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Biomedical subjects

E Gocke

Publications and source records attributed to E Gocke.

At least 37 records · Page 2Linked to original sources
PubMed1993

Photomutagenesis test development: II. 8-Methoxypsoralen, chlorpromazine and sunscreen compounds in chromosomal aberration assays using CHO cells.

Chromosomal changes were analysed in Chinese hamster ovary (CHO) cells treated with 8-methoxypsoralen (8-MOP) or chlorpromazine (CPZ) and irradiated with either a UVA fluorescent tube (emission spectrum ranging from 350 to 400 nm) or a xenon burner (continuous emission spectrum simulating ambient sunlight). In the dark neither 8-MOP nor CPZ was genotoxic by itself. If these compounds were used in combination with UV irradiation the rate of chromosome aberrations was significantly increased. The magnitude of the clastogenic response was dependent on compound concentration and UV dose. The spectral composition also played an important role. Care must be taken to account for spectral changes caused, e.g., by passage of the light through the plastic lid of the container. The possible clastogenicity of two sunscreens was tested with two protocols: (1) cells attached to the culture dish were treated in presence of the sunscreen in the medium or (2) cells were irradiated through a layer of sunscreen solution as a filter. With this a clear UVB-absorbing effect and a decreased frequency of UVAB-induced chromosome aberration was evident with the UVB-absorbing compound Parsol HS but was absent, as expected, with the UVA-absorbing compound Parsol 1789. The presence of the sunscreens in the irradiated cell sample did not cause a significant increase in UV-induced chromosome aberrations.

Animals↗
PubMed1993

Renin inhibitors as an example of presumptive irrelevant positive findings in the Salmonella/mammalian microsome assay (Ames test).

An increase in the number of mutant colonies in the Ames test is generally taken as a strong indication for a genotoxic (e.g., DNA damaging) property of the test compound or its metabolites. However, a few examples are known in which mechanisms usually related to some sort of growth enhancement will lead to increases in mutant frequencies of spontaneous origin. The renin inhibitor Ro 42-5892 increased the number of mutant colonies of strain TA1538 and to a lesser degree of TA98 in the standard plate incorporation assay (Ames test). Since there is no chemical basis for a 'DNA reactivity' of this compound, experiments were performed to obtain information about possible indirect mechanisms of enhancing the number of spontaneous mutant colonies. Circumstantial evidence is presented to attribute the weak activity not to an inherent genotoxic property but rather to an as yet undefined indirect effect on the expression of spontaneous mutants. Since Ro 42-5892 contains a histidine residue it was a reasonable assumption to suspect a growth enhancing property of the test compound. However, none of the strains showed an elevation of the number of revertant colonies or an increase in the density of the background growth. In addition, structurally related non-histidine containing renin inhibitors showed absolutely no increase in the number of revertant colonies. Furthermore, no growth induction (either in liquid or under selective conditions) and no histidine cleave off by a TA1538/TA98 specific metabolism could be shown. A second line of evidence showing parallelism to growth enhancing compounds concerns the comutagenicity of histidine containing renin inhibitors. When Ro 42-5892 was tested in combination with established mutagens, a multiplicative synergism was found. This effect was observed not only in strains TA1538 and TA98 but also in the standard Salmonella tester strains where the spontaneous mutant frequency was not increased by Ro 42-5892. Analogous effects were previously shown for free histidine, isohistidine, phenobarbital and tetracycline and in part explained by molecular mechanisms.

False Positive Reactions↗
PubMed1992

Normal chromosomal aberration frequencies in peripheral lymphocytes of healthy human volunteers exposed to a maximum daily dose of paracetamol in a double blind trial.

