Allium cepa assay based comparative study of selected vegetables and the chromosomal aberrations due to heavy metal accumulation

Irrigation of industrial effluents may end in the bioaccumulation of various toxic metals and consequent genetic changes in contaminated food crops. To test this hypothesis and extent of genetic modifications, Allium cepa test was performed to food crops viz. tomato (Lycopersicum esculentum) and chili (Capsicum annum) as Allium cepa test is a useful tool to assess genetic variations in plants. Prior to A. cepa test, the plants were exposed to various metal concentrations 125–1000 mg/L in the synthetic wastewater. The extracts of harvested plants were used to grow the root of A. cepa following its standard method. The root tips were fixed, stained and examined under compound microscope (almost 300–400 dividing cells) to check the extent of chromosomal variations during various stages of mitosis. The results revealed various chromosomal abnormalities including laggards, stickiness, vagrant chromosomes, binucleated cells, nuclear lesions, giant cells and c-mitosis at different level of treatment. On the whole, aberrations were increasing with the increasing doses along the positive control. In comparison, chili crop had higher level of aberrations depicting the higher chromosomal changes. Lower mitotic index (MI) with increasing level of doses was also describing the hampered cell division due to increased metal stress. The study is showing that the cell division was ceased with increasing metal stress thus increasing the rate of cell aberrations.     

Virus induced chromosomal abnormalities in Chinese hamster lung cell line and human peripheral blood leukocyte culture

Chinese hamster lung (CHL) cells were susceptible to Herpes Simplex type-1 and Chandipura viruses; which induced chromosomal abnormalities in these cells. Chromosomal changes induced in these cells were specific. The cells were refractory to measles virus and chromosomal abnormalities were not detected after inoculation of the virus. On the other hand human peripheral blood (HPB) leukocytes were susceptible to all the 3 viruses studied and exhibited chromosomal abnormalities upon infection. The aberrations induced in HPBL cultures were random. The results suggest that a virus could induce chromosomal changes only in susceptible cells. This is the first report of comparative in vitro study on chromosomes.     

Cytological characterization of transgenic soybean

 Some of the transgenic soybean [Glycine max (L.) Merr.] plants produced by bombarding embryogenic suspension cultures with DNA-coated particles exhibit morphological aberrations, including stunted plant growth, leathery dark green leaves and partialto-total seed sterility. In general, cultures from two Asgrow soybean lines (A2242, A2872) that were maintained for 8 months or longer produced primary transformants with reduced fertility. Cytological examination (mitotic pro-metaphase to metaphase chromosomes) of cells of suspension cultures, of roots from germinating somatic embryos, and of plants (R₀ and R₁) derived from A2242, revealed, besides diploidy (2n=40), various chromosomal aberrations such as deletions, duplications, trisomics and tetraploidy. Diploid transgenic plants with a normal karyotype from A2242 generally exhibited good fertility. No chromosomal abnormalities were observed in A2872-derived plants. However, plants regenerated from relatively old cultures of A2872 (more than 1 year in culture) showed a range of phenotypic abnormalities although they all contained 2n=40 chromosomes. These results indicate that soybean genotypes differ in their susceptibility to chromosomal instability induced by tissue culture. Therefore, chromosome analysis of cell cultures and the plants derived from them can help eliminate chromosomally and genetically abnormal material from gene-transfer experiments. Received: 6 June 1997/Accepted: 9 October 1997     

Alterations in p16 and p53 genes and chromosomal findings in patients with lung cancer: Fluorescence in situ hybridization and cytogenetic studies

