Geh Esmond N, Swertfeger Debi K, Roscoe Isabella, Street Scott E, Bursey Alexiana, Sexmith Hannah, Woollett Laura A, Davidson W Sean, Shah Amy Sanghavi
Division of Endocrinology, Department of Pediatrics, Cincinnati Children's Hospital Medical Center & The University of Cincinnati College of Medicine, Cincinnati, OH, USA.
Department of Pathology and Laboratory Medicine, University of Cincinnati, Cincinnati, OH, USA.
J Lipid Res. 2025 Sep;66(9):100885. doi: 10.1016/j.jlr.2025.100885. Epub 2025 Aug 21.
The entrapment of LDLs by proteoglycans (PGs) in the extracellular matrix of the arterial intima is a key initial step in the development of atherosclerosis. HDLs can interfere with this process, but the underlying mechanism is not fully understood. The aim of this study was to investigate the mechanisms by which HDL inhibits LDL binding to PG. An In-Cell ELISA was used to measure the binding of LDL to PGs in the extracellular matrix synthesized by mouse vascular smooth muscle cells. Fast-protein liquid chromatography, immunoprecipitation, SDS-PAGE, and immunoblotting analysis were performed to characterize how HDL and its apolipoproteins inhibit LDL binding to PGs. HDL and APOA1 inhibited LDL binding to PGs in a dose-dependent manner. Competition experiments showed that HDL did not compete directly with LDL for PG binding. Instead, APOA1 dissociated from HDL and associated with LDL, reducing the ability of LDL to bind PGs. This was demonstrated by separating HDL and LDL using porous filters of different sizes and tracking the movement of either HDL or APOA1. When APOA1 was solidly anchored to HDL particles, HDL lost the ability to affect LDL-PG binding. HDL inhibits LDL binding to PGs through an interaction with its main apolipoprotein, APOA1, specifically, a pool of loosely attached, exchangeable, lipid-free APOA1 on the HDL surface. These findings identify lipid-free APOA1 as a critical mediator of the ability of HDL to reduce LDL retention in the arterial wall and provide new insights into the antiatherogenic properties of HDL.
动脉内膜细胞外基质中的蛋白聚糖（PGs）捕获低密度脂蛋白（LDLs）是动脉粥样硬化发展过程中的关键起始步骤。高密度脂蛋白（HDLs）可干扰这一过程，但其潜在机制尚未完全明确。本研究旨在探究HDL抑制LDL与PG结合的机制。采用细胞内酶联免疫吸附测定法（In-Cell ELISA）来检测LDL与小鼠血管平滑肌细胞合成的细胞外基质中PGs的结合情况。运用快速蛋白液相色谱法、免疫沉淀法、十二烷基硫酸钠 - 聚丙烯酰胺凝胶电泳（SDS-PAGE）及免疫印迹分析，以表征HDL及其载脂蛋白如何抑制LDL与PGs的结合。HDL和载脂蛋白A1（APOA1）以剂量依赖方式抑制LDL与PGs的结合。竞争实验表明，HDL并非直接与LDL竞争PG结合位点。相反，APOA1从HDL上解离并与LDL结合，降低了LDL与PGs结合的能力。通过使用不同孔径的多孔滤器分离HDL和LDL，并追踪HDL或APOA1的移动情况，证实了这一点。当APOA1牢固地锚定在HDL颗粒上时，HDL失去了影响LDL - PG结合的能力。HDL通过与其主要载脂蛋白APOA1相互作用来抑制LDL与PGs的结合，具体而言，是通过HDL表面一组松散附着、可交换的无脂APOA1来实现的。这些发现确定无脂APOA1是HDL降低LDL在动脉壁内潴留能力的关键介质，并为HDL的抗动脉粥样硬化特性提供了新的见解。
