High-Density Lipoprotein Transport Through Aortic Endothelial Cells Involves Scavenger Receptor BI and ATP-Binding Cassette Transporter G1

Abstract

Abstract-Cholesterol efflux from macrophage foam cells is a rate-limiting step in reverse cholesterol transport. In this process cholesterol acceptors like high-density lipoproteins (HDL) and apolipoprotein (apo)A-I must cross the endothelium to get access to the donor cells in the arterial intima. Previously, we have shown that apoA-I passes a monolayer of aortic endothelial cells (ECs) from the apical to the basolateral side by transcytosis, which is modulated by the ATP-binding cassette transporter (ABC)A1. Here, we analyzed the interaction of mature HDL with ECs. ECs bind HDL in a specific and saturable manner. Both cell surface biotinylation experiments and immunofluorescence microscopy of HDL recovered Ϸ30% of the cell-associated HDL intracellularly. Cultivated on inserts ECs bind, internalize, and translocate HDL from the apical to the basolateral compartment in a specific and temperature-dependent manner. The size of the translocated particle was reduced, but its protein moiety remained intact. Using RNA interference, we investigated the impact of SR-BI, ABCA1, and ABCG1 on binding, internalization, and transcytosis of HDL by ECs. HDL binding was reduced by 50% and 30% after silencing of SR-BI and ABCG1, respectively, but not at all after diminishing ABCA1 expression. Knock down of SR-BI and, even more so, ABCG1 reduced HDL transcytosis but did not affect inulin permeability. Cosilencing of both proteins did not further reduce HDL binding, internalization, or transport. In conclusion, ECs transcytose HDL by mechanisms that involve either SR-BI or ABCG1 but not ABCA1. cholesterol show an inverse association with the incidence of coronary artery disease. The cardioprotective effect of HDL and its major apolipoprotein (apo)A-I are, in part, related to the ability to promote the reverse transport of cholesterol from macrophage foam cells in the arterial intima to the liver for excretion into the bile. [1][2][3][4] An early step in the reverse transport of cholesterol is the transfer of excess cholesterol from the lipid-laden macrophages to HDLs. Importantly, the loading of cellular cholesterol to HDL does not take place in the plasma compartment but in the subendothelial space of arteries. 1 Consequently, HDLs must cross the endothelium to get into close proximity to the cholesterol donor cells. This passage is not well understood. 5 The endothelium lines the vasculature as a single layer of endothelial cells (ECs). As a semipermeable barrier, it regulates the flux of liquid, solutes, and cells between blood and interstitial space. Two principal pathways are known for transendothelial macromolecule translocation, the transcellular transport, including transcytosis, and the paracellular transfer between adjacent cells. 6,7 The paracellular pathway is formed by gaps between ECs, but regulated adherence and tight junctions restrict and control the free passage of macromolecules larger than 6 nm. 7 Endothelial transcytosis, which is defined as vesicle-mediated transport of proteins, has been best investigated ...

HDL and Atherothrombotic Vascular Disease

“…There is cell culture evidence that lipid-free apoA-I binds to specific saturable high-affinity binding sites on aortic endothelial cells, which is followed by internalization, transcytosis, and delivery of lipidated apoA-I to the basolateral side . A similar phenomenon was observed when cultivated aortic endothelial cells were treated with mature HDLs (Rohrer et al 2009). However, the mechanisms involved in regulating this endothelial transport of apoA-I and HDLs are not fully understood.…”

“…Induction of ABCA1 expression by a combination of oxysterol and 9-cis-retinoic acid increased the binding and internalization of apoA-I in endothelial cells, whereas knockdown of endothelial ABCA1 by RNA interference reduced the cellular uptake and transcytosis of apoA-I ). Conversely, siRNA-mediated gene silencing experiments revealed that the cell surface binding and transport of HDLs through aortic endothelial cells are highly dependent on SR-BI and ABCG1, but not on ABCA1 (Rohrer et al 2009). Subsequent research has demonstrated expression of the ectopic β-chain of F0F1 ATPase (β-ATPase) on the endothelial cell surface (Cavelier et al 2012).…”

High-Density Lipoproteins

“…Our group found that aortic endothelial cells are able to bind, internalize, and transport both HDLs and lipid-free apoA-I in a specific and saturable manner. 60, 61 Transcytosis of apoA-I through aortic endothelial cells resulted in lipidation of lipid-free apoA-I and was reduced by knockdown of ABCA1 by small interfering RNA, 60,62 providing evidence that ABCA1 is involved in this process. In contrast, binding, uptake, and transendothelial transport of mature HDLs were modulated by ABCG1 and SR-BI, not by ABCA1.…”

“…In contrast, binding, uptake, and transendothelial transport of mature HDLs were modulated by ABCG1 and SR-BI, not by ABCA1. 61 Further research identified a novel mechanism for transport of both apoA-I and HDLs through endothelial cells, in which binding of apoA-I to the β-chain of cell surface F0F1 ATPase expressed on endothelial cells stimulates ATP hydrolysis, and the generated ADP selectively activates P2Y12, leading to internalization and transport of initially lipid-free apoA-I and HDLs. 63 Once apoA-I and HDLs have been loaded with cholesterol, they need to leave the arterial wall to complete reverse cholesterol transport.…”

Genetically Modified Animal Models for Lipoprotein Research

“…Since the fractional catabolic rate for apoB-48-containing lipoproteins is very high compared to that for apoB-100-containing lipoproteins (Li, 1996), concentrations of VLDL and LDL are very low in wild-type mice and rats compared to humans. In reverse cholesterol transport from peripheral tissue to the liver, high-density lipoproteins (HDLs) receive free cholesterol from macrophages through scavenger receptor type B-I (SR-BI) and ABCs such as ABCA1 (Rohrer, 2009). The free cholesterol transported from macrophages is esterified by lecithin:cholesterol acyltransferase (LCAT) in plasma.…”