Purification and functional evaluation of mature neutrophils from human bone marrow.
Purification and functional evaluation of mature neutrophils from human bone marrow.
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DOI：
10.1182/blood.v68.4.853.853
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发表时间：
1986-10
期刊：
影响因子：
20.3
通讯作者：
R. Berkow;R. Dodson
中科院分区：
文献类型：
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作者：
R. Berkow;R. Dodson
人髓细胞的成熟在骨髓中进行，产生成熟的中性粒细胞，释放到外周循环中。先前的报道表明，骨髓中性粒细胞在受到刺激时粘附性降低，吞噬功能受损，硝基蓝四氮唑染料还原减少。由于缺乏一种合适的方法来分离纯化骨髓中性粒细胞，这些研究已通过显微镜技术进行。我们现在报告了一种从10ml骨髓抽吸样本中分离1 × 10(8)个中性粒细胞[条带加多形核白细胞（PMNs）]的方法。通过密度为1.085、1.095和1.10 g/mL的不连续percol梯度离心，可以将富含白细胞的骨髓悬浮液分离成三个白细胞层。通过结合较低的两个白细胞层（M2/3），可以看到由26%条带和63% pmn组成的中性粒细胞群。与外周血pmn相比，这些骨髓中性粒细胞碱性磷酸酶活性较低，摄入油红o包被颗粒减少，受趋化肽Fmet-leu-phe （FMLP）或肿瘤促进因子肉豆酸酯phorbol acetate （PMA）刺激时超氧化物释放受损，nadph依赖性氧化酶活性降低。这些结果表明，骨髓中形态成熟的中性粒细胞在功能上仍处于未成熟状态。
Human myeloid maturation proceeds within the bone marrow and results in a mature neutrophil that is released into the peripheral circulation. Previous reports have indicated that neutrophils from bone marrow demonstrate decreased adherence, impaired phagocytosis, and decreased nitroblue tetrazolium dye reduction when stimulated. Due to lack of a suitable method for isolating purified bone marrow neutrophils, these studies have been performed by microscopic techniques. We now report a method for isolating 1 X 10(8) neutrophils [bands plus polymorphonuclear leukocytes (PMNs)] from 10 mL of bone marrow aspirate sample. By means of a discontinuous Percoll-gradient centrifugation through densities of 1.085, 1.095, and 1.10 g/mL a leukocyte-rich suspension of bone marrow can be separated into three leukocyte layers. By combining the lower two leukocyte layers (M2/3), a population of neutrophils consisting of 26% bands and 63% PMNs is seen. When compared with peripheral blood PMNs, these bone marrow neutrophils had a lower alkaline phosphatase activity, decreased ingestion of Oil Red O-coated particles, impaired superoxide release on stimulation with the chemotactic peptide Fmet-leu-phe (FMLP) or the tumor promotor phorbol myristate acetate (PMA), and less activity of the NADPH-dependent oxidase. These results indicate that morphologically mature neutrophilic cells within the bone marrow exist in a still functionally immature state.