Trenimon: Biochemical,physiological and genetic effects on cells and organisms

The trifunctional alkylating mutagen Trenimon interferes with the genetic material of a variety of organisms and test systems with respect to the induction of point and chromosomal mutations, sister-chromatid exchanges, recombination phenomena and phage induction. Beneath these mutagenic effects several biochemical and cell physiological aspects have been investigated. In this review we discuss chemical and cell physiological effects of Trenimon, aspects of cancer therapy with Trenimon and genetic effects induced by Trenimon. The available data on mutagenic effects of Trenimon are presented according to organisms or test systems. A short discussion on a possible genetic load by therapy with Trenimon in man concludes this review.DNA damage, especially the induction of cross-linkings, seems to represent the common reason for most of the described effects of Trenimon on cells and organisms.     

Evaluation of the co-mutagenicity of ethanol and delta 9-tetrahydrocannabinol with Trenimon.

The mutagenic potential of chronic treatments of male CF-1 mice with ethanol and delta 9-tetrahydrocannibinol (THC), and their comutagenic potential with a known mutagenic agent, Trenimon, were examined. This was accomplished by measuring the frequency of dominant lethal mutations arising from mating of treated males with nontreated females. Adult male mice were treated with 5% (v/v) ethanol as part of a liquid diet (28% ethanol-derived calories) for five weeks; 10 mg/kg body weight (p.o.) THC every two days for five weeks; a single injection of Trenimon (0.125 mg/kg, i.p.) on day 28 of diet treatment; and all combinations of treatments. The control group was pair-fed a liquid diet in which isocaloric sucrose replaced ethanol; these males were also given sesame oil (vehicle for THC) and saline (vehicle for Trenimon) on the same schedule as that for the treated males. Neither body weights nor hematocrits were adversely affected by any treatment. Both ethanol and Trenimon treatments resulted in a small (8-9%; p less than 0.05) decrease in testicular weight. The effect of combined treatment with ethanol and Trenimon was roughly additive. Treatment with THC had no effect on testicular weight. Seminal vesicle weights were not affected by any treatment. Treatments were without significant effect on fertility, as measured by the frequency of males producing pregnancies. Ethanol and Trenimon treatments produced approximately 3- and 7-fold increases, respectively in the frequencies of preimplantational loss over that seen for the control group (7.3%), resulting in significant ethanol and Trenimon effects (p less than 0.001). No interactive effects of ethanol and Trenimon treatments were noted. Frequencies of dead fetuses per pregnancy in the ethanol- and Trenimon-treated groups were increased approximately 2.5- and 4-fold, respectively, over the control value of approximately 16%. However, the effect of combined treatments was not greater than that due to Trenimon alone, resulting in Trenimon and ethanol effects (p less than 0.001) and ethanol-Trenimon interaction (p less than 0.001). The calculated mutation index resulting from each treatment yielded significant (p less than 0.001) ethanol- and Trenimon-induced effects. In contrast to effects of ethanol and Trenimon treatments, THC, given alone, or in combination with ethanol and/or Trenimon, had no effect on either preimplantational loss, fetal mortality or the resulting mutation index. The data suggest that chronic ethanol treatment, at levels resulting in minimal fertility impairment, increases the frequency of dominant lethal mutations. In contrast, chronic treatment with THC, as administered in the present study, appears to be without effect.(ABSTRACT TRUNCATED AT 400 WORDS)     

Lethal and mutagenic action of N-nitroso-N-methylbiuret on the producers of levorin, amphotericin and mycoheptin]

The lethal and mutagenic effect of N-nitrozo-N-methylbiuret (NMB) on the organisms producing levorin, amphotericin B and mycoheptin was studied. The mutagen effect depended on the dose, culture and physiological state of the spores. NMB had a low mutagenic effect on the levorin-producing organism characterized by high activity and genetic homogenicity with respect to the colony morphology and antibiotic production. As for the organisms producing amphotericin B and mycoheptin characterized by high genetic heterogenicity, significant variation of all the features studied was observed on their exposure to the mutagen. Inspite of diverse reaction of the organisms producing levorin, amphotericin B and mycoheptin to the effect of NMB mutants with increased antibiotic production were obtained from the three cultures. The lethal and mutagenic effect of NMB on the mycoheptin-producing organism depended on the process of the spore DNA replication. The spores during the DNA replication period were least sensitive to the lethal effect of the mutagen and most mutable with the respect to the colony morphology. For selection of highly active and stable strains exposure to NMB of the spores of the mycoheptin-producing organism during replication of DNA proved to be more effective than that of the spores during the lag-phase.     

