@article{AdamAhrweilerSahaMoelleretal.1993, author = {Adam, W. and Ahrweiler, M. and Saha-M{\"o}ller, C. R. and Sauter, M. and Sch{\"o}nberger, A. and Epe, B. and M{\"u}ller, E. and Schiffmann, D. and Stopper, Helga and Wild, D.}, title = {Genotoxicity studies of benzofuran dioxetanes and epoxides with isolated DNA, bacteria and mammalian cells}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-63420}, year = {1993}, abstract = {1.2-Dioxetanes, very reactive and high energy molecules. are involved as labile intermediates in dioxygenase- activated aerobic metabolism and in physiological processes. Various toxico1ogica1 tests reveal that dioxetanes are indeed genotoxic. In supercoiled DNA of bacteriophage PM2 they induce endonucleasesensitive sites, most of them are FPG protein-sensitive base modifications (8-hydroxyguanine, fonnamidopyrimidines). Pyrimidinedimersand sites ofbase loss (AP sites) which were probed by UV endonuclease and exonuclease 111 are minor lesions in this system. While the alky1-substituted dioxetanes do not show any significant mutagenic activity in different Salmonella typhimurium strains, heteroarene dioxetanes such as benzofuran and furocoumarin dioxetanes are strongly mutagenic in S. typhimurium strain TA I 00. DNA adducts formed with an intermediary alkyJating agent appear to be responsible for the mutagenic activity of benzofuran dioxetane. We assume that the benzofuran epoxides, generated in situ from benzofuran dioxetanes by deoxygenation are the ultimate mutagens of the latter. since benzofuran epoxides are highly mutagenic in the S. typhimurium strain TAIOO and they form DNA adducts. as detected by the 212Ppostlabelling technique. Our results imply that the type of D NA darnage promoted by dioxetanes is dependent on the structural feature of dioxetanes. Furthermore, the direct photochemical DNA darnage by energy transfer. i.e., pyrimidine dimers, plays a minor role in the genotoxicity of dioxetanes. Instead, photooxidation dominates in isolated DNA. while radical darnage and alkylation prevail in the cellular system.}, subject = {Toxikologie}, language = {en} } @article{JanevskiChohStopperetal.1993, author = {Janevski, J. and Choh, V. and Stopper, Helga and Schiffmann, D. and De Boni, U.}, title = {Diethylstilbestrol alters the morphology and calcium levels of growth cones of PC12 cells in vitro}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-86858}, year = {1993}, abstract = {Diethylstilbestrol (DES) is a synthetic estrogen with carcinogenic properties. DES is known to alter cytoskeletal components, including the organization of actin stress fibres in C6 rat glioma cells. ln a test of the hypothesis that DES disrupts actin Filaments of growth cones in neuron-like cells, DES-induced changes in filopodial lengths were quantified in rat pheochromocytoma (PC12) cells in vitro. DES significantly altered growth cone morphology, with collapse of growth cone filopodia and neurite retraction invariably occurring at a concentration of 10 MikroM. At 5 MikroM DES, transient reductions in total filopodiallengths occurred. At DES concentrations of 0.1 nM and 1 nM, reductions in total filopodiallengths occurred in a fraction of growth cones. Evidence exists which shows that growth cone activity and morphology are intimately linked to Ieveis of intracellular, free calcium and that DES increases such levels. Measurements of free intracellular calcium levels by fluorescence microscopy, at times concurrent with the DES-induced reduction in total filopodial lengths, showed that calcium levels were indeed significantly increased by 10 MirkoM DES. Labelling of filamentaus actin (f-actin) with FITC-phalloidin showed that the f-actin distribution in growth cones exposed to DES could not be differentiated from the distribution found in spontaneously retracting growth cones. Tagether with evidence which showed that growth cone motility was not affected, the results are taken to indicate that DES, rather than acting directly on the cytoskeleton, exerts its effects indirectly, by a calcium-induced destabilization of actin filaments in the growth cone.}, subject = {Calcium}, language = {en} } @article{KirchnerStopperPappetal.1993, author = {Kirchner, S. and Stopper, Helga and Papp, T. and Eckert, I. and Yoo, H. J. and Vig, B. K. and Schiffmann, D.