HUMAN NEUTROPHIL HETEROGENEITY IDENTIFIED USING FLOW MICRO-FLUOROMETRY TO MONITOR MEMBRANE-POTENTIAL
HUMAN NEUTROPHIL HETEROGENEITY IDENTIFIED USING FLOW MICRO-FLUOROMETRY TO MONITOR MEMBRANE-POTENTIAL
复制标题
DOI：
10.1172/jci110356
复制
发表时间：
1981-01-01
影响因子：
15.9
通讯作者：
GALLIN, JI
中科院分区：
文献类型：
--
作者：
SELIGMANN, B;CHUSED, TM;GALLIN, JI
以往的研究表明，中性粒细胞硝基蓝四唑染料减少响应内毒素和玫瑰花结的IgG包被的红细胞表明外周血中性粒细胞的功能异质性。流动显微荧光法和膜电位敏感的荧光染料3- 3“-dipentyloxacarbocyanine在这里被用来评估中性粒细胞的异质性激活后，由各种刺激。来自正常受试者的未受刺激的中性粒细胞表现出单峰分布的荧光，这表明所有的细胞具有相同的静息膜电位。随着中性粒细胞老化（> 5小时），一些细胞失去荧光，产生双峰分布。在新鲜细胞的研究中，促分泌剂佛波醇肉豆蔻酸酯乙酸酯（20 ng/ml）刺激荧光的均匀损失（表观去极化）。化学引诱物f-Met-Leu-Phe（0.1 μ M）引起嗜中性粒细胞呈现并维持（> 30分钟）双峰荧光分布，其中65 ± 0.001 μ M的荧光分布在中性粒细胞中。5%的中性粒细胞首先降低，然后增加荧光（明显的去极化/部分复极化），35 ± 0.5%的中性粒细胞首先降低，然后增加荧光（明显的去极化/部分复极化）。5%的细胞表现出荧光增加（表观超极化）或无变化。在刺激前用细胞松弛素B处理中性粒细胞，导致细胞对f-Met-Leu-Phe的反应均匀。使用慢性肉芽肿性肉芽肿病患者的中性粒细胞进行的其他研究表明，这种缺陷均匀地影响了所有这些中性粒细胞，这些中性粒细胞表现出异常的膜电位反应。需要调查的中性粒细胞功能的异质性的生理意义证明。在大量细胞群研究中观察到的f-Met-Leu-Phe诱导的膜电位变化是2种不同反应的总和。
Previous studies of neutrophil nitro-blue tetrazolium dye reduction in response to endotoxin and rosetting of IgG-coated erythrocytes have suggested functional heterogeneity of peripheral blood neutrophils. Flow microfluorometry and the membrane potential-sensitive fluorescent dye 3-3''-dipentyloxacarbocyanine were used here to assess the heterogeneity of neutrophils upon activation by a variety of stimuli. Unstimulated neutrophils from normal subjects exhibited a unimodal distribution of fluorescence, suggesting that all the cells possessed the same resting membrane potential. As neutrophils aged (> 5 h), some cells lost fluorescence producing a bimodal distribution. In studies with fresh cells, the secretagogue phorbol myristate acetate (20 ng/ml) stimulated a uniform loss of fluorescence (apparent depolarization). The chemoattractant f-Met-Leu-Phe (0.1 .mu.M) caused the neutrophils to assume and maintain (for > 30 min) a bimodal fluorescence distribution in which 65 .+-. 5% of the neutrophils first decreased and then increased fluorescence (apparent depolarization/partial repolarization), and 35 .+-. 5% of the cells exhibited either an increase in fluorescence (apparent hyperpolarization) or no change. Treatment of neutrophils with cytochalasin B before stimulation caused the cells to respond homogeneously to f-Met-Leu-Phe. Additional studies using neutrophils from patients with chronic granulomatus granulomatous disease, which exhibit abnormal membrane potential responses, indicated that this defect affected all such neutrophils uniformly. The need to investigate the physiological significance of the heterogeneity of neutrophil function is demonstrated. The f-Met-Leu-Phe-induced changes in membrane potential observed in bulk population cell studies are the summation of 2 different responses.