The rabbit pulmonary cytochrome P450 arachidonic acid metabolic pathway: characterization and significance.
The rabbit pulmonary cytochrome P450 arachidonic acid metabolic pathway: characterization and significance.
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DOI：
10.1172/jci117904
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发表时间：
1995-05
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影响因子：
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通讯作者：
D. Zeldin;Jonathan D. Plitman;Jun Kobayashi;Robert F. Miller;James R. Snapper;J. Falck;John L. Szarek;R. Philpot;Jorge H. Capdevila
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文献类型：
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作者：
D. Zeldin;Jonathan D. Plitman;Jun Kobayashi;Robert F. Miller;James R. Snapper;J. Falck;John L. Szarek;R. Philpot;Jorge H. Capdevila
细胞色素 P450 在兔肝脏和肾脏中将花生四烯酸代谢为几种独特的生物活性化合物。由兔肺匀浆制备的微粒体组分通过细胞色素 P450 途径代谢花生四烯酸，产生顺式环氧二十碳三烯酸 (EET) 及其水合产物、vic-二羟基二十碳三烯酸、中链顺反共轭二烯醇以及 19- 和 20- 羟基二十碳四烯酸。使用针对纯化的 CYP2B4 制备的多克隆抗体进行的抑制研究表明，对花生四烯酸环氧化物的形成具有 100% 的抑制作用。纯化的 CYP2B4 在 NADPH-细胞色素 P450 还原酶和细胞色素 b5 存在下重建，代谢花生四烯酸，主要产生 EET。使用气相色谱/质谱法在肺匀浆中检测到 EET，为体内花生四烯酸的肺细胞色素 P450 环氧化提供了证据。这些肺 EET 的手性分析表明优先选择 14(R),15(S)-、11(S),12(R)- 和 8(S),9(R)-EET 对映体。在支气管肺泡灌洗液中检测到 EET 和 vic-二羟基二十碳三烯酸。在微摩尔浓度下，甲基化 5,6-EET 和 8,9-EET 在体外显着松弛组胺收缩的豚鼠肺门支气管。相反，20-羟基二十碳四烯酸引起收缩至接近最大张力。我们得出的结论是，CYP2B4（一种丰富的兔肺细胞色素 P450 酶）是主要的组成性肺花生四烯酸环氧化酶，并且这些本地产生的具有生物活性的类二十烷酸可能参与维持肺内的稳态。
Cytochrome P450 metabolizes arachidonic acid to several unique and biologically active compounds in rabbit liver and kidney. Microsomal fractions prepared from rabbit lung homogenates metabolized arachidonic acid through cytochrome P450 pathways, yielding cis-epoxyeicosatrienoic acids (EETs) and their hydration products, vic-dihydroxyeicosatrienoic acids, mid-chain cis-trans conjugated dienols, and 19- and 20-hydroxyeicosatetraenoic acids. Inhibition studies using polyclonal antibodies prepared against purified CYP2B4 demonstrated 100% inhibition of arachidonic acid epoxide formation. Purified CYP2B4, reconstituted in the presence of NADPH-cytochrome P450 reductase and cytochrome b5, metabolized arachidonic acid, producing primarily EETs. EETs were detected in lung homogenate using gas chromatography/mass spectroscopy, providing evidence for the in vivo pulmonary cytochrome P450 epoxidation of arachidonic acid. Chiral analysis of these lung EETs demonstrated a preference for the 14(R),15(S)-, 11(S),12(R)-, and 8(S),9(R)-EET enantiomers. Both EETs and vic-dihydroxyeicosatrienoic acids were detected in bronchoalveolar lavage fluid. At micromolar concentrations, methylated 5,6-EET and 8,9-EET significantly relaxed histamine-contracted guinea pig hilar bronchi in vitro. In contrast, 20-hydroxyeicosatetraenoic acid caused contraction to near maximal tension. We conclude that CYP2B4, an abundant rabbit lung cytochrome P450 enzyme, is the primary constitutive pulmonary arachidonic acid epoxygenase and that these locally produced, biologically active eicosanoids may be involved in maintaining homeostasis within the lung.