Biosynthesis, uptake, and degradation of anandamide and palmitoylethanolamide in leukocytes
Biosynthesis, uptake, and degradation of anandamide and palmitoylethanolamide in leukocytes
复制标题
DOI：
10.1074/jbc.272.6.3315
复制
发表时间：
1997-02-07
影响因子：
4.8
通讯作者：
DiMarzo, V
中科院分区：
文献类型：
--
作者：
Bisogno, T;Maurelli, S;DiMarzo, V
Anandamide（花生四烯酰乙醇酰胺，AnNH）和棕榈酰乙醇酰胺（PEA）已被提议分别作为中枢和外周大麻素受体的生理配体。这两种受体均在免疫细胞中表达，包括巨噬细胞和肥大细胞/嗜碱性粒细胞，最近报道了 AnNH 和 PEA 的免疫调节和/或抗炎作用。现在，我们通过证明 AnNH 和 PEA 的生物合成、摄取和降解发生在白细胞中，为这些作用提供了生化依据。在离子霉素刺激下，J774 巨噬细胞和 RBL-2H3 嗜碱性粒细胞可能通过水解相应的 N-酰基磷脂酰乙醇胺（也存在于内源性磷脂中）产生 AnNH 和 PEA。 RBL-2H3 细胞的免疫攻击也导致 AnNH 和 PEA 释放。通过双放射性标记实验和同位素稀释气相色谱/电子轰击质谱法测定了离子霉素刺激后产生的 AnNH 和 PEA 的化学结构和量。两种细胞系通过温度依赖性、可饱和和化学失活机制迅速从培养基中隔离两种酰胺。一旦被嗜碱性粒细胞摄取，AnNH 和 PEA 就会竞争相同的失活酶，催化它们水解成乙醇胺。这种酶在 RBL 细胞匀浆的微粒体和 10,000 x g 级分中均被发现，并且表现出相似的抑制和温度/pH 依赖性特征，但相对于神经元“anandamide 酰胺水解酶”，PEA 的亲和力显着更高。在巨噬细胞和嗜碱性粒细胞中 AnNH 和 PEA 的生物合成和失活机制的发现支持了先前提出的作为这两种长链免疫/炎症反应的局部调节剂的作用。酰基乙醇酰胺。
Anandamide (arachidonoylethanolamide, AnNH) and palmitoylethanolamide (PEA) have been proposed as the physiological ligands, respectively, of central and peripheral cannabinoid receptors. Both of these receptors are expressed in immune cells, including macrophages and mast cells/basophils, where immunomodulatory and/or anti-inflammatory actions of AnNH and PEA have been recently reported. We now provide biochemical grounds to these actions by showing that the biosynthesis, uptake, and degradation of AnNH and PEA occur in leukocytes. On stimulation with ionomycin, J774 macrophages and RBL-2H3 basophils produced AnNH and PEA, probably through the hydrolysis of the corresponding N-acylphosphatidylethanolamines, also found among endogenous phospholipids. Immunological challenge of RBL-2H3 cells also caused AnNH and PEA release. The chemical structure and the amounts of AnNH and PEA produced upon ionomycin stimulation were determined by means of double radiolabeling experiments and isotope dilution gas chromatography/ electron impact mass spectrometry. Both cell lines rapidly sequestered the two amides from the culture medium through temperature-dependent, saturable and chemically inactivable mechanisms. Once uptaken by basophils, AnNH and PEA compete for the same inactivating enzyme which catalyzes their hydrolysis to ethanolamine. This enzyme was found in both microsomal and 10,000 x g fractions of RBL cell homogenates, and exhibited similar inhibition and temperature/pH dependence profiles but a significantly higher affinity for PEA with respect to neuronal ''anandamide amidohydrolase.'' The finding of biosynthetic and inactivating mechanisms for AnNH and PEA in macrophages and basophils supports the previously proposed role as local modulators of immune/inflammatory reactions for these two long chain acylethanolamides.