Targeted Cleavage of Matrix Molecules in a Neocartilage

新软骨中基质分子的靶向裂解

• 批准号: 8011409
• 负责人: JACK L LEWIS
• 金额: $ 16.57万
• 依托单位: UNIVERSITY OF MINNESOTA
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-09-10 至 2012-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8011409
• 关键词: Animals; Biochemical; Candidate Disease Gene; Cartilage; Cartilage Matrix; Chondrocytes; Cleaved cell; Collagen; Collagen Fibril; Connective Tissue; Consensus; Degenerative polyarthritis; Dermatan Sulfate; Development; Digestion; Disease; Elements; Enzymes; Evaluation; Extracellular Matrix; Extracellular Matrix Proteins; Failure; Fracture; Genes; Genetic; Genome; Glues; Goals; Growth; Indium; Knockout Mice; Laboratories; Location; LytA enzyme; Measures; Mechanics; Methods; Minor; Modeling; Motivation; Peptide Hydrolases; Pharmaceutical Preparations; Prevention; Process; Property; Proteins; Proteoglycan; Research; Site; Specificity; Staging; System; Techniques; Therapeutic Intervention; Thrombin; Tissue Engineering; Tissues; Translating; Validation; Work; aggrecan; cartilage cell; chondroitinase B; decorin; disability; drug development; experience; fibromodulin; interest; mouse model; overexpression; prevent; public health relevance; repaired; single molecule; success; tool; treatment strategy

DESCRIPTION (provided by applicant): The goal of this project is to develop a method and model in which a chosen extracellular matrix protein of a tissue grown from cartilage cells (neocartilage) can be cleaved at a specific site. Addition of a translated thrombin protease recognition site will be inserted into the genetic sequence of an extracellular matrix molecule, the genetically modified molecule expressed by chondrocytes, and a neocartilage generated. The genetically modified protein could be cleaved by the addition of thrombin at any point in the development of the tissue. The immediate application is to allow evaluation of candidate structural molecules, molecules that carry load, by comparison of the mechanical properties of the neocartilage before and after cleaving the molecule. If the candidate molecule is a structural molecule, the mechanical properties should be reduced after cleavage of that molecule. The motivation for developing the method is to understand the mechanical failure process in osteoarthritis, where extracellular matrix molecules are digested by lytic enzymes and subsequently the tissue fails under relatively normal loads. Current strategies for disrupting the disease process involve search for agents to block critical molecules in the process. A significant challenge in this approach has been identifying the critical molecules to block. The traditional approach to identifying structural molecules is by digesting candidate molecules with an exogenous enzyme and measuring mechanical properties before and after. The problem with this approach is that there are very few enzymes available that are single molecule specific. The proposed method would allow digestion (cleavage) of a chosen molecule with specificity, greatly enlarging the number of matrix molecules that could be evaluated as candidate structural molecules. The method may have other applications in tissue engineering and drug release. The approach would be a valuable tool in understanding the OA disease process and as a guide for therapeutic intervention. PUBLIC HEALTH RELEVANCE: Osteoarthritis (OA) is the largest cause of disability, but there is poor understanding of the mechanisms of the disease process, particularly how cartilage is degraded. This project will develop methods to identify how load is transmitted through cartilage, and which specific load carrying molecules might be damaged during the OA process. Identification of these molecules would allow development of drugs that could retard the damage and provide a disease modifying treatment to OA.

描述（由申请人提供）：该项目的目标是开发一种方法和模型，在这种方法和模型中，从软骨细胞（新软骨）生长的组织中选择的细胞外基质蛋白可以在特定部位切割。添加一个翻译的凝血酶蛋白酶识别位点，将其插入细胞外基质分子的基因序列中，该基因修饰分子由软骨细胞表达，并生成新软骨。这种基因修饰蛋白可以在组织发育的任何阶段通过加入凝血酶进行切割。直接的应用是通过比较分子切割前后新软骨的机械性能来评估候选结构分子，即承载载荷的分子。如果候选分子是结构分子，则该分子裂解后力学性能应降低。开发该方法的动机是了解骨关节炎的机械失效过程，其中细胞外基质分子被裂解酶消化，随后组织在相对正常的负荷下失效。目前破坏疾病过程的策略包括寻找阻断该过程中关键分子的药物。这种方法面临的一个重大挑战是确定要阻断的关键分子。鉴定结构分子的传统方法是通过外源性酶消化候选分子并测量前后的力学性质。这种方法的问题是，很少有酶是单分子特异性的。所提出的方法将允许消化（裂解）一个选定的分子特异性，大大增加基质分子的数量，可以评估作为候选结构分子。该方法在组织工程和药物释放方面也有应用前景。该方法将是了解OA疾病过程的宝贵工具，并作为治疗干预的指导。