}, title = {Cytogenetic changes in primary, immortalized and malignant mammalian cells}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-63439}, year = {1993}, abstract = {Some chromosomes in transformed rat cells and somatic cell hybrids fail to display the presence of kinetochore proteins as detected by antikinetochore antibodies. Suchchromosomes (K- Chromosomes) may constitute a novel mechanism for the genesis of aneuploidy. Wehave analyzed primary~ immortalized and malignant marnmalian cells for the presence of kinetochore proteins and micronuclei. Our resuJts suggest a correlation of the K- chromosome and micronucleus frequency with the variability in chromosome number. Upon in situ hybridization with the minor satellite and alpha satellite sequences some Kchromosomes showed a signal. This indicates that the observed lack of kinetocbores is not necessarily due to a lack of centromeric DNA. We conclude that dislocated K- chromosomes may become incorporated into micronuclei which are prone to loss. Such events would be associated with the generation of aneuploidy.}, subject = {Toxikologie}, language = {en} } @article{StopperEckertSchiffmannetal.1994, author = {Stopper, Helga and Eckert, I. and Schiffmann, D. and Spencer, D. L. and Caspary, W. J.}, title = {Is micronucleus induction by aneugens an early event leading to mutagenesis?}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-63390}, year = {1994}, abstract = {This study was designed to investigate a previously unidentified potential mechanism for mutation induction as well as to clarify a biological comequence of micronucleus formation. We compared the induction of micronuclei with mutation inductioo as measured by trißuorothymidine (TFI') resistance in mouse L5178Y cells using four aneugens: colcemid, diethylstilbestrol, griseofulvin and vioblastine. AU four compounds induced micronuclei which appeared in the first cell cycle after treatment. More than 85\% of the micronuclei induced by each compound stained positive for the presence of kinetochores implying that the micronuclei contained wbole cbromosomes. However, these same compounds were unable to induce TFf resistance under tbree different treatment regimes. We concluded that tbese compounds, under conditions where tbey induce primarily kinetochore positive micronuclel, were not able to induce mutations. Thus, the induction of micronuclei containing wbole chromosomes barborlog a select.able gene is not an early event leadlog to mutations in these cells.}, subject = {Toxikologie}, language = {en} } @article{StopperKirchnerSchiffmannetal.1994, author = {Stopper, Helga and Kirchner, S. and Schiffmann, D. and Poot, M.}, title = {Cell cycle disturbance in relation to micronucleus formation induced by the carcinogenic estrogen diethylstilbestrol}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-82250}, year = {1994}, abstract = {In addition to its tumor-promoting activity in honnone-receptive tissue, the carcinogenic estrogen diethylstilbestrol (DES) has been found to induce cell transformation, aneuploidy and micronucleus formation in mammalian cells. The majority of these micronuclei contained whole chromosomes and were fonned during mitosis. Here a possible relationship between a disturbance in cell cycle progression and micronucleus fonnation is investigated by exposing Syrian hamster embryo (SHE) cells to DES. Continuous bromodeoxyuridine labeling followed by bivariate Hoechst 33258/ethidium bromide flow cytometry was employed for analysis of cell cycle transit and related to the time course of micronucleus formation. Treatment of SHE cells with DES resulted in delayed and impaired cell activation (exit from the GO/G 1 phase), impaired S-phase transit and, mainly, G2-phase traverse. Cells forming micronuclei, on the other hand, were predominantly in G2 phase during DES treatment. These results suggest that impairment of Sand G2 transit may involve a process ultimately leading to micronucleus formation.}, subject = {Toxikologie}, language = {en} } @article{StopperKoerberSchiffmannetal.1993, author = {Stopper, Helga and K{\"o}rber, C. and Schiffmann, D. and Caspary, W. J.}, title = {Cell-cycle dependent micronucleus formation and mitotic disturbances induced by 5-azacytidine in mammalian cells}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-63411}, year = {1993}, abstract = {5-Azacytidine was originally developed to treat human myelogenous leukemia. However, interest in this compound has expanded because of reports of its ability to affect cell differentiation and to alter eukaryotic gene expression. In an ongoing attempt to understand the biochemical effects of this compound, we examined the effects of 5-azacytidine on mitosis and on micronucleus formation in mammalian cells. In L5178Y mouse cells, 5-azacytidine induced micronuclei at concentrations at which we and others have already reported its mutagenicity at the tk locus. Using CREST staining and C-banding studies, we showed that the induced micronuclei contained mostly chromosomal fragments although some may have contained whole chromosomes. By incorporating BrdU into the DNA of SHE cells, we determined that micronuclei were induced only when the compound was added while the cells were in S phase. Microscopically visible effects due to 5-azacytidine treatment were not observed until anaphase of the mitosis following treatment or thereafter. 