TY - JOUR A1 - Klotz, Karl-Norbert A1 - Jesaitis, A. J. T1 - Neutrophil chemoattractant receptors and the membrane skeleton N2 - Signal transduction via receptors for N-formylmethionyl peptide chemoattractants (FPR) on human neutrophils is a highly regulated process which involves participation of cytoskeletal elements. Evidence exists suggesting that the cytoskeleton and/or the membrane skeleton controls the distributJon of FPR in the plane of the plasma membrane, thus controlling the accessibility of FPR to different proteins in functionally distinct domains. In desensitized cells, FPR are restricted todomains which are depleted of G proteins but enriched in cytoskeletal proteins such as actin and fodrin. Thus, the G protein signal transduction partners of FPR become inaccessible to the agonist-occupied receptor, preventing cell activation. The mechanism of interaction of FPR with the membrane skeleton is poorly understood but evidence is accumulating that suggests a direct binding of FPR (and other receptors) to cytoskeletal proteins such as actin. KW - Toxikologie Y1 - 1994 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-60471 ER - TY - JOUR A1 - Klotz, Karl-Norbert A1 - Jesaitis, A. J. T1 - Physical coupling of N-formyl peptide chemoattractant receptors to G protein is not affected by desensitization N2 - Desensitization of N-formyl peptide chemoattractant receptors (FPR) in human neutrophils results in association of these receptors to the membrane skeleton. This is thought to be the critical event in the lateral segregation of receptors and guanyl nucleotide-binding proteins (G proteins) within the plane of the plasma membrane resulting in an interruption of the signaling cascade. In this study we probed the interaction of FPR with G protein in human neutrophils that were desensitized to various degrees. Human neutrophils were desensitized using the photoreactive agonist N-formyl-met-leu-phelys- N\(^\epsilon\)-[\(^{125}\)I]2(p-azidosalicylamido )ethyl-1 ,3 '-dithiopropionate (/MLFK-[\(^{125}\)I]ASD). The interaction if FPR with G protein was studied via a reconstitution assay and subsequent analysis of FPR-G protein complexes in sucrose density gradients. FPR-G protein complexes were reconstituted with solubilized FPR from partially and fully desensitized neutrophils with increasing concentrations of Gi purified from bovine brain. The respective EC\(_{50}\) values for reconstitution were similar to that determined for FPR from unstimulated neutrophils (Bommakanti RK et al., J Bio[ Chem 267: 757~7581, 1992). We conclude, therefore, that the affinity of the interaction of FPR with G protein is not affected by desensitization, consistent with the model of lateral segregation of FPR and G protein as a mechanism of desensitization. KW - Toxikologie KW - chemotactic receptors KW - G proteins KW - N-formyl peptides KW - signal transduction KW - receptor-G protein coupling Y1 - 1994 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-60483 ER - TY - JOUR A1 - Klotz, Karl-Norbert A1 - Jesaitis, A. J. T1 - The interaction of N-formyl peptide chemoattractant receptors with the membrane skeleton is energy-dependent N2 - Desensitization of N-fonnyl peptide chemoattractant receptors (FPR) in human neutrophils is thought to be achieved by lateral segregation of receptors and G proteins within the plane of the plasma membrane resulting in an interruption of the signalling cascade. Direct coupling of FPR to membrane skeletal actin appears to be the basis of this process~ however, the molecular mechanism is unknown. In this study we investigated the effect of energy depletion on formation of FPR-membrane skeleton complexes. In addition the effect of the protein kinase C inhibitor stauroporine and the phosphatase inhibitor okadaic acid on coupling of FPR to the membrane skeletonwas studied. Human neutrophils were desensitized using the photoreactive agonist N-formy1-met-leu-phe-1ys-N'[\(^{125}\)I]2(p-azidosalicylamido)ethyl-1,3'-dithiopropionate (fMLFK-[\(^{125}\)I]ASD) after ATP depletion with NaF or after incubation with the respective inhibitors. The interaction of FPR with the membrane skeleton was studied by Sedimentation of the membrane skeleton-associated receptors in sucrose density gradients. Energy depletion of the cells markedly inhibited the formation of FPR-membrane skeleton complexes. This does not appear tobe related to inhibition of protein phosphorylation due to ATP depletion because inhibition of protein kinases and phosphatases bad no significant effect on coupling of FPR to the membrane skeleton. We conclude, therefore, that coupling of FPR to the membrane skeleton is an energy,dependent process which does not appear to require modification of the receptor protein by phosphorylation. KW - Toxikologie KW - Chemotactic receptors KW - G proteins KW - N-formyl peptides KW - signal transduction KW - desensitization KW - membrane skeleton KW - receptor-G protein coupling. Y1 - 1994 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-60499 ER - TY - JOUR A1 - Klotz, Karl-Norbert A1 - Keil, R. A1 - Zimmer, F. J. A1 - Schwabe, U. T1 - Guanine nucleotide effects on 8-cyclopentyl-1,3-[\(^3\)H]dipropylxanthine binding to membrane-bound and solubilized A\(_1\) adenosine receptors of rat brain N2 - The effects of guanine nucleotides on binding of 8-cyclopentyl-1,3-[\(^3\)H]dipropylxanthine [\(^3\)H]DPCPX), a highly selective A\(_1\) adenosine receptor antagonist, have been investigated in rat brain membranes and solubilized A\(_1\) receptors. GTP, which induces uncoupling of receptors from guanine nucleotide binding proteins, increased binding of [\(^3\)H]DPCPX in a concentration-dependent manner. The rank order of potency for different guanine nucleotides for increasing [\(^3\)H]DPCPX bindingwas the same as for guanine nuc1eotide-induced inhibition of agonist binding. Therefore, a role for a guanine nucleotide binding protein, e.g., G\(_i\), in the regulation of antagonist binding is suggested. This was confirmed by inactivation ofGi by N-ethylmaleimide (NEM) treatment of membranes, which resulted in an increase in [\(^3\)H]DPCPX binding similar to that seen with addition of GTP. Kinetic and equilibrium binding studies showed that the GTP- or NEM-induced increase in antagonist binding was not caused by an affinity change of A\(-1\) receptors for [\(^3\)H]DPCPX but by an increased Bmu value. Guanine nucleotides had similar effects on membrane-bound and solubilized receptors, with the effects in the solubilized system being more pronounced. In the absence of GTP, when rnost receptors are in a high-affinity state for agonists, only a few receptors are labeled by [\(^3\)H]DPCPX. It is suggested that [\(^3\)H]DPCPX binding is inhibited when receptors are coupled to G\(_i\). Therefore, uncoupling of A\(_1\) receptors from G\(_i\) by guanine nucleotides or by inactivation of G\(_i\) with NEM results in an increased antagonist binding. Key Words: Adenosine receptors-8 -Cyclopentyl-1,3-eH]dipropylxanthine-Antagenist binding-Guanine nucleotide effects. Klotz K.-N. et al. Guanine nucleotide etfects on 8-cyclopentyl-1 ,3-eH]dipropylxanthine binding to membrane-bound and solubilized A1 adenosine receptors of rat brain. J. Neurochem. 54, 1988-1994 (1990). KW - Toxikologie Y1 - 1990 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-60369 ER - TY - JOUR A1 - Klotz, Karl-Norbert A1 - Krotec, K. L. A1 - Gripentrog, J. A1 - Jesaitis, A. J. T1 - Regulatory interaction of N-formyl peptide chemoattractant receptors with the membrane skeleton in human neutrophils N2 - The cytoskeleton and/or membrane skeleton has been implicated in the regulation of N-formyl peptide receptors. The coupling of these chemotactic receptors to the membrane skeleton was investigated in plasma membranes from unstimulated and desensitized human neutrophils using the photoreactive agonist N-formyl-met-leu-phelys-N\(^6\)-[\(^{125}\)I]2(p-azidosalicylamido)ethyl-1,3'-dithiopropionate (fMLFK-[\(^{125}\)I]ASD). When membranes of unstimulated cells were solubilized in Triton-X 100, a detergent that does not disrupt actin filaments, only 50% of the photoaffinity-labeled receptors were solubilized sedimenting in sucrose density gradients at a rate consistent with previous reports. The remainder were found in the pellet fraction along with the membrane skeletal actin. Solubilization of the membranes in the presence of p-chloromercuriphenylsulfonic acid, elevated concentrations of KCI, or deoxyribonuclease I released receptors in parallel with actin. When membranes from neutrophils, desensitized by incubation with fMLFK-e 251]ASD at 15°C, were solubilized, nearly all receptors were recovered in the pellet fraction. lncubation of cells with the Iigand at 4°C inhibited desensitization partially and prevented the conversion of a significant fraction of receptors to the form associated with the membrane skeletal pellet. ln these separations the photoaffinity-labeled receptors not sedimenting to the pellet cosedimented with actin. Approximately 25% of these receptors could be immunosedimented with antiactin antibodies suggesting that N-formyl peptide receptors may interact directly with actin. These results are consistent with a regulatory role for the interaction of chemotactic N-formyl peptide receptors with actin of the membrane skeleton. KW - Toxikologie Y1 - 1994 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-60466 ER - TY - JOUR A1 - Klotz, Karl-Norbert A1 - Lohse, M. J. T1 - The glycoprotein nature of A\(_1\) adenosine receptors N2 - A\(_1\) adenosine receptors from different tissues and species we~e photoaffinity labelled and then the carbohydrate content was examined by both enzymatic and chemical treatment. