CD44 MEDIATED CATABOLISM OF HYALURONAN BY CHONDROCYTES

描述：(改编自申请人的摘要)：骨性关节炎是一种 以关节退行性变为表现的严重衰弱疾病 软骨衬里关节腔。维护健康的软骨 取决于平衡合成和降解过程中涉及的 软骨细胞外基质的周转。软骨细胞外 基质主要由胶原蛋白和蛋白多糖组成。 到另一种基质大分子透明质酸(HA)的单丝。这 羟基磷灰石则通过以下途径直接结合或锚定到软骨细胞膜 与一种HA受体蛋白相互作用，最近被鉴定为CD44。多数 软骨分解代谢的研究主要集中在细胞外。 通过蛋白水解酶和蛋白水解酶抑制剂处理基质成分。 然而，参与软骨细胞外降解的酶(S) HA从未被记录在案，也没有HA的其他细胞机制 分解代谢。这种知识是缺乏的，尽管HA扮演着 在软骨细胞外基质的组织中起中心作用。 初步研究表明，CD44受体介导内吞作用。 细胞内降解的透明质酸。透明质酸的结合和内化 被HA六糖(HA结合的竞争性抑制剂)阻断，如 以及抗CD44抗体。HA的小降解片段也是 在细胞内检测到，并抑制这些片段的生成 由溶酶促进剂，氯喹。分解代谢细胞介体 上调CD44的表达，增加软骨细胞的能力 在细胞内堆积透明质酸。因此，这是第一次有一种机制 软骨细胞对透明质酸的分解代谢正在进化。的具体目标 这项建议是为了：(I)确定HA内吞作用的确切比率， 包括大小不一的透明质酸和用蛋白多糖片段装饰的透明质酸 和连接蛋白；(Ii)确定CD44的表达水平如何影响 透明质酸内吞的比率。第二个目标将涉及使用 CD44在软骨细胞培养和软骨细胞培养中的反义抑制作用 完整的软骨组织切片；以及(Iii)CD44和CD44的变化 在人的软骨中，在衰老和疾病过程中表达的异构体。

DESCRIPTION: (Adapted from the Applicant's Abstract): Osteoarthritis is a severe debilitating disease manifested by degeneration of articular cartilage-lining joint cavities. The maintenance of healthy cartilage depends on balancing the synthetic and degradative processes involved in turnover of cartilage extracellular matrix. Cartilage extracellular matrices are composed predominately of collagen and proteoglycans tethered to single filaments of another matrix macromolecule, hyaluronan (HA). This HA, in turn, is bound or anchored directly to the chondrocyte membrane via interaction with an HA receptor protein, recently identified as CD44. Most studies of cartilage catabolism have centered on the extracellular processing of matrix components via proteases and protease inhibitors. However, enzymes(s) involved in the extracellular degradation of cartilage HA have never been documented, nor have other cellular mechanisms for HA catabolism. This knowledge is lacking despite the fact that HA plays a central role in the organization of cartilage extracellular matrix. Preliminary studies have shown that CD44 receptors mediate the endocytosis of HA for intracellular degradation. The binding and internalization of HA is blocked by HA hexasaccharides (competitive inhibitors of HA binding), as well as anti-CD44 antibodies. Small degradation fragments of HA are also detected intracellularly and the generation of these fragments is inhibited by the lysosomotropic agent, chloroquine. Catabolic cell mediators up-regulate CD44 expression, resulting in increased capacity of chondrocytes to accumulate intracellular HA. Thus for the first time, a mechanism for the catabolism of HA by the chondrocyte is evolving. The Specific Aims of this proposal are to: (i) determine the exact rates of HA endocytosis, including HA of various sizes and HA "decorated" with proteoglycan fragments and link protein; (ii) determine how the level of expression of CD44 affects the rate of HA endocytosis. This second aim will involve the use of antisense inhibition of CD44 in both chondrocyte cultures, and cultures of intact cartilage tissue slices; and (iii) characterize changes in CD44 and isoform expression in human cartilage that occur during aging and disease.