Center for Theoretical Problems of Physico-Chemical Pharmacology, Russian Academy of Sciences, Moscow, Russia.
Dmitry Rogachev National Medical Research Center of Pediatric Hematology, Oncology and Immunology, Moscow, Russia.
BMC Biol. 2024 May 20;22(1):115. doi: 10.1186/s12915-024-01912-2.
Thromboinflammation is caused by mutual activation of platelets and neutrophils. The site of thromboinflammation is determined by chemoattracting agents release by endothelium, immune cells, and platelets. Impaired neutrophil chemotaxis contributes to the pathogenesis of Shwachman-Diamond syndrome (SDS). In this hereditary disorder, neutrophils are known to have aberrant chemoattractant-induced F-actin properties. Here, we aim to determine whether neutrophil chemotaxis could be analyzed using our previously developed ex vivo assay of the neutrophils crawling among the growing thrombi.
Adult and pediatric healthy donors, alongside with pediatric patients with SDS, were recruited for the study. Thrombus formation and granulocyte movement in hirudinated whole blood were visualized by fluorescent microscopy in fibrillar collagen-coated parallel-plate flow chambers. Alternatively, fibrinogen, fibronectin, vWF, or single tumor cells immobilized on coverslips were used. A computational model of chemokine distribution in flow chamber with a virtual neutrophil moving in it was used to analyze the observed data.
The movement of healthy donor neutrophils predominantly occurred in the direction and vicinity of thrombi grown on collagen or around tumor cells. For SDS patients or on coatings other than collagen, the movement was characterized by randomness and significantly reduced velocities. Increase in wall shear rates to 300-500 1/s led to an increase in the proportion of rolling neutrophils. A stochastic algorithm simulating leucocyte chemotaxis movement in the calculated chemoattractant field could reproduce the experimental trajectories of moving neutrophils for 72% of cells.
In samples from healthy donors, but not SDS patients, neutrophils move in the direction of large, chemoattractant-releasing platelet thrombi growing on collagen.
血栓炎症是由血小板和中性粒细胞的相互激活引起的。血栓炎症的部位取决于内皮细胞、免疫细胞和血小板释放的趋化剂。中性粒细胞趋化作用受损导致 Shwachman-Diamond 综合征（SDS）的发病机制。在这种遗传性疾病中，已知中性粒细胞的趋化剂诱导的 F-肌动蛋白特性异常。在这里，我们旨在确定是否可以使用我们之前开发的在不断生长的血栓中爬行的中性粒细胞的体外测定法来分析中性粒细胞趋化性。
成年和儿科健康供体，以及儿科 SDS 患者被招募进行研究。在纤维蛋白原、纤维连接蛋白、vWF 或固定在载玻片上的单个肿瘤细胞的存在下，通过荧光显微镜可视化在富含凝血酶的全血中的血栓形成和粒细胞运动。在涂有纤维胶原蛋白的平行板流动室中。使用在其中移动的虚拟中性粒细胞的计算模型来分析观察到的数据。
健康供体中性粒细胞的运动主要发生在胶原蛋白上生长的血栓或肿瘤细胞周围的方向和附近。对于 SDS 患者或除胶原蛋白以外的涂层，运动特征为随机性和明显降低的速度。将壁剪切速率增加到 300-500 1/s 导致滚动中性粒细胞的比例增加。模拟白细胞趋化性运动的随机算法在计算的趋化剂场中可以再现 72%的运动中性粒细胞的实验轨迹。
在健康供体的样本中，但不是 SDS 患者，中性粒细胞在胶原蛋白上生长的大的、释放趋化剂的血小板血栓的方向上移动。
