Frequency Of Polyploid Cells Research Articles

Cytogenetic disorders in young cattle vaccinated against trichophytosis

This article continues a series of works by the authors devoted to most of the karyopathological effects of vaccines used in agriculture. The authors of the test cytogenetic analysis of the impact caused by the LTF130 vaccine from an attenuated culture of the fungus Trichophyton verrucosum TF-130 L VGNKI in peripheral blood lymphocytes of healthy 30-day-old healthy Holstein Colonial calves. The experiment was conducted on the material and technical basis of two departments: veterinary genetics and biotechnology, epizootiology and microbiology, FSBEI HE “Novosibirsk State Agrarian University”. The authors found that the spectrum of induced cytogenetic disorders included polyploid, hyper- and hypoploid cells, chromatid and chromosome breaks, and single and paired fragments of chromosomes, regardless of the time since the introduction of the vaccine. They did not differ from the spectrum of spontaneously occurring mutations in this species. It was noted that among cells with a changed number of chromosomes, polyploid cells with tetraploid sets (79.3–91.9%) and aneuploid cells with hypoploid sets (45.7–74.3%) of chromosomes predominated. The authors found a decrease in the frequency of polyploid cells seven days after vaccination of calves against trichophytosis to 1.6%, control - 3.4 ± 0.57% (P < 0.01). An increase in the frequency of hyperploid cells 14 days after vaccination of calves against trichophytosis up to 3.69% was revealed due to trisomy (2.91 ± 0.74%, P < 0.05) and tetrasomy (0.78 ± 0.39%, P < 0.01). An increase in the frequency of cells with chromosome aberrations was found 2 and 7 days after vaccination of calves against trichophytosis, respectively, to 5.74 and 5.24% due to breaks (4.49 ± 1.03 and 3.31 ± 0.66%, P < 0.01–0.01) and paired fragments of chromosomes (1.38 ± 0.43%, Р < 0.05).

CHROMOSOMAL PROFILE OF LARGE WHITE AND LAND BREED PIGS

This article presents the results of studies of karyotypic variability of pigs of two breeds - large white and landrace.The research was carried out in the Department of Animal Genetics and Biotechnology of the Institute of Animal Breeding and Genetics named after M.V. Zubtsia of the National Academy of Sciences. Karyotype analysis was performed on preparations of metaphase chromosomes obtained from peripheral blood lymphocytes according to the generally accepted method. The following cytogenetic indicators were included in the analysis: the frequency of aneuploid and polyploid cells, the frequency of cells with structural aberrations of chromosomes. Statistical processing of experimental data was carried out using Office Excel 2003.A comparative analysis of the karyotypes of two groups of pigs of the Great White and Landrace breeds shows the statistical reliability of the difference between the experimental groups of pigs in terms of the frequency of aberrant cells.The spectrum of numerical violations of the karyotype in all groups is represented by poly- and aneuploid cells. A higher frequency of polyploid cells is observed in pigs of the landrace breed compared to individuals of the large white breed.Almost twice as many aneuploid cells were recorded in landraces than in large white pigs).Analysis of the karyotypes of the studied groups of pigs revealed various chromosomal aberrations - associations and translocations of chromosomes, chromosomal breaks and fragments. Chromosome breaks in both groups of examined pigs occur with the same frequency - almost 3%. In the karyotypes of individual sows, we found two different centric fusions of chromosomes: in sow No. 7659 of the landrace breed, a Robertsonian translocation of rob 15/17, and in sow No. 2817 of the Big White breed, an association of chromosomes 16 and 17. These pigs were diagnosed with a reduced level of reproductive capacity, the cause of which, obviously, is a violation of the separation of chromosomes in meiosis. Among the studied animals, we did not find individuals with reciprocal translocations.In our study, in landraces, the frequency of premature separation of the centromeric regions of chromosomes in mitosis is 35,5% more frequent than in individuals of a large white breed, which indicates an unbalanced genotype of the animals of this studied group and indicates the peculiarities of selection work with them.The analysis of the pedigrees of the sows that were selected in the group for this study confirmed this assumption: the animals were obtained by interbreeding.Almost twice the frequency of polyploid cells confirms their belonging to animals of the meat type of productivity.Therefore, the analysis of polymorphism of karyotypic traits of pigs of the two studied groups of pigs of the Great White and Landrace breeds shows that the cytogenetic parameters of the studied pigs vary in the following range: the frequency of aberrant cells - from 5.11±0.05 to 7.63±0.17; frequency of polyploid cells - from 4.50±1.6 to 7.84±2.6; aneuploid - from 3.0±1.8 to 5.6±2.9; the frequency of chromosome breaks is from 2.8±1.3 to 2.9±1.7. It is obvious that the level of chromosomal instability is dominated by landrace pigs, the reason for which, in our opinion, is the peculiarities of breeding work with this breed.

