Cholesterol Hydroperoxide Trafficking: Impairment of Macrophage Cholesterol Efflux with Implications for Atherogenesis under Oxidative Stress

Abstract

This study describes a type of “stealthy” delivery of cholesterol 7α/β hydroperoxides (7‐OOH) via a natural cholesterol trafficking pathway. In vascular macrophages this pathway normally delivers cholesterol to mitochondria to initiate export signaling when its level becomes excessive. Because 7‐OOHs are structurally similar to cholesterol, they can be recognized by steroidogenic acute regulatory (StAR) proteins and trafficked to mitochondria alongside cholesterol.Oxidative stress associated with cardiovascular disease can produce various oxidized lipids, including cholesterol oxides such as 7‐OOH, 7‐hydroxide (7‐OH), and 7‐ketone (7=O). Unlike 7=O and 7‐OH, 7‐OOH is redox‐active, giving rise to the others via potentially toxic free radical reactions. In the present study we tested the novel hypothesis that under oxidative stress conditions, StAR family proteins not only deliver cholesterol to/into mitochondria of vascular macrophages, but also 7‐OOH, which induces peroxidative damage that impairs early stage reverse cholesterol transport. Stimulation of human monocyte‐derived THP‐1 macrophages with dibutyryl‐cAMP resulted in substantial upregulation of StarD1 and plasma membrane ABCA1 which exports cholesterol to acceptors such as ApoA1. siRNA‐induced StarD1 knockdown (kd) prior to stimulation had no effect on StarD4, but reduced ABCA1 upregulation, linking the latter to StarD1 functionality. Mitochondria in stimulated StarD1‐kd cells internalized 7‐OOH slower than non‐stimulated controls and underwent less 7‐OOH‐induced lipid peroxidation and membrane depolarization, as probed with C11‐BODIPY and JC‐1, respectively. Major functional consequences of 7‐OOH exposure were (i) loss of mitochondrial CYP27A1 activity, (ii) reduced 27‐hydroxycholesterol (27‐OH) output, and (iii) down‐regulation of ABCA1 and ABCG1. Consistently, 7‐OOH‐challenged macrophages exported less cholesterol to apoA‐I or HDL than did non‐challenged controls. StarD1‐mediated 7‐OOH transport was also found to be highly cytotoxic, whereas 7=O and 7‐OH were minimally toxic. These findings support our hypothesis that 7‐OOH found in oxidized low density lipoprotein gets delivered to macrophage mitochondria and promotes atherogenesis through impairment of cholesterol homeostasis.Support or Funding InformationThis work was supported by Polish National Science Center grant 2014/13/B/NZ3/00833 (WK) and by National Institutes of Health grants HL85677 (AWG) and HL58012 (DS). Support of American Heart Association Fellowship 14PRE185800221 (ACC) is also acknowledged