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of the labelled membrane receptors shows that neuraminidase treatment alters the electrophoretic mobility of the receptor band indica ting the presence of terminal neurandnie acids. Neuraminidase digestion does not influence the binding characteristics of the receptor. The totally deglycosylated receptor protein obtained by chemical treatment has an apparent molecular weight Of 32,000. KW - Toxikologie Y1 - 1986 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-60231 ER - TY - JOUR A1 - Klotz, Karl-Norbert A1 - Lohse, M. J. A1 - Schwabe, U. T1 - Chemical modification of A\(_1\) adenosine receptors in rat brain membranes - evidence for histidine in different domains of the ligand binding site N2 - Chemical modification of amino acid residues was used to probe the ligand recognition site of A\(_1\) adenosine receptors from rat brain membranes. The effect of treatment with group·specific reagents on agonist and antagonist radioligand binding was investigated. The histidine-specific reagent diethylpyrocarbonate (DEP) induced a loss of binding of the agonist R-N\(^6\)-[\(^3\)H]phenylisopropyladenosine ([\(^3\)H]PIA), which could be prevented in part by agonists, but not by antagonists. DEP treatment induced also a loss of binding of the antagonist [\(^3\)H]8- cyclopentyl-1 ,3-dipropylxanthine ([\(^3\)H]DPCPX). Antagonists protected A\(_1\) receptors from this inactivation while agonists did not. This result provided evidence for the existence of at least 2 different histidine residues involved in ligand binding. Consistent with a modification of the binding site, DEP did not alter the affinity of [\(^3\)H]DPCPX, but reduced receptor number. From the selective protection of [\(^3\)H] PIA and [\(^3\)H]DPCPX binding from inactivation, it is concluded that agonists and antagonists oocupy different domains at the binding site. Sulfhydryl modifying reagents did not influence antagonist binding, but inhibited agonist binding. This effect is explained by modification of tbe inhibitory guanine nucleotide binding protein. Pyridoxal 5-phosphate inactivated both [\(^3\)H]PIA and [\(^3\)H]DPCPX binding, but the receptors could not be protected from inactivation by ligands. Therefore, no amino group seems to be located at the Iigand binding site. In addition, it was shown that no further amino acids witb polar side chains are present. The absence of bydrophilic amino acids frout the recognition site of the receptor apart from histidine suggests an explanation for the lack of hydrophilic ligands with high affinity for A\(_1\) receptors. KW - Toxikologie Y1 - 1988 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-60295 ER - TY - JOUR A1 - Klotz, Karl-Norbert A1 - Lohse, M. J. A1 - Schwabe, U. T1 - Characterization of the solubilized A\(_1\) adenosine receptor from rat brain membranes N2 - A\(_1\) adenosine receptors from rat brain membranes were solubilized with the zwitterionic detergent 3-[3-( cholamidopropyl)dimethylammonio]-1-propanesulfonate. The solubilized receptors retained all the characteristics of membrane-bound A\(_1\) adenosine receptors. A high and a low agonist affinity state for the radiolabelled agonist (R)-\(N^6\)-[\(^3\)H]phenylisopropyladenosine([\(^3\)H]PJA) with K\(_D\) values of 0.3 and 12 nM, respectively, were detected. High-affinity agonist binding was regulated by guanine nucleotides. In addition agonist binding was still modulated by divalent cations. The solubilized A\(_1\) adenosine receptors could be labelled not only with the agonist [\(^3\)H]PIA but also with the antagonist I ,3-diethyi-8-[\(^3\)H]phenylxanthine. Guanine nucleotides did not affect antagonist binding as reported for membrane-bound receptors. These results suggest that the solubilized receptors are still coupled to the guanine nucleotide binding protein N; and that all regulatory functions are retained on solubilization. Key Words: A1 adenosine receptors - Solubilization- Rat brain membranes. Klotz K.-N. et al. Characterization of the solubilized A1 adenosine receptor from rat brain membranes. J. Neurochem. 46, 1528-1534 (1986). KW - Toxikologie KW - A1 adenosine receptors KW - solubilization KW - rat brain membranes Y1 - 1986 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-60222 ER - TY - JOUR A1 - Klotz, Karl-Norbert A1 - Lohse, M. J. A1 - Schwabe, U. A1 - Cristalli, G. A1 - Vittori, S. A1 - Grifantini, M. T1 - 2-Chloro-N\(^6\)-[\(^3\)H]cyclopentyladenosine ([\(^3\)H]CCPA) - a high affinity agonist radioligand for A\(_1\) adenosine receptors N2 - The tritiated analogue of 2-chloro-N6-cyclopentyladenosine (CCPA), an adenosine derivative with subnanomolar affinity and a 10000-fold selectivity for A1 adenosine receptors, has been examined as a new agonist radioligand. [3H]CCP A was prepared with a specifi.c radioactivity of 1.58 TBqjmmol ( 43 Ci/mmol) and bound in a reversible manner to A1 receptors from rat brain membranes with a high affinity K0 -value of 0.2 nmol/1. In the presence of GTP a K0 -value of 13 nmol/1 was determined for the low affinity state for agonist binding. Competition of several adenosine receptor agonists and antagonists for [3H]CCPA binding to rat brain membranes confrrmed binding to an A1 receptor. Solubilized A1 receptors bound [3H]CCPA with similar affinity for the high affinity state. At solubilized receptors a reduced association rate was observed in the presence of MgC12, as has been shown for the agonist [ 3H]N6-phenylisopropyladenosine ([3H]PIA). [3H]CCPA was also used for detection of A1 receptors in rat cardio myocyte membranes, a tissue with a very low receptor density. A K0 -value of 0.4 nmol/1 and a Bmax-value of 16 fmol/ mg protein was determined in these membranes. In human platelet membranes no specific binding of [3H]CCPA was measured at concentrations up to 400 nmoljl, indicating that A2 receptors did not bind [3H]CCPA. Based on the subnanomolar affinity and the high selectivity for A1 receptors [ 3H]CCPA proved to be a useful agonist radioligand for characterization of A 1 adenosine receptors also in tissues with very low receptor density. KW - Toxikologie KW - Adenosine receptors KW - Radioligauds KW - agonists Y1 - 1989 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-60328 ER - TY - JOUR A1 - Klotz, Karl-Norbert A1 - Vogt, H. A1 - Tawfik-Schlieper, H. T1 - Comparison of A\(_1\) adenosine receptors in brain from different species by radioligand binding and photoaffinity labelling N2 - Radioligand binding to A\(_1\) adenosine receptors at brain membranes from seven species was investigated. The antagonist 8-cyclopentyl-1 ,3-[\(^3\)H]dipropylxanthine ([\(^3\)H]DPCPX) bound with affinities between 0.17 nM in sheep brain and 2.1 nM in guinea pig brain. Competition of several antagonists for [\(^3\)H]DPCPX binding showed that the most potent compounds were DPCPX with K\(_i\) values of 0.05 nM in bovine brain and 1.1 nM in guinea pig brain and xanthine amine congener (XAC) with K\(_i\) values of 0.03 nM in bovine brain and 5.5 nM in guinea pig brain. The differences in affinity of the agonist radio Iigand 2-chloro-N\(^6\) -[\(^3\)H]cyclopen tyladenosine ([\(^3\)H]CCP A) were less pronounced, rauging from a K\(_D\) value of 0.12 nM (hamster brain) to 0.42 nM (guinea pig brain). Agonist competition for [\(^3\)H]DPCPX binding of photoaffinity labelling, however, exhibited marked species differences. N-Ethylcarboxamidoadenosine (NECA) and S-N\(^6\)-phenylisopropyladenosine (S-PIA) showed 20 to 25-fold different K\(_D\) values in different species. NECA had a particularly high affinity in guinea pig brain and was only two-fold less potent than R-PIA. Thus, the difference from the "classical" A\(_1\) receptor profile (R-PIA > -NECA > S-PIA) is not sufficient to speculate that A\(_1\) receptor subtypes may exist that are coupled to different effector systems. Our data show that these difference can easily be explained by species differences. KW - Toxikologie KW - A1 adenosine receptors KW - Species differences KW - Radioligand binding KW - Photoaffinity labelling Y1 - 1991 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-60388 ER - TY - JOUR A1 - Koch, R. A1 - Deger, A. A1 - Klotz, Karl-Norbert A1 - Schenzle, D. A1 - Krämer, H. A1 - Kelm, S. A1 - Müller, G. A1 - Rapp, R. A1 - Weber, U. T1 - Characterization of solubilized insulin receptors from rat liver microsomes. Existence of two receptor species with different binding properties N2 - Insulin receptors were solubilized from rat liver microsomes by the nonionic detergent Triton X-100. After gel filtration of the extract on Sepharose CL-6B, two insulin-binding species (peak I and peak li) were obtained. The structure and binding properties of both peaks were characterized. Gel filtration yielded Stokes radii of 9.2 nm (peak I) and 8.0 nm (peak Il). Both peaks were glycoproteins. At 4°C peak 1 showed optimal insulin binding at pH 8.0 and high ionic strength. In contrast, peak li bad its binding optimum at pH 7.0 and low ionic strength, where peak I bindingwas minimal. For peak I the change in insulin binding under different conditions of pH and ionic strength was due to a change in receptor affinity only. For peak 11 an additional change in