This proposal aims to provide a fundamental understanding of cholesterol crystal nucleation from biliary vesicles and plasma lipoproteins in the context of gallstone disease and atherosclerosis, respectively. Cholesterol crystals are precursors to gallstones; inhibiting cholesterol nucleation offers a potential means of preventing stones. Crystals are also considered to be hallmarks of advanced atherosclerotic plaques, which typically become life-threatening only after rupture, followed by thrombosis and occlusion of an artery. Given that plaque stability correlates with the presence of crystals, inhibiting cholesterol nucleation offers a potential means of maintaining stability of atherosclerotic lesions and protecting against myocardial infarction. Development of preventive, clinical strategies based on inhibiting cholesterol nucleation requires a fundamental understanding of the nucleation mechanism. Molecular details concerning cholesterol nucleation are lacking, both in blood and in bile. However, recent work has shed important new insights, and kinetic and mechanistic studies of cholesterol nucleation are now feasible. This proposal is designed to test the hypotheses that 1) nanodomains of laterally phase-separated cholesterol constitute an equilibrium phase within lipid membranes; these nanondomains are distinct from cholesterol-rich domains and rafts 2) cholesterol nanodomains can act as crystal nucleation sites in non-equilibrium (i.e., in vivo) systems, and 3) aggregation of either vesicles or low density lipoproteins (LDL) facilitates collisions of cholesterol nanodomains in adjacent membranes; whereas aggregation alone is sufficient to induce nucleation from vesicles, nucleation from LDL requires uptake by macrophages.
