RENAL MICROVASCULAR ENDOTHELIAL CELL DIFFERENTIATION

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The glomerular and tubular capillary units are specialized filtration apparati having complex matrix organization and cell-cell interactions. Dysfunction of these units is noted in a variety of renal diseases. While a good deal is known about glomerular podocyte and tubular epithelial cell development, junction complex formation, slit diaphragm formation, function and response to injury, much less is known about the development, maintenance and the response to injury of tubular and glomerular endothelial cells (EC). In general, EC dynamics during development and in response to injury are thought to be modulated, in part, by the composition and organization of the surrounding extracellular matrix (ECM) and soluble factors in the local environment. furthermore, this dynamic interaction of EC with themselves and with ECM is thought to involve several classes of cell adhesion molecules (CAMs) such as PECAM-1, substratum adhesion molecules (SAMs) such as the integrins and cell junctional associated molecules (JAMs) such as ZO1. In this proposal we will examine the hierarchical (spatiotemporal) interrelationships of selected moieties of these three protein families in the processes of tube formation by microvascular endothelial cells in vitro and in the fetal and newborn rat kidney. Specifically, we will examine the roles of PECAM-1 in initiating and modulating the processes of in vitro and in vivo angiogenesis. The interactions of the cytoplasmic domain of PECAM-1 with elements of the filamentous and cortical membrane cytoskeleton will also be investigated. We will utilize affinity chromatography with recombinant proteins; stable transfection and over-expression of the cytoplasmic domain and truncation mutants of this PECAM-1 domain in endothelial cells; functional PECAM-1 antibodies and soluble PECAM-1 antigen in inhibition studies; and utilize immunochemical and immunohistochemical methods to assess the localizations and organization of PECAM-1 on the endothelial cell surface.