receptor number was found. Both peaks yielded non-linear Scatchard plots under most of the buffer conditions examined. At their binding optima at 4 oc the high affinity dissociation constants were 0.50 nM (peak I) and 0.55 nM (peak II). Sodium dodecyl sulfatejpolyacrylamide gel electrophoresis of peak I revealed five receptor bands with Mr 400000, 365000, 320000, 290000, and 245000 under non-reducing conditions. For peak II two major receptor bands with M\(_r\) 210000 and 115000 were found. The peak II receptor bands were also obtained aftermild reduction of peak I. After complete reduction both peaks showed one major receptor band with M\(_r\) 130000. The reductive generation of the peak II receptor together with molecular mass estimations suggest that the peak I receptor is the disulfide-linked dimer of the peak II receptor. Thus, Triton extracts from rat liver microsomes contain two receptor species, which are related, but differ considerably in their size and insulin-binding properties. KW - Toxikologie Y1 - 1986 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-60215 ER - TY - JOUR A1 - Kugler-Steigmeier, M. E. A1 - Friederich, U. A1 - Graf, U. A1 - Lutz, Werner K. A1 - Maier, P. A1 - Schlatter, C. T1 - Genotoxicity of aniline derivatives in various short-term tests N2 - Various substituted aniline derivatives were tested for genotoxicity in several short-term tests in order to examine the hypothesis that a Substitution at both ortho positions (2,6-disubstitution) could prevent genotoxicity due to steric hindrance of an enzymatic activation to electrophilic intermediates. In the Salmonellajmicrosome assay, 2,6-dialkylsubstituted anilines and 2,4,6-trimethylaniline (2,4,6-TMA) were weakly mutagenic in strain TA100 when 20% S9 mixwas used, although effects were small compared to those of 2,4-dimethylaniline and 2,4,5-trimethylaniline (2,4,5-TMA). In Drosophila me/anogaster, however, 2,4,6-TMA and 2,4,6-trichloroaniline (TCA) were mutagenic in the wing spottestat 2-3 times lower doses than 2,4,5-TMA. In the 6-thioguanine resistance test in cultured fibroblasts, 2,4,6-TMA was again mutagenic at lower doses than 2,4,5-TMA. Two methylene-bis-aniline derivatives were also tested with the above methods: 4,4'-methylene-bis-(2-chloroaniline) (MOCA) was moderately genotoxic in al1 3 test systems whereas 4,4'-methylene-bis-(2-ethyl-6-methylaniline) (MMEA) showed no genotoxicity at all. DNA binding sturlies in rats, however, revealed that both MOCA and MMEA produced DNA adducts in the liver at Ievels typically found for moderately strong genotoxic carcinogens. These results indicate that the predictive value of the in vitro test systems and particularly the Salmonellajmicrosome assay is inadequate to detect genotoxicity in aromatic amines. Genotoxicity seems to be a general property of aniline derivatives and does not seem to be greatly influenced by substitution at both ortho positions. KW - Toxikologie KW - Aniline derivatives KW - Genotoxicity KW - Short-term tests KW - Covalent DNA binding Y1 - 1989 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-60857 ER - TY - JOUR A1 - Lohse, M. J. A1 - Böser, S. A1 - Klotz, Karl-Norbert A1 - Schwabe, U. T1 - Affinities of barbiturates for the GABA-receptor complex and A\(_1\) adenosine receptors: A possible explanation of their excitatory effects N2 - The effects of barbiturates on the GABA·receptor complex and the A\(_1\) adenosine receptor were studied. At the GABA-receptor complex the barbiturates inhibited the binding of [\(^{35}\)S]t-butylbicyclophosphorothionate [\(^{35}\)S]TBPT) and enhanced the binding of [\(^3\)H]diazepam. Kinetic and saturation experiments showed that both effects were allosteric. Whereas all barbiturates caused complete inhibition of [\(^{35}\)S]TBPT binding, they showed varying degrees of maximal enhancement of [\(^3\)H]diazepam binding; (±)methohexital was idenafied as the most efficacious compound for this enhancement. At the A\(_1\) adenosine receptor all barbiturates inhibited the binding of [\(^3\)H]N\(^6\)-phenylisopropyladenosine (\(^3\)H]PIA) in a competitive manner. The comparison of the effects on [\(^3\)H]diazepam and [\(^3\)H]PIA binding showed that excitatory barbiturates interact preferentially with the A\(_1\) adenosine receptor, and sedative/anaesthetic barbiturates with the GABA-receptor complex. It is speculated that the interaction with these two receptors might be the basis of the excitatory versus sedative/ anaesthetic properties of barbiturates. KW - Toxikologie KW - GABA-receptor complex KW - Adenosine receptors KW - Barbiturates Y1 - 1987 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-60250 ER - TY - JOUR A1 - Lohse, M. J. A1 - Elger, B. A1 - Lindenborn-Fotinos, J. A1 - Klotz, Karl-Norbert A1 - Schwabe, U. T1 - Separation of solubilized A\(_2\) adenosine receptors of human platelets from non-receptor [\(^3\)H]NECA binding sites by gel filtration N2 - Human platelet membranes were solubilized with the zwitterionic detergent CHAPS (3-[3-(cholamidopropyl)dimethylammonio]- 1-propanesulfonate) and the solubilized extract subjected to gel ftltration. Binding of the adenosine receptor agonist [\(^3\)H]NECA (5'-N-ethylcarboxamidoadenosine) was measured to the eluted fractions. Two [\(^3\)H]NECA binding peaks were eluted, the first of them with the void volume. This first peak represented between 10% and 25% of the [\(^3\)H]NECA binding activity eluted from the column. It bound [\(^3\)H]NECA in a reversible, saturable and GTPdependent manner with an affinity of 46 nmol/1 and a binding capacity of 510 fmol/mg protein. Various adenosine receptor ligands competed for the binding of [\(^3\)H]NECA to the frrst peak with a pharmacological proftle characteristic for the A\(_2\) adenosine receptor as determined from adenylate cyclase experiments. In contrast, most adenosine receptor ligands did not compete for [\(^3\)H]NECA binding to the second, major peak. These results suggest that a solubilized A\(_2\) receptor-Gs protein complex of human platelets can be separated from other [\(^3\)H]NECA binding sites by gel filtration. This allows reliable radioligand binding studies of the A2 adenosine receptor of human plate1ets. KW - Toxikologie KW - A2 Adenosine receptor KW - Human platelets KW - Radioligand binding KW - Adenylate cyclase Y1 - 1988 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-60309 ER - TY - CHAP A1 - Lohse, M. J. A1 - Klotz, K.-N. A1 - Schwabe, U. A1 - Christalli, G. A1 - Vittori, S. A1 - Grifantini, M. T1 - Pharmacology and Biochemistry of Adenosine Receptors N2 - Adenosine modulates a variety of physiological functions via membrane-bound receptors. These receptors couple via G proteins to adenylate cyclase and K+channels. The A1 subtype mediates an inhibition of adenylate cyclase and an opening of K+-channels, and the A2 subtype a Stimulation of adenylate cyclase. Both subtypes have been characterized by radioligand binding. This has facilitated the development of agonists and antagonists with more than 1000-fold A1 selectivity. A1-selective photoaffinity labels have been used for the biochemical characterization of A1 receptors and the study of their coupling to adenylate cyclase. Such selective ligands allow the analysis of the involvement of adenosine receptors in physiological functions. Selective interference with adenosine receptors provides new pharmacological tools and eventually new therapeutic approaches to a number of pathophysiological states. KW - Adenosinrezeptor KW - Pharmakologie KW - Toxikologie Y1 - 1988 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-86251 ER - TY - JOUR A1 - Lohse, M. J. A1 - Klotz, Karl-Norbert A1 - Diekmann, E. A1 - Friedrich, K. A1 - Schwabe, U. T1 - 2',3'-Dideoxy-N\(^6\)-cyclohexyladenosine: an adenosine derivative with antagonist properties at adenosine receptors N2 - Tbe 2',3'-dideoxy analogue of the potent A\(_1\) receptor agonist, N\(^6\)-cyclohexyladenosine (CHA), was synthesized as a potential antagonist for the A\(_1\) adenosine receptor. In sturlies on adenylate cyclase 2',3'-dideoxy-N\(^6\)-cyclohexyladenosine (ddCHA) did not show agonist properties at A\(_1\) or at A\(_2\) receptors. However, it antagonized the inhibition by R-PIA of adenylate cyclase activity of fat cell membranes via A\(_1\) receptors with a K\(_i\) value of 13 \(\mu\)M. ddCHA competed for the binding of the selective A1 receptor antagonist, [\(^3\) HJ8-cyclopentyl-1,3-dipropylxantbine ([\(^3\)H]DPCPX), to rat brain membranes with a K\(_i\) value of 4.8 \(\mu\)M; GTP did not affect the competition curve. In contrast to the marked stereoselectivity of the A\(_1\) receptor for the cx- and the natural ß-anomer of adenosine, the cx-anomer of ddCHA showed a comparable affinity for the A\(_1\) receptor (K\(_i\) value 13.9 \8\mu\)M). These data indicate that the 2'- and 3'-hydroxy groups of adenosine and its derivatives are required foragonist activity at and high affinity binding to A\(_1\) adenosine receptors and for the distinction between the cx- and ß-forms. KW - Toxikologie KW - Adenosine receptors KW - Adenylate cyclase KW - Adenosine receptor antagonists Y1 - 1988 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-60282 ER - TY - JOUR A1 - Lohse, M. J. A1 - Klotz, Karl-Norbert A1 - Jakobs, K. H. A1 - Schwabe, U. T1 - Barbiturates are selective antagonists at A\(_1\) adenosine receptors N2 - Barbiturates in pharmacologically relevant . concentrations inhibit binding of (R)-\(N^6\)-phenylisopropyl[\(^3\)H]adenosine ([\(^3\)H]PIA) to solubilized A\(_1\) adenosine receptors in a concentration-dependent, stereospecific, and competitive manner. K\(_i\) values are similar to those obtained for membrane-bound receptors and are 31 \(\mu\)M for ( ± )-5-(1 ,3-dimethyl)-5-ethylbarbituric acid [( ± )DMBB] and 89 \(\mu\)M for ( ± )-pentobarbital. Kinetic experiments demoostrate that barbiturates compete directly for the binding site of the receptor. The inhibition of rat striatal adenylate cyclase by unlabelled (R)-\(N^6\)-phenylisopropyladenosine [(R)-PIA] is antagonized by barbiturates in the same concentrations that inhibit radioligand binding. The Stimulation of adenylate cyclase via A\(_2\) adenosine receptors in membranes from NIE 115 neuroblastoma cells is antagonized only by 10-30 times higher concentrations of barbiturates. lt is concluded that barbiturates are selective antagonists at the A1 receptor subtype. In analogy to the excitatory effects of methylxanthines it is suggested that A\(_1\) adenosine receptor antagonism may convey excitatory properties to barbiturates. Key Words: Adenosine receptors-Barbiturates - Adenylate cyclase-Receptor solubilization-[3H]PIA binding-N1E 115 cells. Lohse M. J. et al. Barbiturates are selective antagonists at A1 adenosine receptors. KW - Toxikologie KW - adenosine receptors KW - barbiturates KW - adenylate cyclase KW - receptor solubilization KW - N1E 115 cells KW - [3H]PIA binding Y1 - 1985 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-60187 ER - TY - JOUR A1 - Lohse, M. J. A1 - Klotz, Karl-Norbert A1 - Lindenborn Fotinos, J. A1 - Reddington, M. A1 - Schwabe, U. A1 - Olsson, R. A. T1 - 8-Cyclopentyl-1,3-dipropylxanthine (DPCPX) - a selective high affinity antagonist radioligand for A\(_1\) adenosine receptors N2 - The properties of 8-cyclopentyl-1,3-dipropylxanthine (DPCPX) as an antagonist ligand for A\(_1\) adenosirre receptors were examined and conipared with other radioligands for this receptor. DPCPX competitively antagonized both the inhibition of adenylate cyclase activity via A\(_1\) adenosirre receptors and the stimulationvia A\(_2\) adenosirre receptors. The K\(_i\)-values of this antagonism were 0.45 nM at the A\(_1\) receptor of rat fat cells, and 330 nM at the A\(_2\) receptor of human platelets, giving a more than 700-fold A\(_1\)-selectivity. A similar A\(_1\)-selectivity was determined in radioligand binding studies. Even at high concentrations, DPCPX did not significantly inhibit the soluble cAMPphosphodiesterase activity of human platelets. [\(^3\)H]DPCPX (105 Ci/mmol) bound in a saturable manner with high affinity to A\(_1\) receptors in membranes of bovine brain and heart, and rat brain and fat cells (K\(_D\) -values 50-190 pM). Its nonspecific binding was about 1% of total at K\(_D\) , except in bovine myocardial membranes (about 10%). Binding studies with bovine myocardial membranes allowed the analysis of both the high and low agonist affinity states of this receptor in a tissue with low receptor density. The binding properties of [\(^3\)H]DPCPX appear superior to those of other agonist and antagonist radioligands for the A\(_1\) receptor. KW - Toxikologie KW - Adenosine receptors KW - Adenylate cyclase KW - Phosphodiesterase KW - Xanthines KW - Radioligands Y1 - 1987 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-60246 ER - TY - JOUR A1 - Lohse, M. J. A1 - Klotz, Karl-Norbert A1 - Schwabe, U. A1 - Cristalli, G. A1 - Vittori, S. A1 - Grifantini, M. T1 - 2-Chloro-N\(^6\)-cyclopentyladenosine: a highly selective agonist at A\(_1\) adenosine receptors N2 - 2-Chloro-N\(^6\)-cyclopentyladenosine (CCPA) was synthesized as a potential high affinity ligand for At adenosine receptors. Binding of [\(^3\)H]PIA to A1 receptors of rat brain membranes was inhibited by CCP A with a Ki-value of 0.4 nM, compared to a Ki-value of 0.8 nM for the parent compound N\(^6\)-cyclopentyladenosine (CPA). Binding of [\(^3\)H]NECA to A\(_2\) receptors of rat striatal membranes was inhibited with a Ki-value of 3900 nM, demonstrating an almost 10,000-fold A\(_1\)-selectivity of CCPA. CCP A inhibited the activity of rat fat cell membrane adenylate cyclase, a model for the A\(_1\) receptor, with an IC\(_{50}\)-value of 33 nM, and it stimulated the adenylate cyclase activity of human platelet membranes with an EC\(_{50}\)-value of 3500 nM. The more than 100-fold A\(_1\)-selectivity compares favourably with a 38-fold selectivity of CPA. Thus, CCPA is an agonist at A\(_1\) adenosine receptors with a 4-fold higher selectivity and 2-fold higher affinity than CPA, and a considerably higher selectivity than the standard At receptor agonist R-N\(^6\) -phenylisopropyladenosine (R-PIA). CCP A represents the agonist with the highest selectivity for A\(_1\) receptors reported so far. KW - Toxikologie KW - Adenosine receptors KW - Adenylate cyclase Y1 - 1988 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-60279 ER - TY - JOUR A1 - Lohse, M. J. A1 - Maurer, K. A1 - Klotz, Karl-Norbert A1 - Schwabe, U. T1 - Synergistic effects of calcium-mobilizing agents and adenosine on histamine release from rat peritoneal mast cells N2 - 1 Adenosine and its metabolically stable analogue N.etbyl-carboxamidoadenosine (NECA) enhance histamine release from rat peritoneal mast cells when tbese are stimulated by calciummobilizing agents. NECA and adenosine shift the concentration-response curve of tbe calcium ionophore A23187 to lower concentrations. 2 The potencies of NECA or adenosinein enhancing A23187-induced histamine release are dependent on the Ievel of stimulated release in tbe absence of adenosine analogues. At high Ievels of release their potencies are up to 20 times higher than at low Ievels. Consequently, averaged concentration-response curves of adenosine and NECA for enhancing bistamine release are shallow. 3 The adenosine transport blocker S-(p-nitrobenzyl)-6-thioinosine (NBTI) has no effect by itself at low Ievels of stimulated histamine release, but abolishes the enhancing effect of adenosine. At high Ievels of release, however, NBTI alone enhances the release of histamine. 4 lt is concluded that adenosine and calcium reciprocally enhance the sensitivity of the secretory processes to the effects of the other agent. The Ievels of intracellular adenosine obtained by trapping adenosine inside stimulated mast cells are sufficient to enhance histamine release substantially, suggesting that this effect may play a physiological and pathophysiological role. KW - Toxikologie Y1 - 1989 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-60346 ER - TY - JOUR A1 - Lutz, Werner K. T1 - Dose-response relationship for chemical carcinogenesis by genotoxic agents N2 - No abstract available KW - Toxikologie Y1 - 1991 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-60766 ER - TY - JOUR A1 - Lutz, Werner K. T1 - Dose-response relationship and low dose extrapolation in chemical carcinogenesis [commentary] N2 - Data supporting various dose-respome relationships in chemical carcinogenesis are summarized. General principles are derived to explain the relationships between exposure dose, JI>NA adduct Ievel, induction of genetic changes, and tumor incidence. Some mechanistic aspects of epigenetic carcinogens (stimulation of ceU division and maldlfl'erentlation) are analyzed in a similar way. In a bomogeneous pnpulation, non-linearities are frequent. They are due to pbenomena of induction or saturation of enzymatic activities and to the multi-step nature of carcinog~: if a carcinogen acce1erates more than one step, the SUperposition of the dose- response curves for the indJvidual steps can result in an exponential relationship. A fourth power of the dose was the maximum seen in animals (fonnaldehyde). At the lowest dose Ievels, a proportionality between dose and tumor induction is postulated independent of the mechanism of action if the carcinogen aceeierotes the endogenous proass responsible for spootaneous tumor formation. Low-dose thresholds are expected only for situations where the carcinogen acts in a way that has no endogenous counterpart. Epidemiologfcal studies in humans show linear dose- response curves in all but two investigations. The difference from the strongly nonlinear slopes ·seen in animal studies could be due to the heterogeneity of the human population: if the individual sensitivity to a carcinogen is governed by a large number of genetic and Iife-style factors, the non-linea.rities will tend to cancel each other out and the dose- response curve becomes 'quasi-linear'. KW - Toxikologie Y1 - 1990 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-60789 ER - TY - JOUR A1 - Lutz, Werner K. T1 - Endogenous genotoxic agents and processes as a basis of spontaneous carcinogenesis N2 - A list ofendogenaus DNA·damaging agents and processes is given. Endogenaus e/ectrophiles are found with the cosubstrates of physiological transfer reactions (S-adenosylrnethionine for methylation, A TP for phosphorylation, NAD\(^+\) for ADP-ribosylation, acetyl CoA for acetylation). Aldehyde groups (glyceraldehyde- 3-phosphate, formaldehyde, open forms of reducing sugars, degradation products of peroxidation) or alkylating degradation products derived from endogenaus nitrose compounds represent additional possibilities. Radical-forming reactions include leakage of the superoxide anion radical from terminal cytochromes and redox cycles, hydroxyl radical formation by the Fenton reaction from