Apolipoprotein B (apoB) is the major atherogenic apolipoprotein. High plasma levels of apoB lipoproteins are a primary causal factor of atherosclerosis. To date, it is still poorly understood why apoB-lipoproteins are atherogenic. The overall goal of this project is to develop novel mouse models to determine whether the proteoglycan (PG)-binding property of apoB is required for apoB-lipoproteins to promote atherosclerosis and to explore the therapeutic potential of targeting the apoB-PG interaction to intervene in atherosclerosis. Three major hypotheses will be tested. First, we hypothesize that the apoB N-terminus contains the principal PG binding site of apoB-48 lipoproteins (LpB48) and that disruption of this binding site will greatly reduce the atherogenicity of LpB-48. Using recombinant adenoviruses, mouse LpB48 with or without mutations in the putative PG-binding sites will be produced in apoE-null mice. The effect of these mutations on PG-binding of LpB48 will be assessed. The desirable mutations will then be introduced into mouse apoB gene (Apob) by gene targeting. The atherogenic potential of the PG-binding-defective LpB-48 will be determined in apoE-null mice. Our preliminary data indicated that the principal PG-binding site of LpB-48 is located within the N terminal 50 residues of the apoB protein. We have also prepared an ApoB-targeting construct for introducing mutations to this region.
Our second aim i s to test the hypothesis that LpB with increased affinity for PG binding have increased atherogenicity. We will create a novel high affinity PG-binding site in mouse apoB-48 and use adenovirus to produce in vivo recombinant LpB-48 to validate the functionality of this """"""""engineered"""""""" PG-binding site. This new apoB-48 PG-binding domain will then be """"""""knocked"""""""" into the mouse ApoB by gene targeting. The atherogenic potential of the engineered LpB-48 will be determined in apoE-null mice.
The third aim i s to test the hypothesis that transgenic expression of an apoB N-terminal region by vascular smooth muscle cells (VSMC) will block LpB binding to arterial PG's and inhibit atherogenesis. Transgenic mice will be produced using a VSMC-specific promoter to direct over-expression of the N-terminal apoB in VSMC. Experimental atherosclerosis will be examined in the resultant mice. These studies are likely to produce direct in vivo experimental data concerning the role of PG-binding property of apoB in the pathogenesis of atherosclerosis. Such information will advance our understanding of mechanisms of atherosclerosis and, in turn, facilitate developing new therapeutic strategies to treat atherosclerotic diseases. ? ?

National Institute of Health (NIH)
National Heart, Lung, and Blood Institute (NHLBI)
Research Project (R01)
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Pathology A Study Section (PTHA)
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Srinivas, Pothur R
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Washington University
Internal Medicine/Medicine
Schools of Medicine
Saint Louis
United States
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