MECHANISM OF CHONDROPROGENITOR CELL CONDENSATION

软骨祖细胞凝聚机制

基本信息

批准号：
6150540
负责人：
ROCKY S TUAN
金额：
$ 21.5万
依托单位：
THOMAS JEFFERSON UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
1999
资助国家：
美国
起止时间：
1999-02-01 至 2004-01-31
项目状态：
已结题

项目摘要

The condensation of chondroprogenitor cells is one of the earliest steps in endochondral skeletal development. In the developing limb bud, the core of condensing mesenchymal cells subsequently differentiate into chondrocytes to give rise to the cartilage anlage. The cellular condensation step is crucial to chondrogenesis, since both in vivo and in vitro perturbations of condensation inhibit mesenchymal chondrogenesis. We have previously demonstrated that N-cadherin, a cell-cell adhesion protein, is functionally involved in limb mesenchymal condensation and chondrogenesis. Cadherins are membrane glycoproteins which mediate cell adhesion in a Ca2+-dependent, homotypic manner, and have been postulated to act as morphoregulatory molecules during development. Cadherins function as a complex with the cytosolic, alpha-, -beta- and gamma-catenins, and the complex is generally believed to function in adhesion-mediated signaling. In this proposal, we aim to further examine the mechanism and function of N-cadherin-mediated mesenchymal cell interactions in chondrogenesis, using two systems of mesenchymal chondrogenesis: 1) chick embryonic limb mesenchyme, which is primed to undergo chondrogenesis when cultured at high density; and 2) the murine multipotential cell line, C3H1OT1/2, which will undergo chondrogenesis when plated as high density micromass and treated with bone morphogenetic protein-2 (BMP-2). The high density requirement of both systems underscores the importance of intimate cell-cell interactions. The specific aims are: 1) To determine the subcellular site of N-cadherin-mediated activity by examining the effects of expressing structural variants of N-cadherin; 2) To examine the functional role of beta-catenin in cellular condensation and chondrogenesis by analyzing its subcellular distribution, phosphorylation status, and association with N-cadherin, and the effect of expressing its structural variants; 3) To demonstrate that N- cadherin-mediated cell adhesion is pivotally involved in the effects of chondroinductive influences, including the activity of TGF-beta superfamily members, on mesenchymal cells; and 4) To test the hypothesis that N-cadherin expression/activity is a functional marker of chondroprogenitor cells residing in non skeletal tissues. These proposed studies will provide valuable information on the early events of mesenchymal chondrogenesis, and the cellular and molecular mechanisms of cell-cell interactions in general. Investigating the basic mechanism of cartilage development is highly relevant to understanding the etiology of congenital skeletal deformities and the biology of skeletal repair.

软骨源细胞的凝结是最早的步骤之一在软骨骨骼发育中。在发育中的肢体芽中凝结间充质细胞的核心随后分化为软骨细胞引起软骨的厌恶。细胞凝结步骤对软骨发生至关重要，因为体内和凝结的体外扰动抑制间质软骨发生。我们以前已经证明了N-钙粘蛋白，一个细胞细胞粘附蛋白在功能上与肢体间充质有关凝结和软骨发生。钙粘蛋白是膜糖蛋白以Ca2+依赖性，同型方式介导细胞粘附，并且假设在此期间充当形态调节分子发展。钙粘蛋白充当胞质，α-的复合物 -beta-和gamma-catenins，通常认为该建筑群在粘附介导的信号传导中的功能。在此提案中，我们的目标是进一步检查N-钙粘蛋白介导的机制和功能在软骨发生中的间充质细胞相互作用，使用两个系统间充质软骨生成：1）雏鸡胚胎肢体间充质，其中在高密度培养时，可以进行软骨生成；和 2）鼠多能细胞系C3H1OT1/2，将经历当将软骨生成为高密度微量时，并用骨形态发生蛋白-2（BMP-2）。高密度要求这两个系统都强调了亲密细胞细胞的重要性互动。具体目的是：1）确定亚细胞通过检查N-钙粘蛋白介导的活性的部位表达N-钙粘蛋白的结构变体； 2）检查 β-catenin在细胞缩合和软骨发生通过分析其亚细胞分布，磷酸化状态以及与N-钙粘蛋白的关联，效果表达其结构变体； 3）证明n- 钙粘蛋白介导的细胞粘附至关重要地参与软骨诱导的影响，包括TGF-beta的活性超家族成员，在间充质细胞上； 4）检验假设 N-钙粘蛋白的表达/活性是驻留在非骨骼组织中的软骨元基因细胞。这些拟议的研究将提供有关早期事件的宝贵信息间充质软骨生成以及细胞和分子机制一般的细胞 - 细胞相互作用。研究基本机制软骨开发与理解先天性骨骼畸形的病因和骨骼生物学维修。