Background: Chromosomal aberrations and instability of gene(s) are two factors related to the genetic instability of cancer cells. A loss of the tumor-suppressor function of the genes p16 and p53 is the most common event leading to the development of human cancers. Carcinoma of the lung is the leading cause of cancer deaths in the world. Chromosomal abnormalities in lung cancer may provide a valuable clue to the identification of target loci and culminate in a successful search for the major genes. The aim of this study was to investigate (i) alterations of the p16 and p53 genes and (ii) chromosomal aberrations in patients with small cell and non-small cell lung cancer by fluorescence in situ hybridization (FISH) and cytogenetic studies. Methods: We carried out cytogenetic analysis by Giemsa-banding in 18 cases. FISH probes for the p16 and p53 genes were also used on interphase nuclei to screen the alterations in these genes in lung cancer (LC).Results: We observed a high frequency of losses of the p16 – in 8/18 (44%) – and p53 – in 7/18 (39%) – genes in the cases with LC. A total of 18 patients showed predominantly numerical and structural aberrations. Among these two types, structural aberrations predominated and usually consisted of deletions, breaks, and fragilities in various chromosomes. Both structural and numerical changes were observed in almost all patients. Chromosomes 3 and 1 were found to be most frequently involved in structural abnormalities, followed by chromosomes 6, 9, and 8. Autosomal aneuploidies were also observed to be the most frequent (chromosomes 22, 19, 18, 20, 9, and 17), followed by those of the X and Y chromosomes. The expression of fragile sites was also found to be significantly higher in seven chromosomal regions: 3p14, 1q21, 1q12, 6q26, 9q13, 8q22, and 8q24. Conclusion: Our data confirmed that DNA damage and genomic instability may be factors contributing to the mutation profile and development of lung cancer. The patients who developed lung cancer showed a high frequency of loss of both p16 and p53, in addition to chromosomal aberrations. Tobacco could be a major carcinogenic factor in lung-cancer progression. The loss of p16 and p53, and increased incidence of autosomal aneuploidy and chromatid breaks, along with other chromosomal alterations, can contribute to the progression of the disease.     

Chromosomal aberrations and SCEs in Allium cepa root-tip cells treated with caffeine and pyronin Y

The effectiveness of caffeine and pyronin Y in the induction of both chromosomal aberrations and sister-chromatid exchanges (SCEs) in root meristematic cells of A. cepa was studied.The rate of SCEs proved to be increased when 5-bromo-2′-deoxyuridine- (BrdU) substituted chromosomes were allowed to replicate in thymidine (dT) for a second S period simultaneously with caffeine or pyronin Y. In contrast, only caffeine was able to induce aberrations in BrdU-substituted chromosomes, while pyronin Y seemed to be ineffective at the doses employed.     

Results of a Cytogenetic Study of Populations with Different Radiation Risks in the Semipalatinsk Region

A cytogenetic study was conducted for the first time on human populations neighboring the Semipalatinsk nuclear test site (STS) and exposed to ionizing radiation for a long period of time. In populations with the extreme and maximum radiation risks, high frequencies of radiation-induced chromosomal markers, including acentric fragments (1.99 ± 0.10 per 100 cells), dicentrics (0.23 ± 0.01), ring chromosomes (0.38 ± 0.14), and stable chromosomal aberrations (1.17 ± 0.02), were found. These frequencies significantly exceeded those in control populations. The spectrum of chromosomal aberrations and the frequencies of the aberrations of different types in persons living in the areas with the highest radionuclide contamination confirmed the mutagenic effect of radiation on chromosomes in the human populations studied.     

Results of a cytogenetic study of populations with different radiation risks in the Semipalatinsk region

A cytogenetic study was conducted for the first time on human populations neighboring the Semipalatinsk nuclear test site (STS) and exposed to ionizing radiation for a long period of time. In populations with the extreme and maximum radiation risks, high frequencies of radiation-induced chromosomal markers, including acentric fragments (1.99 +/- 0.10 per 100 cells), dicentrics (0.23 +/- 0.01), ring chromosomes (0.38 +/- 0.14), and stable chromosomal aberrations (1.17 +/- 0.02), were found. These frequencies significantly exceeded those in control populations. The spectrum of chromosomal aberrations and the frequencies of the aberrations of different types in persons living in the areas with the highest radionuclide contamination confirmed the mutagenic effect of radiation on chromosomes in the human populations studied.     

A study of the cytogenetics of the human cell strain WI-38

Summary Cultures of the human diploid cell strain WI-38 were subcultivated under conditions which would meet the requirements proposed for the use of this strain as a substrate for the preparation of viral vaccines and would be in keeping with efficient production procedures. For chromosomal analysis, the cultures were combined in three groups at low, intermediate, and high passage levels, the latter being beyond those recommended for vaccine production. At all passage numbers, the incidence of aneuploid cells was low and constant up to those passages where the finite life span was approached and the population doubling time became markedly prolonged. At all passage levels, the incidence of gaps was higher than that of breaks but there was no significant increase of either of these abnormalities with continuous subcultivation. Among structural abnormalities dicentrics, despiralizations and deletions predominated. A significant increase in polyploidy occurred in the highest passage numbers, although the ratio of polyploidy to endoreduplication was constant throughout the series. Neither heteroploid transformation nor nonrandom chromosomal aberrations were observed. Nor was there correlation between observed aberrations and their location on the chromosomes. The incidence of hypodiploidy was lower than reported by other investigators. At the cellular level, no morphological changes could be associated with the distribution of chromosomal aberrations. This study was assisted by funds provided by Canadian Public Health Research Grant 605-7-710 of the National Health Grants Programme.     

