Biomedical subjects

Karl Walter Bock

Publications and source records attributed to Karl Walter Bock.

13 recordsLinked to original sources

PubMed2006

Activation of coupled Ah receptor and Nrf2 gene batteries by dietary phytochemicals in relation to chemoprevention.

The Ah receptor (AhR) is a ligand-activated transcription factor and member of the bHLH/PAS (basic helix-loop-helix/Per-Arnt-Sim) family of chemosensors and developmental regulators. It represents a multifunctional molecular switch involved in regulation of endo- and xenobiotic metabolism, in vascular development and in dioxin-mediated toxicities. Recently, the oxidative stress-protecting Nrf2 has been shown to be a downstream target of the AhR [Miao W, Hu L, Scrivens PJ, Batist G. Transcriptional regulation of NF-E2 p45-regulated factor (NRF2) expression by the aryl hydrocarbon receptor-xenobiotic response element signaling pathway. J Biol Chem 2005;280:20340-8]. This finding offers the possibility that distinct but partially overlapping AhR and Nrf2 gene batteries of Phase II xenobiotic-metabolizing enzymes can be synergistically activated by a number of phytochemicals, acting as selective or mixed activators of target genes. In addition, it is conceivable that AhR-mediated oxidative/electrophile stress may be attenuated by coupled Nrf2 activation. The commentary discusses potentials and limitations of (i) selective Nrf2 and of (ii) synergistic AhR plus Nrf2 activation by phytochemicals in efforts towards chemoprevention of cancer and degenerative diseases, and describes clinical trials providing the expectation that chemopreventive measures may favorably modulate unavoidable endo- and exogenous toxin exposures in high risk populations.

Animals↗

PubMed2006

Ah receptor: dioxin-mediated toxic responses as hints to deregulated physiologic functions.

The aryl hydrocarbon receptor (AhR) is a ligand-activated transcription factor and member of the bHLH/PAS (basic Helix-Loop-Helix/Per-Arnt-Sim) family of chemosensors and developmental regulators. It represents a multifunctional molecular switch regulating endo- and xenobiotic metabolism as well as cell proliferation and differentiation. Physiologic functions of the AhR are beginning to be understood, including functions in vascular development, and in detoxification of endo- and xenobiotics. The AhR is also recognized as the culprit for most toxic responses observed after exposure to dioxins and related compounds such as 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). The non-metabolizable AhR agonist TCDD has to be distinguished from the myriad of metabolizable agonists present as dietary contaminants and plant constituents as well as endogenous toxins. The hypothesis is emerging that the diverse tissue-specific, TCDD-mediated toxicities are due to sustained and inappropriate AhR activation leading to deregulated physiologic functions. In support of this hypothesis recent observations in the context of some TCDD-mediated toxic responses are discussed, such as chloracne, cleft palate, thymus involution and in particular carcinogenesis. Major open questions are addressed, such as ligand-independent AhR activation by phosphorylation and the large differences in species-dependent susceptibility to toxic responses. Though important issues remain unresolved, the commentary is intended to stimulate efforts to understand dioxin-mediated toxic responses with emphasis on carcinogenesis in comparison with AhR-mediated physiologic functions.

Animals↗

PubMed2005

Ah receptor- and TCDD-mediated liver tumor promotion: clonal selection and expansion of cells evading growth arrest and apoptosis.