endogenaus hydrogen peroxide, and the formation of lipid peroxides. Genetic instability by spontaneaus deaminations and depurinations as well as replicative instability by tautomer errors andin the presence of mutagenic metal ions represent a third important dass of endogenaus genotoxic processes. The postulated endogenaus genotoxicity could form the mechanistic basis for what is called 'spontaneous' tumor incidence and explain the possibility of an increased tumor incidence after treatment of animals with non-genotoxic compounds exhibiting tumor-promoting activity only. Individual differences are expected to be seen also with endogenaus DNA damage. The presence of endogenaus DNA darnage implies that exogenaus DNAcarcinogen adducts give rise to an incremental darnage which is expected to be proportional to the carcinogen dose at lowest Ievels. An increased tumor risk due to exposure to exogenaus genotoxic carcinogens could therefore be assessed in terms of the background DNA damage~ for instance in multiples of the mean Ievel or of the interindividual variability in a population. KW - Toxikologie KW - Endogenous genotoxicity KW - Electrophiles KW - Radicals KW - Genetic instability KW - DNA damage KW - Spontaneous tumours KW - Carcinogen risk Individual susceptibili Y1 - 1990 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-60816 ER - TY - JOUR A1 - Lutz, Werner K. T1 - In vivo covalent binding of organic chemicals to DNA as a quantitative indicator in the process of chemical carcinogenesis N2 - The covalent binding of chemical carcinogens to DNA of mammalian organs is expressed per unit dose, and a 'Covalent-Binding Index', CBI, is defined. CBI for various carcinogens span over 6 orders of magnitude. A similar range is observed for the carcinogenic potency in long-term bioassays on carcinogenicity. For the assessment of a risk from exposure to a carcinogen, the total DN A darnage can be estimated if the actual dose is also accounted for. A detailed description is given for planning and performing a DNA-binding assay. A complete literature survey on DNA binding in vivo (83 compounds) is given with a calculation of CBI, where possible, 153 compounds are listed where a covalent binding to any biological macromolecule has been shown in vivo or in vitro. Recent, so far unpublished findings with aflatoxin Mh macromolecule- bound aflatoxin Bh ·diethylstilbestrol, and 1,2-epithiobutyronitrile are included. A comparison of CBI for rat-liver DNA with hepatocarcinogenic potency reveals a surprisingly good quantitative correlation. Refinements for a DN A-binding assay are proposed. Possibilities and Iimitations in the use of D NA binding in chemical carcinogenesis are discussed extensively. KW - Toxikologie Y1 - 1979 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-61122 ER - TY - JOUR A1 - Lutz, Werner K. T1 - Investigation of the potential for binding of di(2-ethylhexyl)phthalate (DEHP) to rat liver DNA in vivo N2 - It was the aim of this investigation to determine whether or not covalent binding of di(2-ethylhexyl) phthalate (DEHP) to rat liver DNA could be a mechanism of action contributing to the observed induction of liver tumors after lifetime feeding of rodents with high doses of DEHP. DEHP radiolabeled in different positionswas administered orally to female F344 rats with or without pretreatment for 4 weeks with 1% unlabeled DEHP in the diet. Livu DNA was isolated after 16 hr and analyzed for radioattivity. Administration of [\(^{14}\)C]carboxylate unabeled DEHP resulted in no measurable DNA radioactivity. With DEHP [\(^{14}\)C]· and [\(^{3}\)H]. labeled in the alcohol moiety as well as with 2-ethyl[1-\(^{14}\)C]hexanol, radioactivity was clearly measurable in the DNA. HPLC analysis of enzyme-degraded DNA relvealed that the normal nucleosides had incorporated radiolabel whereas no radioactivity was detectable in those fractions where the carcinogen-modified nucleoside adducts are expected. A quantitative evaluation of the negative data in terms of a Iimit of detection for a covalent binding Index (CBJ) indicates that covalent interaction with DNA is highly unlikely to be the mode of tumorigenic action of DEHP in rodents. KW - Toxikologie Y1 - 1986 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-60957 ER - TY - JOUR A1 - Lutz, Werner K. T1 - Quantitative evaluation of DNA binding data for risk estimation and for classification of direct and indirect carcinogens N2 - Investigation of covalent DNA binding in vivo provided evidence for whether a test substance can be activated to metabolites able to reach and react with DNA in an intact organism. Fora comparison of DNA binding potencies of various compounds tested under different conditions, a normalization of the DNA lesion with respect to the dose is useful. A covalent binding index, CBI = (\(\mu\)mol chemical bound per mol DNA nucleotide )/(mmol chemical administered per kg body weight) can be determined for each compound. Whether covalent DNA binding results in tumor formation is dependent upon additional factors specific to the cell type. Thus far, all compounds which bind covalently to liver DNA in vivo have also proven tobe carcinogenic in a long-term study, although the liver was not necessarily the target organ for tumor growth. With appropriate techniques, DNA binding can be determined in a dose range which may be many orders of magnitude below the dose Ievels required for significant tumor induction in a long-term bioassay. Rat liver DNA bindingwas proportional to the dose of aflatoxin B1 afteroral administration of a dose between 100 \(\mu\)g/kg and 1 ng/kg. The lowest dose was in the range of generat human daily exposures. Demonstration of a lack of liver DNA binding (CBI<0.1) in vivo for a carcinogenic, nonmutagenic compound is a strong indication for an indirect mechanism of carcinogenic action. Carcinogens of this class do not directly produce a change in gene structure or function but disturb a critical biochemical control mechanism, such as protection from oxygen radicals, control of cell division, etc. Ultimately, genetic changes are produced indirectly or accumulate from endogenaus genotoxic agents. The question of why compounds which act via indirect mechanisms are more likely to exhibitanonlinear rangein the dose-response curve as opposed to the directly genotoxic agents or processes is discussed. KW - Toxikologie KW - Chemical carcinogenesis KW - Mechanism of action KW - Quantitative risk assessment KW - Genotoxicity KW - Dose-response relationship KW - Aflatoxin B1 Y1 - 1986 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-60967 ER - TY - JOUR A1 - Lutz, Werner K. T1 - Endogenous formaldehyde does not produce detectable DNA-protein crosslinks in rat liver N2 - Formaldehydeis an electrophilic molecule able to crosslink DNA and protein. It has been found to induce tumors in the nasal epithelium in rodents. The safety margin between the maximum tolerated FA concentration in the work place and the concentration found to be tumorigenic in animal studies is very small. Because FA is produced endogenously as a result of a variety of oxidative demethylations, the assessment of the tumor risk from exogenaus FA exposure has tobe related quantitatively to the level of DNA-protein crosslinks induced by endogenaus FA generation. It is reported here that the high level of endogenaus FA formed in the liver after a large dose of methanol or of aminopyrine did not lead to any observable increase in DNA-protein crosslinks. Using positive and negative control data from in vitro incubations of liver homogenate with FA or methanol it is estimated that the endogenous level of DNA damage in the liver must be more than three orders of magnitude below the damage observed at tumorigenic concentrations for the rat nose. The fact that FA is formed endogenously cannot, therefore, be used to claim that exogenous FA merely leads to a negligible increase in DNA damage. KW - Toxikologie Y1 - 1986 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-60972 ER - TY - CHAP A1 - Lutz, Werner K. T1 - Quantitative evaluation of DNA-binding data in vivo for low-dose extrapolations N2 - no abstract available KW - Toxikologie Y1 - 1987 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-80079 ER - TY - CHAP A1 - Lutz, Werner K. T1 - Structural characteristics of compounds that can be activated to chemically reactive metabolites: use for a prediction of a carcinogenic potential N2 - Many mutagens and carcinogens act via covalent interaction of metabolic intermediates with DNA in the target cell. This report groups those structural elements which are often found to form the basis for a metabolism to such chemically reactive metabolites. ~mpounds which are chemically reactive per se and which do not require metabolic activation form group 1. Group 2 compri~es of olefins and aromatic hydrocarbons where the oxidation via an epoxide can be responsible for the generation of reactive species. Aromatic amines, hydrazines, and nitrosamirres form group 3 requiring an oxidation of a nitrogen atom or of a carbon atom in alpha position to a nitrosated amine. Group 4 compounds are halogenated hydrocarbons which can either give rise to radicals or can form an ·olefin (group 2) upon dehydrohalogenation. Group 5 compounds depend upon some preceding enzymatic activity either not available in the target cell or acting on positions in the molecule which are not directly involved in the subsequent