Bidirectional fusion of the heart-forming fields in the developing chick embryo.

It is generally thought that the early pre-tubular chick heart is formed by fusion of the anterior or cephalic limits of the paired cardiogenic fields. However, this study shows that the heart fields initially fuse at their midpoint to form a transitory "butterfly"-shaped, cardiogenic structure. Fusion then progresses bi-directionally along the longitudinal axis in both cranial and caudal directions. Using in vivo labeling, we demonstrate that cells along the ventral fusion line are highly motile, crossing future primitive segments. We found that mesoderm cells migrated cephalically from the unfused tips of the anterior/cephalic wings into the head mesenchyme in the region that has been called the secondary heart field. Perturbing the anterior/cranial fusion results in formation of a bi-conal heart. A theoretical role of the ventral fusion line acting as a "heart organizer" and its role in cardia bifida is discussed.

[Teratology and epidemiology in the study of environmental teratogens]. / La teratología y la epidemiología en el estudio de los teratógenos ambientales.

The processes of cellular migration, cellular differentiation and cellular multiplication are studied, since these are the basic developmental processes upon which teratogenic agents act resulting in congenital malformations. We also carefully analyze the interactions between teratogen-embryo in order to establish adequate parameters for analysis of environmental teratogens, as well as experimental teratogenesis and epidemiology. Information on the pathogenesis of congenital malformations obtained from experimental teratology in an adequate biological model, can be extrapolated to the human. The etiology of congenital malformations resulting from environmental teratogens can only be elucidated through epidemiology, since there is species specificity. Such a study must fulfill the following prerequisites: diagnosis of the congenital malformation, ruling out genetic factors in the family tree and determination of the exact time of exposure to the possible teratogen during the pregnancy.

Temporal and spatial asymmetries in the initial distribution of mesenchyme cells in the atrioventricular canal cushions of the developing chick heart.

BACKGROUND: We investigated potential early asymmetries in the distribution of mesenchymal cells within the inferior and superior AV cushions in the developing chick heart. METHODS: Chick embryos stages 16-20 HH were fixed, embedded in polyacrylamide, and the cell nuclei stained with propidium iodide. Cells counts were determined within the cardiac jelly of the atrioventricular canal (AV) by laser confocal microscopy in coronal planes spanning its entire length. RESULTS: Our data show at the different stages studied, 16-20 HH, that the inferior AV cushion invariably contains more cells than the superior AV cushion. In the inferior cushion, the cell distribution is bimodal, i.e., the proximal and distal regions have more mesenchymal cells than the middle part of the AV canal. In the superior cushion, there is a increasing gradient of mesenchymal cells along the longitudinal axis from the atrium to the ventricle. CONCLUSIONS: Our findings reveal that the temporal and spatial characteristics of mesenchyme formation in the inferior vs. superior AV cushion are different. This asymmetry suggests several potential hypotheses: (1) the distribution of the inducer molecule or its receptor has a distribution similar to that of mesenchymal cells, (2) the extracellular matrix has a differential composition or regionally-specific physical associations, (3) the endocardium is heterogeneous with respect to transformation capacity, or (4) these patterns result from an earlier inductive event. The potential importance of the observed asymmetries in the distribution of AV mesenchyme is discussed relative to localization patterns of molecules critical to successful cardiac morphogenesis and remodeling.

Primitive interventricular septum, its primordium, and its contribution in the definitive interventricular septum: in vivo labelling study in the chick embryo heart.