Genetic control and mechanisms of salt and hyperosmotic shock resistance in cyanobacteria

Exposure to high concentrations of environmental NaCl exerts two stress effects on living cells, increasing the osmotic pressure and the concentration of inorganic ions. Salt stress dramatically suppresses the photosynthetic activity in cells of phototrophic organisms, such as cyanobacteria. During salt adaptation, cyanobacterial cells accumulate osmoprotectors, export excessive Na+ with the help of Na+/H+ antiporters, and actively absorb K+ with the help of K+-transporting systems. These physiological processes are accompanied by induction or suppression of several genes involved in salt adaptation. The review considers the main mechanisms responsible for the resistance of cyanobacterial cells to salt and hyperosmotic stresses. Special emphasis is placed on recent achievements in studying the genetic control of salt resistance and regulation of gene expression during adaptation of cyanobacteria to salt and hyperosmotic stresses.     

Induced Dominant Lethal Mutations and Cytotoxic Effects in Germ Cells of DROSOPHILA MELANOGASTER with Trenimon, Pdmt and Sodium Monofluorophosphate

Male and female Drosophila melanogaster with special sex chromosome or special autosome constitutions were fed with the mutagenic chemicals Trenimon (2,3,5-trisethyleneimino-1,4-benzoquinone) and PDMT (1-phenyl-3, 3-dimethyltriazene) and with the toxic substance Na2PO3F (sodium monofluorophosphate). The frequency of dominant lethality was recorded among the progeny. The results clearly show that dominant lethality is dose dependent for Trenimon- or PDMT-treated chromosomes in mature sperm and mature oocytes, and an increased amount of chromosomal material per nucleus yields an enhanced lethality. In contrast, a pure toxic effect of Na2PO3F on mature oocytes was demonstrated with one type of female. --With the stocks of Drosophila used, it is possible to distinguish between mutagenic and toxic effects of chemicals on the germ cells. Therefore, dominant lethality can be used as a simple and quick screening test for chemical mutagens.     

Evolution and development — the nematode vulva as a case study

To understand how morphological characters change during evolution, we need insight into the evolution of developmental processes. Comparative developmental approaches that make use of our fundamental understanding of development in certain model organisms have been initiated for different animal systems and flowering plants. Nematodes provide a useful experimental system with which to investigate the genetic and molecular alterations underlying evolutionary changes of cell fate specification in development, by comparing different species to the genetic model system Caenorhabditis elegans. In this review, I will first discuss the different types of evolutionary alterations seen at the cellular level by focusing mainly on the analysis of vulva development in different species. The observed alterations involve changes in cell lineage, cell migration and cell death, as well as induction and cell competence. I then describe a genetic approach in the nematode Pristionchus pacificus that might identify those genetic and molecular processes that cause evolutionary changes of cell fate specification.     

Genetic Control and Mechanisms of Salt and Hyperosmotic Stress Resistance in Cyanobacteria

Exposure to high concentrations of environmental NaCl exerts two stress effects on living cells, increasing the osmotic pressure and the concentration of inorganic ions. Salt stress dramatically suppresses the photosynthetic activity in cells of phototrophic organisms, such as cyanobacteria. During salt adaptation, cyanobacterial cells accumulate osmoprotectors, export excessive Na⁺ with the help of Na⁺/H⁺ antiporters, and actively absorb K⁺ with the help of K⁺-transporting systems. These physiological processes are accompanied by induction or suppression of several genes involved in salt adaptation. The review considers the main mechanisms responsible for the resistance of cyanobacterial cells to salt and hyperosmotic stresses. Special emphasis is placed on recent achievements in studying the genetic control of salt resistance and regulation of gene expression during adaptation of cyanobacteria to salt and hyperosmotic stresses.     