Paracetamol (acetaminophen) has been examined for mutagenic potential in numerous studies: gene mutation tests consistently gave negative results while in vitro chromosomal aberration tests showed equally consistently positive effects. In vivo studies for chromosome breaking activity gave clearly negative, equivocal or weakly positive results. In particular two reports have indicated that human volunteers taking a maximum daily dose of paracetamol (3 x 1000 mg over 8 h) exhibited significantly elevated frequencies of chromatid breaks in their peripheral lymphocytes 24 h later. In the one study evaluating the time course, levels returned to normal between 3 and 7 days later. We performed a carefully controlled double-blind study in which volunteers were pre-screened for normal liver function, they all were non-smoking and their diet and environmental exposures were controlled during the study. Cell-cycle kinetics were monitored and paralleled and a placebo group was included. Although a larger number of cells than in the other studies was analysed we were unable to reproduce their findings. No significant increases in structural chromosome aberrations (CA) were found either when the paracetamol group (male, female or both) post-dosing values were compared with pre-dosing values, or when treated groups at any sampling time were compared with the placebo groups. There was not even any evidence that individuals responded to the clastogenic potential of paracetamol or that a group response may have been masked by non-responders. In conjunction with the recently published results of the NTP bioassay, showing no carcinogenic activity in mice and no carcinogenic activity in rats except an increase of mononuclear cell leukaemia in female rats which is of doubtful relevance, the study presented here argues that paracetamol does not pose an unacceptable (if any) genotoxic/carcinogenic risk to man.

Acetaminophen↗
PubMed1992

Phenobarbital: does the positive result in TA1535 indicate genotoxic properties?

The liver carcinogen phenobarbital (PB) causes a weak but reproducible increase of the mutant frequency in the Ames test, strain TA1535, without S9. Since there is no obvious chemical basis for a "DNA reactivity" of this compound experiments were performed to obtain information about possible indirect mechanisms of enhancing the number of spontaneous mutant colonies. In the course of the study strong synergistic and comutagenic effects of PB when given in combination with Na-azide or 2-aminoanthracene (2AA) were observed. Not only TA1535 but the complete set of tester strains was responsive. However, PB did not enhance the effects of other mutagens such as 4-nitroquinoline N-oxide or 2-nitrofluorene. It is argued that in strain TA1535 the fixation and expression of spontaneously occurring DNA lesions is amenable to modulation by PB similar to that of Na-azide or 2AA induced lesions. Thus in the usual sense, PB is not genotoxic in the Ames test. Methapyrilene, another liver carcinogen with an assumed nongenotoxic mode of action, showed almost identical properties in these experiments.

Animals↗
PubMed1991

Mechanism of quinolone mutagenicity in bacteria.

Gyrase inhibitors of the quinolone type are genotoxic in bacteria. A functioning excision-repair system is needed to show the mutagenic activity in the Ames tester strains. The antibiotics do not interact with DNA directly but induce the gyrase enzyme to cleave the DNA with protein covalently bound at the site-specific double-strand scission. A prominent site of action is shown to be located at a distance of about 80 bp from the hisG428 sequence at which mutations are scored in the responsive strain TA102. Since the interaction of quinolones with the mammalian counterparts of gyrase, topoisomerase II, is lower by orders of magnitude, it is argued that the bacterial screening tests are of little relevance for predicting effects in mammalian cells.

Anti-Infective Agents↗
PubMed1988

Plasmid copy number and mutant frequencies in S. typhimurium TA102.

Tetracycline and chloramphenicol increase the number of mutant colonies of strain TA102, which carries the reverting gene on the plasmid pAQ1. Determination of the plasmid content by agarose gel analysis shows that the increase of the mutant colony number is paralleled closely by an increase of the number of pAQ1 plasmids per cell, indicating that the two compounds do not increase the frequency of mutants "per gene," but only enhance the number of the genes at which mutations can occur. Thus, not considering the molecular processes could result in mistakenly attributing the increase in the number of mutants per plate (respective to the number of mutants per cell) to a mutagenic activity of the antibiotics.

Chloramphenicol↗
PubMed1988

The 'antimutagenic' effect of cinnamaldehyde is due to a transient growth inhibition.

Addition of cinnamaldehyde to the selective medium causes a reduction in the number of revertant colonies of S. typhimurium or E. coli when the cells have been mutagenized with 4NQO but not when they have been mutagenized with MNNG. Toxicity of the cinnamaldehyde exposure depends largely on the status of growth and/or nutrient supply of the cells. We present evidence that simple growth inhibition due to lack of nutrients mimics the effect of cinnamaldehyde in 4NQO- and MNNG-treated cells. This argues that the reduction of mutant colonies is due to a transient growth retardation caused by cinnamaldehyde exposure, which presumably allows the cells to repair 4NQO-induced damage--but not MNNG-induced damage--via a more error-free pathway.

