Hentzen Eric, McDonough Daniel, McIntire Larry, Smith C Wayne, Goldsmith Harry L, Simon Scott I
Speros Martel Section of Leukocyte Biology, Baylor College of Medicine, Houston, TX, USA.
Ann Biomed Eng. 2002 Sep;30(8):987-1001. doi: 10.1114/1.1511240.
Recently, we reported that tethering and rolling of neutrophils in shear flow over a substrate of E-selectin signals activation of beta 2-integrins and firm adhesion via an intracellular signaling pathway involving phosphorylation of p38 MAP kinase. In the current study the objective was to examine the molecular mechanisms and shear dependence underlying activation and adhesion of beta 2-integrin during shear-induced collisions between human neutrophils and murine B cells (300.19) transfected to express either E-selectin or L-selectin. Three separate parameters of cell activation were assessed over the time course of application of a defined shear field to heterotypic cell suspensions in a cone-plate viscometer. These were the two-body collision doublet lifetime and capture efficiency, surface upregulation of CD11b/CD18, and tyrosine phosphorylation of p38 MAP kinase. The data indicate that neutrophil adhesion to E-selectin expressing 300.19 cells occurs with a fourfold higher efficiency of firm adhesion than do collisions with L-selectin or parent control cells. Visual analysis of aggregation in a transparent cone-plate rheoscope revealed that the lifetime and efficiency of doublet formation increased fourfold as the applied shear stress increased. Neutrophil tethering via E-selectin was associated with rapid activation as indicated by upregulation of surface CD11b/CD18 and phosphorylation of p38 MAP kinase within seconds of application of shear. Activation greatly exceeded that observed for neutrophils sheared alone or with B cells expressing L-selectin. A distinct dependence of activation on the magnitude of the shear rate suggests a coupling between the fluid mechanical effects of shear and signaling of neutrophil adhesion.
最近，我们报道了中性粒细胞在E-选择素底物上的剪切流中系留和滚动通过涉及p38丝裂原活化蛋白激酶磷酸化的细胞内信号通路激活β2整合素并实现牢固黏附。在当前研究中，目的是检查在人中性粒细胞与转染以表达E-选择素或L-选择素的小鼠B细胞（300.19）之间的剪切诱导碰撞过程中，β2整合素激活和黏附的分子机制及剪切依赖性。在锥板粘度计中对异型细胞悬液施加确定的剪切场的时间过程中，评估了细胞激活的三个独立参数。这些参数是双体碰撞双峰寿命和捕获效率、CD11b/CD18的表面上调以及p38丝裂原活化蛋白激酶的酪氨酸磷酸化。数据表明，中性粒细胞与表达E-选择素的300.19细胞的黏附发生牢固黏附的效率比与L-选择素或亲本对照细胞的碰撞高四倍。在透明锥板流变仪中对聚集进行的视觉分析显示，随着施加的剪切应力增加，双峰形成的寿命和效率增加了四倍。如在施加剪切后数秒内表面CD11b/CD18的上调和p38丝裂原活化蛋白激酶的磷酸化所示，通过E-选择素的中性粒细胞系留与快速激活相关。这种激活大大超过了单独剪切的中性粒细胞或与表达L-选择素的B细胞一起剪切时观察到的激活。激活对剪切速率大小的明显依赖性表明剪切的流体力学效应与中性粒细胞黏附信号之间存在耦合。
