Department of Cardiovascular Physiology, Heidelberg University, 69120 Heidelberg, Germany.
Internal Medicine III, University Clinics Heidelberg, 69120 Heidelberg, Germany.
Proc Natl Acad Sci U S A. 2018 Jun 12;115(24):E5556-E5565. doi: 10.1073/pnas.1801366115. Epub 2018 May 23.
Monocyte extravasation into the vessel wall is a key step in atherogenesis. It is still elusive how monocytes transmigrate through the endothelial cell (EC) monolayer at atherosclerosis predilection sites. Platelets tethered to ultra-large von Willebrand factor (ULVWF) multimers deposited on the luminal EC surface following CD40 ligand (CD154) stimulation may facilitate monocyte diapedesis. Human ECs grown in a parallel plate flow chamber for live-cell imaging or Transwell permeable supports for transmigration assay were exposed to fluid or orbital shear stress and CD154. Human isolated platelets and/or monocytes were superfused over or added on top of the EC monolayer. Plasma levels and activity of the ULVWF multimer-cleaving protease ADAMTS13 were compared between coronary artery disease (CAD) patients and controls and were verified by the bioassay. Two-photon intravital microscopy was performed to monitor CD154-dependent leukocyte recruitment in the cremaster microcirculation of ADAMTS13-deficient versus wild-type mice. CD154-induced ULVWF multimer-platelet string formation on the EC surface trapped monocytes and facilitated transmigration through the EC monolayer despite high shear stress. Two-photon intravital microscopy revealed CD154-induced ULVWF multimer-platelet string formation preferentially in venules, due to strong EC expression of CD40, causing prominent downstream leukocyte extravasation. Plasma ADAMTS13 abundance and activity were significantly reduced in CAD patients and strongly facilitated both ULVWF multimer-platelet string formation and monocyte trapping in vitro. Moderate ADAMTS13 deficiency in CAD patients augments CD154-mediated deposition of platelet-decorated ULVWF multimers on the luminal EC surface, reinforcing the trapping of circulating monocytes at atherosclerosis predilection sites and promoting their diapedesis.
单核细胞渗出到血管壁是动脉粥样硬化形成的关键步骤。单核细胞如何穿过动脉粥样硬化易发生部位的内皮细胞 (EC) 单层仍然难以捉摸。血小板通过 CD40 配体 (CD154) 刺激后在腔 EC 表面沉积的超大 von Willebrand 因子 (ULVWF) 多聚体连接，可能有助于单核细胞穿出。用于活细胞成像的平行板流动室或用于迁移测定的 Transwell 可渗透支架中生长的人 EC 暴露于流体或轨道剪切力和 CD154。将人分离的血小板和/或单核细胞在 EC 单层上或上方超滤液或添加。比较了冠心病 (CAD) 患者和对照组血浆 ULVWF 多聚体切割蛋白酶 ADAMTS13 的水平和活性，并通过生物测定进行了验证。进行双光子活体显微镜检查以监测 ADAMTS13 缺陷型与野生型小鼠隐窝微循环中 CD154 依赖性白细胞募集。CD154 诱导的 ULVWF 多聚体-血小板串在 EC 表面形成，即使在高剪切力下也捕获单核细胞并促进穿过 EC 单层的迁移。双光子活体显微镜检查显示，CD154 诱导的 ULVWF 多聚体-血小板串主要在小静脉中形成，这是由于 EC 强烈表达 CD40，导致下游白细胞渗出明显。CAD 患者的血浆 ADAMTS13 丰度和活性明显降低，并强烈促进 ULVWF 多聚体-血小板串的形成和体外单核细胞捕获。CAD 患者的中度 ADAMTS13 缺乏症增强了血小板修饰的 ULVWF 多聚体在腔 EC 表面的 CD154 介导沉积，增强了循环单核细胞在动脉粥样硬化易发生部位的捕获，并促进了它们的穿出。
