Clarke Deborah L, Belvisi Maria G, Hardaker Elizabeth, Newton Robert, Giembycz Mark A
Thoraic Medicine, National Heart and Lung Institute, Imperial College London, London, UK.
Mol Pharmacol. 2005 Feb;67(2):383-93. doi: 10.1124/mol.104.006486. Epub 2004 Nov 4.
8-Isoprostanes are bioactive lipid mediators formed via the nonenzymatic peroxidation of arachidonic acid by free radicals and reactive oxygen species. However, their cognate receptors, biological actions, and signaling pathways are poorly studied. Here, we report the effect of a variety of E- and Falpha-ring 8-isoprostanes on the release of granulocyte/macrophage colony-stimulating factor (GM-CSF) and granulocyte colony-stimulating factor (G-CSF) from human airway smooth muscle (HASM) cells stimulated with interleukin-1beta (IL-1beta). The elaboration of GM-CSF and G-CSF by IL-1beta was inhibited and augmented, respectively, in a concentration-dependent manner by 8-iso-prostaglandin (PG) E(1) and 8-iso-PGE(2), but not by 8-iso-PGF(1alpha), 8-iso-PGF(2alpha), and 8-iso-PGF(3)alpha. AH 6809 (6-isopropoxy-9-oxoxanthine-2-carboxylic acid), an EP(1)-/EP(2)-/DP-receptor blocking drug, antagonized the inhibitory effect of 8-iso-PGE(1) and 8-iso-PGE(2) on GM-CSF output with an affinity consistent with an interaction at prostanoid receptors of the EP(2)-subtype. In contrast, the facilitation by 8-iso-PGE(1) and 8-iso-PGE(2) of G-CSF release was unaffected by AH 6809 and the selective EP(4)-receptor antagonist L-161,982 [4'-[3-butyl-5-oxo-1-(2-trifluoromethyl-phenyl)-1,5-dihydro-[1,2,4]triazol-4-ylmethyl]-biphenyl-2-sulfonic acid (3-methyl-thiophene-2-carbonyl)-amide]. However, when used in combination, AH 6809 and L-161,982 displaced 5-fold to the right the 8-iso-PGE and 8-iso-PGE concentration-response curves. The opposing (1)effect of E-ring (2)8-isoprostanes on GM-CSF and G-CSF release was mimicked by 8-bromo-cAMP and abolished in cells infected with an adenovirus vector encoding an inhibitor protein of cAMP-dependent protein kinase (PKA). Together, these data demonstrate that E-ring 8-isoprostanes regulate the secretion of GM-CSF and G-CSF from HASM cells by a cAMP- and PKA-dependent mechanism. Moreover, antagonist studies revealed that 8-iso-PGE(1) and 8-iso-PGE(2) act solely via EP(2) -receptors to inhibit GM-CSF release, whereas both EP(2)- and EP(4)-receptor subtypes positively regulate G-CSF output.
8-异前列腺素是通过自由基和活性氧对花生四烯酸进行非酶促过氧化反应而形成的生物活性脂质介质。然而，它们的同源受体、生物学作用和信号通路的研究较少。在此，我们报告了多种E环和Fα环8-异前列腺素对白细胞介素-1β（IL-1β）刺激的人气道平滑肌（HASM）细胞释放粒细胞/巨噬细胞集落刺激因子（GM-CSF）和粒细胞集落刺激因子（G-CSF）的影响。8-异前列腺素（PG）E1和8-异前列腺素E2以浓度依赖性方式分别抑制和增强IL-1β诱导的GM-CSF和G-CSF的释放，但8-异前列腺素F1α、8-异前列腺素F2α和8-异前列腺素F3α则无此作用。AH 6809（6-异丙氧基-9-氧代黄嘌呤-2-羧酸）是一种EP1/EP2/DP受体阻断药物，拮抗8-异前列腺素E1和8-异前列腺素E2对GM-CSF释放的抑制作用，其亲和力与在EP2亚型前列腺素受体上的相互作用一致。相比之下，AH 6809和选择性EP4受体拮抗剂L-161,982 [4'-[3-丁基-5-氧代-1-(2-三氟甲基苯基)-1,5-二氢-[1,2,4]三唑-4-基甲基]-联苯-2-磺酸（3-甲基噻吩-2-羰基）-酰胺]对8-异前列腺素E1和8-异前列腺素E2促进G-CSF释放的作用无影响。然而，联合使用时，AH 6809和L-161,982使8-异前列腺素E1和8-异前列腺素E2的浓度-反应曲线向右移动5倍。8-溴-cAMP模拟了E环8-异前列腺素对GM-CSF和G-CSF释放的相反作用，并且在用编码cAMP依赖性蛋白激酶（PKA）抑制蛋白的腺病毒载体感染的细胞中这种作用被消除。总之，这些数据表明E环8-异前列腺素通过cAMP和PKA依赖性机制调节HASM细胞中GM-CSF和G-CSF的分泌。此外，拮抗剂研究表明，8-异前列腺素E1和8-异前列腺素E2仅通过EP2受体抑制GM-CSF释放，而EP2和EP4受体亚型均正向调节G-CSF的分泌。