BACKGROUND: Because the studies on the embryological development of the primitive interventricular septum have been done with postmortem material, we do not know the site within the cardiac tube and the developmental stage at which the primordium appears and its anatomical manifestation in the mature heart. Consequently, we do not know its real contribution to the constitution of the definitive interventricular septum. METHODS: With this purpose, we selected an adequate biological model, the chick embryo heart and the in vivo labelling technique. We placed a label of gelatin India ink in the ventral fusion line of both cardiac primordia at the level of the interventricular grooves in the straight tube heart (stage 9+HH), and we traced the ink up to the mature heart (stage 36HH). We made histological sections of some hearts, of the zone where the label was found to investigate the first morphological manifestation of the primitive interventricular septum. We also made microdissections and scanning electron microscopic studies. RESULTS: The label placed at stage 9+HH in the ventral fusion line of both cardiac primordia, at the level of the interventricular grooves, was found at stage 14HH in the greater curvature of the looped heart, opposite the left interventricular groove. This label at stage 17HH was found in the apical trabecular region of the first cardiac septum (8-shaped septum) and in the mature heart (stage 36HH) in the definitive interventricular septum at the limit between the basal and the medial third of the definitive interventricular septum. CONCLUSIONS: Firstly, the primordium of the primitive interventricular septum appears at stage 9+HH, in the ventral fusion line of both cardiac primordia at the level of the interventricular grooves. Secondly, its first morphological manifestation takes place at stage 17HH, and it forms the apical trabeculated region of the first cardiac septum (8-shaped septum). Thirdly, the primitive interventricular septum gives origin to the middle and apical third of the definitive interventricular septum.

Identification of an autocrine signaling pathway that amplifies induction of endocardial cushion tissue in the avian heart.

Endocardial cushion tissue is formed by an epithelial-mesenchymal transformation of endocardial cells, a process which results from an inductive interaction between the myocardium and endocardium within the atrioventricular (AV) and outflow tract (OT) regions of the heart. We report here that a protein previously found to be required for myocardially induced transformation of endocardial cells in vitro, ES/130, is highly expressed within the AV and OT regions not only by myocardial cells, but also by the endocardium and its mesenchymal progeny. Given these findings and others, we have tested the hypothesis that endocardial cushion tissue secretes factors which autoregulate its transformation to mesenchyme. Endocardial cushion tissue was cultured and its conditioned growth medium was harvested and applied to nontransformed endocardial cells maintained in the absence of the inductive myocardium. This treatment resulted in endocardial cell invasion into three-dimensional collagen gels plus increased expression of proteins associated with endocardial cell transformation in vivo. Whereas endocardial cushion tissue was found to express ES/130 protein in vivo and in vitro, minimal detection of ES/130 in its conditioned growth medium was observed in immunoblots. Attempts to inhibit the mesenchyme-promoting activity of the conditioned medium with ES/130 antisense were unsuccessful. However, strong intracellular ES/130 expression was detected in endocardial cells, and this expression correlated with the ability of endocardial cells to transform. For example, the minority of endocardial cultures that failed to transform in response to conditioned medium treatment also failed to undergo increased expression of ES/130. These observations are interpreted to suggest that (i) endocardial cushion tissue secretes factors that promote its transformation to mesenchyme, and (ii) while endocardial cushion tissue appears to signal through secretion of factors other than or in addition to ES/130, intracellular ES/130 expression nevertheless may be a target endocardial cell response required for endocardial cell transformation.

Living morphogenesis of the ventricles and congenital pathology of their component parts.

Living morphogenetic studies show that each definitive ventricle is constructed from different primitive cardiac segments, and each has its specific anatomical features. These ventricular segments are the atrioventricular junction; the primitive inlet segment, part of the primary heart tube, which initially provides the inlets of each ventricle; the primitive outlet segment, which gives rise to both ventricular outlets; and the apical trabeculated regions of the right and left ventricles which grow from the primary heart tube, respectively. In this review, we describe regional pathology based on the relationship of these primitive ventricular components. We propose that the abnormal morphogenesis of one of these segments gives origin to regional ventricular pathology. For example, abnormal embryogenesis of the atrioventricular canal produces malformations of the atrioventricular junctions, such as double inlet ventricle, absence of one atrioventricular connection, and straddling and overriding atrioventricular valves. Similarly, abnormal morphogenesis of the primitive outlet segment gives rise to malformations of the subarterial region of each ventricle, along with the valves guarding these vessels. The principal anatomical features of these malformations of the ventricular inlets and outlets are described, and their possible morphogenesis is discussed. Due to the fact that the apical trabeculated region of each ventricle arises from a separate primitive segment, each ventricle can be identified according to the pattern of its apical trabeculations. This feature is crucial in the elucidation of complex congenital pathology, such as discordant atrioventricular connections.