Kitagawa S, Johnston R B
J Immunol. 1985 Nov;135(5):3417-23.
In an attempt to understand better the molecular basis for the enhanced respiratory burst of activated macrophages (M phi), we investigated the relationship between stimulus-induced changes in membrane potential and release of superoxide anion (O2-) in mouse peritoneal M phi. Resident M phi and M phi elicited by injection of lipopolysaccharide (LPS-M phi) or obtained from animals infected with bacille Calmette-Guérin (BCG-M phi) were used. LPS-M phi and BCG-M phi showed more pronounced changes in membrane potential (depolarization) and greater release of O2- on contact with phorbol myristate acetate (PMA) than did resident macrophages. The lag time between addition of stimulus and onset of release of O2- was reduced in activated compared with resident cells. Membrane potential changes began 60 to 90 sec before release of O2- could be detected in each cell type. The dose-response curves for triggering of membrane potential changes and O2- release by PMA were identical. The magnitude of membrane potential changes and of O2- release in LPS-M phi and BCG-M phi declined progressively during in vitro culture, and values on day 3 approached those in resident macrophages ("deactivation"). Extracellular glucose was required for effective stimulated change in membrane potential and O2- release. These findings indicate that membrane potential changes are closely associated with O2- -releasing capacity in macrophages, and that the systems that mediate membrane potential changes and production of O2- develop or decline concomitantly during activation or deactivation of the cells. Although the plasma membrane was highly depolarized by high extracellular K+ or by the sodium ionophore gramicidin, O2- release was not induced by these maneuvers, indicating that changes in membrane potential by themselves are not sufficient to trigger the respiratory burst in macrophages. Release of O2- was not impaired in buffers in which Na+ was completely replaced with equimolar concentrations of K+ or choline+; thus, induction or maintenance of the respiratory burst in M phi does not require an influx of Na+.
为了更好地理解活化巨噬细胞（M phi）呼吸爆发增强的分子基础，我们研究了小鼠腹腔M phi中刺激诱导的膜电位变化与超氧阴离子（O2-）释放之间的关系。使用了驻留M phi以及通过注射脂多糖诱导产生的M phi（LPS-M phi）或从感染卡介苗的动物中获得的M phi（BCG-M phi）。与驻留巨噬细胞相比，LPS-M phi和BCG-M phi在与佛波酯（PMA）接触时显示出更明显的膜电位变化（去极化）和更大的O2-释放。与驻留细胞相比，活化细胞中刺激添加与O2-释放开始之间的延迟时间缩短。在每种细胞类型中，在可检测到O2-释放之前60至90秒开始出现膜电位变化。PMA触发膜电位变化和O2-释放的剂量反应曲线相同。LPS-M phi和BCG-M phi中膜电位变化和O2-释放的幅度在体外培养期间逐渐下降，第3天的值接近驻留巨噬细胞中的值（“失活”）。有效的刺激诱导的膜电位变化和O2-释放需要细胞外葡萄糖。这些发现表明膜电位变化与巨噬细胞中O2-释放能力密切相关，并且在细胞活化或失活期间，介导膜电位变化和O2-产生的系统同时发育或衰退。尽管质膜被高细胞外K+或钠离子载体短杆菌肽高度去极化，但这些操作并未诱导O2-释放，这表明膜电位变化本身不足以触发巨噬细胞中的呼吸爆发。在Na+完全被等摩尔浓度的K+或胆碱+替代的缓冲液中，O2-释放未受损；因此，M phi中呼吸爆发的诱导或维持不需要Na+内流。
