Controlling chondrocyte matrix degradation and repair in 3-D culture.

控制 3D 培养中的软骨细胞基质降解和修复。

基本信息

批准号：
8213730
负责人：
Christopher B Raub
金额：
$ 5.22万
依托单位：
UNIVERSITY OF CALIFORNIA, SAN DIEGO
依托单位国家：
美国
项目类别：
财政年份：
2010
资助国家：
美国
起止时间：
2010-02-01 至 2013-01-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8213730
关键词：
3-Dimensional Adherent Culture Adult Affect Alginates American Apoptosis Arthritis Biocompatible Cartilage Cartilage injury Cattle Cells Chemicals Chondrocytes Collagen Data Defect Degenerative polyarthritis Deposition Development Dimensions Disease Dose Elements Embryo Engineering Environmental Risk Factor Event Extracellular Matrix Gene Targeting Generations Genes Genetic Genetic Determinism Genetic Programming Glycosaminoglycans Goals Growth Harvest Hypertrophy Image In Vitro Inflammatory Injury Knock-out Knockout Mice Knowledge Laboratories MAP Kinase Gene MAP2K6 gene MAPK14 gene MAPK8 gene MAPK9 gene MEKs Mechanics Mentors Methods Microscopy Mission Modeling Mus Mutation NF-kappa B NFKB Signaling Pathway National Institute of Arthritis and Musculoskeletal and Skin Diseases Operative Surgical Procedures Pathway interactions Peptide Hydrolases Pharmacologic Substance Phenotype Population Procedures Process Production Property Proteoglycan Relative (related person)Research Signal Pathway Signal Transduction Solutions Stress Structure System TNF gene Techniques Thinking Time Tissue Engineering Tissues Training Transgenic Organisms Trauma Tumor Necrosis Factor-alpha United States National Institutes of Health Weight-Bearing state aggrecan articular cartilage base career cartilage development cartilage repair crosslink experience gene therapy human TNF protein in vitro Model knockout gene mathematical model multidisciplinary novel osteochondral repair osteochondral tissue programs repaired response scaffold second harmonic skills tool two-photon

项目摘要

DESCRIPTION (provided by applicant): This proposal's long-term objectives are: 1) to provide basic knowledge about how manipulation of Inflammatory signaling pathways can affect the mechanical properties of chondrocyte created neo-matrix, and 2) to create a tissue-engineered surgical solution to restore osteochondral defects. These objectives will be through two specific aims: construction and analysis of tissue-engineered cartilage using 1) genetically-modified mouse chondrocytes with knockout of MKK/MEK3; and 2) pharmacologic inhibition of key signaling pathways (p38 and JNK MARK) that regulate proteoglycan and collagen deposition by chondrocytes in tissue-engineered cartilage. Analysis will encompass mechanical characterization, collagen, crosslink, and GAG content, multiphoton microscopy, and mathematical modeling of the tissue. The goals of this proposal are relevant to the mission of the NIH and NIAMS. Functional engineered cartilage would greatly alleviate cartilage damage which affects ~10% of the population (1). Information gained about the genetic determinants of cartilage neo-matrix formation would increase understanding of cartilage degradation in rheumatoid and osteoarthritis. This proposal has tissue engineering, multiphoton imaging, and mathematical modeling components, and is a multidisciplinary effort to advance basic knowledge of chondrocyte genetic programming in disease and repair processes. Chondrocytes from mice with knockouts and mutations of TNF-alpha and IL-1beta signaling elements will be harvested and seeded within alginate scaffolds to allow neo-matrix development. Tissue mechanical properties, collagen, proteoglycan, and crosslink content will be assessed during culture under normal and transgenic phenotype, and with or without pharmacologic inhibition of the same signaling pathways, known to affect extracellular matrix production (2). Multiphoton imaging will be used to assess collagen deposition and network structure (using signal from second harmonic generation) and collagen crosslink formation (through two-photon autofluorescence signal). Data from the tissue-engineered cartilage will be fit to a cartilage growth mixture model, previously developed to assess native cartilage (3). Every year, cartilage Injury affects nearly 1 million Americans resulting in 200,000 procedures. Injury often leads cryptically to osteoarthritis (1). Tissue-engineered cartilage is a promising method to examine chondrocyte-controlled cartilage degradation and repair. Understanding these processes in vitro would help clinicians control and repair cartilage through pharmaceutical, genetic, and surgical manipulations.

描述（由申请人提供）：该提案的长期目标是：1）提供关于炎症信号通路的操纵如何影响软骨细胞创建的新基质的机械特性的基本知识，以及2）创建组织工程外科手术修复骨软骨缺损的解决方案。这些目标将通过两个具体目标实现：使用 1) 敲除 MKK/MEK3 的转基因小鼠软骨细胞构建和分析组织工程软骨； 2) 对关键信号通路（p38 和 JNK MARK）的药理学抑制，这些信号通路调节组织工程软骨中软骨细胞的蛋白聚糖和胶原蛋白沉积。分析将包括机械特性、胶原蛋白、交联和 GAG 含量、多光子显微镜和组织的数学模型。该提案的目标与 NIH 和 NIAMS 的使命相关。功能性工程软骨将极大地减轻影响约 10% 人口的软骨损伤 (1)。获得有关软骨新基质形成的遗传决定因素的信息将增加对类风湿和骨关节炎中软骨退化的了解。该提案包含组织工程、多光子成像和数学建模组成部分，是一项多学科努力，旨在推进疾病和修复过程中软骨细胞遗传编程的基础知识。来自具有 TNF-α 和 IL-1β 信号元件敲除和突变的小鼠的软骨细胞将被收获并接种在藻酸盐支架内，以允许新基质的发育。将在正常和转基因表型下的培养过程中评估组织机械特性、胶原蛋白、蛋白聚糖和交联含量，并在有或没有药物抑制相同信号通路（已知会影响细胞外基质产生）的情况下进行评估 (2)。多光子成像将用于评估胶原蛋白沉积和网络结构（使用二次谐波产生的信号）以及胶原蛋白交联形成（通过双光子自发荧光信号）。来自组织工程软骨的数据将适合软骨生长混合物模型，该模型之前是为评估天然软骨而开发的 (3)。每年，软骨损伤影响近 100 万美国人，导致 20 万例手术。损伤常常会神秘地导致骨关节炎 (1)。组织工程软骨是检查软骨细胞控制的软骨退化和修复的一种有前景的方法。了解这些体外过程将有助于临床医生通过药物、遗传和外科手术控制和修复软骨。