The apparent ubiquity of epoxide hydratase in rat organs
Abstract
Using the recently developed sensitive assay with [3H] benzo [a] pyrene 4,5-oxide as substrate, epoxide hydratase was shown to be present in 26 rat (Sprague-Dawley) organs and tissues investigated. Only blood showed no detectable activity, which indicates that the low enzyme activity found in some organs is not due to the presence of blood components in the tissues. In earlier studies with a less sensitive assay, epoxide hydratase activity was detected only in rat liver and kidney but not in organs such as muscle, spleen, heart and brain. Epoxide hydratase was also measured in 6 organs of the mouse (NMRI). The distribution pattern was quantitatively quite different in the two species. The sp. act. in the rat were in the order liver > testis > kidney > lung > intestine ∼- skin. In the mouse, very surprisingly, testis had the highest specific epoxide hydratase activity. Moreover, the order of sp. act. in the mouse organs was remarkably different from that in the rat, namely testis > liver > lung > skin > kidney > intestine. The fact that the sp. act. in kidney was much lower than in lung or skin is most striking. Pretreatment of rats with Aroclor 1254 (a mixture of polychlorinated biphenyls) increased the epoxide hydratase activity in the liver to 175 per cent of the control level. However, the enzyme activity in the 13 extrahepatic tissues investigated was not significantly changed. In organs possessing sufficiently high enzyme levels, epoxide hydratase activity was also measured with styrene oxide as substrate. The ratio of the sp. act. of the two substrates was very similar in rat liver, kidney, lung and lestis. This supports the assumption that in these organs a single enzyme is responsible for the hydration of both substrates—as was earlier shown by several methods for the rat liver.
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Cited by (126)
Distribution of soluble and microsomal epoxide hydrolase in the mouse brain and its contribution to cerebral epoxyeicosatrienoic acid metabolism
2009, NeuroscienceEpoxide hydrolases comprise a family of enzymes important in detoxification and conversion of lipid signaling molecules, namely epoxyeicosatrienoic acids (EETs), to their supposedly less active form, dihydroxyeicosatrienoic acids (DHETs). EETs control cerebral blood flow, exert analgesic, anti-inflammatory and angiogenic effects and protect against ischemia. Although the role of soluble epoxide hydrolase (sEH) in EET metabolism is well established, knowledge on its detailed distribution in rodent brain is rather limited. Here, we analyzed the expression pattern of sEH and of another important member of the EH family, microsomal epoxide hydrolase (mEH), in mouse brain by immunohistochemistry. To investigate the functional relevance of these enzymes in brain, we explored their individual contribution to EET metabolism in acutely isolated brain cells from respective EH −/− mice and wild type littermates by mass spectrometry. We find sEH immunoreactivity almost exclusively in astrocytes throughout the brain, except in the central amygdala, where neurons are also positive for sEH. mEH immunoreactivity is abundant in brain vascular cells (endothelial and smooth muscle cells) and in choroid plexus epithelial cells. In addition, mEH immunoreactivity is present in specific neuronal populations of the hippocampus, striatum, amygdala, and cerebellum, as well as in a fraction of astrocytes. In freshly isolated cells from hippocampus, where both enzymes are expressed, sEH mediates the bulk of EET metabolism. Yet we observe a significant contribution of mEH, pointing to a novel role of this enzyme in the regulation of physiological processes. Furthermore, our findings indicate the presence of additional, hitherto unknown cerebral epoxide hydrolases. Taken together, cerebral EET metabolism is driven by several epoxide hydrolases, a fact important in view of the present targeting of sEH as a potential therapeutic target. Our findings suggest that these different enzymes have individual, possibly quite distinct roles in brain function and cerebral EET metabolism.
Genetic effects and biotoxicity monitoring of occupational styrene exposure
2009, Clinica Chimica ActaStyrene is a commercially important chemical widely used in the manufacture of synthetic rubber, resins, polyesters, and plastics. The highest levels of human exposure to styrene occurs in occupational settings, especially during the production of reinforced plastic products, which involve manual lay-up or spray-up operations. In these settings, absorption of styrene occurs mainly through inhalation and, to a minor extent, via skin contact.
A variety of biological markers (biomarkers) have been developed for genotoxic agents. Three types of biomarkers are identified: biomarkers of exposure, of effect and of susceptibility. Biomarkers of exposure measure the chemical itself or its metabolites in body fluids. Biomarkers of effect measure indicators of damage by the exposure. Biomarkers of effect are generally pre-clinical indicators of abnormalities and the most frequently used in genotoxicity assessment are sister chromatid exchanges (SCEs), chromosomal aberrations (CAs) and micronuclei (MN). More recently, the use of the single cell gel electrophoresis assay (SCGE), or comet assay has been proposed as a useful biomarker for early effects.
