MacGlashan D W
Johns Hopkins Asthma and Allergy Center, Baltimore, MD 21224, USA.
J Leukoc Biol. 1995 Aug;58(2):177-88. doi: 10.1002/jlb.58.2.177.
Human basophils, stimulated with either anti-IgE antibody or formyl-methionine-leucine-phenylalanine, were examined by two measures of the cell response that may reflect degranulation. Flow cytometric measurement of either of these two measures, changes in forward scatter intensity (an indirect measure of the basophil size) or changes in the intensity of acridine orange-loaded cells (which labels basophil granules), allowed an assessment of the distribution of single cell responses. With regard to the latter technique, structures that appeared to be basophil granules were shown to metachromatically label with low concentrations of acridine orange, which has little or no effect on histamine release. During stimulation these labeled granules were lost, leading to decreased fluorescence. Changes in either the forward scatter parameter or acridine orange labeling occurred on the same time scale as histamine release, differentiating these measures of the basophil response from early signal transduction events. Challenging basophils with a combination of phorbol myristate acetate and ionomycin caused 100% histamine release and allowed a measurement of the maximum change in forward scatter intensity or loss of acridine orange labeling. The flow cytometric distributions after this treatment were then compared with the distributions obtained by challenging cells with several concentrations of anti-IgE antibody or formyl-methionine-leucine-phenylalanine, which induced various submaximal responses. These flow cytometric distributions demonstrated that single cells could be found in intermediate states of activation, i.e., the response of all cells was graded according to the strength of the stimulus. These studies lead to the general conclusion that all aspects of the basophil response, including those late events in the basophil response we have studied here, as well as early events that we have studied previously, are graded in a continuous manner, according to the magnitude of the stimulus.
用抗IgE抗体或甲酰甲硫氨酸-亮氨酸-苯丙氨酸刺激人嗜碱性粒细胞后，通过两种可能反映脱颗粒的细胞反应测量方法进行检测。这两种测量方法中的任何一种，即前向散射强度的变化（嗜碱性粒细胞大小的间接测量）或吖啶橙负载细胞强度的变化（标记嗜碱性粒细胞颗粒），通过流式细胞术测量，可评估单细胞反应的分布。关于后一种技术，似乎是嗜碱性粒细胞颗粒的结构显示用低浓度吖啶橙进行异染性标记，而吖啶橙对组胺释放几乎没有影响。在刺激过程中，这些标记的颗粒丢失，导致荧光降低。前向散射参数或吖啶橙标记的变化与组胺释放发生在同一时间尺度上，从而将这些嗜碱性粒细胞反应的测量方法与早期信号转导事件区分开来。用佛波酯肉豆蔻酸酯和离子霉素联合刺激嗜碱性粒细胞可导致100%组胺释放，并可测量前向散射强度的最大变化或吖啶橙标记的丢失。然后将这种处理后的流式细胞术分布与用几种浓度的抗IgE抗体或甲酰甲硫氨酸-亮氨酸-苯丙氨酸刺激细胞所获得的分布进行比较，这些刺激诱导了各种次最大反应。这些流式细胞术分布表明，单细胞可处于激活的中间状态，即所有细胞的反应根据刺激强度分级。这些研究得出的一般结论是，嗜碱性粒细胞反应的所有方面，包括我们在此研究的嗜碱性粒细胞反应中的那些晚期事件，以及我们先前研究的早期事件，都根据刺激的大小以连续的方式分级。
