Zebrafish models of vascular inflammation and atherosclerosis

本提案的目的是建立斑马鱼（Danio rerio）模型，适用于动脉粥样硬化某些过程的体内研究。人们普遍认为，高胆固醇血症导致动脉内脂质积聚，从而诱发血管炎症，引发动脉粥样硬化，动脉粥样硬化是人类心脏病发作和中风的主要原因。因此，目前动脉粥样硬化的实验动物研究使用小鼠和兔子，其中基因改造和高脂肪饮食相结合可诱导极端高胆固醇血症和动脉粥样硬化的快速发展。在这些研究中使用小鼠和兔子的一个重要限制是，动脉粥样硬化病变的显微镜检查只能在死后进行。使用斑马鱼的一个重要优势是其幼虫的光学透明性。我们建议使用斑马鱼来研究动脉粥样硬化的某些方面，将能够监测活体动物动脉的病理过程。

The aim of this proposal is to establish zebrafish (Danio rerio) models suitable for in vivo studies of certain processes of atherogenesis. It is widely accepted that hypercholesterolemia leads to lipid accumulation in arteries, which in turn induces vascular inflammation and initiates the development of atherosclerosis, the major cause of heart attacks and stroke in humans. Hence, current experimental animal studies of atherosclerosis use mice and rabbits in which a combination of genetic modification and high-fat diet induce extreme hypercholesterolemia and rapid development of atherosclerosis. An important limitation of using mice and rabbits in these studies is that microscopic examination of atherosclerotic lesions is possible only postmortem. An important advantage of using zebrafish is the optical transparency of its larvae. We propose that using zebrafish for studying certain aspects of atherogenesis will enable monitoring the pathologic processes in arteries in a live animal. Specifically, in Aim 1 we will establish a model of vascular inflammation and atherosclerosis in which feeding zebrafish a high-cholesterol diet causes hypercholesterolemia, lipoprotein oxidation and lipid accumulation in the vascular wall. Using adult zebrafish, we will develop a novel animal model to study in vivo lipoprotein oxidation. The optical transparency of zebrafish larvae will enable monitoring the kinetics of lipid accumulation and macrophage recruitment into the vascular wall. Transgenic zebrafish with fluorescent proteins expressed in endothelial cells and in macrophages will be used in these studies. Developing the hypercholesterolemic larvae model will allow us to set up in vivo functional assays to measure the activity of matrix metalloproteinases, cathepsins and phospholipases in live zebrafish larvae (Aim 2). To demonstrate the utility of this unique model we will address a specific mechanism, critical in the development of atherosclerosis. In Aim 3, we will test the importance of lipoprotein oxidation in vascular lipid accumulation and inflammation. Specifically, antioxidants will be added to the fish diet or dissolved in the fish tank water and their effects on lipid and macrophage deposition in the vasculature of transgenic zebrafish, and specific enzyme activities will be studied. In summary, we propose that establishing zebrafish models of vascular inflammation and atherosclerosis will help significantly enhance mechanistic studies of atherosclerosis as well as the design and screening of new therapies.