MECHANISM OF SICKLE ERYTHROCYTE/ENDOTHELIAL ADHESION

我们建议检验这样一种假设，即镰状红细胞与 血管内皮细胞启动或参与局部血管- 以镰状细胞病为特征的闭塞危象。具体来说，我们 提示不同来源的内皮细胞之间的粘附性不同 血管床，这种粘附性在危机时期会增加，以及 内皮细胞激动剂会增加粘附性。 我们之前曾报道过高分子量的冯·威勒布兰德 因子(VWF)多聚体增加镰状红细胞与脐血的黏附 静脉内皮细胞。我们认为内皮细胞刺激 由于化学或机械压力而周期性发生，并导致 高分子量vWF多聚体的局部释放。初步 数据表明，当内皮细胞 受到内毒素或凝血酶的刺激。我们假设这些 多聚体与小静脉内的镰状红细胞和内皮细胞结合 在体内，导致血管阻塞。最近的数据表明， 悬浮在自体血浆中的镰状红细胞更容易粘连到 微血管内皮细胞多于脐静脉内皮细胞 细胞。这表明，红细胞黏附的机制是 取决于内皮细胞的位置。基于这些结果 我们认为体内的镰状红细胞黏附与复合体有关。 红细胞表面、血管内皮细胞和红细胞的变化 血管内环境。 以下参数被视为 镰刀状红细胞黏附：(I)内皮细胞起源(大或小 血管)；(Ii)病人临床情况(危急或稳定)；(Iii) 红细胞年龄；(Iv)局部壁切应力；(V)暴露时间 内皮细胞对激动剂的作用；以及(Vi) 镰刀状红细胞、刺激内皮细胞和分泌vWF 倍增器。这些参数对镰刀状红细胞黏附的影响 将在平行板流室中研究对内皮细胞的影响 粘附性试验。EPI-荧光视频显微镜与数字图像 加工将被用来量化镰状红细胞对 生理流下刺激和非刺激内皮细胞 条件。提出了一种红细胞再循环系统的研制方案。 允许红细胞、激动剂和内皮细胞持续接触 在流下。将进行抗体研究以确定粘附性 内皮细胞和红细胞上的受体。 血管内皮细胞、血浆和血管内皮细胞 在体外导致粘附性增加的红细胞将识别 与血红蛋白S聚合和红细胞病态无关的因素 在体内可促进或参与血管闭塞。

We propose to test the hypothesis that sickle red cell adhesion to endothelial cells initiates or contributes to the localized vaso- occlusive crises characteristic of sickle cell disease. Specifically, we suggest that adhesion varies between endothelial cells from different vascular beds, that adhesion will increase during crisis periods, and that endothelial cell agonists will increase adhesion. We have previously reported that high molecular weight von Willebrand factor (vWF) multimers increase sickle red cell adhesion to umbilical vein endothelial cells. We suggest that endothelial cell stimulation occurs periodically due to chemical or mechanical stresses and leads to localized release of high molecular weight vWF multimers. Preliminary data suggest that adhesion is increased when endothelial cells are stimulated with endotoxin or thrombin. We hypothesize that these multimers bind to sickle erythrocytes and endothelial cells in venules in vivo, leading to vascular obstruction. Recent data indicates that sickle red cells suspended in autologous plasma are more adhesive to microvascular endothelial cells than to umbilical vein endothelial cells. This suggests that the mechanism or red cell adhesion is dependent upon the location of the endothelium. Based on these results we propose that sickle red cell adhesion in vivo is related to complex changes in the erythrocyte surface, the endothelium, and the intravascular milieu. The following parameters are regarded as important determinants of sickle erythrocyte adherence: (i) endothelial cell origin (large or small vessel); (ii) patient clinical condition (crisis or steady-state); (iii) red cell age; (iv) local wall shear stress; (v) the time of exposure of endothelial cells to agonists; and (vi) the duration of contact between sickle erythrocytes, stimulated endothelial cells, and secreted vWF multimers. The influence of these parameters on sickle red cell adhesion to endothelial cells will be investigated in parallel-plate flow chamber adhesion assays. Epi-fluorescence videomicroscopy and digital image processing will be employed to quantify sickle red cell adhesion to stimulated and unstimulated endothelial cells under physiological flow conditions. Development of a red cell recirculation system is proposed to allow red cells, agonists, and endothelial cells continuous contact under flow. Antibody studies will be implemented to determine adhesive receptors on endothelial cells and red cells. Characterization of alterations in the endothelium, plasma, and erythrocytes responsible for increased adhesion in vitro will identify factors not related to hemoglobin S polymerization and red cell sickling which could contribute to or participate in vaso-occlusion in vivo.