formation of electrophilic atoms. Examples for each group are taken from the "List of Chemieals and Irrdustrial Processes Associated with Cancer in Humans" as compiled by the International Agency for the Research on Cancer, and it is shown that 91% of the organic carcinogens would have been detected on the basis of structural elements characteristic for group 1-5. As opposed to this very high sensitivity, the specificity ( the true negative fraction) of using this approach as a short-term test for carcinogenicity is shown to be bad because detoxification pathways have so far not been taken into account. These competing processes are so complex, however, that either only very extensive knowledge about pharmacokinetics, stability, and reactivity will be required or that in vivo systems have to be used to predict, on a quantitative basis, the darnage expected on the DNA. DNA-binding experiments in vivo are presented with benzene and toluene to demonstrate one possible way for an experimental assessment and it is shown that the detoxification reaction at the methyl group available only in toluene gives rise to a reduction by at least a factor of forty for the binding to rat liver DNA. This quantitative approach available with DNA-binding tests in vivo, also allows evaluation as to whether reactive metabolites and their DNA binding are always the most important single activities contributing to the overall carcinogenicity of a chemical. With the example of the livertumor inducing hexachlorocyclohexane isomers it is shown that situations will be found where reactive metabolites are formed and DNA binding in vivo is measurable but where this activity cannot be the decisive mode of carcinogenic action. It is concluded that the lack of structural elements known to become potentially reactive does not guarantee the lack of a carcinogenic potential. KW - Toxikologie KW - Structureactivity relationship KW - Reactive intermediates KW - Metabolic activation KW - DNA Binding KW - Covalent binding index KW - Carcinogens KW - Benzene Y1 - 1984 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-80105 ER - TY - JOUR A1 - Lutz, Werner K. A1 - Brändle, E. A1 - Zbinden, G. T1 - Effect of gum Arabic on aminopyrine demethylation in rats N2 - Stimulation of aminopyrine demethylation induced in rats by oral or i.p. administration of phenobarbital was partially inhibited in animals receiving daily treatments of 2 x 200 mg/kg gum Arabic p.o. KW - Toxikologie Y1 - 1978 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-61146 ER - TY - JOUR A1 - Lutz, Werner K. A1 - Büsser, M. T. A1 - Sagelsdorff, P. T1 - Potency of carcinogens derived from covalent DNA binding and stimulation of DNA synthesis in rat liver N2 - ~n order to investigate the role of the stimu~ation of ceU division for the initiation (and possi:bly promotion) of live·r tumors by chemical carcinogens, the incorporation of radiolabeUed thymidine into liver DNA was dete:rmined in male rats. Single doses of various level!s of af.latoxin 81, benzidine and carbon tetrachloride (aU known to be genotoxic via DNA binding} did not affect cell division, whereas several hepatoca:rcinogens known not to bind to DNA (alphaHCH, dofibrate, and 2,3;7,8-t!etrachlorodiibenzo~p~dioxin) gave rise to a dosedependent stimulation of Ii ver DNA synthesis within 24 h. An equation combining the infl.uences of mitotic stimu:lation, expressed as dose required to double the contro~ Ievei of DNA synthesis, and DNA binding potency, exp:ressed as t.he Covalent Binding Index, correliated weil with the cardnogenk potency for both dasses of hepatocardnogens. KW - Toxikologie Y1 - 1984 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-61026 ER - TY - JOUR A1 - Lutz, Werner K. A1 - Deuber, R. A1 - Caviezel, M. A1 - Sagelsdorff, P. A1 - Friederich, U. A1 - Schlatter, C. T1 - Trenbolone growth promotant: covalent DNA binding in rat liver and in Salmonella typhimurium, and mutagenicity in the Ames test N2 - DNA binding in vivo: (6,7-\(^3\)H]ß-trenbolone (ß-TBOH) was administered p.o. and i.p. to rats. After 8 or 16 h, DNA was isolated from the livers and purified to constant specific radioactivity. Enzymatic digestion to deoxyribonucleotides and separation by HPLC revealed about 90% ofthe DNA radioactivity eluting in the form of possible TBOH-nucleotide adducts. The extent of this genotoxicity, expressed in units of the Covalent Binding Index, CBI = (~mol TBOH bound per mol nucleotide)/(mmol TBOH administered per kg body weight) spanned from 8 t~ 17, i. e. was in the range found with weak genotoxic carcmogens. Ames test: low doses of ß-TBOH increased the number of revertants in Salmonella strain TAl 00 reproducibly and m a dose-dependent manner. The mutagenic potency was 0.2 revertants per nmol after preincubation of the bacteria (20 min at 37° C) with doses between 30 and 60 \(\mu\)g per plate (47 and 94 \(\mu\)g/ml preincubation mixture). Above this dose, the number of revertants decreased to control values, accompanied by a reduction in survival. The addition of rat liver S9 inhibited the mutagenicity. DNA binding in vitro: calf thymus DNA was incubated with tritiated ß-TBOH with and without rat liver S9 Highest DNA radioactivities were determined in the absence of the "activation" system. Addition of inactive S9 (without cofactors) reduced the DNA binding by a factor of up to 20. Intermediate results were found with active S9. DNA binding in Salmonella: ß-TBOH was irreversibly bound to DNA isolated from S. typhimurium TA100 after incubation of bacteria with [\(^3\)H]ß-TBOH. Conclusions: Covalent DNA binding appears to be the mechanism of an activation-independent ("direct") mutagenicity of TBOH which is not easily detected because of the bactericidal activity. The genotoxicity risk arising from exposure of humans to trenbolone residues in meat was estimated using the in vivo data and compared to that from the exposure to unavoidable genotoxins aflatoxin B1 and dimethylnitrosamine. It ts concluded that trenbolone residues represent only a low genotoxic risk. KW - Toxikologie KW - Trenbolone KW - Anabolieagent KW - DNA binding KW - Genotoxicity KW - Ames test KW - Salmonella typhimurium Y1 - 1988 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-60897 ER - TY - JOUR A1 - Lutz, Werner K. A1 - Früh, P. U. A1 - Simon, W. T1 - Microcalorimetric determination of ΔH0, ΔG0 and ΔS0 for the interaction of the carrier antibiotics nigericin and monensin with sodium and potassium ions N2 - The thermodynainic parameters ΔH0, ΔG0 and ΔS0 - and thereby the equilibrium constants - for the complexation of the carrier antibiotics nigericin and monensin with sodium and potassium ions in methanol at 25°C have been determined by microcalorimetry. Tbc results are discussed in terms of the nature of the interaction between ligands and cations. KW - Toxikologie Y1 - 1971 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-61218 ER - TY - JOUR A1 - Lutz, Werner K. A1 - Jaggi, W. A1 - Lüthy, J. A1 - Sagelsdorff, P. A1 - Schlatter, C. T1 - In vivo covalent binding of aflatoxin B\(_1\) and aflatoxin M\(_1\) to liver DNA of rat, mouse and pig N2 - [\(^{14}\)C] Aflatoxin B\(_1\) (AFB\(_1\)) was isolated from cultures of Aspergillus parasiticus grown on [1-\(^{114}\)C] sodium acetate. Covalent binding of AFB1 to liver DNA of rat and mouse was determined 6-8 h afteroral administration. The effectiveness of covalent binding, expressedas DNA binding per dose in the units of a 'Covalent Binding Index' (CBI), (\(\mu\)mol aflatoxin/mol DNA nucleotides)/(mmol aflatoxin/kg animal), was found to be 10 400 for rats and 240 for mice. These CBI partly explain the different susceptibility of the two species for the incidence of hepatic tumors. The corresponding values for pig liver DN A, 24 and 48 h after oral administration, were found to be as high as 19 100 and 13 300. DNA-binding has not so far been reported for this species although it could represent an appropriate animal model for studies where a human-like gastrointestinal tract physiology is desirable. Aflatoxin M \(_1\) ( AFM\(_1\)) is a metabolite found in the milk of cows that have been fed AFB\(_1\)-contaminated diet. [\(^{14}\)C] AFM\(_1\) was also found to be produced by cultures of A. parasiticus giving a yield of about 0.3% of the total aflatoxins. A test for covalent binding to rat liver DN A revealed a CBI of 2100 shoWing that AFM\(_1\) must also be regarded as a strong hepatocarcinogen. It is concluded that AFB\(_1\) contaminations should be avoided in dairy feed. KW - Toxikologie Y1 - 1980 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-61097 ER - TY - JOUR A1 - Lutz, Werner K. A1 - Jaggi, W. A1 - Schlatter, C. T1 - Covalent binding of diethylstilbestrol to DNA in rat and hamster liver and kidney [Short Communication] N2 - No abstract available KW - Toxikologie KW - Carcinogenesis KW - Covalent binding index - Diethylstilbestrol KW - DNA binching KW - Estrogen KW - Hormone Y1 - 1982 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-61066 ER - TY - JOUR A1 - Lutz, Werner K. A1 - Maier, P. T1 - Genotoxic and epigenetic chemical carcinogenesis: one process, different mechanisms N2 - Chemieals that induce cancer in an intact organism are called carcinogens. This term does not differentiale between their various modes of action. In this review, Werner Lutz and Peter Maier make a mechanistic distinction between carcinogens that alter the genetic information and carcinogens that