The induction of chromosome aberrations in mouse dictyate oocytes by X-rays and chemical mutagens.

Oocytes were collected from female mice and matured in vitro to Metaphase I during the first or third week after treatment with a dose of 400 rad X-rays, 1.6 mg/kg triethylenemelamine (TEM) or 75 mg/kg isopropylmethanesulphonate (IPMS). In week 1 the mean number of oocytes per female was similar for all treatments but in week 3 irradiated females yielded fewer oocytes than the chemically treated or control females. In week 1 the proportion of oocytes maturing was smaller in irradiated females than in the other groups but in week 3 was similar in all groups.Structural chromosome aberrations, scored in the Metaphase I oocytes, were of the chromatid or isochromatid type and involved gaps, breaks, fragments, intrachanges and interchanges. Aberration frequency did not increase with time after either of the chemical mutagens but after irradiation was higher in the third week than in the first week. The aberration yield from IPMS-treated females was similar at both sampling times, while a lower yield was recorded in week 3 following TEM treatment than in week 1.     

Comparison of radiation-and chemically-induced dominant lethal mutations in male mice

Ionizing radiation-induced dominant lethal mutations in all spermatogenic stages. After irradiation of male mice with 200 R the yield of induced mutations in early spermatids was twice the yield in spermatozoa, late spermatids, and spermatocytes. After irradiation with 400 R or 800 R the spermatocytes were the most sensitive stage for the induction of dominant lethal mutations. The frequency of radiation-induced dominant lethal mutations in postspermatogonial stages was dose-dependent. The yield of dominant lethal mutations in spermatogonia was independent of the dose.     

Micronuclei in mouse bone-marrow cells. A simple in vivo model for the evaluation of drug-induced chromosomal aberrations

For studying, in vivo, chromosomal damage in bone-marrow cells of CD mice the following compounds were used: Trenimon®; Endoxanm® (cyclophosphamide); triethylenemelamine (TEM); methyl methanesulfonate (MMS); ethyl methanesulfonate (EMS); mitomycin C; colchicine; N-methyl-N′-nitro-N-nitrosoguanidine (MNNG) and caffeine. In a first set of experiments the compounds were given twice intraperitoneally with an interval of 24 h. In a second set, effects on bone marrow were studied after 2 i.v. or p.o. administrations of TEM or EMS. All compounds except MNNG and caffeine produced bone-marrow depression and micronuclei, depending on the dose. For the active compounds an interesting difference was revealed by a comparison of the lowest effective dose (as measured by micronuclei formation) with the lethal dose. Trenimon, TEM, cyclophosphamide and MMS (some of which are used in human chemotherapy in similar mg/kg doses) were active on mouse bone-marrow at very low doses compared with their lethal doses. On the other hand, colchicine, mitomycin C and EMS exhibited an effect only at doses very close to, or within, the toxic range. Different routes of administration of either TEM or EMS produced similar effects.The results indicate that the test is especially suitable for initial large-scale screening of suspected chromosomal mutagens and spindle poisons. In addition, the use of the relationship between doses required to induce micronuclei and lethal doses in mice provides a practical measure of the relative potencies of such compounds.     

H-gene (histocompatibility) mutations induced by triethylenemelamine in the mouse

Based on a simple dermal-graft procedure, the H-test for histo-compatibility mutations screens the mouse “H-system” of not less than 29 and perhaps considerably more than 100 loci that are scattered throughout the genome. Graft-tests with normally heterozygous or hemizygous loci permit scoring mutations as gains, losses, and gains+losses. Tests with normally homozygous loci screen only for gains, but further analysis can detect accompanying losses. Assayed by the H-test, triethylenemelamine (TEM) increased the spontaneous mutation rate per generation by approximately four- to five-fold in the case of BALB/c spermatogonia and F₁ hybrid oocytes (BALB/c females × C57BL/6 males), but had a much smaller effect, if any, on the rate of C57BL/6 spermatogonia. Single intraperitoneal doses of 2.4–4.0 mg/kg were given.     

