Goldman D W, Enkel H, Gifford L A, Chenoweth D E, Rosenbaum J T
J Immunol. 1986 Sep 15;137(6):1971-6.
Peripheral blood polymorphonuclear leukocytes (PMNL) isolated from rabbits after an i.v. injection of endotoxin exhibited decreased chemotactic migration in response to leukotriene B4 (LTB4) and C5a, but not N-formyl-methionyl-leucyl-phenylalanine (fMLP), after endotoxin treatment. The binding of radiolabeled LTB4, fMLP, and C5a to isolated PMNL was assessed in order to determine whether altered receptor expression could account for the observed functional changes. Control PMNL expressed binding sites for fMLP, LTB4, and C5a similar to those previously characterized from human PMNL. Control PMNL expressed a single class of 14,600 +/- 2700 receptors for fMLP with a mean dissociation constant (Kd) of 2.0 +/- 0.6 nM at 0 degrees C, whereas two subclasses of binding sites were expressed for LTB4: 10,300 +/- 6800 high-affinity and 85,600 +/- 53,000 low-affinity binding sites per PMNL with mean Kd for LTB4 of 0.75 +/- 0.43 nM and 70 +/- 58 nM (mean +/- SD, n = 5), respectively. Control PMNL bound [125I]-C5a in a dose-dependent and saturable manner at 24 degrees C. At saturating concentrations of C5a, PMNL obtained from control rabbits bound 270,000 +/- 50,000 molecules of [125I]-C5a with half-maximal binding occurring at [125I]-C5a concentrations of 5.5 +/- 1.9 nM. The binding of LTB4 and C5a to PMNL obtained 24 hr after an i.v. injection of endotoxin was markedly decreased compared with control PMNL. PMNL from endotoxin-treated rabbits exhibited 68% fewer high-affinity binding sites per PMNL for LTB4 and a 51% decrease in the amount of [125I]-C5a bound at saturating concentrations compared with control PMNL. There was no significant change in the Kd of the high-affinity binding sites for LTB4, no change in the Kd and number of the low-affinity binding sites for LTB4, and a small decrease in the apparent Kd for C5a to 3.3 +/- 1.1 nM. Even though the pretreatment with i.v. endotoxin did not alter chemotactic or degranulation responses elicited by fMLP, the endotoxin pretreatment induced an eightfold increase in the receptor density without altering the Kd for fMLP. Decreased receptor expression could account in large part for the decreased chemotactic responsiveness towards C5a and LTB4 induced by LPS. The finding that a substantial increase in receptors for fMLP need not be accompanied by a comparable functional change suggests that decreased efficiency in receptor coupling to intracellular biochemical events may also result from i.v. endotoxin.
静脉注射内毒素后从兔体内分离出的外周血多形核白细胞（PMNL），在内毒素处理后，对白三烯B4（LTB4）和C5a的趋化性迁移反应降低，但对N-甲酰甲硫氨酰-亮氨酰-苯丙氨酸（fMLP）的反应未降低。为了确定受体表达的改变是否可以解释观察到的功能变化，评估了放射性标记的LTB4、fMLP和C5a与分离出的PMNL的结合情况。对照PMNL表达的fMLP、LTB4和C5a结合位点与先前从人PMNL中鉴定的相似。对照PMNL在0℃时表达一类单一的14,600±2700个fMLP受体，平均解离常数（Kd）为2.0±0.6 nM，而LTB4表达两个结合位点亚类：每个PMNL有10,300±6800个高亲和力和85,600±53,000个低亲和力结合位点，LTB4的平均Kd分别为0.75±0.43 nM和70±58 nM（平均值±标准差，n = 5）。对照PMNL在24℃时以剂量依赖性和饱和方式结合[125I]-C5a。在C5a饱和浓度下，从对照兔获得的PMNL结合270,000±50,000个[125I]-C5a分子，半数最大结合发生在[125I]-C5a浓度为5.5±1.9 nM时。与对照PMNL相比，静脉注射内毒素24小时后获得的PMNL对LTB4和C5a的结合明显减少。与对照PMNL相比，内毒素处理兔的PMNL每个PMNL的LTB4高亲和力结合位点减少68%，在饱和浓度下结合的[125I]-C5a量减少51%。LTB4高亲和力结合位点的Kd没有显著变化，LTB4低亲和力结合位点的Kd和数量没有变化，C5a的表观Kd略有下降至3.3±1.1 nM。尽管静脉注射内毒素预处理没有改变fMLP引起 的趋化或脱颗粒反应，但内毒素预处理使受体密度增加了八倍，而没有改变fMLP的Kd。受体表达降低在很大程度上可以解释LPS诱导的对C5a和LTB4趋化反应性降低。fMLP受体大量增加不一定伴随着相应功能变化这一发现表明，静脉注射内毒素也可能导致受体与细胞内生化事件偶联效率降低。