Abstract PR15: Hypoxia, polyploidy, neuropeptide Y, and Ewing sarcoma bone metastases: Is there a link?

Abstract Ewing sarcoma (ES) is a pediatric tumor driven by EWS-ETS fusion proteins that carries dismal prognosis in its metastatic form. Nevertheless, the factors triggering metastatic processes in ES remain unknown. Even though ES is not considered genomically unstable, the presence of tumor cells with complex karyotypes and high chromosome number is one of a few known adverse prognostic factors, suggesting an acquired chromosomal instability (CIN) as a driver of ES progression. Moreover, worse patient survival correlates with tumor hypoxia, which associates with multiple metastases and dissemination to the bone. Interestingly, hypoxia is also a factor up-regulating neuropeptide Y (NPY) and its Y5 receptor (Y5R) in ES cells. Such an up-regulation has been shown to activate growth-promoting and prometastatic functions of the NPY in these tumors. Previously we have shown that: 1) metastatic progression of ES associates with increase in frequency of polyploid cells and CIN; 2) this effect is triggered by tumor hypoxia; 3) tumor hypoxia promotes formation of bone metastases and the large polyploid cells accumulate in the areas of bone invasion; 4) overactivation of NPY/Y5R pathway leads to cytokinesis defects and polyploidy; and 5) expression of NPY and Y5R increases in bone, but not soft tissue metastases. Based on these observations we hypothesize that polyploid tumor cells arising in hypoxic tumors are responsible for ES bone metastases and thereby poor clinical outcome observed in ES patients with hypoxic tumors. We also propose NPY/Y5R as a trigger of these processes. Thus, the goal of our study was to test the metastatic potential of ES polyploid cells and determine if blocking NPY/Y5R pathway will prevent their formation. To this end, we performed in vitro purification of the 4N-G1 phase cell population from hypoxic SK-ES-1 cell line using FUCCI technology. The progeny of these cells, SK-ES1-4N cells, was characterized for its tumorigenic and metastatic potential in vitro and in vivo. Using soft agar and proliferation assays in standard and low attachment plates, we have shown that under hypoxic conditions SK-ES1-4N cells have enhanced anchorage-independent growth and capability to form colonies. In an orthotopic xenograft model, tissues and cells derived from primary tumors and metastases exhibited broader range of cell and nuclear sizes and increased ploidy, as compared to the tumors from original SK-ES-1 cells, confirming their high CIN. Strikingly, in this model the SK-ES1-4N xenografts metastasized almost exclusively to bones. Bone lesions constituted 83% of total metastases in mice bearing SK-ES1-4N tumors, while only 25% of metastases from the original SK-ES1 xenografts were localized to the bones. To determine if the increase in CIN accompanying ES progression can be prevented by blocking NPY/Y5R pathway, the SK-ES1 primary tumors were treated with Y5R antagonists, CGP 71683. The Y5R blockade decreased frequency of polyploid cells in the large, hypoxic tumors to the levels observed in nonhypoxic ES. Altogether, our findings support the role for polyploidy in ES bone metastasis and implicate the hypoxia-induced activation of the NPY/Y5R axis as its potential trigger. This abstract is also being presented as Poster B34. Citation Format: Akanksha Mahajan, Sung-Hyeok Hong, Susana Galli, Congyi Lu, Shiya Zhu, Jason Tilan, Luciane Cavalli, Joanna Kitlinska. Hypoxia, polyploidy, neuropeptide Y, and Ewing sarcoma bone metastases: Is there a link? [abstract]. In: Proceedings of the AACR Conference on Advances in Sarcomas: From Basic Science to Clinical Translation; May 16-19, 2017; Philadelphia, PA. Philadelphia (PA): AACR; Clin Cancer Res 2018;24(2_Suppl):Abstract nr PR15.

