CELL--ECM INTERACTIONS IN MYOCARDIAL FORM AND FUNCTION

细胞：细胞外基质（ECM）的相互作用发挥了关键作用， 在早期心脏形态发生中， 鸟类模型。 这些相互作用在一个 如果我们要开始研究 先天性心脏病的发病机制 我们建议， 在哺乳动物胚胎（小鼠和大鼠）中， 主要已知成分的发展外观 细胞外基质（I、III、IV型胶原，透明质酸， 层粘连蛋白，纤维连接蛋白，基底膜硫酸乙酰肝素 蛋白聚糖），因为它们与心前细胞迁移有关， 心脏形态发生 A）我们将使用整个胚胎安装在 与用特异性抗体标记的连续切片的组合 ECM的每个组成部分，以定义发育外观 这些分子的三维分布 到心前中胚层 B）我们将在现场使用 与相关ECM mRNA的cDNA探针杂交 成分（胶原蛋白I，III，IV，纤连蛋白，层粘连蛋白）定位 细胞的时空分布 影响心前区细胞群， 早期心脏形态发生 此外，我们将研究 心前间质的免疫组织化学标记物， 将这些细胞与周围的侧板中胚层区分开 在迁移和分化的过程中， 利用这些细胞迁移的独特特性 成肌细胞，我们将使用单克隆抗体针对 胚胎肌球蛋白、结蛋白中间丝和成肌细胞 特异性细胞表面表位，以：a）鉴定心前区 间充质作为侧板中胚层的亚群，B） 定义心前细胞迁移的精确区域，和c） 确定这些表位表达的序列， 心前细胞分化 其他单克隆抗体 将针对心前间质发展，如果 需要，为了识别心脏病的早期阶段， 形态发生 最后，我们将在原位研究细胞- 参与心前细胞迁移的ECM相互作用， 分化和早期收缩功能在哺乳动物 胚胎 我们将培养整个胚胎， 在抑制剂存在下通过心脏循环的原肠胚形成 正常细胞：ECM相互作用。 将对胚胎进行评估， 所导致的结构改变和治疗效果将 通过心肌的连续运动分析来量化 收缩功能 这将提供对 ECM组分对正常结构的相对贡献 功能发展。

Cell:extracellular matrix (ECM) interactions play a critical role in early cardiac morphogenesis but what is known has been derived from avian models. Elucidation of these interactions in a mammalian model is essential if we are to begin to investigate the mechanisms of congenital heart disease. We propose to delineate, in the mammalian embryo (mouse and rat), the distribution and developmental appearance of the major known components of the extracellular matrix (collagen types I, III, IV, hyaluronic acid, laminin, fibronectin, basement membrane heparan sulfate proteoglycan) as they relate to precardiac cell migration an early cardiac morphogenesis. A) We will employ whole embryo mounts in combination with serial sections labelled with antibodies specific to each component of the ECM to define the developmental appearance and three dimensional distribution of these molecules in relation to the precardiac mesoderm. B) We will then utilize in situ hybridization with cDNA probes for the mRNA's of relevant ECM components (Collagen I,III,IV, fibronectin, laminin) to localize the temporal and geographical distribution of cells capable of influencing the precardiac cell population during the period of early cardiac morphogenesis. In addition, we will examine immunohistochemical markers for precardiac mesenchyme that will distinguish these cells from the surrounding lateral plate mesoderm at progressive stages in their migration and differentiation. Exploiting the unique character of these cells as migrating myoblasts, we will use monoclonal antibodies directed against embryonic myosins, desmin intermediate filaments, and myoblast specific cell surface epitopes to: a) identify the precardiac mesenchyme as a subpopulation of the lateral plate mesoderm, b) define the precise domain of precardiac cell migration, and c) determine the sequence in which these epitopes are expressed during precardiac cell differentiation. Additional monoclonal antibodies directed against precardiac mesenchyme will be developed, if needed, in order to recognize earlier stages of cardiac morphogenesis. Finally, we will investigate, in situ, the cell- ECM interactions involved in precardiac cell migration, differentiation, and early contractile function in the mammalian embryo. We will culture whole embryos during the period from gastrulation through cardiac looping in the presence of inhibitors of normal cell:ECM interaction. Embryos will be evaluated for resulting structural alteration and the effects of treatment will be quantified through sequential motion analysis of myocardial contractile function. This should provide evaluation of the relative contributions of the ECM components to normal structural and functional development.