The Ah receptor (AhR) has been characterized as a ligand-activated transcription factor which belongs to the bHLH/PAS (basic helix-loop-helix/Per-Arnt-Sim) family of chemosensors. Transgenic mouse models revealed adaptive and developmental functions of the AhR in the absence of exogenous ligands. Use of persistent agonists such as dioxins including 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) and related compounds demonstrated that the AhR mediates a plethora of species- and tissue-dependent toxicities, including chloracne, wasting, teratogenicity, immunotoxicity, liver tumor promotion and carcinogenicity. However, molecular mechanisms underlying most aspects of these toxic responses as well as biological functions of the AhR are currently unknown. Previous studies of liver tumor promotion in the two-stage hepatocarcinogenesis model indicated that TCDD mediates clonal expansion of 'initiated' preneoplastic hepatocytes, identified as enzyme-altered foci (EAF) by inhibiting apoptosis and bypassing AhR-mediated growth arrest. In contrast, the Ah receptor has been shown in cell models to stimulate growth arrest and apoptosis. Possible underlying mechanisms of these AhR responses are discussed, including enhanced metabolism of retinoic acid which attenuates TGFbeta-mediated apoptosis and interaction of the Ah receptor with the hypophosphorylated retinoblastoma tumor suppressor protein. The discrepancy between in vivo findings in EAF and AhR functions may be solved by hypothesizing that sustained activation of the Ah receptor generates a strong selective pressure in liver treated with genotoxic carcinogens leading to selection and expansion of clones evading growth arrest and apoptosis. Models are discussed which may facilitate verification of this hypothesis.

Animals↗

PubMed2005

Serotonin glucuronidation by Ah receptor- and oxidative stress-inducible human UDP-glucuronosyltransferase (UGT) 1A6 in Caco-2 cells.

Caco-2 cells are a widely used model in drug development to study intestinal drug transport and metabolism. Recently, serotonin (5-hydroxytryptamine, 5-HT) has been characterized as a highly selective substrate of human UDP-glucuronosyltransferase UGT1A6 [Krishnaswamy S, Duan SX, von Moltke LL, Greenblatt DJ, Court MH. Validation of serotonin (5-hydroxytryptamine) as an in vitro substrate probe for human UDP-glucuronosyltransferase (UGT) 1A6. Drug Metab Disp 2003; 31:133-9], an isoform which conjugates planar phenols and is inducible by Ah receptor agonists and by oxidative/electrophile stress. To gain more insight into intestinal 5-HT disposition, uptake and metabolism of this neurotransmitter was studied in Caco-2 cell monolayers. It was found that 5-HT was taken up from the basolateral and to a lesser extent from the apical surface. It was mainly excreted basolaterally as 5-HT glucuronide. 5-HT UGT activity and UGT1A6 mRNA were induced by Ah receptor agonists and by oxidative stress generated by tert-butylhydroquinone and by isomeric thymoquinone, a potential antitumor agent and constituent of Nigella sativa seeds, commonly used as a condiment in the Middle East. While UGT1A6 induction was clearly detectable in NAD(P)H:quinone oxidoreductase 1 (NQO1)-deficient Caco-2 cells, it was not induced in NQO1-efficient HT-29 colon adenocarcinoma cells. The results suggest that--in addition to its detoxification function--intestinal UGT1A6 contributes to intestinal homeostasis of 5-HT from dietary sources and from release by enterochromaffin cells.

Antioxidants↗

PubMed2005

UDP-glucuronosyltransferase 1A6: structural, functional, and regulatory aspects.

Glucuronidation, catalyzed by two families of UDP-glucuronosyltransferases (UGTs), represents a major phase II reaction of endo- and xenobiotic biotransformation. UGT1A6 is the founding member of the rat and human UGT1 family. It is expressed in liver and extrahepatic tissues, such as intestine, kidney, testis, and brain, and conjugates planar phenols and arylamines. Serotonin has been identified as a selective endogenous substrate of the human enzyme. UGT1A6 is also involved in conjugation of the drug paracetamol (acetaminophen) and of phenolic metabolites of benzo[a]pyrene (together with rat UGT1A7 and human UGT1A9). High interindividual variability of human liver protein levels is due to a number of influences, including genetic, tissue-specific, and environmental factors. Evidence shows that homo- and heterozygotic expression of UGT1A6 alleles markedly affects enzyme activity. HNF1 may be responsible for tissue-specific UGT1A6 expression. Multiple environmental factors controlling UGT1A6 expression have been identified, including the pregnane X receptor, the constitutive androstane receptor, the aryl hydrocarbon receptor, and Nrf2, a bZIP transcription factor mediating stress responses. However, marked differences have been noted in the expression of rat and human UGT1A6. Regulatory factors have been studied in detail in the human Caco-2 colon adenocarcinoma cell model.