A comparison of mutation induction by 14 MeV fast neutrons and 137 Cs -rays in meiotic spermatocytes in the silkworm

Barley seeds were treated with methyl methanesulphonate (MMS) and ethyl methanesulphonate (EMS), stored at 15% water content and washed for 16–24 h. These treatments resulted in an increase of toxic and genetic effects. In teh DNA of embryos of such stored MMS- and EMS-treated seeds, a strong enhancement of the amount of single-strand breaks and/or alkali-labile sites took place. In contrast, the amount of alkylated sites, particularly of 7-methylguanine, was somewhat lower. It can be that the depurination and/or backbone breakage, which proceeds during the storage period, is responsible for the enhancement of toxic and genetic effects, whereas the influence of the alkylation of DNA during the storage period by the unreacted residual mutagen is negligible.     

Cytological analysis of storage effects on various types of complete and mosaic change induced in drosophila chromosomes by some chemical mutagens

Structural chromosomal changes induced by ethyleneimine (EI) in Drosophila spermatozoa and the effect of storage on the relative frequencies of mosaic and complete changes were studied cytologically, using salivary gland chromosomes. The pattern of EI-induced changes in unstored spermatozoa differes from that induced by X-rays by (1) a higher proportion of mosaics, (2) a higher proportion of repeats and deficiencies and a lower one of translocations and inversions, (3) a lower proportion of interchromosomal changes and a lower proportion of breaks located in heterochromatin. All of these characteristic feautures that distinguish mutagenic effects of EI from those of X-rays are present also after treatment with triethylene melamine (TEM) and formaldehyde food (FF), but the differences between EI and X-rays are on the whole smaller than those between X-rays and TEM or FF.Effect of storage on the frequencies of EI-induced changes is similar to that found when TEM of FF were used as mutagens: the whole pattern of changes became more like that found after irradiation. The only qualitative difference between EI and TEM of FF is the way in which storage affects the frequencies of mosaics.The peculiar pattern of EI-, TEM-, or FF-induced changes and the effect of storage is attributed to the high proportion of pre-breakage lesions with delayed fixation of breaks.Residual-mutagen hypothesis, as the possible cause of storage effects is also discussed.     

Assessment of mutagenicity induced by MMS and DES in Capsicum annuum L.

Seeds of Capsicum annuum L. var. G4 were subjected to different concentrations of methyl methane sulphonate (MMS) and diethyl sulphate (DES). The effects of different mutagenic treatments on meiosis, chiasma frequency, and pollen fertility have been studied in M₁ generation. Various types of meiotic aberrations such as univalent, multivalent, stickiness, bridge, laggards, cytomixis etc. were observed in all the treatments. However, the MMS treatments proved to be more effective in inducing meiotic aberrations as compared to DES. Moreover, the frequency of meiotic aberrations was at its maximum at metaphase followed by anaphase and telophase stages. As the concentrations increase, reduction in chiasma frequency and pollen fertility was observed in all the treatments and, MMS again was found to be more effective than DES treatments.     

Thiocyclam does not induce structural chromosome aberrations in human lymphocytes in vitro

Thiocyclam (trade name Evisect) is a broad-spectrum nereistoxin analogue insecticide used widely for agricultural applications. The aim of this investigation was to determine its genotoxic effects in the chromosome aberration (CA) test and determining of mitotic index (MI), using lymphocytes from peripheral blood samples of healthy human donors. A negative and a positive control (MMC) were also included. Chromosomal analyses of the metaphase plates of the samples treated with 14 different concentrations (from 0.1 to 120 μg/ml) of thiocyclam, indicating the lack effect on chromosomes. Thus thiocyclam is not genotoxic but highly toxic on cell proliferation in human lymphocytes.     

Comparison of the level of sister chromatid exchanges and chromosome aberrations induced by chemical mutagens in vitro and in vivo

The paper presents results of a study of a dose dependence of induction of SCE and chromosomal aberrations at the exposure of human lymphocytes in vitro and bone marrow cells of mice in vivo to 5 alkylating chemicals. The efficiency of SCE induction in vitro is found to be 300-30 times as high as that of arising of chromosomal aberrations. The same regularity is observed in experiments in vivo, but the ratio is reduced to 60-20 times.     