Molecular microbiology and pathogenesis of Helicobacter and Campylobacter updated: a meeting report of the 11th conference on Campylobacter,Helicobacter and related organisms

The genome analysis of the gastrointestinal pathogens Helicobacter and Campylobacter has stimulated a wealth of new research activities, which are presented every 2 years at the international conferences on Campylobacter, Helicobacter and Related Organisms (CHRO). Both organisms represent excellent models for the identification of new molecular mechanisms involved in pathogenesis, host response and physiological adaptation in course of acute and chronic infectious diseases. The investigation of their global distribution, pronounced genetic and antigenic diversity as well as the molecular mechanisms allowing long-term persistence in hostile and unusual microbial habitats, is a challenge for scientists of many different disciplines world-wide. With a focus on the molecular microbiology aspects, this review summarizes recent trends in Helicobacter and Campylobacter research by highlighting selected presentations at the 11th CHRO conference. The topics include the discovery of new virulence factors, functional analysis of protein secretion systems, host signalling pathways, adaptation to stress conditions, global gene regulation, and genetic variability.     

Epithelial cells in tissue culture

In this review the characteristics of established renal and intestinal epithelial cell lines are described by summarizing the accumulated literature about specific properties retained by the cells in tissue culture. Furthermore, brief examples are given for the use of cultured epithelia as model systems to study epithelial transport and metabolic functions, epithelial cell polarity, and aspects of the differentiation and maturation of epithelia by physiological, biochemical and genetic, or cell and molecular biological approaches.     

Bacterial systems for carcinogenicity testing

During the past 30 years, bacterial test systems have been extensively refined in their ability to detect not only mutagenic agents but, in many cases, carcinogenic ones as well. Since many carcinogens are known to be activated within the mammalian body, major improvements in bacterial test systems were made when representative parts of mammalian metabolism were included as part of the test protocol. Presently, systems of great simplicity and convenience are available for the efficient detection of gene mutations, lysogenic induction of prophages, and differential DNA repair. These qualities render bacterial systems potentially useful in distinguishing between carcinogens and non-carcinogens, in characterizing induced mutation spectra, and possibly in quantifying mutagenic potency that may be used to predict tumor-initiating potency. Sensitive strains of Salmonella typhimurium. Escherichia coli and Bacillus subtilis with altered DNA-repair capacities have been constructed which accurately identify many carcinogens. Comparative studies have shown that techniques using these strains can be standardized to some extent and that the majority of carcinogens are active in all adequately sensitive genetic systems. Because of this redundancy, it may be sufficient to employ only one standardized set of tester strains and methodology. However, serveral classes of known carcinogens are undetected or underestimated when assayed in standard testing procedures. Some of these chemicals can be efficiently recognized as mutagens upon varying the methodology, the genetic endpoint, or the mammalian activation system. Thus, to modify and adjust the experimental protocol to the particular type of chemical under study and to calibrate the system with appropriate carcinogenic and non-carcinogenic reference compounds is advisable. It is noteworthy that chemical carcinogens which probably act by non-genotoxic mechanisms thus far remain undetected in bacterial tests. Newly developed systems which measure specific types of genetic events, such as transpositions of DNA segments and derepression of genes, presently are being tested for their ability to detect such carcinogens. A final matter of growing concern is the increasing number of environmental chemicals that are found to be mutagenic in bacteria but for which information about carcinogenic activity in vivo is insufficient. The possible use of bacteria for quantifying mutagenic potency and extrapolating this information to tumor-initiating potency can be envisaged in three ways: (i) direct extrapolation from standard in vitro tests, (ii) indirect extrapolation making use of an in vitro/in vivo comparison of induced effects (the parallelogram method) as devised by Sobels [138] on the basis of identical dose (to DNA), and (iii) host-mediated assays to assess mutagenic potency of carcinogens in selected organs of mammals...     

The genetic toxicology of 2-amino-N6-hydroxyadenine in eukaryotic organisms: significance for genetic risk assessment

A collaborative study was designed to assess the mutagenicity of 2-amino-N6-hydroxylaminopurine (AHA) in a wide variety of eukaryotic assays systems in terms of potency and specificity. Earlier studies in Salmonella and Neurospora had shown that AHA was an extremely potent mutagen which appeared to cause predominantly AT to GC base-pair transitions. This discovery was viewed as an unusual opportunity to explore the general utility of different eukaryotic assay systems for genetic risk assessment. The objective was to determine whether AHA would show comparable potency and specificity in those eukaryotic organisms used to evaluate mutagenic potential of environmental chemicals for the human population. The data presented in this report show that AHA was mutagenic in all the eukaryotic assays utilized; however, the level of effect was found to be assay system-dependent. In addition, in assays where other base analogs were used as positive controls, differences in relative potency were observed from those obtained in the earlier studies with Salmonella and Neurospora. When alkylating agents were used as positive controls in the higher eukaryotic assays, AHA was found to have a mutagenic potency comparable to ethylnitrosourea (ENU), ethyl methanesulfonate (EMS) or methyl methanesulfonate (MMS) for many of the assays. With regard to mutagenic specificity, AHA appears to induce gene/point mutations in eukaryotic organisms, resulting predominantly from base-pair substitutions, predominantly AT to GC base-pair transitions; however, there was some unexplained variation in the ratio of these base-pair transitions and other transitions and transversions as a function of assay system. In addition, studies on the induction of micronuclei have shown that AHA induces chromosomal damage at high concentrations and low levels of survival.     

