Previous work indicates the versican, the major vascular interstitial chondroitin sulfate proteoglycan (CSPG), and the glycosaminoglycan, hyaluronan (HA), which interacts with versican, accumulate in the extracellular matrix (ECM) in early atherosclerotic and restenotic lesions. We have also shown that the synthesis of these two molecules is highly regulated by specific growth factors and cytokines and is modulated when arterial smooth muscle cells (ASMC) are stimulated to migrate and proliferate. Inhibition of the interaction of versican and HA with receptors at the cell surface blocks ASMC proliferation and migration. One consequence of versican accumulation in the ECM is the trapping and retention of lipoproteins. We have shown that versican interacts with apoB and apoE-containing lipoproteins and this interaction is enhanced by factors that influence chondroitin sulfate chain biosynthesis. These observations have led us to continue to explore the importance of versican/HA complexes in the ECM in the regulation of ASMC phenotype, and in the retention of lipoproteins in the vascular wall. We hypothesize that modifications in versican/HA synthesis are required for ASMC proliferation and migration, and that these modifications influence other properties of ASMC, such as the ability to remodel the ECM during different phases of atherogenesis. We further postulate that the accumulation of versican within the ECM is partly responsible for the extracellular accumulation of lipoproteins. We propose to examine these hypotheses in four aims.
The first aim will address questions concerning the role of versican and versican isoform expression in the regulation of ASMC phenotype and matrix remodeling. In this work we will express versican isoforms and mini-genes that contain various domains of versican, and examine the effect on cell proliferation, migration and matrix protein deposition.
The second aim i ncludes experiments designed to examine the expression of HA synthases by ASMC, and the role of HA, both as an extracellular component of the matrix and as a newly discovered intracellular molecule, in ASMC proliferation, migration and ECM remodeling.
The third aim i s designed to extend our studies concerning the interaction of lipoproteins with versican chondroitin sulfate side chains. We propose to determine the specific oligosaccharide sequences that are responsible for the binding of ApoB and ApoE-contain lipoproteins to versican, and determine factors that induce changes in chondroitin sulfate chain structure that give rise to versican forms with differing affinities for lipoproteins. In the fourth aim, we will use new reagents to examine the presence of versican variants and proteolytic cleavage products in naturally-occurring atherosclerotic lesions, investigate the effect of altered expression of versican variants and HA synthases on lesion progression by a cell-mediated gene transfer approach in the injured rat carotid neointimal model, and use targeted over-expression of versican variants in a fat-fed transgenic mouse model to examine the role of versican expression on the accumulation of lipoproteins in vascular lesions in vivo. Manipulation of versican and HA metabolism should influence the progression and/or regression of atherosclerotic lesion development.
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