5-Azacytidine did not induce micronuclei via interference with formation of the metaphase chromosome arrangement in mitosis, a common mechanism leading to aneuploidy. SupravitalUV microscopy revealed that chromatid bridges were observed in anaphase and, in some cases, were sustained into interphase. In the first mitosis after 5-azacytidine treatment we observed that many cells were unable to perform anaphase separation. All of these observations indicate that 5-azacytidine is predominantly a clastogen through its incorporation into DNA.}, subject = {Toxikologie}, language = {en} } @article{StopperKoerberSpenceretal.1993, author = {Stopper, Helga and K{\"o}rber, C. and Spencer, D. L. and Kirchner, S. and Caspary, W.J. and Schiffmann, D.}, title = {An investigation of micronucleus and mutation induction by oxazepam in mammalian cells}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-63404}, year = {1993}, abstract = {Tbe benzodiazepines are a class of d.rugs that are widely used in the treatment of various psychiatric disorders. One member of um ~' oxazepam, is also a common metabolite of sevmd other benzod.iazepines. Since the evidence for the genetic toxicity and carcinogenic properties of these compounds is incol:lsb1ent, we investigated the oxazepam-induced fonnation of micronuclei in Syrian Hamster embryo fibroblast (SHE) cells, human amniotic fluid fibroblast-like (AFFL) cells and LS178Y mouse cells. A dose-dependent increase in micronucleus fractions was found in all tbree ceU llnes. The time course of micronucleus induction in L5178Y cells showed a maximum at 5 h after treatment, suggesting that the micronuclei were fonned in the first mitosis after treatment. Kinetochore staining (CREST -antiserum) revealed the presence of kinetochores in -SO\% of the micronuclei in aU tbree ceU types. ThJs resu1t was further confinned by in situ bybridization in LS178Y cells and indicates tbe presence of wbole Chromosomes or centric fragments as weU as acentric fragments in the oxazepam-induced micronuclei. The LS178Y cells did not show a mutagenic response to oxazepam at any of the doses or expression times used.}, subject = {Toxikologie}, language = {en} } @article{StopperPechanSchiffmann1992, author = {Stopper, Helga and Pechan, R. and Schiffmann, D.}, title = {5-azacytidine induces micronuclei in and morphological transformation of Syrian hamster embryo fibroblasts in the absence of unscheduled DNA synthesis}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-63443}, year = {1992}, abstract = {lt is known that 5-azacytidine (5-AC) induces tumors in several organs of rats and mice. The mechanisms of these effects are still poorly understood although it is known that 5-AC can be incorporated into DNA. Furthermore, it can inhibit DNA methylation. The known data on its clastogenic andjor gene mutation-inducing potential are still controversial. Therefore, we have investigated the kinds of genotoxic effects caused by 5-AC in Syrian hamster embryo (SHE) fibroblasts. Three different endp6ints (micronucleus formation, unscheduled DNA synthesis (UDS) and cell transforrnation) were assayed under similar conditions of metabolism and dose at target in this cell system. 5-AC induces morphological transformation of SHE cells, but not UDS. Therefore, 5-AC does not seem to cause repairable DNA lesions. Furthermore, our studies revealed that 5-AC is a potent inducer of mkronuclei in the SHE system. Immunocytochemical analysis revealed that a certain percentage of these contain kinetochores indicating that 5-AC may induce both clastogenic events and numerical chromosome changes.}, subject = {Toxikologie}, language = {en} } @article{TasStopperKoscheletal.1991, author = {Tas, P. and Stopper, Helga and Koschel, K. and Schiffmann, D.}, title = {Influence of the carcinogenic oestrogen diethylstilboestrol on the intracellular calcium level in C6 rat glioma cells}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-63459}, year = {1991}, abstract = {The ~fthetic oes~rog~n diethylsti~boestrol (DES) causes a dose-dependent elevation of the cytoplasuuc Ca concentratton m C6 rat ghoma cells. This Ca2+ rise is caused neither by Ca2+ influx nor ~-r release from the ~a2 + stores of the endoplasmic reticulum. Therefore it seems likely that DES mob!hzes Ca2+ from a nutochondrial source. The DES-induced Ca2+ signal is remarkably similar to the one mduced by the. tumou~ promotor ~hapsigargin. As this compound causes leakage of calcium from the endoplasmt~ rettculum tt ~ms posstble that DES induces a similar leakage from mitochondrial Ca2+ stores. It remaans to be estabhshed whether the DES-mediated rise in intracellular calcium is causally related to the tumour-promoting properties of this compound}, subject = {Toxikologie}, language = {en} }