CELL--ECM INTERACTIONS IN MYOCARDIAL FORM AND FUNCTION

细胞与细胞外基质（ECM）的相互作用起着关键作用

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.