ABSTRACT The first steps in atherosclerosis are the transendothelial movement and subendothelial accumulation of lipoprotein lipid. Endothelial cell (EC) transcytosis of LDL involves two receptors, scavenger receptor-BI (SR-BI) and activin-like kinase 1 (ALK1). We showed that ECs also internalize undigested chylomicrons via SR-BI and process them in lysosomes, leading to storage of some lipid as lipid droplets and the release of small extracellular vesicles (sEVs) that cause lipid accumulation in macrophages. While many have considered chylomicrons as non-atherogenic and too large to cross the EC barrier, this concept is now outdated with the understanding that lipoprotein entry into the artery is a receptor-mediated process. The overall goal of Project 3 (P3) is to determine how EC chylomicron uptake affects EC biology, delivers lipids to the artery, and accelerates atherosclerosis. In Aim 1, we propose to determine how triglyceride-rich lipoproteins (TRLs) from the liver and intestines are internalized and either processed within ECs or transcytosed. To do this, we will use mice with selective knockout of liver and intestinal microsomal triglyceride transfer protein (MTP) obtained from Dr. Hussain (P1) to determine lipoprotein characteristics that determine their interaction with SR-BI and ALK1. We will also determine how lipoproteins created with liver FIT2 knockout (P2) affect EC inflammation and the production and composition of EC released sEVs. Our preliminary data show that the N-terminal region of apoB has separate ligand binding regions for ALK1 and SR-BI and this aim will determine why and how lipoprotein uptake via each of these receptors affects ECs. In Aim 2, we propose in vivo studies to determine whether postprandial lipemia leads to EC inflammation and whether the site of origin of these TRLs determines their effects on arterial ECs. We provide preliminary data suggesting that chylomicrons that accumulate in lipoprotein lipase (LpL) deficient mice increase atherosclerosis. We will use N-terminal fragments of apoB to reduce lipoprotein uptake into ECs to determine role(s) of ALK1 and SR-BI in EC inflammation and atherosclerosis. Completion of the proposed studies promises to alter our view of the relationship of chylomicrons to vascular disease, determine whether TRLs from liver and intestine have similar effects on ECs, and define a novel approach to atherosclerosis prevention.