression in human cartilage (immunohistochemical studies) and, as appropriate, the possible co-localization of IGF with BP-3 and FN (by immunoelectron microscopy); to examine selected interactions of IGFBP-3 and IGF in vitro; and to test isolated chondrocytes for the presence of high affinity IGFBP-3 association proteins (putative receptors); (2) changes in the levels and synthesis of the components of the IGF axis (IGFBPs, IGFs, IGFIR), as well as in the putative IGFBP-3 anchor, fibronectin, during OA; and (3) the function of the IGFBPs, as assessed by their effects on matrix synthesis and cell division. The overall function of resident IGFBPs will be determined during OA by culturing human cartilage slices and examining the role of IGFBPs by addition of IGFs and various IGF analogs that have been mutated to selectively lose their capacity to bind IGFBPs, or to lose only their ability to bind to the IGF receptor. In addition, antisense oligodeoxynucleotides will be used to block IGFBP-3, as well as blocking antibodies following permeabilization of the cartilage matrix. The long-term goal is to design plausible protocols to shift the IGF axis towards a repair mode so as to define therapeutic candidates for human OA management.