The luminal plasmalemma of continuous and fenestrated microvascular endothelia (obtained from bovines and rats) will be mapped at the macromolecular level by selecting promising antigens (glycoproteins and proteoglycans) from cultured cells for the generation of monoclonal or polyclonal antibodies. The latter will be used to localize the cognate antigens to specific differentiated plasmalemmal microdomains, e.g., plasmalemmal vesicles, transendothelial channels, apertured fenestrae and intercellular junctions. Immunocytochemical tests will be carried out in the intact animal (rat) to find out whether the antigen distribution is the same in situ as in culture. Since we have evidence suggesting that albumin is transported across the endothelium by receptor-mediated transcytosis, we'll attempt to identify and characterize this receptor, and to extend the inquiry to the transcytosis of other plasma proteins. Antibodies to the putative receptor will be tested to find out whether they block ligand binding and transport. Experiments will be carried out on the mesenteric microvasculature of the frog in an attempt to localize the sites of exit (small pores) for albumin and other smaller proteins (e.g., peroxidases). Albumin (or peroxidases) conjugated to photoactivatable groups will be perfused in individual capillaries, crosslinked to surrounding structures (by light exposure) seconds after the beginning of perfusion, and localized close to its exit sites by immunocytochemistry, using a tagged antibody (or a peroxidatic reaction). The mesenteric microvasculature will also be used to investigate the ability of pericytes to control local blood flow. Work on the contraction-associated proteins of pericytes will be expanded to new antigens, especially vascular smooth muscle-specific Alpha-actin, to find out how close these cells come to regular vascular smooth muscle. TEM and SEM work on endothelial surface fine structure will be continued and extended to antigen localization via tagged (or untagged) antibodies.

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
Method to Extend Research in Time (MERIT) Award (R37)
Project #
5R37HL017080-18
Application #
3485518
Study Section
Pathology A Study Section (PTHA)
Project Start
1977-03-01
Project End
1992-02-29
Budget Start
1991-03-01
Budget End
1992-02-29
Support Year
18
Fiscal Year
1991
Total Cost
Indirect Cost
Name
University of California San Diego
Department
Type
Schools of Medicine
DUNS #
077758407
City
La Jolla
State
CA
Country
United States
Zip Code
92093
Predescu, Sanda A; Predescu, Dan N; Timblin, Barbara K et al. (2003) Intersectin regulates fission and internalization of caveolae in endothelial cells. Mol Biol Cell 14:4997-5010
Stan, Radu-Virgil (2002) Structure and function of endothelial caveolae. Microsc Res Tech 57:350-64
Stan, R V; Arden, K C; Palade, G E (2001) cDNA and protein sequence, genomic organization, and analysis of cis regulatory elements of mouse and human PLVAP genes. Genomics 72:304-13
Predescu, S A; Predescu, D N; Palade, G E (2001) Endothelial transcytotic machinery involves supramolecular protein-lipid complexes. Mol Biol Cell 12:1019-33
Stan, R V; Kubitza, M; Palade, G E (1999) PV-1 is a component of the fenestral and stomatal diaphragms in fenestrated endothelia. Proc Natl Acad Sci U S A 96:13203-7
Stan, R V; Ghitescu, L; Jacobson, B S et al. (1999) Isolation, cloning, and localization of rat PV-1, a novel endothelial caveolar protein. J Cell Biol 145:1189-98
Roberts, W G; Delaat, J; Nagane, M et al. (1998) Host microvasculature influence on tumor vascular morphology and endothelial gene expression. Am J Pathol 153:1239-48
Predescu, D; Predescu, S; McQuistan, T et al. (1998) Transcytosis of alpha1-acidic glycoprotein in the continuous microvascular endothelium. Proc Natl Acad Sci U S A 95:6175-80
Stan, R V; Roberts, W G; Predescu, D et al. (1997) Immunoisolation and partial characterization of endothelial plasmalemmal vesicles (caveolae). Mol Biol Cell 8:595-605
Predescu, S A; Predescu, D N; Palade, G E (1997) Plasmalemmal vesicles function as transcytotic carriers for small proteins in the continuous endothelium. Am J Physiol 272:H937-49

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