interfere with epigenetic processes. They considercardnogenesis tobe an ongoing, part1y unavoidable process which is based on a succession of mutations, most likely in stem cells, leading to autonomaus cellular growth regulation. Chemical carcinogens either induce such changes through mutations (genotoxic carcinogens) or they aceeierate the accumulation of critica1 spontaneaus mut11tions (epigenetic carcinogens). Examples are given for both classes of carcinogens, and for the processes that act at genoto:tic/nuclear 11nd epigenetic/mitotic Ievels. KW - Toxikologie Y1 - 1988 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-60884 ER - TY - JOUR A1 - Lutz, Werner K. A1 - Poetzsch, J. A1 - Schlatter, J. A1 - Schlatter, C. T1 - The real role of risk assessment in cancer risk management N2 - Rtgulatory aclio11s Iaken to reduu tht risk of harmfultffects of exposure to chemieals ofltn arenot commensurDtt with the toxicologicDf risk SJsstS&ment. A numbtr of factors relating to psychology, sociology, economics Dntl politics rather than science and medicine afftct tht final decision. Wemer Lutz and colleagues illustratt the situation using tht feuktmia-indudng chtmiCJJI benzene as an examplt. KW - Toxikologie Y1 - 1991 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-60730 ER - TY - JOUR A1 - Lutz, Werner K. A1 - Schlatter, C. T1 - Saccharin does not bind to DNA of liver or bladder in the rat [Short Communication] N2 - No abstract available KW - Toxikologie Y1 - 1977 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-61194 ER - TY - JOUR A1 - Lutz, Werner K. A1 - Schlatter, C. T1 - Mechanism of the carcinogenic action of benzene: irreversible binding to rat liver DNA N2 - No abstract available KW - Toxikologie Y1 - 1977 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-61208 ER - TY - JOUR A1 - Lutz, Werner K. A1 - Schlatter, C. T1 - A closed inhalation system for pharmacokinetic and metabolism studies of volatile compounds with small laboratory animals N2 - In the inhalation system described an animal can be kept in the same atmosphere of a 2-liter desiccator for up to 24 h. The expired carbon dioxide is adsorbed with soda lime and the resulting reduced pressure is balanced by a supply of oxygen also used for the inflow of the chemical to be investigated. Urine and faeces can be collected ~eparately and the system allows a periodical control of the concentration of the chemical by sampling the air with needle and syringe. KW - Toxikologie Y1 - 1978 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-80145 ER - TY - CHAP A1 - Lutz, Werner K. A1 - Schlatter, C. T1 - Extrapolation of carcinogenicity data to low doses with a dose-response study of the binding of benzo(a)pyrene to rat liver DNA N2 - The binding of tritiated benzo(a)pyrene (BP) to liver DNA of 25 adult male rats (SIV 50) has been determined 50 h after a single intraperitoneal injection of doses between 40 ug/kg and 4; mg/kg. The dose-response relations~ ip is linear up to i mg/kg, shows a sigmoid step towards 2 mg/kg and a shallow linear. slope above that value. TlJe 0 bserved bin ding ranges from 1.7 to 180 nmoles BP per mole DNA phosphate. The non-linearity between 1 and 2 mg/kg could be explained 0):1 the basis of an induction of metabolizing enzymes. A pure1y mathematical extrapolation of therumour incidence from a carcinogenic dose (1 x 40mg/kg for a 20% hepatoma incidence in newborn mice) to human exposure levels (aboilt 0.1 ug/kg per day) would never have followed a step like the on~ found in our experiments. Our dose-effect study therefore shows how carcinogenitity data could be extrapolated in a biologically founded way to low doses. KW - Toxikologie Y1 - 1978 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-80157 ER - TY - JOUR A1 - Lutz, Werner K. A1 - Schlatter, J. T1 - Chemical carcinogens and overnutrition in diet-related cancer [commentary] N2 - The intake of known dietary carclnogens was compiled and the cancer risk was estlmated on the basis of carcinogenic potencies in animals as derived from the Carcinogenic Potency Database by Gold and co-workers. The total cancer risk was compared with the number of cancer cases attributed by epidemiologists to dietary factors (one-third of all cancer cases, i.e. -80 000 per one million Jives). Except for alcohol, the known dietary carcinogens could not account for more than a few bundred cancer cases. Tbis was seen both with tbe DNA-reactive carcinogens (beterocyclic aromatic amines, polycyclic aromatic hydrocarbons, N-nitroso compounds, estragole, aflatoxin B., ethyl carbamate, to name the most important factors) as wen as with those carclnogens wbich have not been shown to react with DNA (e.g. caffelc acid and the carcinogeruc metals arsenic and cadmium). Residues and contaminants turned out to be negligible. Among the various pmsibilities to explain the discrepancy we investigated the roJe of ovemutritlon. Dietary restriction in animals is weil known for its strong reducing effect on spontaneous tumor formation. These data can be used to derive a carcinogenic potency for excess macronutrients: tbe tumor incidence seen with the restrlcted animals is taken as a control value and the increased tumor incidence in the animals fed ad libitum is attributed to the additional feed iotake. For excess standard diet in rats, a carcinogenic potency TD50 of 16 glkg/day was deduced from a recent study. Ovemutrition in Switzerland, estimated to be 5.5 kcallkg/day, was converted to excess food (1.9 g/kg/day) and tbe cancer incidence was calculated. The result, 60 000 cancer cases per one million Jives, is provocatively close to the number of cases not explained by the known dietary chemical carcinogens. Mechanistic studies will be required to test our hypothesis and investigate the role of different types of macronutrients in ovemutrition. KW - Toxikologie Y1 - 1992 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-60712 ER - TY - JOUR A1 - Lutz, Werner K. A1 - Viviant, A. A1 - Schlatter, C. T1 - Nonlinear dose-response relationship for the binding of the carcinogen benzo(a)pyrene to rat liver DNA in vivo N2 - Wlth radioactive compound of high specific activity, the binding of carcinogene to DNA can be measured wlth doses that are ineffective ln long-term studies. The binding of tritiated benzo(a )pyrene to liver DNA of adult male rats has been determined 50 hr after a singie l.p. injection of doses between 40 1'9/kg and 4 mg/kg. The doseresponse relationship is linear up to 1 mg/kg, shows a step towards 2 mg/kg, and gives a shallow linear slope above that value. The observed binding ranges from 1.7 to 180 nmoles benzo(a)pyrene per mole DNA phosphate. The nonlinearity could be due to an induction of metabolizing enzymes. The microsomal aryl hydrocarbon hydroxylase activity increases significantly 24 hr after a single dose of 4 mg/kg and 48 hr after doses of 2 and 4 mg/kg, but no induction Ia found with 1 mg/kg. The binding from an equimolar dose is 35 times lower than the one found on mouse skin DNA and 300 times lower than that of N,Ndlmethylnitrosamine in rat liver. A good correlatlon exiats to the respective tumor formation in long-term studles. KW - Toxikologie Y1 - 1978 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-61179 ER - TY - JOUR A1 - Lutz, Werner K. A1 - Winkler, F. K. A1 - Dunitz, J. D. T1 - Crystal structure of the antibiotic monensin similarities and differences betweeen free acid and metal complex N2 - The structure of monensin, C36H620 11 , has been deterrnined by X-ray analysis of its crystalline monohydrate (orthorhombic, a = 15.15, b = 23.61, c = 10.65 A, Z = 4, space group P212121). Phases were assigned by direct methods, malring use of the 'tangent formula'. Although the conformation of the free acid resembles that of the silver salt in being cyclic, there are differences in the hydrogen bonding pattern. These featurcs are discussed in relation to the cornplexation of metal ions by m.onensin. KW - Toxikologie Y1 - 1971 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-61228 ER - TY - JOUR A1 - Marinovich, M. A1 - Lutz, Werner K. T1 - Covalent binding of aflatoxin B\(_1\) to liver DNA in rats pretreated with ethanol JF - Experientia N2 - Male Fischer F-344 rats were given ethanol in the drinking water and/or by single oral administration. Following this, the animals received p.o. 100 ng/kg of the hepatocarcinogen eHJaflatoxin BI (AFBI)' 24 h later, the level of DNA-bound AFBI was determined in the liver and was found not to be affected by any type of ethanol pretreatment. A cocarcinogenic effect of ethanol in the liver is therefore unlikely to be due to an effect on the metabolic activation and inactivation processes governing the formation of DNA-binding AFBI metabolites. KW - Toxikologie KW - Carcinogenesis KW - DNA KW - covalent binding KW - aflatoxin KW - ethanol Y1 - 1985 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-55237 VL - 41 IS - 10 SP - 1338 EP - 1340 ER - TY - JOUR A1 - Meier, I. A1 - Shephard, S. E. A1 - Lutz, Werner K. T1 - Nitrosation of aspartic acid, aspartame, and glycine ethylester. Alkylation of 4-(p-nitrobenzyl)pyridine (NBP) in vitro and binding to DNA in the rat N2 - In a colorimetric assay using 4-( p-nitrobenzyl)pyridine (NBP) as a nucleophilic scavenger of alkylating agents, the nitrosation and alkylation reactions were investigated for a number of amino acids and derivatives. The alkylating activity increased with the square of the nitrite