Plant regeneration from callus culture of Curcuma aromatica and in vitro detection of somaclonal variation through cytophotometric analysis

Callus cultures initiated from shoot base explants of Curcuma aromatica Salisb. were maintained on Murashige and Skoog (MS) media supplemented with 2 mg dm−3 2,4-dichlorophenoxyacetic acid alone or with 0.5 mg dm−3 kinetin. Plantlets were regenerated from 60 and 180-d-old callus on MS media supplemented with 3 mg dm−3 benzyladenine and 0.5 mg dm−3 α-naphthalene acetic acid. Approximately 8–10 plantlets were produced after 30–40 d of culture per 50 mg of callus inoculated. Out of 113 regenerants analyzed 85 plants were exclusively diploid and 28 were predominantly diploid revealing presence of polyploid nuclei. Frequency of polyploid cells were more in regenerants obtained from 180-d-old callus then from 6-d-old callus which might be attributed to the ageing of callus.

Hepatic endopolyploidy as a cellular consequence of age‐specific selection for rate of development in mice

Endopolyploidy is the generation of polyploid cells by DNA replication without subsequent cell division and is correlated with hypertrophic growth or growth via cell size. Thus, selection that alters growth may also change onset and frequency of endopolyploidy as a correlated response. We search for endopolyploidy in the liver in response to age-specific restricted index selection for the rate of development. Polyploidy changes over ontogeny are described in five mouse lines: two selected for divergence in early growth (0-10 days of age), two selected for divergence in late growth (28-56 days of age), and one randombred control. Polyploid cell frequency within each line increased as ontogeny continued, as expected from previous research. However, selection for altered growth clearly plays a role in the frequency and onset of polyploid cells. Lines selected for divergence in early growth have polyploidy differences after weaning that are not seen in adult mice. However, lines selected for divergence in late growth are divergent in frequency of polyploid cells, starting near sexual maturity and continuing into adulthood.

Cytotoxicity and genotoxicity of low doses of mercury chloride and methylmercury chloride on human lymphocytes in vitro

Mercury is a xenobiotic metal that is a highly deleterious environmental pollutant. The biotransformation of mercury chloride (HgCl2) into methylmercury chloride (CH3HgCl) in aquatic environments is well-known and humans are exposed by consumption of contaminated fish, shellfish and algae. The objective of the present study was to determine the changes induced in vitro by two mercury compounds (HgCl2 and CH3HgCl) in cultured human lymphocytes. Short-term human leukocyte cultures from 10 healthy donors (5 females and 5 males) were set-up by adding drops of whole blood in complete medium. Cultures were separately and simultaneously treated with low doses (0.1 to 1000 microg/l) of HgCl2 and CH3HgCl and incubated at 37 degrees C for 48 h. Genotoxicity was assessed by chromosome aberrations and polyploid cells. Mitotic index was used as a measure of cytotoxicity. A significant increase (P < 0.05) in the relative frequency of chromosome aberrations was observed for all concentrations of CH3HgCl when compared to control, whether alone or in an evident sinergistic combination with HgCl2. The frequency of polyploid cells was also significantly increased (P < 0.05) when compared to control after exposure to all concentrations of CH3HgCl alone or in combination with HgCl2. CH3HgCl significantly decreased (P < 0.05) the mitotic index at 100 and 1000 microg/l alone, and at 1, 10, 100, and 1000 microg/l when combined with HgCl2, showing a synergistic cytotoxic effect. Our data showed that low concentrations of CH3HgCl might be cytotoxic/genotoxic. Such effects may indicate early cellular changes with possible biological consequences and should be considered in the preliminary evaluation of the risks of populations exposed in vivo to low doses of mercury.

The correlation between the chromosome variation in callus and genotype of explants of Arabidopsis thaliana.

Twelve callus lines of Arabidopsis thaliana were derived from four types of explants excised from diploid plants of two ecotypes (Columbia and Wilna) and autotetraploid plants of the Wilna ecotype. Cytogenetic analysis of the chromosome variation in particular callus lines was carried out for primary culture and callus during 5 months of culture. Ploidy levels of interphase nuclei were estimated by counting the number and size of chromocentres and nuclei of interphase cells. The first polyploid cells in all callus lines were observed during callogenesis. In primary culture the ploidy level ranged between 2 and 15x (10-75 chromosomes). The frequency of polyploid cells was higher in the 5-month old callus culture, but the ploidy level was the same. In the callus lines derived from autotetraploid plants, cells with reduced chromosome number appeared quite frequently along with diploid and polyploid cells.

