Localization and functional interactions of fibronectin and associated basement membrane proteins during embryonic heart development



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Texas Tech University


The early embryonic heart is composed of two, cylindrical epithelial layers, an inner endothelium and an outer myocardium. The cardiac jelly (CJ), an a cellular accumulation of extra cellular matrix (ECM), fills the space between the two layers. All cardiac endothelial cells (EC) do not follow an identical course of differentiation. Some of the EC of the atrio ventricular (AV) and outflow tract (OT) regions undergo an epithelial-mesenchymal transition to form mobile cardiac mesenchymal (CM) cells while in other regions (e.g., ventricle), EC differentiate along the lines of a typical vascular endothelium. The mechanisms controlling the biphasic differentiation of EC and the subsequent migration of CM cells are poorly understood.

Although the CJ lies between two epithelia and is spatially equivalent to a basement membrane (BM), it has not traditionally been considered to be organized into a BM-like structure. The potential significance of this observation lies in the possibility that BM, or their individual components (i.e., fibronectin (FN), laminin (LH), type IV collagen, and heparan sulfate proteoglycan (HSPG)), may function as the regulatory site o f " epithelial differentiation and morphogenesis.

The temporal changes in the localization and the functional involvement of the BM components FN, LM, type IV collagen, and HSPG were investigated with respect to (1) EC attachment and differentiation and (2) CM cell attachment and migration. A cryofixation technique was developed in order to determine the in situ immunohistochemical distribution of the BM components in the CJ. Results indicated that the CJ exists IJS the fusion between a larger, myocardially derived BM and an attenuated, endothelial associated BM. Except for FN, the individual BM components were not all present during early stages. Instead, they appeared in a sequential manner, suggesting that all components of an adult-type B'A are not required to initiate the assembly of a structural and functional BM during development. In the AV and OT, FN lippeared as a progressively expanding gradient of material with the greatest density nearer the myocardium.

An in vitro collagen gel bioassay was employed to directly test whether FN or other BM components play a role in EC and CM cell attachment, differentiation, and migration. Biochemical analysis and results from experiments using probes directed against the BH components indicated that mora than one mechanism of attachment, involving FN and/or HSPG, exist during EC development and CM cell migration.