Department of Biomedical Engineering, University of California Davis, Davis, CA 95616, USA.
Sci Signal. 2018 Mar 13;11(521):eaal4289. doi: 10.1126/scisignal.aal4289.
Global bursts in free intracellular calcium (Ca) are among the most conspicuous signaling events in immune cells. To test the common view that Ca bursts mediate rearrangement of the actin cytoskeleton in response to the activation of G protein-coupled receptors, we combined single-cell manipulation with fluorescence imaging and monitored the Ca concentration in individual human neutrophils during complement-mediated chemotaxis. By decoupling purely chemotactic pseudopod formation from cell-substrate adhesion, we showed that physiological concentrations of anaphylatoxins, such as C5a, induced nonadherent human neutrophils to form chemotactic pseudopods but did not elicit Ca bursts. By contrast, pathological or supraphysiological concentrations of C5a often triggered Ca bursts, but pseudopod protrusion stalled or reversed in such cases, effectively halting chemotaxis, similar to sepsis-associated neutrophil paralysis. The maximum increase in cell surface area during pseudopod extension in pure chemotaxis was much smaller-by a factor of 8-than the known capacity of adherent human neutrophils to expand their surface. Because the measured rise in cortical tension was not sufficient to account for this difference, we attribute the limited deformability to a reduced ability of the cytoskeleton to generate protrusive force in the absence of cell adhesion. Thus, we hypothesize that Ca bursts in neutrophils control a mechanistic switch between two distinct modes of cytoskeletal organization and dynamics. A key element of this switch appears to be the expedient coordination of adhesion-dependent lock or release events of cytoskeletal membrane anchors.
细胞内游离钙（Ca）的爆发是免疫细胞中最显著的信号事件之一。为了验证 Ca 爆发介导 G 蛋白偶联受体激活后肌动蛋白细胞骨架重排的普遍观点，我们将单细胞操作与荧光成像相结合，监测了人中性粒细胞在补体介导的趋化作用过程中单个细胞的 Ca 浓度。通过将纯趋化性伪足形成与细胞-基底粘附解耦，我们表明，过敏毒素（如 C5a）的生理浓度诱导非粘附性人中性粒细胞形成趋化性伪足，但不会引发 Ca 爆发。相比之下，病理或超生理浓度的 C5a 常引发 Ca 爆发，但在这种情况下，伪足突起停滞或反转，有效地阻止了趋化性，类似于与败血症相关的中性粒细胞麻痹。在纯趋化作用中，伪足延伸过程中细胞表面积的最大增加比已知的粘附性人中性粒细胞扩展其表面的能力小 8 倍。由于测量到的皮层张力增加不足以解释这种差异，我们将这种有限的可变形性归因于在没有细胞粘附的情况下，细胞骨架产生突起力的能力降低。因此，我们假设 Ca 爆发控制了中性粒细胞中两种不同的细胞骨架组织和动力学模式之间的机制转换。这种转换的一个关键要素似乎是细胞骨架膜锚定的粘附依赖性锁定或释放事件的便利协调。
