Apolipoprotein A1 and High-density Lipoprotein as a Cargo and Regulator of Endothelial Transcytosis

Lipoproteins are circulating lipid carriers that regulate lipid homeostasis; low density lipoprotein (LDL) provides cholesterol, while high density lipoprotein (HDL) removes cholesterol from peripheral tissue. An imbalance of these processes leads to dyslipidemias and atherosclerosis, the accumulation of LDL in the arteries. To reach the target tissue, lipoproteins cross a protective monolayer of endothelial cells through transcytosis. Current understanding of transcytosis is limited and further complicated because endothelial cells vary depending on the tissue and vessel of origin. Indeed, brain endothelial cells are the most impermeant due to limited endocytic trafficking. In this dissertation, I investigate the relationship of HDL with the endothelium under physiological and pathophysiological states. Specifically, I aimed to elucidate the mechanism of HDL internalization in the brain endothelium in one study while determining the role of HDL in atherosclerosis in the other. In the brain endothelium, I found that HDL internalization is partially mediated by Scavenger receptor B1 (SR-B1) in a cholesterol-dependent, caveolae- and clathrin- independent manner. Furthermore, HDL internalization is independent of PDZK1 and the production of nitric oxide, which belong to the signaling pathway stimulated by the association of HDL with SR-B1. Together this suggests that HDL internalization is dependent on SR-B1 and cholesterol through brain- specific mechanisms. Given that atherosclerosis is also due to SR-B1-mediated transcytosis of LDL across coronary endothelial cells, I aimed to determine whether ApoA1, the protein found in HDL can prevent LDL transcytosis and how this is affected by the lipid state of ApoA1 as well as the natural mutant called Milano ApoA1. The Milano mutation results in half the plasma HDL levels, yet carriers do not develop atherosclerosis - the mechanism of which is not clear. I observed that ApoA1 can decrease LDL internalization and transcytosis by competition for SR-B1, while lipid-associated ApoA1 is more effective in inhibiting LDL internalization. The Milano protein is a slightly better inhibitor of LDL internalization but is comparable to wild-type ApoA1 in inhibiting LDL transcytosis and associating with SR-B1. Together this suggests that HDL is athero-protective by also inhibiting LDL transit across the endothelium, a process not affected by the Milano mutation.Ph.D