Griseofulvin: A potential agent of chromosomal segregation in cultured cells

The fungicidal compound griseofulvin (GF) induces abnormalities in nuclear division in mammalian cells cultured in vitro. For these properties it has been studied as a potential agent of chromosomal segregation. A marked effect on the dynamics of chromosomal complements was observed both on diploid and heteroploid cell lines, including hybrids produced by fusion. After treatment for three days with doses ranging from 40 to 60 μg/ml, according to the cell type, a tendency to a doubling of the chromosomal set was evident. When cells were allowed to recover in normal medium in the absence of GF a scattering of the distribution of the chromosomal numbers occured. After removal of the drug a selective advantage of the double chromosomal complements was observed on prolonged cultures. The possibility of using GF to induce chromosomal segregation for linkage studies and for chromosomal assignment is discussed.     

Comparison of the effects of radiation and deoxyribonucleic acids on the mitosis in roots ofVicia faba

Differences as well as similarities in the action of ionizing radiation and deoxyribonucleic acids from various sources on mitosis in root cells ofVicia faba were established. The time course of occurrence of aberrations were examined. Whereas in irradiated broad beant the maximum percentage of aberrations was observed immediately after irradiation, the aps plication of non-isologous DNA was followed by maximum aberrations after 8–16 hours. As all the time-intervals studied, an incraasad number of aberrations was found during metaphase-as compared with anaphases, both after irradiation and after application of DNA. A comparison of isologous, homologous and heterologous DNA as inductors of chromosomal aberrations supported our previous findings and showed that the efficiency of DNA depends on the genetic difference between donor and acceptor. During a study of distribution of aberrations between large and small chromosomes of meristematic cells ofVicia faba, at various time-intervals it was obsarved that after irradiation the distribution of aberrations between individual chromosomes is proportional to their total length, whereas the effect of heterologous DNA is mostly in the damage to small chromosomes. It was also found that aftar irradiation mostly chromatid aberrations are formed at shorter time-intervals and only later chromosomal aberrations will appear. On the other hand, heterologous DNA brings about in all time-intervals a predominance of chromatid aberrations.     

Studies on the chromosomal rearrangements induced by the male recombination factor 31.1 MRF in Drosophila melanogaster

A cytological analysis was carried out on the salivary-gland chromosomes of third-instar larvae deriving from appropriate crosses of the 31.1/CyL⁴ strain with the dp b cn bw and dp b cn bw; ve strains.(1) The 31.1 MRF induced chromosomal rearrangements in both male and female germ cells. (2) The aberrations that occurred in the male germ cells were of three types — inversions, duplications and deficiencies — whereas those that occurred in the female germ cells were of all types. (3) The distributions of the break-points among the 4 major autosomal arms between the 1F₂ and 2AF₂ larvae were different. (4) The total distribution of the break-points along the polytene chromosomes showed that the factor induced breaks in all major chromosomes (X, II, III), and that 51.81% of them occurred in sites showing late replication and/or ectopic pairing. (5) A relationship existed between the frequencies of the second chromosomal aberrations and male recombination.A comparison of the break-points induced by the factor was made with those found in the rare endemic inversions of the same natural population from which the 31.1 MRF was isolated. The possibility that the factor acts in the natural population is discussed.     

Interphase in situ hybridization to disaggregated and intact tissue specimens of prostatic adenocarcinomas

A comparative study was performed of interphase in situ hybridization (ISH) to deparaffinized 4-m tissue sections and nuclear suspensions from eight prostatic adenocarcinomas, as well as one normal prostatic control. Whole nuclear suspensions were derived from the same tumor areas to evaluate differences of ISH to truncated versus whole nuclei. DNA probes specific for the centromeres of chromosome 1, 7, 8, 10, and Y were used for detection of numerical chromosomal changes and aneuploidy. In six adenocarcinomas chromosome aberrations (+7, +8, –8, –10, –Y) were seen. However, ISH to sections revealed focal aberrations (–10, –Y) in four cases that could not be distinguished in the suspensions. Chromosomal alterations occurring in larger tumor areas were also detected in the nuclear suspensions. Chromosome copy number changes, especially gains, were better discriminated in the nuclear suspensions. The rate of ISH aneuploidy seen in nuclear suspensions corresponded with that observed in the tissue sections (P<0.01). Ploidy patterns as assessed by ISH to sections and nuclear suspensions were in concordance with DNA flow cytometry (bothP<0.001). We conclude that both section and suspension ISH were able to accurately detect aneuploidy and numerical chromosomal aberrations occurring in larger histological areas. However, section ISH was also capable of revealing (small) focal cytogenetic abnormalities, due to a precise analysis of only target cells. Focal abnormalities were not detected by suspension ISH, probably due to an admixture of non-aberrant tumor cells and stromal elements.     

