Membrane cholesterol and vascular inflammation

ABSTRACT Our long-term objective is to define cellular pathways that regulate lipid flux and to elucidate their impact on cardiovascular disease. Although the connection between hypercholesterolemia and vascular inflammation has been appreciated for decades, the mechanistic underpinnings of this association remain incompletely understood. This application builds on recent work that defined mechanisms of nonvesicular lipid transport and identified pathways to regulate PM cholesterol abundance in experimental systems. We showed that the Aster transporter family facilitates the movement of cholesterol from PM to ER, and that loss of Aster function leads to PM cholesterol accumulation. But the role this transporter in vascular physiology is completely unknown. Preliminary observations have revealed that Aster-A is responsive to inflammatory activation of endothelial cells (ECs), and suggested that loss of Aster function modulates vascular inflammation through control of PM cholesterol availability. We have further discovered that cholesterol binds directly to endothelial PM proteins, including ICAM-1 and VCAM-1, pointing to a novel mechanism by which changes in PM lipid composition may determine inflammatory responses. Specific Aim 1 will identify novel cholesterol-responsive proteins in the vasculature using chemical biology. Provocative early results have revealed that multiple cell surface proteins involved in vascular inflammation bind cholesterol directly and are responsive to changes in PM cholesterol. We will define the spectrum of cholesterol-binding proteins in ECs in response to hypercholesterolemia and in settings of vascular inflammation. Specific Aim 2 will define molecular mechanisms whereby membrane cholesterol regulates inflammatory signaling in vascular cells. We will determine how changes in PM accessible cholesterol affect signaling by membrane inflammatory receptors and elucidate how the physical interaction of cholesterol with endothelial adhesion molecules regulates their abundance. Specific Aim 3 will elucidate the impact of accessible PM cholesterol on vascular inflammation and cardiovascular disease. We will use Aster-A loss-of-function models to manipulate accessible PM cholesterol in cultured ECs and in animals.