Effect of microcystin-LR and cyanobacterial extract from polish reservoir of drinking water on cell cycle progression, mitotic spindle, and apoptosis in CHO-K1 cells

Microcystin-LR is a cyanobacterial toxin possessing a potent tumor-promoting activity mediated through inhibition of protein phosphatases PP1 and PP2A. Because these enzymes are involved in fundamental cell processes, we decided to examine the influence of microcystin-LR on cell cycle progression, onset of anaphase, segregation of chromosomes by the mitotic spindle, and apoptosis in Chinese hamster ovary (CHO-K1) cells. Cells were incubated with 25, 50, and 100 μM of pure microcystin-LR and a cyanobacterial extract for 14, 18, and 22 h. Giemsa staining of cells treated with these toxins revealed a dose- and time-dependent increase of mitotic indices, accumulation of abnormal G 2/M figures with hypercondensed chromosomes, abnormal anaphases with defective chromosome separation, and polyploid cells. Because spindle checkpoint is a fundamental regulatory mechanism that assures the onset of anaphase and subsequent exit from mitosis, we examined the spindle organization in microcystin-treated cells. The majority of the mitotic cells showed monopolar and multipolar mitotic spindles (multiple asters). Microtubule bundles were present in interphase cells. Our results indicate that microcystin-LR induces apoptosis and necrosis in a dose- and time-dependent manner and that the frequency of dead cells cells is positively correlated with the frequency of polyploid cells.

The relation between cytogenetic instability in cattle and heavy metals level in Kuzbass environment

The water and feed samples were analysed on the farms of ecologically pure (N 1) and polluted (N 2) areas in Kuzbass. The presence of Cd (0.002 mg/l) was revealed in water samples taken in the polluted area The Hg. Cu and Pb concentrations in the hay from area N2 exceeded those from area N1 by 1.4, 1.9 and 3.3 times respectively. The Cu and Hb contents in haylage from the pure area were 10 times lower than those from the polluted area. The evaluation of cytogenetic status was done in Black-and-White cows raised in the investigated areas II was established that the frequency of polyploid cells in blood lymphocytes of the animals from the polluted area was 1.3 time higher than that in the control group (P<0.01). The frequency of single chromosome fragments was registered 2.3 times more often in the cows from area N 2 as well (P<0.05).

Bovine embryos contain a higher proportion of polyploid cells in the trophectoderm than in the embryonic disc.

The frequency of polyploid cells in the embryonic disc (ED) and in the trophectoderm (TE) was assessed in 50 in vitro produced bovine embryos fixed at days 7-8 post insemination (pi) and in 20 in vitro produced embryos that were transferred to uteri of recipients at day 7 and then recovered and fixed at day 12 pi. Separation of TE and ED cells was obtained by microdissection and the frequency of polyploid cells was determined by interphase cytogenetic analysis using fluorescence in situ hybridization (FISH) with chromosome 6- and chromosome 7-specific probes. The results show that 96% of day 7 embryos contain polyploid cells in the TE, whereas only 58% contain polyploid cells in the ED. In day 12 embryos 85% of TE and 40% of ED preparations contain polyploid cells. Statistical analysis revealed that the frequency of polyploid cells was significantly higher in the TE than in the ED in embryos containing less than 25% polyploid cells (n = 65). The few embryos (n = 5), which contained more than 25% polyploid cells, did not show this difference. Further, it was revealed that the level of polyploidy on day 7-8 was significantly higher than on day 12, both in the TE (two-fold) and in the ED (seven-fold).

Pig chromosome aberrations after vaccination against classical swine fever in field trials

Chromosomal aberrations were observed after vaccination against classical swine fever (CSF) in a previous study done on experimental pigs. To determine if the same effect occurs in farm animals, field trials were done with immunized pigs. The cytogenetic analysis was made from lymphocyte cultures of pigs sampled in three farms of Rı́o Cuarto region on six different periods: one, pre-vaccination (control) and in five post-vaccination (days 3, 7, 10, 15 and 22). Vaccine inoculation induced significant increase of cell frequency with chromosomal aberrations from days 3 to 10, when maximal frequencies of 6.78, 10.36 and 7.21% were observed in farms A, B and C, respectively. Afterwards, a decrease was recorded, reaching values not significantly different from controls. Mean frequencies of cells with chromosomal aberrations were 2.8±0.3, 5.9±0.4, 3.1±0.3%, for A, B and C farms, respectively. Cytogenetic effect was clastogenic, being chromosome breaks the first type of alteration to increase from the day 3 post-vaccination onwards, followed by chromatid exchanges and cells with multiple abnormalities. Chromosome pulverization was the last type to show increment, reaching a top value on day 10, after that it started to diminish gradually. There was no difference in the frequency of polyploid cells among sampling dates meaning that this type of alteration would not be induced by vaccination. Results of this field study confirm the mutagenic capacity of live virus vaccines against CSF and indicate that the evolution of chromosome alterations on the five evaluated post-vaccination periods was similar at different environments.

