Bass D A, Grover W H, Lewis J C, Szejda P, DeChatelet L R, McCall C E
J Clin Invest. 1980 Dec;66(6):1265-73. doi: 10.1172/JCI109978.
Previous studies of the biochemistry and physiology of eosinophils have relied upon cells obtained from patients with eosinophilia (EE). It is unknown whether such cells might have been activated or partially exhausted by the pathological state causing eosinophilia. We examined cell surface charge, membrane transport of deoxyglucose, activation of lyso-somal acid phosphatase, and oxidative metabolism to provide a profile to compare EE with purified normal eosinophils (NE) and normal neutrophils. Eosinophils or neutrophils were obtained in >95% purity from normal individuals and patients with eosinophilia of diverse etiologies. Cell surface charge was determined by electrophoretic mobility in micromoles per second per volt per centimeter. Normal eosinophils demonstrated a surface charge of 2.46+/-0.03. Stimulation of the cells by zymosan-activated serum (ZAS) reduced the surface charge to 1.82+/-0.02. In contrast, the charge of "resting" EE was already reduced (1.89+/-0.05) and was not altered by ZAS. Resting and stimulated neutrophils had a charge of 1.98+/-0.01 and 1.69+/-0.02, respectively. Uptake of [(3)H]2-deoxyglucose has been shown to reflect carrier-facilitated hexose transport in granulocytes. Deoxyglucose uptake by resting NE and NE stimulated by ZAS was 2.40+/-0.40 and 5.44+/-0.39 (cpm x 10(-3)/2 x 10(5) eosinophils), respectively. Resting and stimulated EE demonstrated deoxyglucose uptake of 7.55+/-0.58 and 15.3+/-0.6, respectively.Lysosomal acid phosphatase was determined by an electron microscopic cytochemical technique. In normal eosinophils and neutrophils, lysosomal acid phosphatase in mature cells is held in a latent form. Normal eosinophils demonstrated weakly positive acid phosphatase activity in 7.8+/-1.2% of the specific granules. Normal eosinophils, stimulated by opsonized staphylococci or the calcium ionophore A23187, develop rapid activation of acid phosphatase in approximately 80% of the granules throughout the cells. Resting EE were usually already activated and demonstrated acid phosphatase in 48.6+/-8.6% of the granules (range, 2-95% granules positive; significant activation was observed in preparations in EE from 11 of 15 patients). Oxidative metabolism was monitored by measurement of the hexose monophosphate shunt (HMPS) (metabolism of 1-[(14)C]glucose to (14)CO(2)). Previous studies demonstrated that resting EE have an HMPS activity which is nearly that of stimulated neutrophils, yet EE remain capable of further 7-10-fold increase when stimulated by opsonized zymosan. In contrast, the HMPS of NE (resting and stimulated) was not significantly different from that of neutrophils. Thus eosinophils obtained from patients with eosinophilia appear significantly activated when compared with normal eosinophils by the criteria of surface charge, activation of lysosomal acid phosphatase, membrane hexose transport, and hexose monophosphate shunt activities.
先前关于嗜酸性粒细胞生物化学和生理学的研究依赖于从嗜酸性粒细胞增多症（EE）患者获取的细胞。尚不清楚这些细胞是否可能已被导致嗜酸性粒细胞增多的病理状态激活或部分耗竭。我们检测了细胞表面电荷、脱氧葡萄糖的膜转运、溶酶体酸性磷酸酶的激活以及氧化代谢，以提供一个特征图谱，用于将EE与纯化的正常嗜酸性粒细胞（NE）和正常中性粒细胞进行比较。从正常个体和不同病因的嗜酸性粒细胞增多症患者中获取纯度>95%的嗜酸性粒细胞或中性粒细胞。细胞表面电荷通过电泳迁移率来确定，单位为微摩尔每秒每伏特每厘米。正常嗜酸性粒细胞的表面电荷为2.46±0.03。用酵母聚糖激活的血清（ZAS）刺激细胞后，表面电荷降至1.82±0.02。相比之下，“静息”EE的电荷已经降低（1.89±0.05），且不受ZAS影响。静息和受刺激的中性粒细胞电荷分别为1.98±0.01和1.69±0.02。已证明[³H]2-脱氧葡萄糖的摄取反映了粒细胞中载体介导的己糖转运。静息NE和受ZAS刺激的NE对脱氧葡萄糖的摄取分别为2.40±0.40和5.44±0.39（每分钟计数×10⁻³/2×10⁵个嗜酸性粒细胞）。静息和受刺激的EE对脱氧葡萄糖的摄取分别为7.55±0.58和15.3±0.6。溶酶体酸性磷酸酶通过电子显微镜细胞化学技术测定。在正常嗜酸性粒细胞和中性粒细胞中，成熟细胞中的溶酶体酸性磷酸酶以潜伏形式存在。正常嗜酸性粒细胞在7.8±1.2%的特异性颗粒中显示出弱阳性酸性磷酸酶活性。用调理过的葡萄球菌或钙离子载体A23187刺激正常嗜酸性粒细胞后，细胞内约80%的颗粒中酸性磷酸酶迅速激活。静息EE通常已经被激活，在48.6±8.6%的颗粒中显示出酸性磷酸酶（范围为2 - 95%的颗粒呈阳性；在15例患者的EE样本中有11例观察到显著激活）。通过测量磷酸己糖旁路（HMPS）（1-[(¹⁴)C]葡萄糖代谢为¹⁴CO₂）来监测氧化代谢。先前的研究表明，静息EE的HMPS活性几乎与受刺激的中性粒细胞相同，但EE在受到调理过的酵母聚糖刺激时仍能够进一步增加7 - 10倍。相比之下（静息和受刺激的）NE的HMPS与中性粒细胞的无显著差异。因此，根据表面电荷、溶酶体酸性磷酸酶激活、膜己糖转运和磷酸己糖旁路活性的标准，与正常嗜酸性粒细胞相比，从嗜酸性粒细胞增多症患者获取的嗜酸性粒细胞似乎已被显著激活。
