Barzaghi G, Sarau H M, Mong S
Laboratory for Cardiovascular Clinical Pharmacology, Instituto di Ricerche Farmacologiche Mario Negri, Milano, Italy.
J Pharmacol Exp Ther. 1989 Feb;248(2):559-66.
Human monocytic leukemic U-937 cells, when differentiated with dimethylsulfoxide to macrophage-like state, express receptors for platelet-activating factor (PAF). In the differentiated U-937 cells, PAF induced hydrolysis of phosphoinositides and synthesis of inositol phosphates. PAF-induced production of inositol phosphates was rapid, concentration-dependent and was inhibited by a receptor antagonist CV3988, indicating that it was mediated via a specific receptor. In fura-2-loaded, differentiated U-937 cells, PAF induced immediate and concentration-dependent calcium mobilization [( Ca++]i) that was inhibited by CV3988, but not by calcium channel blockers. Addition of an increasing concentration of calcium chelator, ethylene glycol bis(beta-aminoethyl ether)-N,N'-tetraacetic acid, to the medium inhibited a large fraction (approximately 75%) of PAF receptor-induced [Ca++]i mobilization thus suggesting the majority of [Ca++]i mobilization was originated from extracellular milieu and a small portion (approximately 25%) was originated from intracellular sources. The inositol phosphate production induced by PAF, however, was independent from the extracellular calcium and was not inhibited by the addition of ethylene glycol bis(beta-aminoethyl ether)-N,N'-tetraacetic acid. Neither [Ca++]i mobilization or phosphoinositide metabolism in U-937 cells was sensitive to treatment of pertussis toxin, but both types of effects were sensitive to treatment by an inhibitor of phospholipase C, manoalide. These results suggest that in differentiated U-937 cells PAF receptor is coupled through a pertussis toxin-insensitive guanine nucleotide binding protein to a phosphoinositide specific phospholipase C. Inositol-trisphosphate, and possibly diacylglycerol, could be the intracellular messengers for PAF receptor in U-937 cells.
人单核细胞白血病U - 937细胞在用二甲基亚砜分化为巨噬细胞样状态时，表达血小板活化因子（PAF）受体。在分化的U - 937细胞中，PAF诱导磷酸肌醇水解并合成肌醇磷酸。PAF诱导的肌醇磷酸生成迅速，呈浓度依赖性，并被受体拮抗剂CV3988抑制，表明其通过特异性受体介导。在加载fura - 2的分化U - 937细胞中，PAF诱导即刻且浓度依赖性的钙动员[（Ca++]i），该过程被CV3988抑制，但不受钙通道阻滞剂抑制。向培养基中添加浓度递增的钙螯合剂乙二醇双（β - 氨基乙醚）- N，N'-四乙酸可抑制大部分（约75%）PAF受体诱导的[Ca++]i动员，因此表明大部分[Ca++]i动员源自细胞外环境，一小部分（约25%）源自细胞内来源。然而，PAF诱导的肌醇磷酸生成与细胞外钙无关，且不受乙二醇双（β - 氨基乙醚）- N，N'-四乙酸添加的抑制。U - 937细胞中的[Ca++]i动员或磷酸肌醇代谢对百日咳毒素处理均不敏感，但这两种效应均对磷脂酶C抑制剂 manoalide处理敏感。这些结果表明，在分化的U - 937细胞中，PAF受体通过对百日咳毒素不敏感的鸟嘌呤核苷酸结合蛋白与磷酸肌醇特异性磷脂酶C偶联。肌醇 - 1，4，5 - 三磷酸以及可能的二酰基甘油可能是U - 937细胞中PAF受体的细胞内信使。