The third type is a biomarker of susceptibility, which indicates that the individual is vulnerable to the effect of a xenobiotic or to the effects of a group of such compounds. This type of biomarkers are related with individual genetic polymorphisms that could lead to different capacities to activate, detoxify or repair DNA lesions arising from exposure to chemical carcinogens.
Styrene metabolism involves cytochrome P450 enzymes (CYP)-mediated that by oxidation convert styrene to its reactive metabolite styrene-7,8-oxide (SO) which is capable of binding covalently with macromolecules and is considered to be directly responsible for the genotoxic effects of styrene. SO, is mainly hydrolysed to styrene glycol by the microsomal epoxide hydrolase (mEH), and subsequently oxidized by alcohol and aldehyde dehydrogenases to the main urinary metabolites, mandelic acid (MA) and phenylglyoxylic acid (PGA) (major pathway). MA and PGA represent more than 95% of the urinary metabolites of styrene. Further transformation of MA and PGA, via transamination of α-keto- and α-hydroxyacids into the corresponding amino acid, leads to phenylglycine.
Evidence of the carcinogenicity of styrene has been reported in studies with mice. Epidemiologic evidence does not suggest a causal association between styrene and any forms of cancer in humans. However, the possibility of a small elevation of risk for one or more cancers cannot be ruled out.
The International Agency for Research on Cancer (IARC) has designated styrene as possibly carcinogenic to humans (group 2B). Concern about the potential carcinogenicity of styrene stems largely from the ability of its metabolite, SO to bind covalently to DNA and to its activity in a variety of genotoxicity test systems. SO has been classified by IARC in group 2A, probably carcinogenic to humans.
Styrene exposure has been reported to cause an increase in DNA and haemoglobin adducts and in the frequency of CAs; there is less evidence for an association between styrene exposure and the frequency of SCEs.
This article thoroughly reviews all available published data on the genetic effects of styrene and the biotoxicity markers of exposure monitoring.
Epoxide hydrolases: Their roles and interactions with lipid metabolism
2005, Progress in Lipid ResearchThe epoxide hydrolases (EHs) are enzymes present in all living organisms, which transform epoxide containing lipids by the addition of water. In plants and animals, many of these lipid substrates have potent biologically activities, such as host defenses, control of development, regulation of inflammation and blood pressure. Thus the EHs have important and diverse biological roles with profound effects on the physiological state of the host organisms. Currently, seven distinct epoxide hydrolase sub-types are recognized in higher organisms. These include the plant soluble EHs, the mammalian soluble epoxide hydrolase, the hepoxilin hydrolase, leukotriene A4 hydrolase, the microsomal epoxide hydrolase, and the insect juvenile hormone epoxide hydrolase. While our understanding of these enzymes has progressed at different rates, here we discuss the current state of knowledge for each of these enzymes, along with a distillation of our current understanding of their endogenous roles. By reviewing the entire enzyme class together, both commonalities and discrepancies in our understanding are highlighted and important directions for future research pertaining to these enzymes are indicated.
Five Novel Single Nucleotide Polymorphisms in the EPHX1 Gene Encoding Microsomal Epoxide Hydrolase
2003, Drug Metabolism and PharmacokineticsFive novel single nucleotide polymorphisms (SNPs) were found in the EPHX1 gene from 96 Japanese epileptic patients. The detected SNPs were as follows:
- 1)
SNP, MPJ6_EX1009; GENE NAME, EPHX1 ACCESSION NUMBER, NT_004525.12; LENGTH, 25 bases; 5'-CCTCACTTCAGTG/ACTGGGCTTTGCC-3'.
- 2)
SNP, MPJ6_EX1013; GENE NAME, EPHX1; ACCESSION NUMBER, NT_004525.12; LENGTH, 25 bases; 5'-TCCGCAGCCAGGG/CAGGACGACAGCA-3'.
- 3)
SNP, MPJ6_EX1026; GENE NAME, EPHX1; ACCESSION NUMBER, NT_004525.12; LENGTH, 25 bases; 5'-GTTCTCCCTGGAC/TGACCTGCTGACC-3'.
- 4)
SNP, MPJ6_EX1028; GENE NAME, EPHX1; ACCESSION NUMBER, NT_004525.12; LENGTH, 25 bases; 5'-AGGCAGGGGGACG/AGCCAGTCTTGGG-3'.
