PROTEOGLYCAN STRUCTURE, METABOLISM AND ROLE IN TENDON

这个项目的长期目标是了解结构， 一类细胞外基质的代谢、调节和作用 肌腱中的成分-蛋白聚糖。肌腱通常只包含 少量的小蛋白聚糖核心蛋白聚糖。然而，一个聚集体- 在牛深屈肌腱经过的地方形成丰富的纤维软骨 在骨下并在原位经受压缩和拉伸。在 体外实验支持机械负荷导致 增加大蛋白聚糖的合成和积累， 具有其特定的所需的附加压缩刚度的组织 机械环境实验被提议调查长- 肌腱中细胞对施加的长期和短期反应 特定的机械载荷。(1)将建造一个新的装置来装载 离体胎牛肌腱正常拉伸的离散区域 应力、法向压应力或剪应力， 从几分钟到几天不等。不同载荷的影响 环境对蛋白聚糖合成的数量和类型的影响， 结构和周转，以及mRNA表达和定位， 将确定几种基质成分。合成和活化 将测量作为负荷结果的TGF-β的量。(2)共焦 显微镜将用于观察细胞对 组织负荷，如（Ca++]的变化或组织的改变， 细胞骨架酪氨酸磷酸化或酪氨酸 将在不同的负荷方案后评估磷酸酶。(3)的 细胞外基质组织和特定基质成分的作用 在调节中，将确定细胞对组织负荷的反应， 利用酶改变基质完整性和药物改变细胞骨架 organization.这一综合实验计划将首次， 允许将不同的测量机械载荷与相关的 一个组织中的细胞反应。纤维软骨提供压缩 刚度和滑动表面的位置， 属性对肌腱是有用的。不适当的发展 纤维软骨导致与重复运动相关的病理 综合征，改变愈合性能，和风湿性疾病。研究 这一建议将增加对组织水平反应的理解 允许软结缔组织适当地调节它们的 以适应不断变化的机械环境。

The long-term goal of this project is to understand the structure, metabolism, regulation, and roles of one category of extracellular matrix constituent in tendon - the proteoglycans. Tendon generally contains only a small amount of the small proteoglycan decorin. However, an aggrecan- rich fibrocartilage develops where the bovine deep flexor tendon passes under bone and experiences compression as well as tension in situ. In vitro experiments support the hypothesis that mechanical loading leads to increased synthesis and accumulation of large proteoglycans, generating a tissue with the added compressive stiffness needed for its particular mechanical environment. Experiments are proposed to investigate the long- term and short-term responses of cells in tendon to the imposition of specific mechanical loads . (1) A new device will be constructed to load discrete regions of a fetal bovine tendon in vitro with normal tensile stress, normal compressive stress, or shear stress, for time periods ranging from minutes to days. The effects of different loading environments on the amount and type of proteoglycan synthesized, its structure and turnover, as well as on mRNA expression and localization for several matrix constituents, will be determined. Synthesis and activation of TGF-Beta as a result of loading will be measured. (2) Confocal microscopy will be used to visualize immediate cellular responses to tissue loading, such as changes in (Ca++] or altered organization of the cytoskeleton. The activity of tyrosine phosphorylation or tyrosine phosphatase will be assessed after different loading regimens. (3) The role of extracellular matrix organization and specific matrix components in regulation the cellular response to tissue loading will be determined using enzymes to alter matrix integrity and drugs to alter cytoskeletal organization. This integrated experimental plan will, for the first time, allow correlation of distinct measured mechanical loads with relevant cellular responses in one tissue. Fibrocartilage provides compressive stiffness and a gliding surface in precisely the location where such properties are useful to the tendon. The inappropriate development of fibrocartilage leads to pathology associated with repetitive motion syndromes, altered healing properties, and rheumatic disease. The studies of this proposal will increase understanding of tissue-level responses that allow soft connective tissues to appropriately modulate their composition to meet a changing mechanical environment.