Department of Molecular Biophysics and Physiology, Rush University Medical Center, Chicago, Illinois 60612, USA.
Am J Physiol Cell Physiol. 2012 Jan 1;302(1):C286-95. doi: 10.1152/ajpcell.00335.2011. Epub 2011 Oct 19.
Reactive oxygen species (ROS) production by human monocytes differs profoundly from that by neutrophils and eosinophils in its dependence on external media glucose. Activated granulocytes produce vast amounts of ROS, even in the absence of glucose. Human peripheral blood monocytes (PBM), in contrast, are suspected not to be able to produce any ROS if glucose is absent from the media. Here we compare ROS production by monocytes and neutrophils, measured electrophysiologically on a single-cell level. Perforated-patch-clamp measurements revealed that electron current appeared after stimulation of PBM with phorbol myristate acetate. Electron current reflects the translocation of electrons through the NADPH oxidase, the main source of ROS production. The electron current was nearly abolished by omitting glucose from the media. Furthermore, in preactivated glucose-deprived cells, electron current appeared immediately with the addition of glucose to the bath. To characterize glucose dependence of PBM further, NADPH oxidase activity was assessed as hydrogen peroxide (H(2)O(2)) production and was recorded fluorometrically. H(2)O(2) production exhibited similar glucose dependence as did electron current. We show fundamental differences in the glucose dependence of ROS in human monocytes compared with human neutrophils.
活性氧 (ROS) 的产生在人类单核细胞中与中性粒细胞和嗜酸性粒细胞有很大的不同,它依赖于外部介质葡萄糖。激活的粒细胞即使没有葡萄糖也能产生大量的 ROS。相比之下,人类外周血单核细胞 (PBM) 如果培养基中没有葡萄糖,则被怀疑不能产生任何 ROS。在这里,我们比较了单核细胞和中性粒细胞的 ROS 产生,在单细胞水平上进行了电生理测量。穿孔膜片钳测量显示,在用佛波醇肉豆蔻酸酯刺激 PBM 后出现电子电流。电子电流反映了通过 NADPH 氧化酶的电子转移,NADPH 氧化酶是 ROS 产生的主要来源。电子电流几乎被从培养基中去除葡萄糖所消除。此外,在预先激活的葡萄糖剥夺细胞中,当将葡萄糖添加到浴液中时,电子电流立即出现。为了进一步描述 PBM 对葡萄糖的依赖性,评估了 NADPH 氧化酶活性作为过氧化氢 (H(2)O(2)) 的产生,并通过荧光法记录。H(2)O(2) 的产生与电子电流一样表现出相似的葡萄糖依赖性。与人类中性粒细胞相比,我们展示了人类单核细胞中 ROS 在葡萄糖依赖性方面的根本差异。