Differential effects of aneugens and clastogens on incidences of multinucleated cells and of micronucleate cells in Chinese hamster lung (V79) cell line in vitro

Two aneugens, vinblastine (0.025–0.4 μg/ml) and colchicine (0.05–0.4 μg/ml), and two clastogens, mitomycin C (0.05–0.4 μg/ml) and cyclophosphamide (1–16 μg/ml) were applied respectively to the micronucleus test in Chinese hamster lung (V79) cells in vitro, and the frequency of multinucleated cells (Fmu) and that of micronucleate cells (Fmi) in each group were observed. The results showed that at relatively high concentrations, all four agents increased both Fmu and Fmi, but the ratios of Fmu to Fmi in groups of the two aneugens (average of 2.2, 2.8, respectively) were much (10–30 folds) higher than that in groups of the two clastogens (0.09, 0.20). The difference between aneugens and clastogens in the above ratio was much more remarkable than that in areas of micronuclei (only 1.6–2.5 folds for the latter). At relatively low concentrations, the two clastogens increased only Fmi (but not Fmu), while the two aneugens increased only Fmu (but not Fmi). This indicates that the induction of multinuclei by aneugens may be more sensitive than by clastogens, and the induction of micronuclei by clastogens may be more sensitive than by aneugens. So, it is possible for the ratio of Fmu to Fmi to become a simple and sensitive (though indirect) index for distinguishing aneugens from clastogens. Further studies with other mutagens and (or) other cell types will be needed to confirm the deduction. As no difference in frequency of polyploid cells was observed between control group and each treatment, the multinucleation does not seem related to endoreduplication of chromosomes.

Direct assessment of triploid cells in mosaic human fetuses by fluorescence in-situ hybridization.

Villous tissues from 30 spontaneous abortions and the same number of artificial abortions were obtained and analysed for the frequency of polyploid cells. Single cell suspensions were made from these tissues without culture and the ploidy of > 100 cells was analysed. Trisomies of chromosomes 17 and 4 have rarely been reported in villous cells of spontaneous abortions, suggesting that the presence of more than three copies of chromosomes 17 and 4 per cell indicates polyploidy. The number of chromosomes 17 and 4 was detected by fluorescence in-situ hybridization analysis using centromeric probes D17Z1 and D4Z1. Most villous cells from cases of spontaneous and artificial abortions had two D17Z1 or D4Z1 signals per cell, with very small percentages of cells (0.5 +/- 0.4%) showing three signals per cell. However, in four cases of spontaneous abortions, 2-12% of cells had three D17Z1 or D4Z1 signals per cell. This indicates the presence of triploid cells in these cases of spontaneous abortion, at a significantly higher frequency compared to artificial or the remaining 26 cases of spontaneous abortion. In addition, three cases contained 0.2-0.4% of cells showing six signals, indicating that these cells were dividing triploid cells. The low frequency of mosaicism reported here would not be detectable by conventional chromosomal analysis.

A Single Injection of Pegylated Murine Megakaryocyte Growth and Development Factor (MGDF ) Into Mice Is Sufficient to Produce a Profound Stimulation of Megakaryocyte Frequency, Size, and Ploidization