Animals↗

PubMed2005

Nomenclature update for the mammalian UDP glycosyltransferase (UGT) gene superfamily.

Several novel UDP glycosyltransferase (UGT) genes, mainly UDP glucuronosyltransferases, have been identified in the human, mouse and rat genomes and in other mammalian species. This review provides an update of the UGT nomenclature to include these new genes and prevent the confusion that arises when the same gene is given different names. The new genes are named following previously established recommendations, taking into consideration evolutionary relatedness and the names already in general usage in the literature. The mammalian UGT gene superfamily currently has 117 members that can be divided into four families, UGT1, UGT2, UGT3 and UGT8. The 5-exon genes of the UGT1 family each contain a unique first exon, plus four exons that are shared between the genes; the exons 1 appear to have evolved by a process of duplication, leading to the synthesis of proteins with identical carboxyl-terminal and variable amino-terminal domains. Exon-sharing is also seen with the 6-exon UGT2A1 and UGT2A2 genes. However, UGT2A3 and those of the UGT2B (six exons), UGT3 (seven exons) and UGT8 gene families (five or six exons) do not share exons and most likely were derived by a process of duplication of all exons in the gene. Most UGT1 and UGT8 enzymes have been characterized in detail; however, the catalytic functions of the UGT3A enzymes and several UGT2 enzymes remain to be characterized.

Animals↗

PubMed2004

Coordinate regulation of drug metabolism by xenobiotic nuclear receptors: UGTs acting together with CYPs and glucuronide transporters.

Xenobiotic nuclear receptors (PXR, CAR, and the Ah receptor) coordinately induce genes involved in all phases of xenobiotic metabolism including oxidative metabolism, conjugation, and transport. The comment--dedicated to honor the memory of Herbert Remmer, mentor of the author K. W. B.--discusses mechanistic, functional, and evolutionary aspects of xenobiotic nuclear receptors which induce UGTs together with CYPs and glucuronide transporters in human and rodent liver and intestine. Recent findings on regulation of CYPs, UGTs, and transporters suggest that while nuclear receptor signaling induces different CYPs, regulation may converge on single UGTs and transporters. Functional consequences of co-regulation are discussed using examples from the metabolism of xeno- and endobiotics (drugs, bilirubin, bile salts, steroid hormones, and carcinogens). Animal-plant interactions may have been a major driving force in the evolutionary divergence of CYPs and UGTs in mammals and insects as well as in their regulation by nuclear receptors. In addition, regulation by nuclear receptors was probably shaped by the need for homeostatic control of endobiotic signals in the evolution of multicellular organisms.

Animals↗

PubMed2003

Vertebrate UDP-glucuronosyltransferases: functional and evolutionary aspects.

UDP-glucuronosyltransferases (UGTs) represent major phase II drug metabolizing enzymes. They are part of a rapidly growing, sequence similarly based superfamily of UDP-glycosyltransferases, including a number of enzymes, which presumably are functionally unrelated to UGTs. The present commentary discusses evolutionary aspects of the large glycosyltransferase superfamily emphasizing functionally related members which share roles in detoxication and elimination of endo- and xenobiotics. The discussion starts with the two human UGT families and polymorphism frequencies in different populations. These families probably evolved in vertebrates as a result of the struggle against toxic phytoalexins at the hepatogastrointestinal barrier. Co-regulation of some UGTs with other drug metabolizing enzymes may also have evolved in the course of 'animal-plant warfare'. Related UDP-glucosyltransferases evolved in insects. Even in plants and bacteria UDP-glucosyltransferases have been characterized which may be functionally related.

