Platelets augment granulocyte aggregation and cytotoxicity: uncovering of their effects by improved cell separation techniques using Percoll gradients.
Platelets augment granulocyte aggregation and cytotoxicity: uncovering of their effects by improved cell separation techniques using Percoll gradients.
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血小板增强粒细胞聚集和细胞毒性：通过使用 Percoll 梯度改进的细胞分离技术揭示其作用。
DOI：
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发表时间：
2009
期刊：
影响因子：
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通讯作者：
H. S. Jacob
中科院分区：
文献类型：
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作者：
Marc Boogaerts;G. Vercelotti;C. Roelant;S. Malbrain;R. Verwilghen;H. S. Jacob
用于体外研究的粒细胞制备的“标准”技术使用右旋糖苷去除红细胞和Ficoll-Hypaque梯度离心法来提高粒细胞纯度。这个过程很长，在我们手中大约150分钟，并提供严重污染的血小板(约。5个血小板/中性粒细胞)。我们报道了一种用羟乙基淀粉取代右旋糖苷和用Percoll取代Ficoll-Hypaque的技术。准备时间减少约40%，血小板污染减少80%以上。如此制备的粒细胞代谢功能(O2-产生、化学发光、HMP-最大分流)，在运动/吞噬试验中，与标准制剂相同。然而，发现了血小板在粒细胞聚集反应中的增强作用及其对细胞毒性的中介作用。与Percoll制剂(缺乏血小板)相比，Ficoll-Hypaque纯化细胞(富血小板)与FMLP或激活的补体凝集素聚集的程度明显更高，并过度杀死51Cr标记的靶细胞。将纯化的血小板或血小板释放上清液重新加入到后者中，可以复制使用标准制剂的结果。
The 'standard' technique of granulocyte preparation for in vitro studies uses dextran removal of erythrocytes and Ficoll-Hypaque gradient centrifugation to increase granulocyte purity. The procedure is lengthy, approximately 150 min in our hands, and provides granulocytes significantly contaminated with platelets (approx. 5 platelets/PMN). We report a technique that replaces dextran with hydroxy-ethylstarch and Ficoll-Hypaque with Percoll. Preparation time is reduced by approximately 40% and platelet contamination by more than 80%. Granulocytes, so prepared, function metabolically (O2-generation, chemiluminescence, HMP-shunt maxima) and, in motility/phagocytosis assays, identically to 'standard' preparations. However, an augmentatory effect of platelets in granulocyte aggregation responses and their mediation of cytotoxicity is uncovered. Ficoll-Hypaque purified cells (platelet-rich) aggregate to a significantly greater degree with FMLP or activated complement lectins and excessively kill 51Cr-labelled target cells when compared to Percoll-preparations (platelet-poor). Re-addition of purified platelets or of platelet release supernatants to the latter reproduces results using the 'standard' preparations.