HDL functionality: pathophysiological role in cardiovascular disease and as novel metric of the neurodegenerative disorder

Abstract

Cardiovascular diseases (CVD) remain a leading cause of morbidity and the first cause of death globally. The project builds upon the ideas that have been recently developed in the field of cardiovascular disease regarding the cholesterol efflux capacity (CEC) of high-density lipoprotein (HDL), the main acceptor of cell cholesterol in plasma. HDL CEC is thought to reflect the robustness of reverse cholesterol transport from macrophages (mRCT) to the liver for excretion with bile. Inadequate mRCT causes abnormal accumulation of cholesterol and death of macrophages, formation of lipid core in the subendothelium of coronary arteries and eventually rupture of the lesion with resultant occlusion of the artery and heart ischemia. In most recent years, HDL CEC has been identified as a strong inverse predictor of incident cardiovascular disease, independently of HDL-, LDL-cholesterol levels and other major risk factors. The definition of CEC as a biomarker of CVD may improve risk stratification in human population and help to better define the level of risk in subjects that, classified as intermediate/low risk with the currently used parameters, undergo unexpected cardiovascular events. Elucidation on the role of HDL CEC as an index of disease as well as investigations on the pathophysiological mechanisms involved in the process of cholesterol transport from cells to HDL appear as an attractive field of research and constitute the general aim of this PhD project. Enhancement of HDL functions is a putative therapeutic strategy to treat cardiovascular disease and other human disorders. In fact, intravenous administration of synthetic HDL containing human apolipoprotein A-I (apoA-I) causes significant atherosclerosis regression in several experimental and clinical studies. The first part of my thesis project focuses on the evaluation of the effects of a new kind of reconstituted HDL (TN-rHDL) on ex vivo CEC in relationship to atherosclerosis development in a model of hypercholesterolemic rabbit. Main finding was that TN-rHDL is effective in increasing plasma total CEC after 4 hours from a single high-dose infusion. This effect on cholesterol efflux capacity is associated with parameters of plaque stabilization and may explain, at least in part, the reduced atherosclerosis progression observed in treated rabbits. In the second part of my thesis, I investigated the phisiological role of a specific human protein involved in lipid metabolism on HDL functionality and mRCT. Phospholipid transfer protein (PLTP) is a key player in lipoprotein remodeling and both direct and inverse correlations between PLTP activity and atherosclerotic cardiovascular disease have been reported in human studies. A unique in vivo experimental design was used in order to understand the function of PLTP overexpression and deletion on HDL efflux capacity and macrophage reverse cholesterol transport in mice. Our results showed that PLTP overexpression and deletion reduce HDL cholesterol mass and ex vivo plasma cholesterol efflux capacity without adversly affecting the rate of cholesterol efflux from macrophages to plasma and in vivo macrophage cholesterol transport in plasma to the liver. These interesting findings imply a substantial resilience of mRCT in face of drastic changes in HDL metabolism. In the third paragraph, CEC was evaluated in a human population of overweight individuals that underwent diet supplementation with a new kind of wheat flour enriched with bioactive components and functional probiotics. Several foods and dietary supplements have been shown to protect against the development of CVD; in this study, a slight but significant improvement of plasma ABCG1 CEC was observed in humans after three months of innovative food consumption. Despite no change in HDL concentration, a rearrangement of their composition can occur with enhancement of their function. ABCG1-mediated CEC of treated subjects inversely correlates with homocysteinemia, an independent risk factor for coronary disease, while a significant, direct relation was found with plasmatic folic acid, considered as a cardioprotective factor. It can be speculated that consumption of bioactive components within the innovative pasta act simultaneously on the amelioration of subjects inflammatory profile and on HDL functional quality thus “linking” ABCG1 CEC to the levels of such metabolic markers. The last part of my thesis focuses on the development of a method to measure cholesterol efflux of human cerebrospinal fluid. Alterations in lipid metabolism is a common feature underlying atherosclerotic cardiovascular disease (ASCVD) and neurodegenerative disorders. Indeed, experimental evidence suggests that excessive cholesterol in neurons, astrocytes and microglia promotes Aβ accumulation and stimulates Aβ-driven inflammation in Alzheimer’s disease. In particular, we advance the hypothesis that cholesterol efflux to apolipoproteins and HDL for disposal via the cerebrospinal fluid (CSF) is an important mechanism in neurons and glia for management of intracellular cholesterol and prevention of Aβ pathology. Here, we aim to adapt the HDL CEC assay to measure CSF CEC for the purposes of early diagnosis of Alzheimer’s disease and gaining insight into pathogenesis of this condition. Therefore, I characterized cell cholesterol efflux in in vitro cellular models relevant to the pathology of AD and identified biochemical components in CSF that determine CSF CEC variability in a small human cohort. Alltoghether, these results establish a robust method to study human CSF CEC as a putative, novel metric of the neurodegenerative disease.