Role of Small Proteoglycans in the Structure and Biomechanics of Articular Cartilage

关节软骨为关节运动提供了关键的生物力学功能，如承载、能量吸收和润滑。骨关节炎是一种导致软骨功能障碍的慢性疾病，影响着2700多万美国人，是一个重大的社会负担。为了恢复骨关节炎患者的软骨功能，缓解关节疼痛，有必要了解健康软骨的结构和力学性能作为基准。软骨素主要由胶原蛋白和蛋白聚糖（带负电荷的糖链连接的蛋白质）组成。胶原蛋白和大的蛋白聚糖聚集蛋白聚糖对软骨生物力学的直接贡献是众所周知的。 小的蛋白聚糖不直接有助于软骨的生物力学特性，但是，它们可以通过与胶原蛋白和聚集蛋白聚糖相互作用来调节软骨的结构组装。结合新的纳米技术和基因修饰工具，该项目旨在确定软骨中最丰富的小蛋白聚糖，核心蛋白聚糖，如何改变软骨的结构和生物力学。这些结果将通过确定一种控制软骨结构完整性的新分子机制产生更广泛的影响，这可以促进开发恢复软骨功能和减轻骨关节炎软骨退化的新策略。 将建立新的外展计划，以增加少数民族学生对STEM教育和职业道路的参与。具体的外展活动包括为费城市中心的高中教师和学生举办研讨会，以及开发科学博物馆展览。核心蛋白聚糖基因敲除小鼠的软骨中聚集蛋白聚糖含量大幅降低，机械性能受损。这个项目将确定核心蛋白聚糖是否控制聚集蛋白聚糖的结构组装。我们假设核心蛋白聚糖通过与细胞外基质中的聚集蛋白聚糖相互作用以及通过影响软骨细胞聚集蛋白聚糖的合成来调节软骨中聚集蛋白聚糖的存在。这一假设将在长度尺度的层次结构中进行检验。首先，在分子水平上，核心蛋白聚糖和聚集蛋白聚糖之间的粘附将通过基于原子力显微镜（AFM）的分子力光谱来量化。第二，在细胞水平上，将通过生物化学测定和AFM单细胞纳米压痕来评估正常和核心蛋白聚糖敲除软骨细胞的聚集蛋白聚糖的合成和力学。最后，在组织水平，核心蛋白聚糖的表达将在出生后关节生长的不同阶段使用我们的新的诱导型核心蛋白聚糖敲除小鼠模型消融。由此产生的软骨生物力学变化，包括弹性和多孔粘弹性特性，将由我们定制的纳米流变仪进行评估。总之，本项目将阐明小蛋白聚糖核心蛋白聚糖如何调节软骨中聚集蛋白聚糖的组装，从而确保软骨结构的完整性和适当的生物力学功能

Articular cartilage provides key biomechanical functions for joint motion, such as load bearing, energy absorption and lubrication. Osteoarthritis, a chronic disease causing cartilage dysfunction, affects more than 27 million Americans and represents a significant societal burden. To restore cartilage function and alleviate joint pain in osteoarthritis patients, it is necessary to understand the structure and mechanical properties of healthy cartilage as a benchmark. Cartilage is mainly composed of collagen and proteoglycan (protein attached with negatively charged sugar chains). The direct contribution of collagen and the large proteoglycan, aggrecan, to cartilage biomechanics is well understood. Small proteoglycans do not contribute directly to cartilage biomechanical properties; however, they may regulate the structural assembly of cartilage by interacting with both collagen and aggrecan. Combining novel nanotechnology and gene modification tools, this project aims to determine how the most abundant small proteoglycan in cartilage, decorin, modifies the structure and biomechanics of cartilage. The outcomes will have a broader impact by identifying a new molecular mechanism that governs cartilage structural integrity, which can enable the development of novel strategies for restoring cartilage function and attenuating cartilage degradation in osteoarthritis. New outreach programs will be established to increase the participation of minority students in STEM education and career paths. Specific outreach activities include workshops for Philadelphia inner city high school teachers and students, as well as the development of science museum exhibition displays.The cartilage of decorin knockout mice develops substantially reduced aggrecan content and impaired mechanical properties. This project will determine whether decorin governs the structural assembly of aggrecan. We hypothesize that decorin regulates the presence of aggrecan in cartilage through interacting with aggrecan in the extracellular matrix, and through influencing the synthesis of aggrecan by chondrocytes. This hypothesis will be tested at a hierarchy of length scales. First, at the molecular level, adhesion between decorin and aggrecan will be quantified by atomic force microscopy (AFM)-based molecular force spectroscopy. Second, at the cellular level, the synthesis and mechanics of aggrecan by normal and decorin knockout chondrocytes will be evaluated by biochemical assays and AFM single cell nanoindentation. Lastly, at the tissue level, the expression of decorin will be ablated at different phases of post-natal joint growth using our novel inducible decorin-knockout murine model. The resulting changes in cartilage biomechanics, including both elastic and poroviscoelastic properties, will be evaluated by our custom-built nanorheometer. In conclusion, this project will elucidate how the small proteoglycan, decorin, regulates the aggrecan assembly in cartilage, thereby ensuring cartilage structural integrity and proper biomechanical functions