AbstractDespite numerous studies investigating the action of c-mpl ligand, no reports have defined the in vivo changes in megakaryocytopoiesis in response to a single injection of this cytokine. Here we compare the kinetics of the megakaryocytopoietic response in C57Bl/6J mice administered 25 μg/kg or 250 μg/kg of pegylated (PEG) murine megakaryocyte growth and development factor (MGDF ) as a single intravenous injection. Megakaryocytes of mice treated with MGDF had normal ultrastructure, showing a typical distribution of the demarcation membrane system, α-granules, and other cytoplasmic organelles. Megakaryocyte ploidy, size, and frequency were markedly increased with both MGDF doses. Megakaryocyte ploidy was maximally increased from a modal value of 16N to 64N on day 3, with both doses of MGDF. Similarly, a comparable increase in megakaryocyte size occurred in the two MGDF groups. Increased megakaryocyte size was coupled to the increase in megakaryocyte ploidy, and no evidence for independent regulation of megakaryocyte size within individual ploidy classes was apparent. In contrast to megakaryocyte ploidy and size, the increase in megakaryocyte frequency was markedly different with the two doses of MGDF. The proportion of 2N and 4N cells was increased from a baseline of 0.035% to 0.430% by day 4 in mice treated with the higher dose of MGDF, but only to 0.175% in mice administered 25 μg/kg of MGDF. The marked increase in the pool of these immature megakaryocytes translated to a sustained elevation in the frequency of polyploid megakaryocytes (8N cells and greater). In contrast to the sustained increase in the frequency of polyploid cells, the level of polyploidization was downregulated on days 6 to 10, but normalized by day 14. We conclude that a single injection of MGDF is able to expand the megakaryocytic pool in a dose-dependent manner, which, with subsequent maturation, should lead to an increased rate of platelet production.

Ploidy instability of embryogenic cucumber (Cucumis sativus L.) callus culture

Embryogenic callus cultures were established from immature cucumber(Cucumis sativus L.) embryos on E20A (Dumas de Vaulxet al. 1981) or MS (Murashige and Skoog 1962) media supplemented with 6-benzylaminopurine (BAP), α-naphthylacetic acid (NAA) and/or 2,4-dichlorophenoxyacetic acid (2,4-D). Regeneration of plants was observed after a transfer to culture media either without growth regulators or supplemented with kinetin and NAA. Flow cytometry was employed to estimate DNA ploidy levels. Most of cell nuclei in young leaf tissues were found in G1 phase with 2C DNA content. Callus cultures were mixoploid with DNA content ranging from 2C to 32C. The frequency of polyploid cells was increasing with the age of culture and the polyploidization was accompanied by a gradual loss of regeneration ability. Plants regenerated from callus cultures were classified as diploid (57 %), tetraploid (18 %), octoploid (4 %) and mixoploid (2n/4n, 4 %) and (4n/8n, 17 %). The results of this study confirmed a close link between the polyploidization and the loss of totipotencyin vitro. Tetraploid plants obtained in this study have a potential to be used in interspecific crosses where their tetraploid status could help in overcoming existing breeding barriers due to differences in chromosome number.

Colchicine-induced polyploidy in Citrus embryogenic cultures, somatic embryos, and regenerated plantlets

Two morphologically distinct Citrus sinensis (L.) Osbeck embryogenic cultures were grown for 4 or 12 weeks on Murashige and Tucker (MT) medium amended with 0.01% or 0.10% colchicine. Ploidy level frequencies were determined in treated cultures and untreated controls, as well as somatic embryos and regenerated plantlets, by chromosome counts. Mortality was widespread after treatment. The frequency of polyploid cells and the number of ploidy classes was increased by colchicine treatment; the frequency of non-diploid cells in untreated cultures was less than 10%. One line partially reverted back to diploidy, but the other line remained cytogenetically unstable; however, colchicine-induced cytogenetic variation was attenuated through the regeneration process. Although 2x, 3x, 4x, 6x, and 8x cells and somatic embryos were observed, only diploid and tetraploid plantlets were regenerated from treated cultures. Untreated cultures yielded only diploid regenerants.

Investigations into the induction of aneuploidy and polyploidy in mammalian cells by the anti-tussive agent noscapine hydrochloride

