Synthesis and metabolism of leukotrienes in γ-glutamyl transpeptidase deficiency

Abstract

Leukotrienes (LTs) are active lipid mediators derived in the 5-lipoxygenase pathway. LTC 4 , the primary cysteinyl LT, is cleaved by ␥ -glutamyl transpeptidase (GGT), resulting in LTD 4 . We studied the synthesis and metabolism of LTs in three patients with GGT deficiency. LTs were analyzed in urine, plasma, and monocytes after HPLC separation by enzyme immunoassays, radioactivity detection, and electrospray tandem mass spectrometry. Analysis of LTs in urine revealed increased concentrations of LTC 4 (12.8-17.9 nmol/mol creatinine; controls, Ͻ 0.005 nmol/mol creatinine), whereas LTE 4 was below the detection limit ( Ͻ 0.005 nmol/ mol creatinine; controls, 32.2 ؎ 8.6 nmol/mol creatinine). In plasma of one patient, LTC 4 was found to be increased (17.3 ng/ml; controls, 9.6 ؎ 0.4 ng/ml), whereas LTD 4 and LTE 4 were below the detection limit ( Ͻ 0.005 ng/ml). LTB 4 was found within normal ranges. In contrast to controls, the synthesis of LTD 4 and LTE 4 in stimulated monocytes was below the detection limit ( Leukotrienes (LTs) constitute a group of biologically highly active lipid mediators derived from 20-carbon polyunsaturated fatty acids, predominantly arachidonic acid via the 5-lipoxygenase pathway (1-3). They include the cysteinyl LTs LTC 4 , LTD 4 , and LTE 4 , representing biologically active constituents of the long-known "slow-reacting substance of anaphylaxis" and the dihydroxyeicosatetraenoate LTB 4 .The biosynthesis of LTs is limited to a few types of human cells, including mast cells, eosinophils, basophils, and macrophages. The synthesis of LTs is initiated by cell activation with the release of arachidonic acid from membrane phospholipid by the action of cytosolic phospholipase A 2 . Arachidonic acid then binds to the 5-lipoxygenase-activating protein and is presented to 5-lipoxygenase (4). Calcium-dependent activation of 5-lipoxygenase converts arachidonate via 5-hydroperoxyeicosatetraenoate to 5,6-epoxide LTA 4 , which is unstable and is catalyzed to LTB 4 (5, 6). Alternatively, the conjugation of LTA 4 with glutathione at carbon 6 is mediated by LTC 4 synthase, resulting in the formation of LTC 4 , the primary cysteinyl LT (7). LTC 4 is known to be cleaved by ␥ -glutamyl transpeptidase (GGT), which removes the glutamyl moiety to form LTD 4 (1). LTC 4 conversion to LTD 4 has long been thought to be mediated solely by GGT. The cleavage of glycine from LTD 4 yields LTE 4 (8).Some years ago, a human gene was cloned that appeared to direct the cleavage of LTC 4 . This enzyme was termed ␥ -glutamyl transpeptidase-related (GGT-rel) (9). GGT-rel shares an overall 40% amino acid sequence identity with human GGT and is capable of cleaving the ␥ -glutamyl linkage of LTC 4 , but it is unable to hydrolyze synthetic substrates that are commonly used to assay GGT. GGT-rel is not expressed in the mouse (9).Recently, mice deficient in GGT were developed and used in LT metabolism studies (10-12). These studies un-

The human gamma-glutamyltransferase gene family

“…Other biochemical Wndings reported included increased plasma glutathione levels, low GGT activity, and the presence of -glutamylcysteine and cysteine in urine. Moreover, Mayatepek et al (2004) was able to measure leukotriene levels in monocytes, plasma and urine of three of these patients (the ones reported by Hammond et al and Wright et al) and found that they were completely deWcient in LTD4 synthesis. Phenotypically, patients varied in their symptoms.…”

Unveiling the roles of the glutathione redox system <i>in vivo</i> by analyzing genetically modified mice

“…(79) Three of the patients show deficiencies in LTD 4 that are generated by a GGT-catalyzed reaction from the primary cysteinyl leukotriene LTC 4 . (80) …”

ReviewIs Serum Gamma Glutamyltransferase a Marker of Oxidative Stress?

“…[52] In addition to its gamma glutamyltranspeptidase activity it is also essential for leukotriene biosynthesis and bone metabolism as demonstrated in GGT deficient humans. [53] …”