| Title | The endothelial cell glycocalyx and vascular function |
|---|---|
| Period | 01 / 2001 - unknown |
| Status | Completed |
| Dissertation | Yes |
| Research number | OND1278484 |
| Contact between blood and the luminal endothelial cell surface of the vascular system is mediated by the endothelial cell glycocalyx. Although the early pathogenesis of atherosclerotic vascular disease is associated with loss of proteoglycans from the glycocalyx, studies on its implications for vascular function have been hampered by the biochemical complexity of proteoglycan structure-function relations and technical limitations to measure the contribution of the glycocalyx to the endothelial exchange barrier. Recently developed techniques to study the in vivo contribution of the glycocalyx to the endothelial barrier will allow us to relate atherogenic modification of the glycocalyx to changes in lipoprotein transfer into the vascular wall. In addition, new insight into the relation between proteoglycan structure and protein binding enables us to test for reduced activity of proteoglycan bound antithrombin III (ATIII) and lipoprotein lipase (LPL), leading to increased vascular wall thrombogenicity and elevated plasma levels of atherogenic triglycerides, respectively. Advanced microscopic techniques will be used to identify the contribution of the glycocalyx to the endothelial barrier in skeletal muscle capillaries and coronary arteries during control conditions (C57B1/6 mice), following acute increases in plasma levels of atherogenic lipoproteins (bolus administration of lipoproteins to C57B1/6 mice), and after prolonged lipoprotein exposure (LDLr-/- knockout mice). The effect of atherogenic lipoprotein levels on HSPG-associated ATIII and LPL activity will be studied using wildtype (CHO K1) and glycosaminoglycan deficient (CHO pgsA-745) cultured epithelial cells. Our goal is to demonstrate that glycocalyx modification plays a causal role in atherogenic loss of vascular function. We aim to identify the mechanisms (mechanical barrier, electrostatic repulsion) by which glycocalyx constituents contribute to the permselective nature of the endothelial barrier. In addition we aim to determine whether atherogenic glycocalyx modification impairs the activity of ATIII and LPL. Insight into the mechanisms by which atherogenic conditions alter permselective and protein binding properties of the endothelial cell glycocalyx may lead to new therapies to salvage vascular function when the cardiovascular system is exposed to increased atherogenic stress. Furthermore, the proposed experiments will contribute significantly to our understanding of the role of the glycocalyx in the pathogenesis of atherosclerotic vascular damage and its role in myocardial perfusion and coronary endothelial function, which are main long-term interests of our cardiovascular research group. This research proposal aims to answer two basic questions related to the role of the endothelial cell glycocalyx in vascular function: 1) what is the contribution of atherogenic alteration of glycocalyx thickness to altered permselective properties of the endothelial exchange barrier, and 2) what is the contribution of reduced glycocalyx thickness to loss of proteoglycan associated protein function during the pathogenesis of atherosclerotic disease? - The endothelial cell glycocalyx and vascular wall permeability. The contribution of altered glycocalyx thickness to atherogenic increases in lipoprotein transport across the endothelial lining of blood vessels will be studied in skeletal muscle capillaries using a recently developed wide field microscopic approach (year 1 and 2) as well as in individually perfused larger coronary vessels in collaboration with Prof. Huxley (year 2 and 3), to determine the extent to which observations in capillaries can be extrapolated to larger vessels. To evaluate the relative importance of glycocalyx size and charge density in the transfer of lipoproteins across the vascular endothelial lining, experimental data on transvascular lipoprotein exchange will be analyzed using an mechano-electrochemical model of tracer movement across the glycocalyx developed by Stace and Damiano (year 2 and 3). - The endothelial cell glycocalyx and protein binding. The effect of atherogenic alteration of glycocalyx thickness on proteoglycan dependent protein binding and activity will be studied in a cultured cell system using different Chinese Hamster Ovary (CHO) epithelial cell lines. Recent biochemical insight into the relation between proteoglycan structure and specific protein binding has led to the design of mutant CHO cell lines with specifically altered proteoglycan domains. Wild-type (CHO-K1) and glycocalyx mutant CHO cells (CHO-pgsA-745) will be exposed to atherogenic conditions and its effect on proteoglycan dependent protein binding and activity will be determined in collaboration with experts in biochemical sciences to guarantee optimal use of biochemical techniques and expertise for handling and culturing the CHO cells (collaboration with Prof. Pannekoek, year 1 and 2) studying the interaction of cell surface HSPG with lipoprotein lipase (LPL, Dr. Kastelein, year 2 and 3), and the interaction of HSPG with antithrombin III (ATIII, Prof. Rosenberg, year 2 and 3). |
| Secretariat | Department of Biomedical Engineering & Physics (UvA) |
|---|---|
| Financier | Royal Netherlands Academy of Arts and Sciences - KNAW (KNAW) |
| Supervisor | Prof.dr.ir. J.A.E. Spaan |
|---|---|
| Researcher | Prof.dr. E.T. van Bavel |
| Researcher | Dr. K. Boer |
| Researcher | Prof.dr. H.R. Büller |
| Researcher | Prof.dr. J. Dankelman |
| Researcher | Dr. M. Groening |
| Researcher | Ing. R. Jonges |
| Researcher | Prof.dr. A.G.J.M. van Leeuwen |
| Researcher | Prof.dr. H. Lie |
| Researcher | Prof.dr. H. Pannekoek |
| Researcher | Dr. M. Pfaffendorf |
| Researcher | Prof.dr. J.J. Piek |
| Researcher | Dr.ir. M. Siebes |
| Researcher | Dr. I. Vergroesen |
| Project leader | Prof.dr.ir. H.G. Stassen |
| Project leader | Prof.dr. H. Vink |
| A70000 | Public health and health care |
|---|---|
| D21200 | Biophysics, clinical physics |
| D21300 | Biochemistry |
| D21500 | Histology, cell biology |
| D23210 | Dermatology, venereology, rheumatology, orthopedics |
| D23220 | Internal medicine |
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