The exchange of macromolecules and nutrients across mammalian capillary endothelium is mediated by cytoplasmic vesicles. This mode of blood-tissue exchange may play a significant role in maintaining normal tissue function. Yet, the mechanism, regulation and specificity of the vesicular mode of capillary permeability is poorly understood due to the difficulty in studying this process. We have developed new methodologies which permit the accurate quantitation of vesicular activity in vitro employing a system of freshly isolated capillary endothelium. We have developed techniques for fluorescently labelling numerous macromolecules for the biochemical analysis of vesicular activity as well as the direct visualization of molecular-endothelial cell interaction using the fluorescence microscope. The purpose of this research project is to determine the basic mechanisms which exist for the regulation and specificity of vesicular transport. Using capillary endothelium isolated from fat, muscle, brain and lung we will examine the regulation of vesicular transport by hormones, temperature, cations, vasoactive substances, blood factors, cellular metabolites and endothelial specific growth (angiogenesis) factors. These experiments will determine whether changes in vesicular transport rates play a role in the observation of capillary leakiness observed during inflammation, wounding and tissue neovascularization. We will examine the specificity of vesicular transport. Experiments are designed to determine whether capillary endothelium exhibit specific receptors for hormones and macromolecules which mediate their subsequent transendothelial transport. The specificity of vesicular transport for molecules of different molecular weight, shape and charge will be examined. The interaction of proteins prior to vesicular transport will be studied to determine whether certain proteins can inhibit or stimulate the capillary permeability of other macromolecules. These experiments will add to our understanding of the mechanism of altered endothelial cell permeability observed in diabetes mellitus and atherosclerosis. These physiologic and pathophysiologic studies will expand our knowledge concerning the role of capillary endothelial vesicular activity in normal and diseased states.
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