Protective effects of solvent fractions of Mentha spicata (L.) leaves evaluated on 4-nitroquinoline-1-oxide induced chromosome damage and apoptosis in mouse bone marrow cells

Spearmint leaves (Mentha spicata L.) contain high levels of antioxidants that are known to protect against both exogenous and endogenous DNA damage. In this study, the protective effects of the hexane fraction (HF), chloroform fraction (CF) and ethyl acetate fraction (EAF) in an ethanol extract from M. spicata were evaluated against 4-nitroquinoline-1-oxide (4-NQO) induced chromosome damage and apoptosis in bone marrow cells of Swiss albino mice. Two (EAF; 80 and 160 mg/ kg body weight - bw) or three (HF and CF; 80, 160 and 320 mg/ kg bw) doses of solvent fractions or vehicle control (25% DMSO in water) were administered orally for five consecutive days. Upon the sixth day, 4-NQO was injected intraperitoneally. The animals were killed the following day. Other control groups were comprised of animals treated with either the vehicle control or the various doses of solvent fractions, but with no 4-NQO treatment. 4-NQO induced micro-nucleated polychromatic erythrocytes (MnPCEs) in all the test groups. However, pre-treatment of animals with the solvent fractions significantly reduced the 4-NQO-induced MnPCEs as well as the percentage of apoptotic cells. The reduction of both MnPCE and apoptosis was more evident following the pre-treatment of animals with 160 mg/kg bw EAF.     

Blood chemical changes and renal histological alterations induced by gentamicin in rats

Gentamicin is an effective widely used antibiotic, but the risk of nephrotoxicity and oxidative damage limit its long-term use. Hence, the current study aims to elucidate such hazardous effects. To achieve the study aim male Wistar albino rats (Rattus norvegicus) were exposed to gentamicin to investigate the resultant blood chemical changes and renal histological alterations. In comparison with control rats, gentamicin produced outstanding tubular, glomerular and interstitial alterations that included degeneration, necrosis, cytolysis and cortical tubular desquamation together with mesangial hypercellularity, endothelial cell proliferation and blood capillary congestion. Compared with control animals significant blood chemical changes (P < 0.05) including free radicals, ALT, AST, ALP, serum creatinine and serum urea were recorded in gentamicin-injected animals. The findings revealed that exposure to gentamicin can induce significant histological alterations in the kidney as well as remarkable blood chemical changes that might indicate marked renal failure.     

Effects of caffeine on chromosome aberrations and sister-chromatid exchanges induced by mitomycin C in BrdU-labeled human chromosomes.

The BrdU-Hoechst staining technique has been used in analyzing the effect of caffeine (CAF) on chromosome aberrations and sister-chromatid exchanges (SCEs) induced by mitomycin C (MC). CAF increased the frequency of SCE in MC-treated chromosomes in all specimens. The combination of MC and CAF caused a remarkable increase in all types of chromosome aberrations, but the most startling effect was the appearance of many cells with multiple aberrations (shattered chromosomes). The BrdU-Hoechst technique showed that the shattered chromosomes did not appear in cells that had replicated only once, but did occur in cells which replicated twice in the presence of MC and CAF. The large majority of chromatid breaks observed did not involve areas common to SCE; and the SCE frequency significantly increased in spite of the existence of multiple breaks. This indicates that very few of the breaks are incomplete exchanges and that the mechanism for formation of SCE might be different from that of chromosome breaks. In another experiment, monofunctional-MC (M-MC) had a small effect on SCE rates, though it induced shattered chromosomes with CAF post-treatment. Possible differences in the mechanisms leading to SCE and chromosome breaks are discussed.     

A simple rule for the evolution of contingent cooperation in large groups

Humans cooperate in large groups of unrelated individuals, and many authors have argued that such cooperation is sustained by contingent reward and punishment. However, such sanctioning systems can also stabilize a wide range of behaviours, including mutually deleterious behaviours. Moreover, it is very likely that large-scale cooperation is derived in the human lineage. Thus, understanding the evolution of mutually beneficial cooperative behaviour requires knowledge of when strategies that support such behaviour can increase when rare. Here, we derive a simple formula that gives the relatedness necessary for contingent cooperation in n-person iterated games to increase when rare. This rule applies to a wide range of pay-off functions and assumes that the strategies supporting cooperation are based on the presence of a threshold fraction of cooperators. This rule suggests that modest levels of relatedness are sufficient for invasion by strategies that make cooperation contingent on previous cooperation by a small fraction of group members. In contrast, only high levels of relatedness allow the invasion by strategies that require near universal cooperation. In order to derive this formula, we introduce a novel methodology for studying evolution in group structured populations including local and global group-size regulation and fluctuations in group size.     

