Lipoprotein Metabolism Laboratory, Translational Vascular Medicine Branch, National Heart, Lung, and Blood Institute, NIH, Bethesda, MD, USA.
Lipoprotein Metabolism Laboratory, Translational Vascular Medicine Branch, National Heart, Lung, and Blood Institute, NIH, Bethesda, MD, USA.
Biochim Biophys Acta Mol Cell Biol Lipids. 2020 Feb;1865(2):158537. doi: 10.1016/j.bbalip.2019.158537. Epub 2019 Oct 30.
Highly elevated plasma levels of interleukin-10 (IL-10) are causally associated with "Disappearing HDL Syndrome" and low plasma LDL-cholesterol, but the underlying mechanism is poorly understood. Fluid-phase endocytosis, a process highly dependent on actin dynamics, enables cells to internalize relatively high amounts of extracellular fluids and solutes. We sought to investigate whether IL-10 induces lipoprotein uptake by fluid-phase endocytosis in macrophages.
Macrophages (RAW264.7, Kupffer and human) were incubated with vehicle (PBS) or IL-10 (20 ng/ml) for 7 days. Uptake of HDL, LDL, and/or fluid-phase endocytosis probes (albumin-Alexa680®, 70 kDa FITC-Dextran and Lucifer Yellow, LY) was evaluated by FACS. Intracellular cofilin and phosphorylated cofilin (p-cofilin) levels were determined by immunoblotting. Macrophage uptake of lipoproteins and probes was non-saturable and increased after IL-10 incubation (p < 0.0001). Furthermore, pre-incubation with fluid-phase endocytosis inhibitors (LY294002, Latrunculin A, and Amiloride) significantly reduced uptake (p < 0.05). IL-10 increased the cofilin/p-cofilin ratio (p = 0.021), signifying increased cofilin activation and hence filamentous actin. Consistently, phalloidin staining revealed increased filamentous actin in macrophages after IL-10 treatment (p = 0.0018). Finally, RNA-seq analysis demonstrated enrichment of gene sets related to actin filament dynamics, membrane ruffle formation and endocytosis in IL-10-treated macrophages (p < 0.05). IL-10 did not alter mRNA levels of Ldlr, Vldlr, Scarb1, Cd36 or Lrp1. In primary human monocyte-derived macrophages and murine Kupffer cells, IL-10 incubation also increased uptake of lipoproteins, albumin and LY (p < 0.01).
Interleukin-10 induces the uptake of HDL and LDL by fluid-phase endocytosis by increasing actin-filament rearrangement in macrophages, thus providing a plausible mechanism contributing to "Disappearing HDL Syndrome".
白细胞介素-10（IL-10）的血浆水平升高与“消失的高密度脂蛋白综合征”和低血浆 LDL-胆固醇有关，但潜在机制尚不清楚。液相内吞作用是一种高度依赖肌动蛋白动力学的过程，使细胞能够内化相对大量的细胞外液和溶质。我们试图研究 IL-10 是否通过巨噬细胞中的液相内吞作用诱导脂蛋白摄取。
用载体（PBS）或 IL-10（20ng/ml）孵育巨噬细胞（RAW264.7、Kupffer 和人源）7 天。通过流式细胞术评估 HDL、LDL 和/或液相内吞探针（白蛋白-Alexa680®、70kDa FITC-右旋糖酐和 Lucifer Yellow，LY）的摄取。通过免疫印迹法测定细胞内原肌球蛋白和磷酸化原肌球蛋白（p-cofilin）的水平。脂蛋白和探针的巨噬细胞摄取是非饱和的，并在 IL-10 孵育后增加（p<0.0001）。此外，用液相内吞作用抑制剂（LY294002、Latrunculin A 和阿米洛利）预先孵育可显著减少摄取（p<0.05）。IL-10 增加了 cofilin/p-cofilin 比值（p=0.021），表明 cofilin 激活增加，进而肌动蛋白丝增加。同样，鬼笔环肽染色显示 IL-10 处理后巨噬细胞中的丝状肌动蛋白增加（p=0.0018）。最后，RNA-seq 分析表明，在 IL-10 处理的巨噬细胞中，与肌动蛋白丝动力学、膜皱襞形成和内吞作用相关的基因集富集（p<0.05）。IL-10 并未改变 Ldlr、Vldlr、Scarb1、Cd36 或 Lrp1 的 mRNA 水平。在原代人单核细胞衍生的巨噬细胞和鼠源 Kupffer 细胞中，IL-10 孵育也增加了脂蛋白、白蛋白和 LY 的摄取（p<0.01）。
白细胞介素-10 通过增加巨噬细胞中肌动蛋白丝的重排，诱导 HDL 和 LDL 的液相内吞作用摄取，从而为“消失的高密度脂蛋白综合征”提供了一种合理的机制。