The influence of exogenous DNA of different origin on the mitosis and chromosomes of irradiated meristematic cells ofVicia faba

In this paper we compare the influence of heterologous and isologous DNA on the radiation damage repair of primary root meristematic cells ofVicia faba. Roots, irradiated by exposure of 150 r were cultivated at different time intervals either in tap water, or in a solution of heterologous or isologous DNA. In comparing mitotic activity of meristematic cells it was found that both types of DNA studied enhance the recovery of irradiated cells. The frequency of postmetaphase chromosomal aberrations of irradiated cells was influenced also by post-irradiation action of exogenous DNA. While heterologous DNA exhibited synergical effect with radiation in the sense that it increased the post-irradiation incidence of aberrations in all time intervals studied, isologous DNA had a strong repair effect—the application caused a significant decrease of the percentage of post-metaphase aberrations. Both kinds of DNA caused changes in the relation of chromosome to chromatid aberrations; a higher percentage of chromatid aberrations was registered. The study of the distribution of aberrations between large and small chromosomes ofVicia faba showed that the post-irradiation application of heterologous DNA increases damage of small chromosomes while isologous DNA caused an increased repair ability in this chromosomal group.     

Chromosomal abnormalities in maternal and fetal tissues of magnesium- or zinc-deficient rats.

The effect of dietary deficiency during pregnancy of zinc or magnesium on maternal and fetal chromosomes was studied. Pregnant rats were given a zinc-deficient or a magnesium-deficient diet from the beginning of pregnancy and maternal bone marrow and fetal liver were removed on day 19 of gestation. Chromosome spreads were prepared and metaphases examined for abnormalities. Both magnesium- and zinc-deficient maternal bone-marrow and fetal liver cells showed significantly more chromosomal abnormalities than did those of controls. The chromosomal aberrations occurring in highest incidence in magnesium-deficient animals were terminal deletions and fragments. A higher than normal incidence of "stickiness" was also observed in cells from magnesium-deficient animals. In zinc-deficient animals, on the other hand, the chromosomal aberrations with the highest incidence were gaps and terminal deletions.     

Chromosomal evaluation in a group of Tunisian patients with non-obstructive azoospermia and severe oligozoospermia attending a Tunisian cytogenetic department

Male infertility is the cause in half of all childless partnerships. Numerous factors contribute to male infertility, including chromosomal aberrations and gene defects. Few data exist regarding the association of these chromosomal aberrations with male infertility in Arab and North African populations. We therefore aimed to evaluate the frequency of chromosomal aberrations in a sample of 476 infertile men with non-obstructive azoospermia (n = 328) or severe oligozoospermia (n = 148) referred for routine cytogenetic analysis to the department of cytogenetics of the Pasteur Institute of Tunis. The overall incidence of chromosomal abnormalities was about 10.9%. Out of the 52 patients with abnormal cytogenetic findings, sex chromosome abnormalities were observed in 42 (80.7%) including Klinefelter syndrome in 37 (71%). Structural chromosome abnormalities involving autosomes (19.2%) and sex chromosomes were detected in 11 infertile men. Abnormal findings were more prevalent in the azoospermia group (14.02%) than in the severe oligozoospermia group (4.05%). The high frequency of chromosomal alterations in our series highlights the need for efficient genetic testing in infertile men, as results may help to determine the prognosis, as well as the choice of an assisted reproduction technique. Moreover, a genetic investigation could minimize the risk of transmitting genetic abnormalities to future generations.     

The correlation between the frequency of micronuclei and specific chromosome aberrations in human lymphocytes exposed to microwave radiation in vitro.

Human whole-blood samples were exposed to continuous microwave radiation, frequency 7.7 GHz, power density 0.5, 10 and 30 mW/cm2 for 10, 30 and 60 min. A correlation between specific chromosomal aberrations and the incidence of micronuclei after in vitro exposure was observed. In all experimental conditions, the frequency of all types of chromosomal aberrations was significantly higher than in the control samples. In the irradiated samples the presence of dicentric and ring chromosomes was established. The incidence of micronuclei was also higher in the exposed samples. The results of the structural chromosome aberration test and of the micronucleus test were comparatively analyzed. The values obtained showed a positive correlation between micronuclei and specific chromosomal aberrations (acentric fragments and dicentric chromosomes). The results of the study indicate that microwave radiation causes changes in the genome of somatic human cells and that the applied tests are equally sensitive for the detection of the genotoxicity of microwaves.     