4-Nitroquinoline-1-oxide↗
PubMed1986

Mutagenicity testing experiments with the Cobas Bact.

The feasibility of mutagenicity assays with the Cobas Bact Automatic analyser was explored using selected model mutagens. The reduction of the latency period (the period until the growth of the mutant cells becomes optically measurable) was found to be a valid measure for the mutagenic activity of strong mutagens. For weaker mutagens an evaluation analogous to the fluctuation test seemed the more appropriate approach. The influence of various variables, such as concentration of histidine, size of inoculum, medium composition and S9 concentration, is described. Adaptation of the Cobas Bact system to the differential growth inhibition test is also mentioned.

Animals↗
PubMed1985

Topoisomerase I has a strong binding preference for a conserved hexadecameric sequence in the promoter region of the rRNA gene from Tetrahymena pyriformis.

Topoisomerase I is in situ associated with DNaseI hypersensitive sites located in the promotor and terminator regions of the extrachromosomal rDNA in Tetrahymena thermophila at sites with sequences fitting the motif (sequence in text) Reconstitution experiments with purified topoisomerase I and cloned fragments of rDNA demonstrate that the enzyme exhibits the same binding and cleavage properties on naked DNA. These observations are striking as topoisomerase I previously has been found to exhibit low sequence specificity. The specific binding of the enzyme has an absolute requirement for divalent cations with a preference for Ca2+. The strong binding to the hexadecamer has been characterized by competition experiments, and it has been used to determine the molecular weight of the enzyme.

Animals↗
PubMed1985

A high affinity topoisomerase I binding sequence is clustered at DNAase I hypersensitive sites in Tetrahymena R-chromatin.

Topoisomerase I is associated with DNAase I hypersensitive sites in the nontranscribed spacers flanking the rRNA genes in Tetrahymena thermophila. The endogenous topoisomerase I introduces site and strand specific single-strand cleavages in the rDNA spacers in situ. The cleavages occur base specifically within a hexadecameric sequence element present in two or three direct repeats at the hypersensitive sites. The sequence specificity and polarity of the cleavage reaction are identical when the enzyme is reacted with naked rDNA, indicating that the repetitive element functions as a high-affinity topoisomerase I attraction site in the r-chromatin. The biological mechanism associated with this phenomenon appears to be widespread among eukaryotes, since the topoisomerase I recognition sequence is conserved in the rDNA spacers of phylogenetically remote organisms, such as fungi, slime molds, ciliates, and insects.

Animals↗
PubMed1983

A site and strand specific nuclease activity with analogies to topoisomerase I frames the rRNA gene of Tetrahymena.

Exposure of macronuclear chromatin from Tetrahymena thermophila to sodium dodecyl sulfate causes an endogenous nuclease to cleave the extra-chromosomal rDNA at specific sites. All cuts are single-strand cleavages specific to the non-coding strand. Three cleavages map in the central non-transcribed spacer of the palindromic molecule at positions -1000, -600 and -150 bp with respect to the transcription initiation point. A fourth site is located close to the transcription termination point, while no cleavage is observed in the coding region. The position of each cleavage is in the immediate neighbourhood of DNAse I hypersensitive sites. Additionally, certain DNA sequence motifs are repeated in the region around the cleavages. Upon cleavage induction a protein becomes attached to the rDNA. Our results indicate covalent binding to the generated 3' end, in analogy to the aborted reaction of topoisomerase I.

Animals↗
PubMed1983

Autoradiographic detection of 6-thioguanine-resistant lymphocytes of mice. A novel system in somatic mutagenesis testing.

After treatment of mice with ethylnitrosourea in vivo, induction of 6-thioguanine-resistant splenic lymphocytes was measured in vitro. The lymphocytes were mitogen-stimulated in presence of thioguanine. After a culture period of 30 h, TG resistance was determined by the ability of cells to incorporate [3H]thymidine. The results suggest that at this selection condition the majority of resistant cells stem from somatic mutations. Thus, the system is potentially useful as a screening assay for somatic mutagenesis testing. The problem of misleading results due to the presence of phenocopies was investigated by treatment in vitro with UV radiation or ethylnitrosourea.