Noscapine, a non-narcotic, centrally-acting anti-tussive drug induces polyploidy in Chinese hamster CHL cells; further studies were carried out to investigate whether similar effects could be induced in other rodent cells (Chinese hamster V79) and in human lymphocytes. In both cases, large increases in the frequency of polyploid cells were induced at test concentrations ranging from 15 to 120 micrograms/ml after 24 and 48 h continuous treatment in the absence of S9 mix. In addition, spindle damage was observed in V79 cells and human skin fibroblasts after 24 h treatment with test concentrations of 30 and 60 micrograms/ml. Furthermore, after treatment of human skin fibroblasts there was a marked increase in the proportion of cells containing chromosomes which had become dislocated from the spindle. Treatment of the mouse/human hybrid cell line R3-5 induced a significant increase in the number of 6-thioguanine resistant colonies and it was confirmed cytogenetically that these colonies had arisen due to loss of human chromosome 2. From these experiments it can be concluded that noscapine induces polyploidy in both rodent and human somatic cells, and that this could arise through a direct effect upon spindle structure and/or function. The aneugenic properties of noscapine are less certain and further work is required in this area. Exposure to the drug through its therapeutic use (15mg up to four times daily) could exceed, at least locally within the gastrointestinal (GI) tract, the concentration range shown to be active in these in vitro studies. An immediate topical hazard might exist within the buccal cavity and GI tract, but further confirmation of these in vitro results are required using suitable in vivo systems before definite conclusions can be made regarding any potential hazard associated with the administration of this drug.

Microspectrophotometric DNA analysis in ulcerative colitis with special reference to its application in diagnosis of carcinoma and dysplasia.

The deoxyribonucleic (DNA) content was measured by microspectrophotometry in 100 specimens from 60 patients with ulcerative colitis, including six patients in whom the colitis was associated with carcinoma. Some 23 of 30 (77%) specimens of dysplastic tissue showed aneuploidy or polyploidy, whereas 50 of 53 (94%) specimens of non-dysplastic tissue showed diploidy. The difference was statistically significant (p less than 0.001). Polyploidy was often observed in non-dysplastic mucosa from patients who had carcinoma or dysplasia. In the non-dysplastic patients all samples of inflamed tissue showed diploidy. Some 10% of samples without inflammation, however, also showed polyploidy. A good correlation was found between the frequency of polyploid cells and the grade of dysplasia. Microspectrophotometric measurement of DNA content proved useful in the assessment and diagnosis of dysplasia in ulcerative colitis and could be considered for screening high risk patients.

Does Smooth Muscle Cell Polyploidy Occur in Resistance Vessels of Spontaneously Hypertensive Rats?

The ploidy of smooth muscle cells (SMCs) enzymatically isolated from the aorta and superior mesenteric artery (elastic arteries), caudal artery (small muscular artery) and the small mesenteric arteries and arterioles (mesenteric resistance vessels) of the spontaneously hypertensive (SHR) and Wistar-Kyoto (WKY) rats at ages 12, 26, 32 and 40 weeks was determined by flow cytometric DNA analysis and Feulgen-DNA photometric measurements. Frequency of polyploid cells in the aorta and superior mesenteric artery of the SHR increased from 4.43 +/- 1.35 and 7.58 +/- 1.69%, respectively, at 12 weeks to 31.26 +/- 3.00 and 14.13 +/- 1.30% at 40 weeks. There was a smaller increase in the percentage of polyploid cells in these two vessels of the WKY from 4.73 +/- 0.74 and 5.82 +/- 0.33%, respectively, at 12 weeks to 10.64 +/- 0.17 and 7.68 +/- 0.64% at 40 weeks. The caudal artery and mesenteric resistance vessels showed no significant increase in the percentage of 4N (tetraploid) cells in the SHR from 12 weeks (6.80 +/- 0.92 and 6.10 +/- 0.75%) to 40 weeks (7.83 +/- 0.67 and 7.57 +/- 0.07%). Similarly, there was no significant change in ploidy in these arteries of the WKY. Hence, while polyploidy of SMCs increases in the aorta and superior mesenteric artery of the rat with increasing age and with duration of hypertension, there is no significant change in the number of polyploid cells in smaller vessels such as the caudal artery or mesenteric resistance vessels. Since it is the resistance vessels that are involved in the development and maintenance of hypertension, polyploidy of SMCs in the blood vessel wall appears to hold little relevance to the etiology of this disease. As well, increased incidence of polyploidy is not directly attributable to increases in blood pressure as the caudal artery has a high systolic pressure in the SHR yet the incidence of polyploid cells in this artery does not differ from that of the WKY.

Effect of hormone on chromosome behaviour in callus cultures ofAsparagus racemosus

Chromosomal variability in callus culture ofAsparagus racemosus was comparatively higher in the presence of 2,4-D than of NAA. The frequency of polyploid cells was enhanced with the increase in the concentration of 2,4-D or with the addition of coconut water. Gradual polyploidization with increasing age of the callus has been recorded.