concentration. The nitrosation rate constants for aspartic acid, aspartame, and glycine ethylester ( = precursors C) were 0.08, 1.4 and ~ 0.2, respectively, expressed in terms of the pH-dependent \(k_2\) rate constant of the equation dNOCjdt = \(k_2\) • (C]· [nitrite]\(^2\) • The rates correlated inversely with the basicity of the amino group. The stability of the alkylating activity was astonishingly high, both in acid and at neutral pH. Half-lives of 500, 200, and 30 min were determined for aspartic acid (pH 3.5), aspartame (pH 2.5), and glycine ethylester (pH 2.5). Values of 60, 15, and 2 min; respectively, were found at pH 7. It is concluded that rearrangement of the primary N-nitroso product to the ultimate alkylating agent could be rate-limiting. The potential of nitrosated a-amino acids to bind to DN A in vivo was investigated by oral gavage of radiolabelled glycine ethylester to rats, followed irnmediately by sodium nitrite. DNA was isolated from stomach and liver and analysed for radioactivity and modified nucleotides. No indication of DNA adduct formation was obtained. Based on an estimation of the dose fraction converted from glycine ethylester to the nitroso product under the given experimental conditions, the maximum possible DNA-binding potency of nitroso glycine ethylester is about one order of magnitude below the methylating potency of N-nitrosomethylurea in rat stomach. The apparent discrepancy to the in vitro data could be due to efficient detoxification processes in mammalian cells. KW - Toxikologie KW - Nitrosation KW - Alkylation KW - Amino acids KW - DNA binding Y1 - 1990 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-60804 ER - TY - JOUR A1 - Meier-Bratschi, A. A1 - Lutz, Werner K. A1 - Schlatter, C. T1 - Methylation of liver DNA of rat and mouse by N-nitrosodimethylamine formed in vivo from dimethylamine and nitrite N2 - The extent of formation of N-nitrosodimethylaminc {NDMA) in the stomachs of rats and mice after sirnultancous oral administration of [\(^{14}\)C]dimethylamine and potassium nitrite was determined by measuring the methylation of liver DNA. With doses of around 1 mg dimethylamine hydrochloride/ kg body weight and 50 mg potassium nitrite/kg body weight. 0,8 % of the amine was nitrosated on average. The individual fluctuations ranged from 0.2 to 1.30% in the rat and from 0.2 to 1.9% in the mouse. Simultaneous administration of 50 mg sodium ascorbate (vitamin Cl/kg body weight inhibited the nitrosation by ahout 80% while 50 mg \(\alpha\)-tocopherol acetate [Vitamin E)/kg body weight reduced the nitrosation by about a half. Assuming similar kinctics and conditions of nitrosation in rats and man. a comparison of the formation of NDMA in vivo from dietary dimethylamine and nitrite with the estimated human uptake of preformed N DMA revealed that in vitro formation in the stomach of man is probably negligible. KW - Toxikologie Y1 - 1983 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-61052 ER - TY - JOUR A1 - Nolte, D. A1 - Lorenzen, A. A1 - Lehr, H.-A. A1 - Zimmer, F.-J. A1 - Klotz, Karl-Norbert A1 - Messmer, K. T1 - Reduction of postischemic leukocyte-endothelium interaction by adenosine via A\(_2\) receptor N2 - The adhesion of leukocytes to the endothelium of postcapillary venules hallmarks a key event in ischemia-reperfusion injury. Adenosine has been shown to protect from postischemic reperfusion injury, presumably through inhibition of postischemic leukocyte-endothelial interaction. This study was performed to investigate in vivo by which receptors the effect of adenosine on postischemic leukocyte-endothelium interaction is mediated. The hamster dorsal skinfold model and fluorescence microscopy were used for intravital investigation of red cell velocity, vessel diameter, and leukocyte-endothelium interaction in postcapillary venules of a thin striated skin muscle. leukocytes were stained in vivo with acridine orange (0.5 mg kg\(^{-1}\) min\(^{-1}\) i.v. ). Parameters were assessed prior to induction of 4 h ischemia to the muscle tissue and 0.5 h, 2 h, and 24 h after reperfusion. ·Adenosine, the adenosine A1-selective agonist 2-chloro-N\(^6\) -cyclopentyladenosine (CCPA), the Arselective agonist CGS 21,680, the non-selective adenosine receptor antagonist xanthine amine congener {XAC), and the adenosine uptake blocker S-(p-nitrobenzyl)-6-thioinosine (NBTI) were infused viajugular vein starting 15 min priortorelease of ischemia until 0.5 h after reperfusion. Adenosine and CGS 21,680 significantly reduced postischemic leukocyte-endothelium interaction 0.5 h after reperfusion (p< 0.01), while no inhibitory effect was observed with CCPA. Coadministration of XAC blocked the inhibitory effects of adenosine. Infusion of NBTI alone effectively decreased postischemic leukocyte-endothelium interaction. These findings indicate that adenosine reduces postischemic leukocyte-endothelium interaction via A\(_2\) receptor and suggest a protective role of endogenous adenosine during ischemia-reperfusion. KW - Toxikologie KW - Adenosine receptors KW - Ischemia/reperfusion KW - Leukocyte/endothelium interaction KW - Microcirculation Y1 - 1992 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-60424 ER - TY - JOUR A1 - Ohgaki, H. A1 - Ludeke, B. I. A1 - Meier, I. A1 - Kleihues, P. A1 - Lutz, Werner K. A1 - Schlatter, C. T1 - DNA methylation in the digestive tract of F344 rats during chronic exposure to N-methyl-N-nitrosourea N2 - The formation of \(O^6\)-methyldeoxyguanosine (\(O^6\)-MedGuo) was determined by an immuno-slot-blot assay in DNA of various tissues of F344 rats exposed to N-methyl-N-nitrosourea (MNU) in the drinking waterat 400 ppm for 2 weeks. Although the pyloric region of the glandular stomach is a target organ under these experimental conditions, the extent of DNA methylation was highest in the forestomach (185 \(\mu\)mol \(O^6\)-MedGuojmol guanine). Fundus (91 J.!moljmol guanine) and pylorus (105 J.!moljmol guanine) of the glandular stomach, oesophagus (124 \(\mu\)mol/mol guanine) and duodenum (109 )lmoljmol guanine) showed lower Ievels of \(O^6\) - MedGuo but differed little between each other. Thus, no correlation was observed between target organ specificity and the extent of DNA methylation. This is in contrast to the gastric carcinogen, N-methyl-N'-nitro-N-nitrosoguanidine (MNNG), which preferentially alkylates DNA of the pylorus, the main site of induction of gastric carcinomas by this chemical. In contrast to MNU, the nonenzymic decomposition of MNNG is accelerated by thiol compounds (reduced glutathione, L-cysteine), which are present at much higher concentrations in the glandular stomach than in the forestomach and oesophagus. During chronic exposure to MNNG (80 ppm), mucosal cells immunoreactive to 0 6-MedGuo are limited to the luminal surface [Kobori et al. (1988) Carcinogenesis 9:2271-2274]. Although MNU (400 ppm) produced similar Ievels of \(O^6\)-MedGuo in the pylorus, no cells containing methylpurines were detectable by immunohistochemistry, suggesting a more uniform methylation of mucosal cells by MNU than by MNNG. After a single oral dose of MNU (90 mg/kg) cells containing methylpurines were unequivocally identified using antibodies to \(O^6\)-MedGuo and the imidazole-ring-opened product of 7-methyldeoxyguanosine. In the gastric fundus, their distribution was similar to those methylated by exposure to MNNG, whereas the pyloric region contained immunoreactive cells also in the deeper mucosallayers. After a 2-week MNU treatment, the rate of cell proliferation, as determined by bromodeoxyuridine immunoreactivity, was only slightly enhanced in the oesophagus andin the fundus, but markedly in the forestomach and the pyloric region of the glandular stomach. lt is concluded that the overall extent of DNA methylation, the distribution of alkylated cells within the mucosa and the proliferative response all contribute to the organ-specific carcinogenicity of MNU. KW - Toxikologie KW - Gastric carcinogenesis KW - N-methyl-N-nitrosourea KW - DNA methylation Y1 - 1991 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-60759 ER - TY - JOUR A1 - Parodi, S. A1 - Lutz, Werner K. A1 - Colacci, A. A1 - Mazzullo, M. A1 - Taningher, M. A1 - Grilli, S. T1 - Results of animal studies suggest a nonlinear dose-response relationship for benzene effects N2 - Considering the very large industrial usage of benzene, studies in risk assessment aimed at the evaluation of carcinogenic risk at low Ievels of exposure are important. Animal data can offer indications about what could happen in humans and provide more diverse information than epidemiological data with respect to doseresponse consideration. We have considered experiments investigating metabolism, short·term genotoxicity tests, DNA adduct formation, and carcinogenicity long-term tests. According to the different experiments, a Saturation of benzene metabolism and benzene effects in terms of genotoxicity seems evident above 30 to 100 ppm. Below 30 to 60 ppm the initiating effect ofbenzene seems tobe linear fora large intervaJ ofdosages, at least judging from DNA adduct formation. Potentiallack of a promoting effect of benzene (below 10 ppm) could generate a sublinear response at nontox.ic levels of ex.posure. This possibility was suggested by epidemiological data in humans and is not confirmed or excluded by our observations with animals. KW - Toxikologie Y1 - 1989 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-60843 ER -