This proposal aims to define the contribution of the pulmonary microvascular endothelium to size and charge permselectivity in the lung. The hypothesis to be tested is that anionic glycoproteins in glycosaminoglycans on the surface of the plulmonary microvascular endothelium are organized in discrete domains which may influence the routes taken by macromolecules across the capillary wall. Pathways for neutral and charged macromolecules across the capillary walls will be identified qualitatively by electron microscopy using electron-dense permeability probes of different molecular size and net electrical charge. The experiments will be done in the isolated perfused rat lung preparation to avoid systemic effects resulting from interaction between charged probes and blood constituents. Quantitative assessment of transcapillary movement of permeability probes will be done by morphometric measurements at different time intervals and/or by measurements of lung radioactivity in lungs perfused with radioactive permeability probes. Selective enzymatic removal of glycoproteins or glycosaminoglycans on the endothelium will provide insights in the biochemical nature of the constituents of the charged barrier. Using lectins bound to colloidal gold the distribution of carbohydrate units of different glycoproteins on the microvascular endothelium and adjacent basement membrane will be examined in thin sections of lungs. The overall goal of the project is to identify new structural determinants of lung permeability and gain information for future studies on mechanism of endothelial damage in differnt types of lung injury.
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