Walker B A, Hagenlocker B E, Ward P A
Department of Pathology, University of Michigan Medical School, Ann Arbor 48109.
J Immunol. 1991 May 1;146(9):3124-31.
For superoxide (O2-) responses of human neutrophils stimulated by FMLP, experiments were designed to assess the requirement of extracellular calcium [( Ca2+]o) for priming of O2- responses by platelet-activating factor (PAF), PMA, or ionomycin. Although priming by PMA did not require [Ca2+]o, there was, as expected, a requirement for [Ca2+]o for the optimal priming effects of PAF and ionomycin. The ED50 value for [Ca2+]o in the priming function of PAF was 105 microM. The [Ca2+]o-dependent priming with ionomycin was bimodal with two ED50 values for [Ca2+]o of 90 microM and 3.2 mM. Optimal priming by PAF required at least 4-min exposure of cells to [Ca2+]o. Cells primed by PAF exhibited faster initial rates of O2-production after addition of FMLP, but the duration of O2- production was not prolonged. Whereas PAF-primed responses to FMLP are usually associated with increases in intracellular calcium [( Ca2+]i) after addition of FMLP, two conditions were found in which O2- responses to FMLP in PAF-primed cells occurred in the absence of any detectable increase in [Ca2+]i. When cells were loaded with the calcium chelator, bis-(O-aminophenoxy)-ethane-H,N,N',N'-tetraacetic acid, and then primed with PAF, normal amounts of inositol 1,4,5-trisphosphate were formed, but no increase in [Ca2+]i occurred after addition of FMLP even though the cells exhibited a fully primed O2- response; in Ca2(+)-depleted and ionomycin-permeabilized cells that were primed with PAF and then stimulated with FMLP, O2- was generated in amounts comparable to reference control (primed) cells, but there was suppressed production of inositol 1,4,5-trisphosphate and no increase in [Ca2+]i after addition of FMLP to PAF-primed cells. These data confirm the requirement of [Ca2+]o for optimal priming of neutrophils by PAF and ionomycin (but not cells primed by PMA) and indicate that, under certain conditions, generation of O2- in response to FMLP in PAF-primed neutrophils can occur independent of any increase in [Ca2+]i.
为研究在N-甲酰甲硫氨酰-亮氨酰-苯丙氨酸（FMLP）刺激下人中性粒细胞的超氧阴离子（O2-）反应，设计实验评估细胞外钙（[Ca2+]o）对血小板活化因子（PAF）、佛波酯（PMA）或离子霉素引发O2-反应的必要性。尽管PMA引发反应不需要[Ca2+]o，但正如预期，PAF和离子霉素的最佳引发效应需要[Ca2+]o。PAF引发功能中[Ca2+]o的半数有效剂量（ED50）值为105微摩尔。离子霉素的[Ca2+]o依赖性引发呈双峰，[Ca2+]o的两个ED50值分别为90微摩尔和3.2毫摩尔。PAF的最佳引发需要细胞至少暴露于[Ca2+]o 4分钟。PAF引发的细胞在添加FMLP后表现出更快的初始O2-产生速率，但O2-产生的持续时间未延长。虽然PAF引发的对FMLP的反应通常与添加FMLP后细胞内钙（[Ca2+]i）增加有关，但发现两种情况下，PAF引发的细胞对FMLP的O2-反应在[Ca2+]i无任何可检测增加的情况下发生。当细胞用钙螯合剂双（O-氨基苯氧基）乙烷-N，N，N'，N'-四乙酸加载，然后用PAF引发时，形成正常量的肌醇1，4，5-三磷酸，但添加FMLP后[Ca2+]i未增加，尽管细胞表现出完全引发的O2-反应；在用PAF引发然后用FMLP刺激的Ca2+耗尽和离子霉素通透的细胞中，O2-产生量与参考对照（引发）细胞相当，但添加FMLP到PAF引发的细胞后肌醇1，4，5-三磷酸产生受抑制且[Ca2+]i无增加。这些数据证实了[Ca2+]o对PAF和离子霉素（但不是PMA引发的细胞）引发中性粒细胞的最佳作用是必需的，并表明在某些条件下，PAF引发的中性粒细胞对FMLP的O2-反应可以在[Ca2+]i无任何增加的情况下发生。
