Effect of Versican Mutation on Limb Development In Vitro

Versacan 突变对体外肢体发育的影响

• 批准号: 6497006
• 负责人: ANTHONY A CAPEHART
• 金额: $ 13.95万
• 依托单位: EAST CAROLINA UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2002
• 资助国家: 美国
• 起止时间: 2002-08-01 至 2004-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6497006
• 关键词: cartilage development; cell differentiation; developmental genetics; embryo /fetus; embryogenesis; extracellular matrix; gene expression; gene mutation; histochemistry /cytochemistry; immunocytochemistry; in situ hybridization; intermolecular interaction; laboratory mouse; limbs; muscle; neurogenesis; protein localization; protein structure function; proteoglycan; tissue /cell culture

DESCRIPTION (provided by applicant): The long-term objective of the proposed research is to define the role of cell-extracellular matrix ECM) interactions in differentiation of cartilage, muscle, and nerve in their proper spatial relationships luring embryonic limb development. Recently, a recessive lethal insertional mutation in the Cspg2 gene was identified in a transgenic mouse line, hdf (heart defect). Cspg2 encodes the core protein of the chondroitin sulfate proteoglycan, versican, and hdf mutation results in loss of expression of the mature proteoglycan. Versican has been suggested to play a role in chondrogenesis and cellular patterning. However, little is known regarding details of versican function in the embryonic ECM. The hdf mutation offers a unique opportunity to explore versican function in the limb. The specific aims of this proposal are to first analyze the spatial and temporal expression of versican during limb development in the hdf mouse. LacZ reporter histochemistry, in situ hybridization, and immunohistochemistry will be utilized to determine versican expression during limb development in carefully staged hemizygous hdf and wildtype mice to correlate localization with its hypothesized functions. Second, the role of the mature versican proteoglycan in precartilage aggregation during limb chondrogenesis in vitro will be evaluated. micromass cultures of hdf limb mesenchyme will be utilized to determine if this mutation results in inhibition of prechondrogenic mesenchymal condensation and subsequent cartilage differentiation.

描述（由申请人提供）：所提出的研究的长期目标是确定细胞-细胞外基质（ECM）相互作用在软骨、肌肉和神经分化中的作用，这些分化在诱导胚胎肢体发育的适当空间关系中。最近，在转基因小鼠品系hdf（心脏缺陷）中发现了Cspg 2基因的隐性致死插入突变。cspg 2编码硫酸软骨素蛋白聚糖多功能蛋白聚糖的核心蛋白，hdf突变导致成熟蛋白聚糖的表达丧失。Versican被认为在软骨形成和细胞模式中发挥作用。然而，鲜为人知的是，关于多功能蛋白聚糖在胚胎细胞外基质中的功能的细节。hdf突变提供了一个独特的机会来探索多功能蛋白聚糖在肢体中的功能。本研究的具体目的是首先分析多功能蛋白聚糖在hdf小鼠肢体发育过程中的时空表达。LacZ报告基因组织化学、原位杂交和免疫组织化学将用于确定多功能蛋白聚糖在仔细分期的半合子hdf和野生型小鼠肢体发育过程中的表达，以将定位与其假设的功能相关联。第二，将评估体外肢体软骨形成过程中成熟多功能蛋白聚糖在前软骨聚集中的作用。将利用HDF肢体间充质的微团培养物来确定该突变是否导致抑制软骨形成前间充质凝聚和随后的软骨分化。