The metabolism of leukotriene B4 in Lewis lung carcinoma porcine kidney cells.

Abstract

Leukotriene B 4 [5(S), 12(R)-dihydroxy-6(Z),8(E),10(E),14(Z)-eicosatetraenoic acid, (LTB 4 )] is a product of the 5-lipoxygenase pathway of arachidonic acid metabolism operational in several cell types, including the human polymorphonuclear leukocyte (1, 2). Because of its purported biological function to mediate neutrophil infiltration, there is considerable interest in an understanding of the biosynthesis of this eicosanoid during inflammatory reactions and, in particular, during inflammatory reactions in the lung. Several studies have shown that LTB 4 can be biosynthesized in the challenged human lung and measured in bronchoalveolar lavage fluid (3-5). LTB 4 , like other eicosanoids, is rapidly metabolized in vivo by multiple pathways into biologically inactive products. Certain cells, including the human polymorphonuclear leukocyte. are known to express a specific cytochrome P-450 (CYP4F3) that carries out ω-oxidation of LTB 4 to form 20-hydroxy-LTB 4 (6). (3-Oxidation is known to be a predominant pathway of LTB 4 metabolism in the hepatocyte after the initial ω-oxidation of the methyl terminus and alcohol dehydrogenase-mediated formation of an ω-carboxyl moiety (7, 8). Several other enzymatic pathways have been discovered that transform LTB 4 into inactive products, including the 12-hydroxyeicosanoid dehydrogenase (9) and 10,11-reductase pathways (10-12), which ultimately convert the conjugated triene into a conjugated diene moiety. Most recently, the formation of a glucuronide of LTB 4 has been described (13). Emergence of electrospray tandem mass spectrometry has facilitated the studies of eicosanoid metabolism including the metabolism of LTB 4 (14, 15). This sensitive technique permits direct analysis of metabolites as they are separated and eluted from a high-performance liquid chromatography (HPLC) column. With the implementation of microbore HPLC columns, concentration of metabolites can be maintained at high levels, ensuring maximal sensitivity of this mass spectrometric technique. While electrospray ionization generates abundant negative ions such as the carboxylate anion, tandem mass spectrometry provides structural information necessary to characterize the metabolites. Interest in the production of LTB 4 within pulmonary tissue has increased, but the means by which one can assess in vivo production of LTB 4 in the lung has been problematic because the lung is a difficult organ to sample. One strategy that has been used for the assessment of prostaglandin production in vivo is the measurement of specific prostaglandin metabolites excreted into the urine (16). Rates of endogenous LTC 4 production have been estimated by measuring urinary LTE 4 excretion (17). Therefore, the identification of stable metabolites of LTB 4 as markers of LTB 4 production and a more complete understanding of the ultimate metabolic fate of LTB 4 has become of interest. The metabolism of eicosanoids within the intact organism could involve multiple tissues, for example, an eicosanoid could be metabolized directly in the tissue responsible for its biosynthesis and subsequently enter the circulation and be transported to a secondary site such as the liver or kidney. Some studies, which have suggested that LTB 4 rapidly exits the lung once it enters the pulmonary circulation (18), bring into focus other tissue sites of LTB 4 metabolism before excretion into urine.