Howard T H, Wang D
J Clin Invest. 1987 May;79(5):1359-64. doi: 10.1172/JCI112962.
Formyl-methionylleucylphenylalanine (fMLP) activation of neutrophils causes an increase in intracellular Ca2+, activation of protein kinase C and an increase in F-actin content. To examine the role of Ca2+ and protein kinase C activation as determinants of change in F-actin content of neutrophils, we used the NBD-phallacidin extraction assay to compare the kinetics and extent of change in F-actin content of cells activated with fMLP, the calcium ionophore A23187 or phorbol myristate acetate (PMA). All stimuli increase the F-actin content in a dose-dependent manner; however, the rate of increase is slower and the maximum F-actin content is less for calcium ionophore and PMA than for fMLP-activated cells. The A23187-induced increase in F-actin content, but not that of fMLP, depends upon external free [Ca2+]. In A23187-activated cells, F-actin content increases at [Ca2+]free greater than or equal to 5 microM, is maximal at [Ca2+]free greater than or equal to 10 microM and is negligible at physiologic free [Ca2+] (10(-7)-10(-6) M). Combinations of PMA with A23187 or fMLP inhibit the A23187, but not the fMLP, activated actin polymerization. Comparison and combination of these activators shows that neither Ca2+-dependent activation with A23187 nor activation with PMA alone or in combination mimic the fMLP-induced changes in cytoskeleton organization of neutrophils.
甲酰甲硫氨酰亮氨酰苯丙氨酸（fMLP）激活中性粒细胞会导致细胞内钙离子增加、蛋白激酶C激活以及F-肌动蛋白含量增加。为了研究钙离子和蛋白激酶C激活作为中性粒细胞F-肌动蛋白含量变化决定因素的作用，我们使用NBD-鬼笔环肽提取试验来比较用fMLP、钙离子载体A23187或佛波酯（PMA）激活的细胞中F-肌动蛋白含量的动力学和变化程度。所有刺激均以剂量依赖性方式增加F-肌动蛋白含量；然而，钙离子载体和PMA导致F-肌动蛋白含量增加的速率较慢，且最大F-肌动蛋白含量低于fMLP激活的细胞。A23187诱导的F-肌动蛋白含量增加（但fMLP诱导的增加并非如此）取决于细胞外游离钙离子浓度。在A23187激活的细胞中，当游离钙离子浓度大于或等于5微摩尔时，F-肌动蛋白含量增加，在游离钙离子浓度大于或等于10微摩尔时达到最大值，而在生理游离钙离子浓度（10⁻⁷ - 10⁻⁶摩尔）时可忽略不计。PMA与A23187或fMLP联合使用会抑制A23187激活的肌动蛋白聚合，但不会抑制fMLP激活的肌动蛋白聚合。这些激活剂的比较和联合表明，无论是A23187介导的钙离子依赖性激活，还是单独或联合使用PMA激活，均无法模拟fMLP诱导的中性粒细胞细胞骨架组织变化。