Animals↗

PubMed2003

Contribution of the Ah receptor to the phenolic antioxidant-mediated expression of human and rat UDP-glucuronosyltransferase UGT1A6 in Caco-2 and rat hepatoma 5L cells.

UDP-glucuronosyltransferases (UGTs) represent major phase II enzymes of drug metabolism which are regulated in a tissue-specific manner by endogenous and environmental factors. Among the latter, aryl hydrocarbon receptor (AhR) agonists such as 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) and phenolic antioxidants such as tert-butylhydroquinone (tBHQ) are known to induce the expression of human UGT1A6 in Caco-2 cells. While binding of the TCDD-activated AhR to one xenobiotic response element (XRE) in the 5'-flanking regulatory region of UGT1A6 was characterised previously, the mechanism responsible for tBHQ induction is unknown. Therefore, it was investigated whether antioxidant response elements (AREs) are involved in tBHQ induction of UGT1A6. Transfectants of 3 kb of its regulatory region and its deletion mutants were treated with tBHQ. These studies suggested a region with approximately 2-fold induction, including an ARE-like motif, 15 bp downstream of the previously characterised XRE. Transfectants of the point-mutated ARE-like motif showed marginally reduced response to tBHQ, but surprisingly, loss of response to TCDD, suggesting interference of flanking proteins with the AhR/Arnt complex. Coordinate responses of UGT activity after treatment with TCDD or tBHQ were also observed in rat hepatoma 5L cells, mutants without the AhR and with recomplemented AhR. The results suggest a contribution of the AhR pathway and of proteins binding to the XRE flanking region to the induction of human UGT1A6 by both AhR agonists and phenolic antioxidants.

5' Flanking Region↗

PubMed2003

Frequent co-occurrence of the TATA box mutation associated with Gilbert's syndrome (UGT1A128) with other polymorphisms of the UDP-glucuronosyltransferase-1 locus (UGT1A62 and UGT1A7*3) in Caucasians and Egyptians.

Polymorphisms of drug metabolizing enzymes are frequently associated with diseases and side effects of drugs. Recently, a TATA box mutation of UGT1A1 (UGT1A1*28), a common genotype leading to Gilbert's syndrome, and several missense mutations of other UDP-glucuronosyltransferase 1 (UGT1) family members have been described. Furthermore, co-occurrence of UGT1A1*28 and UGT1A6*2 has been observed. In order to elucidate the basis for co-occurrence of UGT1 mutations, fluorescence resonance energy transfer techniques were developed for rapid determination of polymorphisms of three UGT isoforms (UGT1A1*28, 1A6*2, and 1A7*2/*3). Hundred healthy Caucasians and 50 Egyptians were genotyped. All genotypes followed the Hardy-Weinberg equilibrium. Only three major haplotypes were found, including a haplotype consisting of allelic variants of all three isoforms (29% in Caucasians and 22% in Egyptians), all leading to reduced UGT activity. Frequent haplotypes containing several UGT1 allelic variants should be taken into account in studies on the association between diseases, abnormal drug reactions, and UGT1 family polymorphisms.

Egypt↗

PubMed2002

Conditional expression of a constitutively active aryl hydrocarbon receptor in MCF-7 human breast cancer cells.

In addition to inducing transcription of a battery of target genes encoding drug-metabolizing enzymes, the environmental pollutant 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) is known to induce antiestrogenic responses. However, the mechanisms underlying such complex biologic responses affecting growth and differentiation remain unclear. In the present study we have investigated biological effects of a constitutively active mutant of the aryl hydrocarbon (Ah) receptor (CA-AhR), in particular whether it modulates estrogen receptor function in human MCF-7 breast cancer cells. To this end, the CA-AhR protein was conditionally expressed using the tet repressor. Expression of CA-AhR resulted in constitutive formation of a DNA-binding AhR-aryl hydrocarbon receptor nuclear translocator heterodimeric complex and enhanced expression of the Ah receptor target gene CYP1A1 in the absence of TCDD. Moreover, expression of CA-AhR inhibited estrogen-dependent cathepsin D expression and growth of these cells. Thus, the present model system conditionally expressing the CA-AhR protein provides a novel tool for the investigation of AhR-mediated signaling pathways.

