The basis of our work has been the proposition that understanding the regulation and implications of macrophage apo E production will provide insight into the pathophysiology of the atherosclerotic vessel wall lesion. We have, therefore, proposed investigations to examine the regulation of macrophage apo E production as well as the implications of such production for macrophage biology.
The Specific Aims of this current ampliation are as follows:
Specific Aim # 1 To identify the molecular basis for the response of the macrophage apo E gene to sterols. To accomplish this we will use acute transfection protocols in THP1 macrophages. After transfection analyses have identified important gene sequences, protein binding sites will be investigated as will the identity of protein binding factors.
Specific Aim #2 To characterize the co- translational and post-translational processing of apo E in macrophages. Using subcellular fractionation techniques we will study the distribution and transport of apo E in macrophage cells.
Specific aim #3 To investigate the mechanism by which macrophage apo E expression modulates macrophage cholesterol homeostasis. Here we will investigate the mechanism by which apo E enhances cholesterol efflux from J774 macrophages to HDL3. We will specifically examine three potential mechanisms: a) alteration of subcellular cholesterol transport; b) alteration of plasma membrane composition; or c) alteration of interaction with extracellular cholesterol acceptors. Production of apo E by macrophages in the vessel wall has been shown to be anti-atherogenic. Understanding the mechanism by which aop E exerts its anti-atherogenic effect in the vessel wall will provide important insight into the pathobiology of the vessel wall lesion.
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