预激活的中性粒细胞需要磷脂酸才能实现最大程度的受体激活弹性蛋白酶释放。

Primed neutrophils require phosphatidic acid for maximal receptor-activated elastase release.

作者信息

Tamura D Y, Moore E E, Partrick D A, Johnson J L, Zallen G, Silliman C C

机构信息

Denver Health Medical Center, University of Colorado Health Sciences Center, Denver, Colorado, 80204, USA.

出版信息

J Surg Res. 1998 Jun;77(1):71-4. doi: 10.1006/jsre.1998.5342.

DOI:10.1006/jsre.1998.5342

PMID:9698536

Abstract

BACKGROUND

Priming of neutrophils (PMNs) for protease release is believed to be central to the pathogenesis of PMN-mediated tissue injury observed in ARDS/MOF. Defining the intracellular signaling pathways involved with this excessive protease release may aid in establishing future therapies for ARDS. Phospholipase D (PLD) production of phosphatidic acid (PA) is thought to be pivotal in reactive oxygen species generation but its role in degranulation (i. e., protease release) remains unclear. Our hypothesis was that primed neutrophils require PLD production of PA for maximal activated release of elastase.

METHODS

Isolated human PMNs were incubated with a well described antagonist of PA production, ethanol (ETOH, 100-1000 mg/dL), and then primed (PAF, 200 nM) followed by activation (fMLP, 1 microM). To mimic fMLP receptor-dependent activation, PMNs were primed and then directly activated with exogenous dioctanoyl l-alpha-phosphatidic acid (PA8, 0.5-200 microM). To confirm the importance of PA in elastase release, PA8 was given to primed-activated PMNs after ethanol pretreatment in an attempt to recover the maximal response. Elastase release was measured by the cleavage of AAPV-pNA.

RESULTS

PA blockade with ETOH attenuated PAF-primed/fMLP-activated PMN elastase release in a dose-dependent manner. Exogenous PA8 reproduced maximally primed-activated PMN elastase release, and furthermore, PA8 was able to restore maximal elastase release following ethanol attenuation.

CONCLUSIONS

Elastase release from PAF-primed/ fMLP-activated neutrophils is dependent on PA production. Thus, PA production appears to be involved in both oxidant-dependent and independent mechanisms of neutrophil cytotoxicity and may be a potential therapeutic target in the treatment of hyperinflammatory diseases such as ARDS/MOF.

摘要

背景

中性粒细胞（PMN）蛋白酶释放的启动被认为是急性呼吸窘迫综合征/多器官功能障碍综合征（ARDS/MOF）中PMN介导的组织损伤发病机制的核心。确定与这种过度蛋白酶释放相关的细胞内信号通路可能有助于建立未来治疗ARDS的方法。磷脂酶D（PLD）产生磷脂酸（PA）被认为在活性氧生成中起关键作用，但其在脱颗粒（即蛋白酶释放）中的作用仍不清楚。我们的假设是，预激活的中性粒细胞需要PLD产生PA才能实现弹性蛋白酶的最大激活释放。

方法

将分离的人PMN与一种已充分描述的PA产生拮抗剂乙醇（ETOH，100 - 1000 mg/dL）孵育，然后进行预激活（血小板活化因子，PAF，200 nM），接着进行激活（N-甲酰甲硫氨酸-亮氨酸-苯丙氨酸，fMLP，1 μM）。为模拟fMLP受体依赖性激活，PMN先进行预激活，然后用外源性二辛酰基-L-α-磷脂酸（PA8，0.5 - 200 μM）直接激活。为证实PA在弹性蛋白酶释放中的重要性，在乙醇预处理后的预激活-激活PMN中给予PA8，试图恢复最大反应。通过AAPV-pNA的裂解来测量弹性蛋白酶的释放。