Breast Neoplasms↗

PubMed2002

Tissue-specific regulation of canine intestinal and hepatic phenol and morphine UDP-glucuronosyltransferases by beta-naphthoflavone in comparison with humans.

UDP-glucuronosyltransferases (UGTs) are regulated in a species- and tissue-dependent manner by endogenous and environmental factors. The present study was undertaken to further our knowledge about regulation of UGTs in dogs, a species widely used in preclinical safety evaluation. beta-Naphthoflavone (BNF) was selected as a known aryl hydrocarbon receptor agonist and antioxidant-type inducer. The latter group of inducers is intensively investigated as dietary chemoprotectants against colon cancer. Dog UGTs were investigated in comparison with related human UGTs by examples, (i) expression of dog UGT1A6, the first sequenced dog phenol UGT, and (ii) morphine UGT activities, responsible for intestinal and hepatic first-pass metabolism of morphine. The following results were obtained: (i) dog UGT1A6 was found to be constitutively expressed in liver and marginally increased by BNF treatment. Expression was low in small intestine but ca. 6-fold higher in colon than for example in jejunum. Conjugation of 4-methylumbelliferone, one of the substrates of dog UGT1A6, was also enhanced 7-fold in colonic compared to jejunal microsomes. (ii) Compared to the corresponding human tissues, canine 3-O- and 6-O-morphine UGT activities were found to be >10-fold higher in dog liver and ca. 10-fold lower in small intestinal microsomes. Small intestinal morphine and 4-hydroxybiphenyl UGT activities appeared to be moderately (2- to 3-fold) induced by oral treatment with BNF. (iii) In contrast to dogs, morphine UGT activities were found to be similar in homogenates from human enterocytes and liver. The results suggest marked differences in tissue-specific regulation of canine vs. human hepatic and intestinal phenol or morphine UGTs.

Animals↗

PubMed2002

Influence of t-butylhydroquinone and beta-naphthoflavone on formation and transport of 4-methylumbelliferone glucuronide in Caco-2/TC-7 cell monolayers.

Human Caco-2 cells have been established as a model system for intestinal biotransformation and permeability. When grown on Transwell polycarbonate filters they develop morphologic and biochemical characteristics of enterocytes with well separated apical and basolateral surfaces. In addition, Caco-2/TC-7 cells have proven to be useful to study regulation of human UDP-glucuronosyltransferases (UGTs) by Ah receptor agonists and antioxidant-type inducers such as beta-naphthoflavone (BNF) and t-butylhydroquinone (TBHQ). In the present investigation, formation and transport of 4-methylumbelliferone glucuronide was studied in intact Caco-2 cell monolayers. The following results were obtained: when loaded with 50-200 microM MUF either apically or basolaterally, MUF-GA was the major metabolite which was mostly released (80%) at the basolateral surface, probably via the multidrug resistance protein isoform MRP3; MUF sulfate formation was low (5 +/-2%). Pretreatment of cells with 80 microM TBHQ or 50 microM BNF for 72 hr before addition of 100 microM MUF enhanced basolateral secretion of MUF-GA 1.4- and 1.7-fold, respectively. However, at >200 microM MUF, MUF-GA secretion and induction was smaller, probably due to inhibition of intracellular UGT activity. MRP3 protein was localized to the basolateral surface of Caco-2 cells but was not induced by TBHQ or BNF. The results suggest that MUF-GA is mostly secreted basolaterally in Caco-2 cell monolayers. Treatment with TBHQ or BNF significantly enhanced MUF-GA formation in the intact cell.

Biological Transport↗