The lack of mutagenic properties of patulin and patulin adducts formed with cysteine in Salmonella test systems.

The mutagenic properties of patulin and the patulin adducts formed with cysteine were tested with histidine auxotroph Salmonella typhimurium strains as indicator organisms. The tests were performed by microsomal activation and host-mediated assay. Neither patulin nor patulin--cysteine reaction mixture was mutagenic in these test systems.     

Estimation of genetic effects of chronic exposure to low-dose rate gamma-radiation by cytogenetic methods and DNA-comet assay

The study of genetic effects in CBA/lac mice exposed for 1 year to constant low dose-rate gamma-radiation at a dose-rate 63 cGy/year has been carried out. We have shown the significant increase in the DNA breaks' level in spleen lymphocytes by comet-assay beginning from the total absorbed dose of 20 cGy. It is possible that the DNA breaks' level increase resulted from the structural rearrangement of chromatin or induction of lymphocyte proliferation. The results obtained by micronucleus test have proved that the mutagenic effect of chronic low dose-rate gamma-radiation depends on cell type and respectively on cell proliferation rate, cell differentiation, etc. So, by the end of experiment the significant increase in the frequency of PCE with micronuclei (MN) was observed. However, in contrast, the frequency of NCE with MN was not increased. No significant increase in the percent of lung cells with MN was registered.     

Fluoride: interaction with chemical mutagens in Drosophila

Simultaneous feeding of sodium fluoride and some chemical mutagens to Drosophila has been reported to reduce the yield of induced mutations compared with feeding the mutagens alone. This observation has been interpreted as a genuine case of antimutagenesis in which fluoride specifically inhibits the induction of chromosome breaks. An alternative hypothesis is that the presence of fluoride inhibits the uptake of the mutagen solutions, producing the same effect as an antimutagen, but for a trivial reason. We have tested this hypothesis using radioactive labelled sucrose to measure the uptake of test solutions. The results show that differential uptake can account for the "antimutagenic" effects reported in Drosophila. Comparison of recessive lethal frequencies induced by Trenimon and PDT do not support the hypothesis that fluoride has any genuine antimutagenic action. Antimutagenic effects of fluoride have been reported in other systems. We cannot exclude the possibility of some genuine effects but we consider that these reports should be critically re-examined in the light of our present findings.     

Mutants of the phototrophic bacteria Rhodobacter sphaeroides sensitive to the mutagenic action of long-wave ultraviolet light

The mutagenic action of near ultraviolet (NUV, greater than or equal to 280) nm) on purple phototrophic soil bacteria Rhodobacter sphaeroides: wild strain 2R and 12 mutants obtained earlier sensitive to UV derivates (UVS) was investigated. The mutagenic action of NUV was measured by induction of resistance to tetracycline (Tet) and nalidixic acid (Nal) and reversion of pigment mutants to wild-type phenotype. The NUV light induces the mutations of resistance to Nal and Tet in wild-type strain 2R; the UVS mutants differed greatly in their NUV-induced mutability. Three UVS mutants were characterized by greatly increased mutability in all analysed loci; slight mutability was found in seven mutants. On the basis of the data obtained it has been concluded that the UVS mutants R. sphaeroides can be used as test organisms in estimation of mutagenic activity of NUV. The molecular mechanisms and genetic control of NUV-induced mutagenesis are discussed.     

