IDENTIFICATION OF DIFFERENTIALLY EXPRESSED CRITICAL GENES IN CHONDROCLASTS AND OS

软骨细胞和 OS 中差异表达的关键基因的鉴定

• 批准号: 8111107
• 负责人: HICHAM M DRISSI
• 金额: $ 16.62万
• 依托单位: UNIVERSITY OF CONNECTICUT SCH OF MED/DNT
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-07-15 至 2012-08-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8111107
• 关键词: Address; Adult; Affect; Biological Assay; Bone Growth; Bone Marrow; Bone Matrix; Bone callus; Bone remodeling; Calcified; Candidate Disease Gene; Cartilage; Cartilage Matrix; Cattle; Cell physiology; Cells; Characteristics; Chondrocytes; Clinical; Data; Disease; Environment; Event; Exhibits; Femoral Fractures; Fracture; Fracture Healing; Gene Expression; Gene Expression Profile; Genes; Goals; Harvest; Healed; Histologic; In Vitro; Incubated; Lasers; Lead; Measures; Mediating; Modality; Modeling; Molecular; Molecular Profiling; Mus; Myelogenous; Osteoclasts; Osteogenesis; Pattern; Population; Process; Proteins; RNA; RNA Interference; Radiolabeled; Relative (related person); Reverse Transcriptase Polymerase Chain Reaction; Role; Sampling; Skeletal Development; Slice; Speed; Splenocyte; Staging; Staining method; Stains; System; Testing; Time; Virus; aged; bone; bone cell; cartilage cell; cell type; cellular transduction; design; healing; in vivo; laser capture microdissection; long bone; loss of function; mouse model; novel; novel therapeutics; precursor cell; progenitor; public health relevance; radiotracer; repaired; research study; selective expression

DESCRIPTION (provided by applicant): During endochondral ossification, mineralized cartilage matrix is resorbed prior to bone formation and remodeling. While this cartilage matrix resorption is mediated by cells that exhibit many phenotypic features of osteoclasts, a longstanding question remains as to whether differences exist in the gene expression between these chondroclasts and the better characterized bone resorbing osteoclasts. To address this question we will use our mouse femoral fracture model, in which calcified cartilage matrices in the early callus will be remodeled prior to the remodeling of bony callus, during late stages of healing. While histological evidence has shown distinct identities of chondroclasts and osteoclasts, little information is available regarding the molecular characteristics that differentiate their respective functions. Our goal is to compare the gene expression profiles of TRAP-positive cells resorbing mineralized cartilage matrix (chondroclasts) with that of cells resorbing bone matrix (osteoclasts). We will further determine which of these differentially expressed genes is important for the respective functions of chondrocytes versus osteoclasts. These studies will be performed on cells that are removed from histologic samples of fractured mouse bones. We believe that the matrix and environment surrounding chondroclasts and osteoclasts critically influences the expression profiles of these cells, which almost certainly derive from a common progenitor. Therefore, we have designed our studies to examine differences in gene expression in cells that are removed directly from their in vivo environment so that we can best appreciate differences in their gene expression profiles. Our central hypothesis posits that osteoclasts and chondroclasts have distinct patterns of gene expression, which facilitate their unique functions. We propose the following two Specific Aims to test this hypothesis: Specific Aim1: To examine the molecular differences between laser-captured TRAP-positive cells resorbing cartilage or bone matrices during fracture healing in mice. We hypothesize that TRAP-positive cells resorbing calcified cartilage matrix will exhibit a different gene expression profile than TRAP-positive cells resorbing bone matrix. Specific Aim2: To establish functional differences between chondroclasts and osteoclasts isolated from fractured femurs. We hypothesize that functional differences exist between osteoclast and chondroclasts, which facilitate matrix-specific resorption. Our proposed studies will for the first time demonstrate, using a fracture healing mouse model, whether there are functional differences in the molecular mechanisms regulating cartilage versus bone remodeling during endochondral ossification post-natally. PUBLIC HEALTH RELEVANCE: Skeletal development and fracture repair of long bones require successive cellular events that not only involve cartilage and bone cells, but also cells that remodel cartilage (chondroclasts) and bone (osteoclasts) matrices. Here we propose to identify molecular differences between these two cell types and further examine the critical genes that are required for chondroclasts to mediate cartilage matrix remodeling for proper fracture healing. Clinical evidence is emerging demonstrating the importance of cartilage matrix remodeling for proper skeletal development and normal fracture healing. Data obtained from these studies may help develop novel therapeutic modalities to accelerate fracture repair in young and aged populations.

描述（由申请方提供）：在软骨内骨化过程中，矿化软骨基质在骨形成和重塑之前被再吸收。虽然这种软骨基质的再吸收是由表现出破骨细胞的许多表型特征的细胞介导的，但一个长期存在的问题是，这些破软骨细胞和更好表征的骨再吸收破骨细胞之间的基因表达是否存在差异。为了解决这个问题，我们将使用我们的小鼠股骨骨折模型，其中在愈合的晚期阶段，在早期骨痂中的钙化软骨基质将在骨痂重塑之前重塑。虽然组织学证据显示了破软骨细胞和破骨细胞的不同身份，但关于区分其各自功能的分子特征的信息很少。我们的目标是比较TRAP阳性细胞再吸收矿化软骨基质（破软骨细胞）与细胞再吸收骨基质（破骨细胞）的基因表达谱。我们将进一步确定这些差异表达的基因是重要的软骨细胞与破骨细胞的各自的功能。这些研究将在从小鼠骨折的组织学样本中取出的细胞上进行。我们相信，破软骨细胞和破骨细胞周围的基质和环境严重影响这些细胞的表达谱，这几乎可以肯定来自一个共同的祖先。因此，我们设计了我们的研究，以检查直接从体内环境中取出的细胞中基因表达的差异，以便我们能够最好地了解其基因表达谱的差异。我们的中心假设假定破骨细胞和破软骨细胞具有不同的基因表达模式，这有助于其独特的功能。 我们提出了以下两个具体目标来检验这一假设：具体目标1：研究激光捕获的TRAP阳性细胞在小鼠骨折愈合过程中吸收软骨或骨基质的分子差异。 我们假设，TRAP阳性细胞再吸收钙化软骨基质将表现出不同的基因表达谱比TRAP阳性细胞再吸收骨基质。 具体目标2：建立从骨折股骨分离的破软骨细胞和破骨细胞之间的功能差异。 我们假设破骨细胞和破软骨细胞之间存在功能差异，这有助于基质特异性吸收。 我们提出的研究将首次证明，使用骨折愈合小鼠模型，是否有功能差异的分子机制调节软骨与骨重塑在endochondrialossification出生后。 公共卫生关系：长骨的骨骼发育和骨折修复需要连续的细胞事件，不仅涉及软骨和骨细胞，而且还涉及重塑软骨（破软骨细胞）和骨（破骨细胞）基质的细胞。在这里，我们建议确定这两种细胞类型之间的分子差异，并进一步研究破软骨细胞介导软骨基质重塑以实现适当骨折愈合所需的关键基因。临床证据显示软骨基质重塑对于骨骼发育和骨折愈合的重要性。从这些研究中获得的数据可能有助于开发新的治疗方法，以加速年轻和老年人群的骨折修复。