- 5)
SNP, MPJ6_EX1030; GENE NAME, EPHX1; ACCESSION NUMBER, NT_004525.12; LENGTH, 25 bases; 5'-TGAAAAGTGGGTG/AAGGTTCAAGTAC-3'.
The frequencies were 0.016 for MPJ6_EX1028 (IVS8 + 54G>A) and 0.005 for the other SNPs. The SNP MPJ6_EX1013 (130G > C) results in an amino acid alteration (E44Q). The other three SNPs in the coding region, MPJ6_EX1009 (30G>A), MPJ6_EX1026 (1056C>T), and MPJ6_EX1030 (1239G>A) result in synonymous changes (V10V, D352D, and V413V, respectively).
- 1)
Relative importance of maternal and embryonic microsomal epoxide hydrolase in 7,12-dimethylbenz[a]anthracene-induced developmental toxicity
2002, Biochemical PharmacologyCitation Excerpt :mEH plays a central role in the metabolic transformation of many aliphatic epoxides and arene oxides derived from drugs and environmental chemicals [6,7]. mEH is expressed not only in liver, but also in several extrahepatic tissues including the placenta, uterus, and embryo [8,9]. mEH protein and activity were detected during the period of organogenesis in humans and rats [10,11], suggesting the possibility that mEH is involved in the metabolism of teratogens at the target organ.
Microsomal epoxide hydrolase (mEH) catalyzes the hydrolysis of epoxide intermediates derived from drugs and environmental chemicals. The response of in vivo (embryo) and in vitro (embryo fibroblast) tests were analyzed using mEH-null and wild-type mice to determine the relative role of maternal and embryonic mEH in the developmental toxicity induced by 7,12-dimethylbenz[a]anthracene (DMBA). Embryos derived from DMBA-treated [50 mg/kg, daily from gestational day (GD) 11 to GD 15] dams were analyzed. Although weight (P=0.0009) and crown-rump length (P=0.0003) of wild-type fetuses on GD 18 were significantly lower than those of mEH-null fetuses, respectively, no significant difference was found between mEH-null and heterozygous fetuses of mEH-null dams. Cell viability was decreased to 50% in wild-type mouse embryo fibroblasts (MEFs) treated with 3 μM DMBA, but no significant decrease was found in mEH-null MEFs. DMBA-3,4-diol produced a significant decrease in cell viability and suppressed the proliferation of wild-type MEFs at a 10-fold lower concentration than did DMBA. Although mEH protein was expressed in liver microsomes from wild-type embryos (GD 15), DMBA-3,4-diol was not detected among the DMBA metabolites. However, it was detected in the serum of wild-type pregnant mice treated with DMBA, but not in that of mEH-null mice. These results suggest that maternal mEH plays a major role in DMBA-induced developmental toxicity, and embryonic mEH is less involved in the toxicity.
Mechanism of 7,12-dimethylbenz[a]anthracene-induced immunotoxicity: Role of metabolic activation at the target organ
2001, Japanese Journal of PharmacologyThe polycyclic aromatic hydrocarbon, 7,12-dimethylbenz[a]anthracene (DMBA), is an immunosuppressor as well as a potent organ-specific carcinogen. To understand the organ-specific mechanism of DMBA-induced lymphoid toxicity, aryl hydrocarbon-nonresponsive mice and microsomal epoxide hydrolase (mEH)-null mice were analyzed. DMBA caused a dose-dependent decrease in spleen weights, but not the thymus weights in aryl hydrocarbon-nonresponsive mice. On the other hand, both spleen and thymus weights were decreased to less than a half in wild-type mice exposed to 30 mg /kg of DMBA. In contrast, no decrease was detected in spleen weights of mEH-null mice exposed to up to 100 mg /kg of DMBA, while thymus weights were markedly lower. Responses to the B-cell mitogen lipopolysaccharide and to T-cell mitogen phytohemagglutinin were nearly completely abolished in splenocytes isolated from wild-type mice treated with 100 mg / kg of DMBA. These responses were decreased, but maintained in splenocytes isolated from mEH-null mice treated with DMBA. Two DMBA metabolites dependent on mEH including DMBA-3,4-diol were detected in an HPLC chromatogram of spleen microsomes isolated from wild-type mice, but not those from mEH-null mice. These results suggest the involvement of mEH in splenic activation of DMBA for immunotoxicity and the difference for the DMBA-induced lymphoid toxicity between spleen and thymus.
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This study is part of the Ph.D. Thesis of H. Schmassmann.