Test for positional candidate genes for body composition on pig chromosome 6

One QTL affecting backfat thickness (BF), intramuscular fat content (IMF) and eye muscle area (MA) was previously localized on porcine chromosome 6 in an F₂ cross between Iberian and Landrace pigs. This work was done to study the effect of two positional candidate genes on these traits: H-FABP and LEPR genes. The QTL mapping analysis was repeated with a regression method using genotypes for seven microsatellites and two PCR-RFLPs in the H-FABP and LEPR genes. H-FABP and LEPR genes were located at 85.4 and 107 cM respectively, by linkage analysis. The effects of the candidate gene polymorphisms were analyzed in two ways. When an animal model was fitted, both genes showed significant effects on fatness traits, the H-FABP polymorphism showed significant effects on IMF and MA, and the LEPR polymorphism on BF and IMF. But when the candidate gene effect was included in a QTL regression analysis these associations were not observed, suggesting that they must not be the causal mutations responsible for the effects found. Differences in the results of both analyses showed the inadequacy of the animal model approach for the evaluation of positional candidate genes in populations with linkage disequilibrium, when the probabilities of the parental origin of the QTL alleles are not included in the model.     

Homoeologous chromosome pairing between the A and B genomes of Musa spp. revealed by genomic in situ hybridization

Background and Aims

Most cooking banana and several desert bananas are interspecific triploid hybrids between Musa acuminata (A genome) and Musa balbisiana (B genome). In addition, M. balbisiana has agronomical characteristics such as resistance to biotic and abiotic stresses that could be useful to improve monospecific acuminata cultivars. To develop efficient breeding strategies for improving Musa cultivars, it is therefore important to understand the possibility of chromosome exchange between these two species.

Methods

A protocol was developed to prepare chromosome at meiosis metaphase I suitable for genomic in situ hybridization. A series of technical challenges were encountered, the main ones being the hardness of the cell wall and the density of the microsporocyte''s cytoplasm, which hampers accessibility of the probes to the chromosomes. Key parameters in solving these problems were addition of macerozyme in the enzyme mix, the duration of digestion and temperature during the spreading phase.

Results and Conclusions

This method was applied to analyse chromosome pairing in metaphase from triploid interspecific cultivars, and it was clearly demonstrated that interspecific recombinations between M. acuminata and M. balbisiana chromosomes do occur and may be frequent in triploid hybrids. These results provide new insight into Musa cultivar evolution and have important implications for breeding.     

A strategy for generation and balancing of autosome: Y chromosome translocations

We describe a method for generation and maintenance of translocations that move large autosomal segments onto the Y chromosome. Using this strategy we produced (2;Y) translocations that relocate between 1.5 and 4.8 Mb of the 2^nd chromosome.. All translocations were easily balanced over a male-specific lethal 1 (msl-1) mutant chromosome. Both halves of the translocation carry visible markers, as well as P-element ends that enable molecular confirmation. Halves of these translocations can be separated to produce offspring with duplications and with lethal second chromosome deficiencies . Such large deficiencies are otherwise tedious to generate and maintain.     

A new mouse mutant for the LDL receptor identified using ENU mutagenesis

In an effort to discover new mouse models of cardiovascular disease using N-ethyl-N-nitrosourea (ENU) mutagenesis followed by high-throughput phenotyping, we have identified a new mouse mutation, C699Y, in the LDL receptor (Ldlr), named wicked high cholesterol (WHC). When WHC was compared with the widely used Ldlr knockout (KO) mouse, notable phenotypic differences between strains were observed, such as accelerated atherosclerotic lesion formation and reduced hepatosteatosis in the ENU mutant after a short exposure to an atherogenic diet. This loss-of-function mouse model carries a single base mutation in the Ldlr gene on an otherwise pure C57BL/6J (B6) genetic background, making it a useful new tool for understanding the pathophysiology of atherosclerosis and for evaluating additional genetic modifiers regulating hyperlipidemia and atherogenesis. Further investigation of genomic differences between the ENU mutant and KO strains may reveal previously unappreciated sequence functionality.