Xu Yanhong, Huang Dandan, Lü Shouqin, Zhang Yan, Long Mian
APL Bioeng. 2019 Mar 28;3(1):016104. doi: 10.1063/1.5045115. eCollection 2019 Mar.
Atherosclerosis is caused by chronic inflammation associated with the adhesion of neutrophils and endothelial cells (ECs) that is mediated by their respective cellular adhesive molecules to stiffened blood vessel walls. However, the stiffness dependence of calcium flux on neutrophils remains unclear yet. Here, the effect of substrate stiffness by ECs on neutrophils' calcium spike was quantified when the individual neutrophils that adhered to the human umbilical vascular endothelial cell (HUVEC) monolayer were pre-placed onto a stiffness-varied polyacrylamide substrate (5 or 34.88 kPa) or glass surface. Our data indicated that E-/P-selectins and intercellular adhesion molecule 1 (ICAM-1) on HUVECs and β-integrins, P-selectin glycoprotein ligand 1 (PSGL-1), and CD44s on neutrophils were all involved in mediating neutrophil calcium spike in a stiffness-dependent manner, in which the increase in substrate stiffness enhanced the calcium intensity and the oscillation frequency (spike number). Such stiffness-dependent calcium response is associated with the induced selectin related to β-integrin activation through the Syk/Src signaling pathway, and F-actin/myosin II are also involved in this. Moreover, tension-activated calcium ion channels displayed critical roles in initiating stiffness-dependent calcium spike. These results provide an insight into understanding how the stiffening of vascular walls could regulate the calcium flux of adhered neutrophils, and thus the immune responses in atherosclerosis.
动脉粥样硬化是由慢性炎症引起的，这种炎症与中性粒细胞和内皮细胞（ECs）的黏附有关，而它们之间的黏附是由各自的细胞黏附分子介导至硬化的血管壁上的。然而，中性粒细胞钙通量对硬度的依赖性仍不清楚。在此，当将黏附于人脐血管内皮细胞（HUVEC）单层的单个中性粒细胞预先放置在硬度不同的聚丙烯酰胺底物（5或34.88kPa）或玻璃表面上时，量化了ECs的底物硬度对中性粒细胞钙尖峰的影响。我们的数据表明，HUVECs上的E-/P-选择素和细胞间黏附分子1（ICAM-1）以及中性粒细胞上的β-整合素、P-选择素糖蛋白配体1（PSGL-1）和CD44s均以硬度依赖性方式参与介导中性粒细胞钙尖峰，其中底物硬度的增加增强了钙强度和振荡频率（尖峰数量）。这种硬度依赖性钙反应与通过Syk/Src信号通路诱导的与β-整合素激活相关的选择素有关，F-肌动蛋白/肌球蛋白II也参与其中。此外，张力激活的钙离子通道在启动硬度依赖性钙尖峰中起关键作用。这些结果为理解血管壁硬化如何调节黏附中性粒细胞的钙通量，进而调节动脉粥样硬化中的免疫反应提供了见解。