Animals↗
PubMed1983

Mutagenicity studies with the mouse spot test.

The mammalian spot test, which detects somatic gene mutations in mouse embryos, was investigated with selected chemicals to (a) further validate this test system (ENU, EMS, 2AAF, colchicine) and (b) evaluate the mutagenic potential, in a whole-mammal system, of environmental compounds that had been previously recognized as mutagens in other mammalian or submammalian test systems (1,2-dichloroethane, hydroquinone, nitrofurantoin, o-phenylenediamine, fried sausage extract). Of these substances, ENU, EMS and 2AAF were significantly mutagenic, 1,2-dichloroethane was probably weakly mutagenic. The ENU data were used to estimate the number of pigment precursor cells present at the time of treatment (day 9.25). We also describe in this report the use of a fluorescence microscope for classification of hairs from spots on the coat of C57BL/6JHan X T hybrids.

2-Acetylaminofluorene↗
PubMed1983

Study of artificial flavouring substances for mutagenicity in the Salmonella/microsome, Basc and micronucleus tests.

Seventy-six compounds used as artificial flavouring substances in food products were studied for mutagenic properties by the use of the Salmonella/mammalian microsome test (Ames test), Basc test on Drosophila melanogaster and micronucleus test on mouse bone marrow. The following four compounds were mutagenic in Ames tests: ethyl nitrite, ethyl 3-phenylglycidate, 6-methylquinoline and musk ambrette. Of these, ethyl nitrite and musk ambrette also induced a significant (P less than or equal to 0.01) increase in sex-linked recessive lethal mutations in Drosophila. Two further compounds, ethyl 3-methyl-3-phenylglycidate and 4-n-propylanisole, appeared weakly mutagenic in Drosophila only. The result with 4-n-propylanisole was judged to be of equivocal biological significance. None of the flavouring substances induced micronuclei, i.e. cytogenetic damage in the bone marrow of mice.

Animals↗
PubMed1982

5-Bromodeoxyuridine tablets with improved depot effect for analysis in vivo of sister-chromatid exchanges in bone-marrow and spermatogonial cells.

An improved 5-bromodeoxyuridine (BrdU) tablet technique for observation in vivo of SCE in mouse bone-marrow and spermatogonial cells is described. BrdU tablets were coated with agar as protecting barrier before subcutaneous implantation into mice. In comparison with the original tablets, the agar-coated tablets provided a slower and more uniform delivery of BrdU to the animals. This was corroborated (1) by recovering the undissolved portion of tablets at 1-2-h intervals, and (2) by quantitative determination of the BrdU levels in blood with the help of an analytical HPLC technique. The time required for complete dissolution of the coated tablets was considerably longer than that for the original tablets. This means that the dose of BrdU required for observation of SCE in mouse bone-marrow cells can be reduced accordingly. By using these modified tablets, therefore, undesired effects of high doses of BrdU on mutation (base-line SCE frequency) as well as on cellular replication and proliferation can be diminished. Moreover, the improved depot effect of the modified tablets facilitates the differential labeling of sister chromatids in mouse spermatogonia, a tissue containing cells with a relatively long DNA synthesis period.

Animals↗
PubMed1982

Mutagenicity study of fried sausages in Salmonella, Drosophila and mammalian cells in vitro and in vivo.

The basic extract of pan-fried sausages was studied for mutagenic potential in seven test systems. Mutagenic activity was high in the standard Ames assay in the Salmonella typhimurium strains TA1538 and TA98 in presence of S9 mix. In vivo, in the intrasanguine host-mediated assay with strain TA98 on Aroclor-pretreated mice, the mutagenic activity of the extract was low. A borderline activity was seen in the SCE assay in vitro with V79 Chinese hamster cells in presence of S9 mix. No significant mutagenic action was found in the gene-mutation assay for thioguanine resistance with V79 cells, the Drosophila sex-linked recessive lethal test, the micronucleus test and the mammalian spot test.

Animals↗