Hwang S B
Merck Sharp & Dohme Research Laboratories, Department of Biochemical Regulation, Rahway, New Jersey 07065.
J Biol Chem. 1988 Mar 5;263(7):3225-33.
Due to multiple molecular species of platelet-activating factor (PAF) and the existence of high affinity binding sites in a variety of cells and tissues, possible existence of PAF receptor subtypes has been suggested. This report shows differences between specific PAF receptors in human leukocytes and platelets. Human polymorphonuclear leukocyte membranes showed high affinity binding sites for PAF with an equilibrium dissociation constant (KD) of 4.4 (+/- 0.3) x 10(-10) M. We compared the relative potencies of several PAF agonists and receptor antagonists between human platelet and human leukocyte membranes. One receptor antagonist (Ono-6240) was found to be 6-10 times less potent in inhibiting the specific [3H]PAF receptor binding, PAF-induced GTPase activity, as well as the PAF-induced aggregation in human leukocytes than in human platelets. Mg2+, Ca2+, and K+ ions potentiated the specific [3H]PAF binding in both systems. Na+ and Li+ ions inhibited the specific [3H]PAF binding to human platelets but showed no effects in human leukocytes. K+ ions decreased the Mg2+-potentiated [3H]PAF binding in human leukocytes but showed no effects in human platelets. PAF stimulates the hydrolysis of [gamma-32P] GTP with an ED50 of about 1 nM, whereas the biological inactive enantiomer shows no activity even at 10 microM in both human platelets and human leukocytes. The PAF-stimulated GTPase in human leukocytes can be abolished by the pretreatment of membranes with pertussis toxin and cholera toxin. However, the PAF-stimulated activity of GTPase in human platelets is insensitive to pertussis toxin and cholera toxin. These results suggest that there exists a second type of PAF receptor in human polymorphonuclear leukocytes, which is structurally different from the one characterized in human platelets, and that the guanine nucleotide-binding protein coupled to PAF receptors in human leukocytes is also different from the one in human platelets.
由于血小板活化因子（PAF）存在多种分子形式，且在多种细胞和组织中存在高亲和力结合位点，因此有人提出可能存在PAF受体亚型。本报告显示了人类白细胞和血小板中特异性PAF受体之间的差异。人类多形核白细胞膜显示出对PAF的高亲和力结合位点，平衡解离常数（KD）为4.4（±0.3）×10⁻¹⁰ M。我们比较了几种PAF激动剂和受体拮抗剂在人类血小板膜和人类白细胞膜之间的相对效力。发现一种受体拮抗剂（Ono - 6240）在抑制特异性[³H]PAF受体结合、PAF诱导的GTP酶活性以及PAF诱导的人类白细胞聚集方面，其效力比在人类血小板中低6 - 10倍。Mg²⁺、Ca²⁺和K⁺离子增强了两个系统中特异性[³H]PAF的结合。Na⁺和Li⁺离子抑制了特异性[³H]PAF与人类血小板的结合，但对人类白细胞无影响。K⁺离子降低了Mg²⁺增强的人类白细胞中[³H]PAF的结合，但对人类血小板无影响。PAF刺激[γ - ³²P]GTP水解，半数有效剂量（ED50）约为1 nM，而生物无活性的对映体即使在10 μM时在人类血小板和人类白细胞中均无活性。用百日咳毒素和霍乱毒素预处理膜可消除人类白细胞中PAF刺激的GTP酶活性。然而，人类血小板中PAF刺激的GTP酶活性对百日咳毒素和霍乱毒素不敏感。这些结果表明，人类多形核白细胞中存在第二种PAF受体，其结构与人类血小板中已鉴定的受体不同，并且与人类白细胞中PAF受体偶联的鸟嘌呤核苷酸结合蛋白也与人类血小板中的不同。