Prophage Induction in Lysogenic Escherichia coli with N-Nitroso Compounds and Derivatives

Prophage induction in lysogenic Escherichia coli W1709 (iota) was determined for 29 N-nitroso compounds, 13 of their denitrosated derivatives, and 7 hydroxylamino and hydrazino analogues of nitrosamines. Minimal inducing concentrations of 0.1 to 2.0 mug/ml were demonstrated for eight nitrosamidines, and concentrations of 0.5 to 25.0 mug/ml were shown for six nitrosamides. Weak inducing activities were found with N,N-diethylhydroxylamine oxalate and N-methyl-N-phenylhydrazine sulfate, derivatives of inactive N-nitrosodiethylamine and N-nitrosomethylphenylamine, respectively. Inactive compounds including N-methyl-N-nitroso-p-toluenesulfonamide, 11 nitrosamines, 3 N, N'-dialkyl substituted-N-nitrosoureas, 13 denitrosated derivatives, and 5 hydroxylamino and hydrazino analogues of nitrosamines are listed. Since 7 of the 14 prophage-inducing nitrosamidines and nitrosamides reported thus far have carcinostatic activity in rodent tumor systems, it is concluded that the induction test may provide a useful screen for the detection of potential antitumor compounds. The induction test may also be useful for the detection of responsive N-nitroso compounds which may be potential toxicological hazards in the environment since, of the six active nitrosamides, five have already been reported to produce mutagenic and carcinogenic effects, four produce chromosomedamaging effects, and two produce teratogenic effects. Use of the prophage induction system for detection of biologically active intermediates formed by N-nitroso compounds under physiological conditions is considered.     

A comparison of the CHO/HGPRT+ and the L5178Y/TK+/− mutation assays using suspension treatment and soft agar cloning: Results for 10 chemicals

The mouse lymphoma assay (MLA) and Chinese hamster ovary (CHO) cell assay are sensitive indicators of mutagenicity. The CHO assay has been modified technically to permit treatment in suspension and soft agar cloning comparable to the MLA. This methodology eliminates the risk of metabolic cooperation and the trauma of trypsinization. In addition, a larger population of cells can be treated and cloned for mutant selection. In order to compare the effectiveness of the test systems, 10 chemicals were evaluated for the induction of forward mutations in the CHO and MLA. Several of these chemicals have been reported as clastogenic; therefore, abbreviated colony sizing was performed to gauge the extent of genetic damage to the MLA cells. Both test systems detected benzo[a]pyrene, mitomycin C, acridine orange, and proflavin, and, with the exception of proflavin, more large colonies were present than small colonies. The suspect clastogen, phenytoin, was not mutagenic in the MLA and produced inconclusive results in the CHO. Ethidium bromide, a clastogen and a bacterial mutagen, was not mutagenic in either the MLA or CHO. Four compounds (p-aminophenol, benzoin, methoxychlor, and pyrene) were positive in the MLA, generally inducing a large number of small colonies, while demonstrating no mutagenic activity in the CHO assay. They have also been shown to be generally nongenotoxic in other test systems. Overall, the modified CHO assay did not appear to be better than the MLA for the detection of mutagenic agents. However, the MLA does appear to have lower specificity.Abbreviations AO acridine orange - BAP benzo[a]pyrene - BZN benzoin - CHO Chinese hamster ovary cell assay - DPH diphenylhydantoin - EB ethidium bromide - EMS ethylmethanesulfonate - 3MC 3-methylcholanthrene - MLA mouse lymphoma asay - MMC mitomycin C - MXC methoxychlor - PAP p-aminophenol - PRO proflavin - PYR pyrene     

The Theory of Speciation VIA the Founder Principle

The founder principle has been used to explain many instances of rapid speciation. Advances from theoretical population genetics are incorporated into Mayr''s original founder-effect genetic-revolution model to yield a newer model called the genetic transilience. The basic theoretical edifice lies upon the fact that founder event can sometimes lead to an accumulation of inbreeding and an induction of gametic disequilibrium. This, in turn, causes alleles to be selected more for their homozygous fitness effects and for their effects on a more stable genetic background. Selection occurring in multi-locus systems controlling integrated developmental, physiological, behavioral, etc., traits is particularly sensitive to these founder effects. If sufficient genetic variability exists in the founder population, such multilocus genetic systems can respond to drift and the altered selective forces by undergoing a rapid shift to a new adaptive peak known as the genetic transilience. A genetic transilience is, therefore, most likely to occur when the founder event causes a rapid accumulation of inbreeding without a severe reduction in genetic variability. The implications of this model are then examined for three aspects of the founder-effect genetic-transilience model: the attributes of the ancestral population, the nature of the sampling process used to generate the founders and the attributes of the founder population. The model is used to explain several features of the evolution of the Hawaiian Drosophila, and experimental designs are outlined to test the major predictions of the theory. Hence, this theory of speciation can be tested in the laboratory, using systems and techniques that already exist—a rare attribute of most models of speciation.