A novel system for correcting large-scale chromosomal aberrations: ring chromosome correction via reprogramming into induced pluripotent stem cell (iPSC)

Approximately 1 in 500 newborns are born with chromosomal abnormalities that include trisomies, translocations, large deletions, and duplications. There is currently no therapeutic approach for correcting such chromosomal aberrations in vivo or in vitro. When we attempted to produce induced pluripotent stem cell (iPSC) models from patient-derived fibroblasts that contained ring chromosomes, we found that the ring chromosomes were eliminated and replaced by duplicated normal copies of chromosomes through a mechanism of uniparental isodisomy (Bershteyn et al. 2014, Nature 507:99). The discovery of this previously unforeseen system for aberrant chromosome correction during reprogramming enables us for the first time to model and understand this process of cell-autonomous correction of ring chromosomes during human patient somatic cell reprograming to iPSCs. This knowledge could lead to a potential therapeutic strategy to correct common large-scale chromosomal aberrations, termed “chromosome therapy”.     

Differential chromosomal radiosensitivity within the first G1-phase of the cell cycle of early-dividing human leukocytes in vitro after stimulation with PHA

Summary Human leukocyte cultures were irradiated with 200 R X-rays before the addition of phytohemagglutinin (PHA) in the G₀-stage and at different times up to 25 h within the first G₁-phase of the cell cyle after the addition of PHA. The results of the analysis of chromosomal aberrations show that the frequencies of dicentric chromosomes increase significantly when leukocytes leave the G₀-stage, reaching a maximum yield of aberrations about halfway through the first G₁-phase. After that, toward the end of the G₁-phase, the frequencies of dicentric chromosomes decrease again, to a level similar to that found in the G₀-stage. Different possible explanations for the differential chromosomal radiosensitivity of human leukocytes within the first poststimulation G₁-phase are discussed.     

Effects of the bacterial algicide IRI-160AA on cellular morphology of harmful dinoflagellates

The algicide, IRI-160AA, induces mortality in dinoflagellates but not other species of algae, suggesting that a shared characteristic or feature renders this class of phytoplankton vulnerable to the algicide. In contrast to other eukaryotic species, the genome of dinoflagellates is stabilized by high concentrations of divalent cations and transition metals and contains large amounts of DNA with unusual base modifications. These distinctions set dinoflagellates apart from other phytoplankton and suggest that the nucleus may be a dinoflagellate-specific target for IRI-160AA. In this study, morphological and ultrastructural changes in three dinoflagellate species, Prorocentrum minimum, Karlodinium veneficum and Gyrodinium instriatum, were evaluated after short-term exposure to IRI-160AA using super resolution structured illumination microscopy (SR-SIM) and transmission electron microscopy (TEM). Exposure to the algicide resulted in cytoplasmic membrane blebbing, differing chloroplast morphologies, nuclear expansion, and chromosome expulsion and/or destabilization. TEM analysis showed that chromosomes of algicide-treated K. veneficum appeared electron dense with fibrous protrusions. In algicide-treated P. minimum and G. instriatum, chromosome decompaction occurred, while for P. minimum, nuclear expulsion was also observed for several cells. Results of this investigation demonstrate that exposure to the algicide destabilizes dinoflagellate chromosomes, although it was not clear if the nucleus was the primary target of the algicide or if the observed effects on chromosomal structure were due to downstream impacts. In all cases, changes in cellular morphology and ultrastructure were observed within two hours, suggesting that the algicide may be an effective and rapid approach to mitigate dinoflagellate blooms.     

THE MECHANISMS OF X-RAY EFFECTS ON CELLS

Irradiation of Tradescantia microspores does not increase subsequent sensitivity to x-rays as measured by the frequency of induced chromosomal aberrations curing the nuclear cycle. The slight decrease in sensitivity is to be expected because acentric fragments are less sensitive than the centric chromosomes. The physiological effects of x-rays appear to be of minor importance in causing injury or death of individual cells, and most of the deleterious effects can be attributed to "direct hits" which produce chromosomal alterations. In the reaction of tissues to x-rays the physiological effects may play a more important part.